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Benzyl Alcohol, Benzoic Acid, and its Salts and Ester

CIR EXPERT PANEL MEETING JUNE 27-28, 2011 Administrative

June 3, 2011

Memorandum

To: CIR Expert Panel

From: Wilbur Johnson, Jr. Manager/Lead Specialist

Subject: Draft Report on Benzyl Alcohol, Benzoic Acid, and its Salts and Ester

In 1998, the CIR Expert Panel issued a final safety assessment on Benzyl Alcohol, Benzoic Acid and Sodium Benzoate, concluding that these ingredients are safe for use in cosmetics at concentrations up to 5%, but that the data are insufficient to support the safety of these ingredients in cosmetic products in which inhalation is a primary route of exposure. In June of 2010, the Expert Panel reviewed new inhalation toxicity data on benzyl alcohol and benzoic acid provided by the Council. The Panel decided to reopen its review of this report to consider the new data.

A Scientific Literature Review (SLR) on this group of ingredients was announced on February 24, 2011. Technical comments and use concentration data were received from the Council during the 60-day comment period. Additionally, an inhalation toxicity study on benzyl alcohol and benzoic acid and a RIFM synopsis of data on benzyl benzoate were received in 2010. These data have been added to the draft report for the Expert Panel’s review at this meeting.

A copy of the draft report on these ingredients is included along with the following: CIR report history, Minutes from the June 2010 Panel meeting, Literature search strategy, Data profile, the Council’s technical comments on the SLR (pcpc, in pdf), Transmittal memo for inhalation toxicity study (data 1, in pdf), Inhalation toxicity study (data 2, in pdf), RIFM synopsis on benzyl benzoate (data 3, in pdf), Use concentration data (data 4, in pdf), and 2001 published CIR final report on benzyl alcohol, benzoic acid, and sodium benzoate (finalrep, in pdf). After reviewing the draft report, the Expert Panel needs to determine whether the data included in this report are sufficient for evaluating the safety of benzyl alcohol, benzoic acid, and its salts and ester in cosmetic products.

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CIR History of:

Benzyl Alcohol, Benzoic Acid, and its Salts and Ester

In 1998, the CIR Expert Panel issued a final safety assessment of Benzyl Alcohol, Benzoic Acid and Sodium Benzoate, concluding that they are safe for use in cosmetics up to 5%, but that the data are insufficient to support the safety of these ingredients in which the primary route of exposure is inhalation. The report was published in 2001.

In June, 2010, the Expert panel reviewed new data provided by the Council on the safety of benzyl alcohol and benzoic acid for inhalation use. The Panel decided to reopen its review of this report to consider the new data.

New Data Review, Belsito and Marks Teams/Panel: June 28-29, 2010

The Council provided data in response to the insufficient data determination on the inhalation use of benzyl alcohol, benzoic acid and sodium benzoate. The Personal Care Products Council provided a 4-week inhalation toxicity study (rats) of aerosolized benzyl alcohol and benzoic acid. After reviewing this study, the Expert Panel determined that the final report on the safety assessment of benzyl alcohol, benzoic acid, and sodium benzoate should be reopened to amend the original conclusion. Furthermore, the Panel determined that data available in the original safety assessment, with the addition of the new inhalation toxicity data, were likely sufficient to support the safety of other simple salts of benzoic acid (calcium, magnesium, and potassium benzoate) and the benzyl benzoate ester. Thus, it was agreed that the safety of these ingredients and those reviewed in the original final report would be reviewed in the SLR that will be developed.

SLR in February, 2011

The availability of a scientific literature review (SLR) on this group of ingredients was announced on February 24, 2011. Technical comments and use concentration data from the Council were received during the 60-day comment period.

Draft Report, Belsito and Marks Teams/Panel: June 27-28, 2011

Technical comments on the SLR and use concentration data, received from the Council during the 60-day comment period have been added to the draft report. The draft report also contains an inhalation toxicity study on benzyl alcohol and benzoic acid and studies from a RIFM synopsis of data on benzyl benzoate that were received in 2010.

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Literature Search on Benzyl Alcohol and Related Ingredients*

Ingre- Toxline ChemIDplus Multidatabase DART Household Beilstein Registry Kosmet Napralert RTECS CAplus dients &PubMed (See legend*) Products BZA 1098 1 5 44 1 1 1 5 341 674 1534 (1997 to (1997 2011) to 2011) BA 1610 1 6 293 1 4 1 19 1263 3830 3648 (1997 to (1997 2011) to 2011) SB 226 1 4 21 1 2 1 0 5 13 643 1997 to 2011) CB 6 1 0 0 0 1 1 0 0 1 26

MB 0 1 0 0 0 1 1 0 0 1 12

PB 23 1 1 0 0 1 1 0 1 0 58

BB 331 1 1 11 1 1 1 1 117 31 227 (1997 to 2011) *Data in Table: Total no. publications in search; Multidatabase = HSDB, CCRIS, ITER, IRIS, Gene-Tox, and LacMed;

Searches Performed on 7/9 and 12/2010, and 3/2011 (1 chemical name and 1 CAS No.; Toxline added additional chemical names to search term); Any limitations on years searched indicated. Website list also searched.

Ingredients (BZA) Benzyl Alcohol OR 100-51-6

(BA) Benzoic Acid OR 65-85-0

(SB) Sodium Benzoate OR 532-32-1

(CB) Calcium Benzoate OR 2090-05-3

(MB) Magnesium Benzoate OR 553-70-8

(PB) Potassium Benzoate OR 582-25-2

(BB) Benzyl Benzoate OR 120-51-4

Benzyl Alcohol OR 100-51-6 OR Benzoic Acid OR 65-85-0 OR Sodium Benzoate OR 532-32-1 OR Calcium Benzoate OR 2090-05-3 OR Magnesium Benzoate OR 553-70-8 OR Potassium Benzoate OR 582-25-2 OR Benzyl Benzoate OR 120-51-4

CIR Panel Book Page 3 Distributed for Comment Only - Do Not Quote or Cite Benzyl Alcohol Group Check List for June, 2011. Writer – Wilbur Johnson

Repeated dose

Acute toxicity Irritation Sensitization toxicity Penetration Skin Enhancement Penetration ADME Oral Parenteral Dermal Inhale Oral Parenteral Dermal Inhale Irritation Ocular Animal Dermal Dermal ty Phototoxicity Human Animal Sensitization Human Sensitization toxicity Repro/Devel Genotoxicity Carcinogenici

Irr. Irr

Benzyl alcohol X X X X X X X X X X X X X X X Benzoic acid X X X X X X X X X X X X X X X Sodium benzoate X X X X XX X X XXX Calcium benzoate Magnesium benzoate Potassium X X X X X XX benzoate Benzyl benzoate X X X X X XXX XXXX X XX X

CIR Panel Book Page 4 Transcripts

Distributed for Comment Only - Do Not Quote or Cite Day 1 of the June 28-29, 2010 CIR Expert Panel Meeting – Dr. Marks’ Team DR. MARKS: Okay. So let me see. I'm the one tomorrow who will make the motion, and I'll make the motion to reopen to amend the conclusion and, as we've said, add the four salts. And as you said, Alan, we'll get a draft with DR. MARKS: Okay. Next, Buff book 2, benzyl alcohol, benzoic acid, and sodium benzoate. And that's right the new inhalation data and more, if there is available -- behind – DR. ANDERSEN: Maybe August, for sure in December. DR. ANDERSEN: Ron Hill, you won't understand this. Ron Shank and Tom will understand it. This is one more

(inaudible). We've been here before. Day 1 of the June 28-29, 2010 CIR Expert Panel Meeting – Dr. Belsito’s Team DR. HILL: On the benzyl alcohol? DR. BELSITO: So, moving right along to benzyl alcohol, benzoic acid, and sodium benzoate. So, we had had DR. ANDERSEN: Yes. issues about these primarily due to some inhalation toxicity data. It sort of took an interesting approach. It started DR. HILL: Yeah, okay. I got that sense, honestly, without you even having to say it. (Laughter) off with us worrying about the potential for pericardial reactions in the bronchial tree because of their report of non- DR. MARKS: So, from the memo from Alan there are two questions. One, do we reopen this safety assessment? immunologic contact pericardia from benzoic acid and sodium benzoate. And then we ended up with data To recall, in '98 the CIR expert Panel completed its final assessment of these three ingredients, concluding a "safe suggesting some other effects on the lung, but there were issues with the control population being equally affected for use in cosmetics" up to 5 percent. But it was insufficient to support the safety from an inhalation route. And then potentially viral infections in the colonies. And so now we have data coming back on yet another study to try and benzyl alcohol was also safe for use in hair dyes up to 10 percent. The additional data need in '98 was inhalation assess whether aerosol exposures to benzyl alcohol, benzoic acid, sodium benzoate are adequate to allow us to just toxicity. And we're at the point now, do we have that? And can we amend it? And then also Alan raised the issue go with a safe as used rather than safe except for inhalation. And I don't know, Alan. Should I recluse myself on this of adding, potentially, four additional salts. So -- Ron, Ron? since I'm part of RIFM and RIFM did these studies for us? Or? DR. SHANK: I would recommend reopening to remove the inhalation, the need for inhalation data. What we have DR. ANDERSEN: No, I don't think so. I think data will speak for themselves. was acceptable. And then DR. BELSITO: Okay. So, we have these data. So, what do you think, guys? DR. SLAGA: I agree with that. DR. LIEBLER: My note here is yes. DR. HILL: I'm good with that. DR. SNYDER: Safe as used. DR. MARKS: So I'll move tomorrow we reopen this safety assessment, that we now have inhalation data, that we DR. BELSITO: Safe as used? Okay. can delete that "insufficient" need. And that we add the four salts. And so, basically, if we look on page 46 of the DR. LIEBLER: Under take to amend the safety assessment -- original document -- that's on Panel book page 135 -- the available data are, "insufficient to support the safety of DR. BERGFELD: Yes to open -- these ingredients in which the primary route of exposure is inhalation." We can delete that sentence in the DR. LIEBLER: -- yes, it's safe to use. Yes -- yes, and also is yes, should four additional salts be added as CIR conclusion. Okay. Any – prepares an amended safety assessment. I said yes. DR. ANDERSEN: If we get -- assuming the other team agrees, what you will see next time --and I'm not sure I can DR. BELSITO: Yeah, so did I. The only issue is, I think we need to bring in not only the data that RIFM has promise that that will be August, but what you will see next time is a draft amended safety assessment that supplied that allows us to say safe as used, but remember we had -- we actually have two studies. We had one -- or incorporates all the new inhalation toxicity data, adds the four salts -- plus any available literature on those four was it one study and then we had the teleconference with Unilever in London over how to interpret the study? Or salts. I don't expect that to be massive, but we will look, and if there are any data, we will include those, as well. whether it was two studies, I don't remember. But those studies, whatever they were -- the one where there were DR. MARKS: It's a draft. bronchial alveolar changes and the question mark whether there was a viral infection in the colony -- those studies DR. ANDERSEN: A draft conclusion that simply deletes that sentence. should be brought in and I think explained. No? DR. MARKS: Okay. So this -- is this technically a re-review? DR. ANDERSEN: Well, that's a good question. I -- left to my own devices, I was not going to bring those in. DR. ANDERSEN: No, it isn't because we don't re- review things that were "insufficient," in which the onus is on Because the panel had -- the industry or any other interested party to come in and say, "Here are the data you asked for. Now let's move on to DR. BELSITO: Discounted them. amend it." So we're reopening it, but it's not a re-review. DR. ANDERSEN: Considered those and then reconsidered those and disposed of them as -- DR. SLAGA: So have we reviewed the inhalation data? DR. BELSITO: Flawed. DR. MARKS: Okay. Well, we won't – I -- DR. ANDERSEN: -- being methodologically flawed. MR. BAILEY: That was under the "insufficient data" umbrella. DR. BELSITO: Okay.

1 2

DR. ANDERSEN: And, you know, just -- you weren't going to resolve this question based on those data. Day 2 of the June 28-29, 2010 CIR Expert Panel Meeting DR. BELSITO: I don't have a problem with that. I just wanted to make note that there was data out there that we DR. BERGFELD: The last item is the benzyl alcohol group. Dr. Marks? had looked at, found it inadequate, and thought the studies were flawed. And whether we should bring that in. DR. MARKS: In 1998 the CIR Expert Panel issued a final safety assessment of benzyl alcohol, benzoic acid and But I'm fine not bringing it in. sodium benzoate. Within that conclusion the available are data to support the safety of these ingredients in cosmetic DR. SNYDER: Yeah, I don't -- I think they should be deleted from the report. products in which a primary route of exposure is inhalation. We now have received new data which would ally that DR. ANDERSEN: Yeah, they're not in -- concern and so our team moves to reopen this safety assessment, delete the insufficient sentence in the conclusion, DR. BELSITO: It's not in the report. and then also at the same time add four salts. DR. ANDERSEN: They're not in, and the of the revised report will be these robust data with basically no effected DR. BERGFELD: That's a motion? levels at the highest exposures. DR. MARKS: Correct. DR. BELSITO: Good, okay. DR. BERGFELD: Is there a second? DR. LIEBLER: So I had one question here. In the memo -- in the cover memo, it refers to the council provided use DR. BELSITO: Second. concentration -- oh, I'm sorry. The previous sentence is: If these data are sufficient, the panel determines to reopen, DR. BERGFELD: Is there any further discussion regarding reopening and adding the salts? then there's an opportunity to also consider available data support of the simple salts, benzoic acid -- of benzoic acid DR. BELSITO: Yes. We discussed there were one or two studies that we felt were flawed but raised concerns such as calcium benzoate, magnesium benzoate, potassium benzoate, and benzyl benzoate, which is not a salt. It's a about potential pulmonary issues with inhalation and we discussed whether they should be included in this report de-ester of itself -- and then as updated use concentration for benzyl alcohol, benzoic acid, and sodium benzoate. and then discussed as being flawed. However, I think our team felt that since we clearly felt these studies were So, what about benzyl benzoate? Is that under consideration as well or not? flawed, there was a viral infection in the colony, that it wouldn't really do any justice to include them, that it would DR. ANDERSEN: Yeah, it's -- it was presented as a possible addition. What I thought I had heard is that there's a just confuse the reader and elected not to do so. comfort level with the salts. Is the panel equally comfortable with benzyl benzoate? DR. SNYDER: An additional comment is we have more robust studies with NOAEL. DR. LIEBLER: Just to follow up on that. I am, and I was just wondering why it wasn't – it seemed unclear to me DR. LIEBLER: One other point of clarification. You mentioned the three salts I guess, calcium, magnesium and and I'm just registering that I am comfortable with that. Particularly if you're going to do benzoic acid and benzyl potassium benzoates. Also listed on the memo was benzyl benzoate and I thought that that would be reasonable to alcohol. include as well. It's not a salt. It would be an ester. I don't know if you discussed that or if you have a differing DR. BELSITO: Well, if you're comfortable, Dan, I'm comfortable. view on it. DR. ANDERSEN: Well -- and we have the data for benzyl benzoate use concentrations that have been provided DR. MARKS: That's fine. and they are, if anything, less than the others across the board, so. DR. BERGFELD: Are there any other comments? Seeing none, I'll call for the vote to reopen. All those in favor? DR. BELSITO: Any other comments? Unanimous. DR. SNYDER: I just had one query. So, we received that there was a new comment assay that was performed in results? But I didn't see the data in the new data section. DR. ANDERSEN: Correct. Its genotoxicity really hasn't ever been an issue with these. The comet assay was basically negative. I figured I would simply add it in once we took the next step. But it addressed an issue that wasn't an issue -- wanted you to focus on the inhalation tox stuff. But thank you for bringing it up because I should have just mentioned that upfront. DR. BELSITO: All right. Anything else? If not, we're done.

3 4 CIR Panel Book Page 5 Report

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Draft Report

Benzyl Alcohol, Benzoic Acid, and its Salts and Ester

June 27, 2011

The 2011 Cosmetic Ingredient Review Expert Panel members are: Chair, Wilma F. Bergfeld, M.D., F.A.C.P.; Donald V. Belsito, M.D.; Ronald A. Hill, Ph.D.; Curtis D. Klaassen, Ph.D.; Daniel C. Liebler, Ph.D.; James G. Marks, Jr., M.D.; Ronald C. Shank, Ph.D.; Thomas J. Slaga, Ph.D.; and Paul W. Snyder, D.V.M., Ph.D. The CIR Director is F. Alan Andersen, Ph.D. This report was prepared by Wilbur Johnson, Scientific Analyst/Writer.

CIR Panel Book Page 6 Distributed for Comment Only - Do Not Quote or Cite TABLE OF CONTENTS INTRODUCTION ...... 1 CHEMISTRY...... 1

DEFINITION AND STRUCTURE ...... 1 CHEMICAL AND PHYSICAL PROPERTIES ...... 1 METHOD OF MANUFACTURE ...... 2 UV ABSORPTION ...... 2 ANALYTICAL METHODS ...... 2 IMPURITIES ...... 2 USE ...... 3

COSMETIC ...... 3 NONCOSMETIC ...... 4 TOXICOKINETICS ...... 5

ORAL STUDIES ...... 5 DERMAL STUDIES ...... 6 TOXICOLOGICAL STUDIES ...... 6

ACUTE AND REPEATED DOSE INHALATION TOXICITY ...... 7 ACUTE ORAL TOXICITY ...... 7 ACUTE INTRAPERITONEAL TOXICITY ...... 7 ACUTE DERMAL TOXICITY ...... 8 ACUTE SUBCUTANEOUS TOXICITY ...... 8 ACUTE INTRAMUSCULAR TOXICITY ...... 8 REPEATED DOSE TOXICITY – ORAL STUDIES ...... 8 REPEATED DOSE TOXICITY – DERMAL STUDIES ...... 9 REPEATED DOSE TOXICITY – PARENTERAL STUDIES ...... 9 OCULAR IRRITATION/TOXICITY ...... 10 SKIN IRRITATION AND SENSITIZATION ...... 11 CASE REPORTS ...... 11 CYTOTOXICITY ...... 11 PHOTOTOXICITY ...... 12 PHOTOHEMOLYSIS ...... 12 REPRODUCTIVE AND DEVELOPMENTAL TOXICITY ...... 12 GENOTOXICITY ...... 15

PHOTOGENOTOXICITY ...... 16 EFFECT ON DNA SYNTHESIS ...... 16 CARCINOGENICITY ...... 17 SUMMARY ...... 18

iii

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INTRODUCTION

Data that are relevant to the safety assessment of benzyl alcohol, benzoic acid, and its salts and benzyl ester as used in cosmetics are summarized in this scientific literature review. In cosmetics, these ingredients may function as fragrance ingredients, pesticides, pH adjusters, preservatives, solvents, and/or viscosity decreasing agents in cosmetic products.

In 2001, the Cosmetic Ingredient Review (CIR) published a final report on the safety assessment of benzyl alcohol, benzoic acid, and sodium benzoate with the conclusion that benzyl alcohol, benzoic acid, and sodium benzoate are safe for use in cosmetic formulations at concentrations up to 5%.1 Benzyl alcohol was safe for use in hair dyes at concentrations up to 10%. The available data were insufficient, however, to support the safety of these ingredients in cosmetic products in which a primary route of exposure is inhalation. The Expert Panel also stated that inhalation toxicity data were needed for completion of this safety assessment.

In response to the need for inhalation toxicity data, the Personal Care Products Council provided a 4-week inhalation toxicity study (rats) of aerosolized benzyl alcohol and benzoic acid, and the results of this study were discussed at the June 28-29, 2010 CIR Expert Panel meeting. After reviewing this study, the Expert Panel determined that the final report on the safety assessment of benzyl alcohol, benzoic acid, and sodium benzoate should be reopened to amend the original conclusion. Furthermore, the Panel determined that data available in the original safety assessment, with the addition of the new inhalation toxicity data, were likely sufficient to support the safety of other simple salts of benzoic acid (calcium, magnesium, and potassium benzoate) and the benzyl benzoate ester. Thus, the safety of these ingredients and those reviewed in the original final report is being evaluated in the current report. In addition to the inhalation toxicity data on benzyl alcohol and benzoic acid, data on benzyl alcohol, benzoic acid, or sodium benzoate that became available since the final report was issued in 1998 are also included.

New data reviewed included an Organization for Economic Cooperation and Development (OECD) Screening Information Data Sets (SIDS) for high volume chemicals initial assessment report on benzoic acid, sodium benzoate, potassium benzoate, and benzyl alcohol published by the United Nations Environmental Program Chemicals Branch (UNEP chemicals) in 2001,2 the same year that the CIR Final Safety Assessment on benzyl alcohol, benzoic acid, and sodium benzoate was published. Furthermore, the European Commissions’ International Uniform Chemical Information Database (IUCLID) on benzyl alcohol, benzoic acid, sodium benzoate, and benzyl benzoate was created in the year 2000.3 Data from the OECD SIDS assessment and IUCLID data sets are summarized in the Toxicology section. In the process of selecting new data for inclusion in the current report, efforts were made to eliminate redundancy between OECD SIDS, IUCLID, or other data found in the published literature and data included in the original final report.

CHEMISTRY

Definition and Structure

Definitions, other chemical names, and cosmetic ingredient functions for ingredients reviewed in this safety assessment are included in Table 1. Structural formulas are included in Figure 1.

Chemical and Physical Properties

Chemical and physical properties of benzyl alcohol, benzoic acid and its salts, and the benzyl benzoate ester are included in Table 2.

The photodegradation of benzoic acid aqueous solution in the presence of UV light has been reported. Benzoic Acid absorbs UV light below 300 nm, and its concentration in solution was monitored using reverse-phase high-performance liquid chromatography (HPLC). The mercury lamp used emitted polychromatic radiation between 240 and 540 nm. The photon flow absorbed by the reaction system varied with the concentration of benzoic acid and with the presence of the products of

CIR Panel Book Page 8 Distributed for Comment Only - Do Not Quote or Cite degradation. When the temperature was increased from 25ºC to 50ºC, photodegradation was increased by 20%. Photodegradation products were not mentioned.4

The conversion of benzyl alcohol to benzaldehyde and benzoic acid in aqueous solution has been demonstrated using liquid chromatography/spectrophotometry.5 When benzyl benzoate (in ethanol) was exposed to sunlight for 11 days (at 25ºC), 35 products were isolated from exposed samples and identified using gas chromatography/mass spectrometry. Benzyl alcohol and benzoic acid were among the products identified.6

Method of Manufacture Benzyl Alcohol

Large scale production of benzyl alcohol is achieved by the action of sodium or potassium carbonate on benzyl chloride.1

Benzoic Acid

Benzoic acid is produced via the decarboxylation of phthalic anhydride in the presence of catalysts. Another production method involves the chlorination of toluene to yield benzotrichloride, which is hydrolyzed to benzoic acid.7

Sodium Benzoate

Sodium benzoate is produced by the neutralization of benzoic acid with sodium bicarbonate, sodium carbonate, or sodium hydroxide.8

Potassium Benzoate

Potassium benzoate can be prepared by reacting methyl benzoate with potassium thioacetate.9

Benzyl Benzoate

Benzyl benzoate results from the condensation of benzoic acid and benzyl alcohol. It can also be generated from benzaldehyde via the Tishchenko reaction.10

UV Absorption

The following UV absorption maxima have been reported for salts and an ester of benzoic acid: 228.6 nm (magnesium benzoate), 226.8 nm (potassium benzoate), and 256 nm (benzyl benzoate).11,12

Analytical Methods

Benzoic acid has been analyzed using mass spectroscopy,13 and potassium benzoate has been analyzed using IR spectroscopy.14 Benzyl benzoate has been analyzed using IR and NMR spectroscopy.15,16

Impurities

Benzoic acid can react with ascorbic acid in beverages via metal-catalyzed reduction (cans) or on exposure to solar UV (bottles) to form benzene, but the yield of benzene is not great (< 1 ppb). Limits for heavy metal impurities in food-grade (as Pb, ≤ 10 mg/kg) and pharmaceutical-grade (heavy metals, ≤ 0.001%) potassium benzoate have been established.

Benzoic Acid and Sodium Benzoate

Benzene production from benzoic acid in the presence of a hydroxyl radical generating system has been demonstrated.17

Hydroxyl radical generated by the metal-catalyzed reduction of O2 and H2O2 by ascorbic acid can attack benzoic acid to produce benzene under conditions that are prevalent in products such as canned soda. The yield of benzene, however, was extremely small (< 50 nM or < 1 ppb) under conditions that were selected to approximate relative amounts of ascorbic acid and sodium benzoate in foods or beverages.

CIR Panel Book Page 9 Distributed for Comment Only - Do Not Quote or Cite The effects of UVA exposure on benzene formation were determined in another study. Benzene formation was examined for samples contained in UV stabilized and non-UV stabilized packaging.18 Some of the samples selected for UVA testing included model solutions prepared with 0.04% benzoate and 0.025% ascorbic acid in unbuffered water. The results of 24-h irradiation studies indicated that, under intense UV light, benzene levels increased by as much as 53% in model solutions stored in non-UV stabilized bottles. However, the use of UV stabilized polyethylene terephthalate bottles reduced benzene formation by ~ 13%, relative to the non-UV stabilized bottles. Similar trends were observed following irradiation for 7 days. According to the Food and Drug Administration (FDA), the U.S. beverage industry voluntarily reformulated beverages that were found to contain benzene levels at or above the 5 ng/g maximum contaminant level for drinking water established by the U.S. Environmental Protection Agency.18 Furthermore, in a 2008 survey conducted by FDA, all of the reformulated beverages tested were shown to contain ≤ 1.1 ng/g benzene. Most of the beverages were reformulated by removing or reducing either benzoate or ascorbic acid, or by adding EDTA.19

Benzyl Benzoate, Potassium Benzoate, and Sodium Benzoate

According to the United States Pharmacopoeia, benzyl benzoate contains less than 99% and not more than 100.5% C14H12O2 and potassium benzoate contains not less than 100.5% and not more than 99% C7H5KO2. Limits for water and heavy metals in potassium benzoate are stated as ≤ 1.5% and ≤ 0.001%, respectively.20

The Food Chemicals Codex specification for sodium benzoate includes the following limitations: heavy metals, as Pb (≤ 2 mg/kg), alkalinity, as NaOH (≤ 0.04%), and water (≤ 1.5%).21

USE

Cosmetic

As stated in the International Cosmetic Ingredient Dictionary and Handbook, benzoic acid, benzyl alcohol, sodium benzoate, calcium benzoate, magnesium benzoate, potassium benzoate, and benzyl benzoate may function as fragrance ingredients, pesticides, pH adjusters, preservatives, solvents, viscosity decreasing agents, and pH adjusters in cosmetic products (See Table 1).22

According to information supplied to the Food and Drug Administration (FDA) by industry as part of the Voluntary Cosmetic Registration Program (VCRP) in 2010, the following ingredients were being used in personal care products: benzoic acid, benzyl alcohol, sodium benzoate, potassium benzoate, and benzyl benzoate.23 Uses of calcium benzoate and magnesium benzoate were not reported to the VCRP. These data are summarized in Table 3.

Independent of the VCRP data, a survey of ingredient use concentrations was conducted by the Personal Care Products Council in 2010. The results of this survey indicated that the following ingredients were being used in cosmetic products: benzoic acid (0.000002 to 5%), benzyl alcohol (0.000006 to 10%), sodium benzoate (0.000001 to 1%), calcium benzoate (0.002 to 0.004%), potassium benzoate (0.002 to 0.003%), and benzyl benzoate (0.000005 to 4%). 24 No uses of magnesium benzoate were reported in the Council survey.

Personal care products containing the ingredients reported as being used may be applied to the skin, nails, or hair, or, incidentally, may come in contact with the eyes and mucous membranes. Products containing these ingredients may be applied as frequently as several times per day and may come in contact with the skin, nails, or hair for variable periods following application. Daily or occasional use may extend over many years.

Benzyl alcohol, sodium benzoate, and benzyl benzoate are used in hair sprays, and effects on the lungs that may be induced by aerosolized products containing these ingredients are of concern.

The aerosol properties that determine deposition in the respiratory system are particle size and density. The parameter most closely associated with deposition is the aerodynamic diameter, da, defined as the diameter of a sphere of unit density possessing the same terminal settling velocity as the particle in question. In humans, particles with an aerodynamic diameter

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of ≤ 10µm are respirable. Particles with a da from 0.1 - 10µm settle in the upper respiratory tract and particles with a da < 0.1 µm settle in the lower respiratory tract.25,26

In the European Union, the maximum authorized concentrations for benzoic acid and sodium benzoate as preservatives in cosmetic products are: rinse-off products, except oral care products (2.5%, as acid), oral care products (1.7%, as acid), and leave-on products (0.5%, as acid). For salts of benzoic acid other than sodium benzoate, the maximum authorized concentration is 0.5% (as acid).27

The Scientific Committee on Cosmetic Products and Non-Food Products Intended for Consumers (SCCNFP) was asked to answer the following question: Can benzoic acid and it salts and esters be used safely for non-preservative purposes in cosmetic rinse-off products at a maximum concentration of 2.5% and in cosmetic oral-care products at a maximum concentration of 1.7%. Data were provided to the SCCNFP in support of the use of benzoic acid as a non-preservative. The SCCNFP did not find the submission appropriate for a safety evaluation of benzoic acid and sodium benzoate for the applied “other uses” in cosmetic products.28 Subsequently, a second data set was provided, and these data were considered adequate for arriving at a conclusion on the safety of benzoic acid and sodium benzoate, but not the other salts or esters. Based on these data, the Scientific Committee on Consumer Products (SCCP) issued the following opinion: The SCCP is of the opinion that benzoic acid and sodium benzoate are safe for use for preservative and non-preservative purposes in cosmetic rinse-off products at a maximum concentration of 2.5 % and in cosmetic oral-care products at a maximum concentration of 1.7%, and in leave-on products up to 0.5%.29

Noncosmetic

According to FDA’s Select Committee on Generally Recognized as Safe (GRAS) substances, benzoic acid (21 CFR [Code of Federal Regulations] 184.1021)30 and sodium benzoate (21 CFR 184.1733)31 are classified as GRAS food substances. Sodium benzoate has also been approved by FDA for use as an antimycotic when migrating from food-packaging material (21 CFR 181.23).32 Other CFR citations relating to FDA-approved direct/indirect food additive ingredient uses include: benzyl alcohol (21 CFR 172.515, 175.105, 175.300, and 177.1210)33,34,35,36 ; benzoic acid (21 CFR 150.141, 150.161, 166.110, 175.300, and 177.1390)37,38,39,35,40 ; sodium benzoate (21 CFR 150.141, 150.161, and 166.110)37,38,39 ; calcium benzoate (21 CFR 166.110 and 178.2010)39,41 ; potassium benzoate (21 CFR 166.110 and 177.1210)39,36 ; and benzyl benzoate (21 CFR 172.515 and 175.105).33,34

Benzyl alcohol is an FDA-approved diluent in color additive mixtures for external drug use.42 In 2009, the FDA approved benzyl alcohol lotion, 5%, as a prescription medication for the treatment of head lice (Pediculosis capitis) infestation in patients 6 months of age and older.43 The results of clinical trials supporting benzyl alcohol lotion 5% (UlesfiaTM ) as a safe and effective topical treatment for head lice are included in the section on Skin Irritation and Sensitization later in the report text.44 Benzyl benzoate (10%) has been used in the treatment of scabies in developing countries.45

Benzyl alcohol has been classified by FDA regarding its use in the following types of over-the-counter (OTC) drug products (Category I: generally recognized as safe (S) and effective (E) for the claimed therapeutic indication; Category II: not generally recognized as safe and effective or unacceptable indications; Category III: insufficient data available to permit final classification):46

 anorectal (final monograph: 1 to 4% use concentration range established)  external analgesic (both Category I and final monograph pending)  external analgesic (final monograph: Category III)  oral health care (Category I pending)  oral discomfort care (Category II pending)  oral discomfort care (Category IIISE pending)  pediculicide (final monograph: Category III)  skin protectant (final monograph: Category III)

Similarly, benzoic acid has been classified by FDA regarding its use in the following types of OTC drug products:46

CIR Panel Book Page 11 Distributed for Comment Only - Do Not Quote or Cite  acne (final monograph: Category III)  antifungal (final monograph: Category III)  oral health care - (Category IIIE pending)  skin protectant - (final monograph: Category III)

Sodium benzoate has been classified by FDA regarding its use in menstrual/diuretic OTC drug products (final monograph: Category III), and benzyl benzoate has been approved for use in OTC pedulicides (final monograph: Category II).46

Calcium benzoate, magnesium benzoate, and potassium benzoate are inert non-food ingredients that, under the Federal Insecticide, Fungicide, and Rodenticide Act, are permitted by the Environmental Protection Agency (EPA) for use in nonfood use pesticide products.47 Pesticide products containing benzoic acid as the active ingredient have been registered with EPA for use in the extermination of dust mites.48

Benzyl alcohol has been listed as a chemical in photographic developing systems,49 and, at concentrations of 0.9 to 2%, is commonly used as an antibacterial agent in a variety of pharmaceutical formulations intended for intravenous administration.50 Additionally, benzyl alcohol has been used in parenteral medications commonly administered to critically ill neonates.51

TOXICOKINETICS

Oral Studies

Benzyl alcohol and benzoic acid and its salts are rapidly metabolized and excreted via a common pathway within 24 h. Benzyl alcohol is metabolized to benzoic acid via simple oxidation. Benzoic acid and sodium benzoate are rapidly absorbed from the gastrointestinal tract of mammals, conjugated with glycine in the liver, and then excreted as hippuric acid. The urinary excretion of benzyl benzoate as glycuronate conjugates and hippuric acid in mammals has also been reported.

Benzyl Alcohol, Benzoic Acid, Sodium Benzoate, and Potassium Benzoate

Benzyl alcohol is metabolized to benzoic acid. Both benzoic acid and sodium benzoate are rapidly absorbed from the gastrointestinal tract of mammals and conjugated with glycine in the liver; the resulting hippuric acid is rapidly excreted in the urine.1 Benzyl alcohol and benzoic acid and its sodium and potassium salt can be considered as a single category regarding human health, because they are all rapidly metabolized and excreted via a common pathway within 24 h.2

Benzyl Benzoate

Following oral administration (gavage, 0.5, 1, and 2 ml/kg) of benzyl benzoate to fasted female dogs, urine was collected for 5 to 6 days. All 3 doses resulted in creatinuria and urinary excretion of glycuronate conjugates.52

Following the administration of unspecified oral doses of benzyl benzoate to cats and dogs (number and breed not stated), small amounts of hippuric acid were detected in the 24 h urine. The metabolism of benzyl benzoate to benzoic acid in vivo has also been reported. Additional details for these 2 studies were not provided. 3

In a human subject, urinary excretion of hippuric acid was measured after oral dosing (single dose, 2.02 g) with benzyl benzoate. Urine was collected for 6 h. Approximately 90% of the dose administered was excreted as hippuric acid in the urine. After oral dosing with 1 g, approximately 80% of the administered dose was excreted as hippuric acid. In a second subject, 71% of the administered dose was excreted as hippuric acid during the first 6 h and an additional 14% of the dose was excreted as such during the second 6 h after dosing.52 The rate of hydrolysis of benzyl benzoate (in acetonitrile) by 80% human plasma in vitro was also studied. The t ½ for the in vitro hydrolysis of benzyl benzoate to benzoic acid in human plasma was 19 minutes.52

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In in vitro studies, the percutaneous absorption of benzoic acid through guinea pig skin and the percutaneous absorption of benzyl alcohol and benzyl benzoate through human skin was reported. The percutaneous absorption and urinary excretion of benzyl alcohol and benzyl benzoate was observed following dermal application to rhesus monkeys.

Benzoic Acid

The penetration of benzoic acid through excised dorsal skin (full-thickness and with stratum corneum removed) from guinea pigs was examined. Skin preparations were mounted in a two-chamber diffusion cell. A suspension of excess amount of benzoic acid in saline was added to donor compartments. In full-thickness skin, permeation proceeded with a short lag time. -2 -1 Solubility (Cd, unit = mM) and permeability (Kp, unit = x 10 cm · h ) coefficients for benzoic acid were 32.7 ± 1.6 and 9.01 ± 1.51, respectively. Removal of the stratum corneum by tape stripping and its delipidation using an organic solvent mixture enhanced the skin penetration of benzoic acid.53

Benzyl Alcohol and Benzyl Benzoate

The percutaneous absorption of [7-14C]benzyl alcohol and [7-14C]benzyl benzoate in vivo was evaluated using 4 female Rhesus monkeys (10 to 19 years old). Each chemical was applied (4 µg/cm2, without or with occlusion [glass chamber - G or plastic wrap - P]) to a clipped 1 cm2 area of abdominal skin; the vehicle was acetone (10 to 20 µl/cm2). The animals remained in metabolism chairs/metabolism cages for 5 days. The amount of absorbed compound in the urine was determined by liquid scintillation counting. Mean values (± SEM, 4 animals) for urinary excretion of the administered dose were: 77.4 ± 4.3% (benzyl alcohol) and 65.3 ± 13.4% (benzyl benzoate). Percutaneous absorption values (absorption, as % of applied dose) were as follows: benzyl alcohol (31.6 ± 4.2% [unoccluded]; 56.3 ± 14.5% [P]; 79.9 ± 7.4% [G]) and benzyl benzoate (57.0 ± 10.4% [unoccluded]; 71.2 ± 4.4% [P]; 64.7 ± 10.2% [G]). Over a 5-day period, the urinary excretion of radioactivity was 65.3 ± 13.4% following dermal application of benzyl benzoate and 56.5 ± 7.7% following dermal application of benzyl alcohol. When the acetone vehicle was replaced with a lotion (10 mg/cm2), percutaneous absorption was increased. The increase was significant for benzyl benzoate (P < 0.05), but not benzyl alcohol. There was no apparent correlation between the skin penetration of benzyl alcohol and benzyl benzoate and their octanol-water partition coefficients of 7.4 and 9333, respectively.54

The percutaneous absorption of benzyl benzoate in vitro was evaluated using human epidermis (from abdominal skin) placed in a glass diffusion chamber. Saline (5 ml) was added to the chamber and was in contact with the bottom of the epidermis. Benzyl benzoate (0.2 ml) was applied to the top of the epidermis. Six experiments were performed and the mean value for skin penetration of benzyl benzoate was 0.018± 0.002%.52

The penetration of benzyl alcohol through split-thickness cadaver skin was evaluated using nonoccluded Franz diffusion cells. Benzyl alcohol (spiked with 14C radiolabel) in ethanol was applied to the skin at doses ranging from 0.9 to 10,600 µg/cm2. After 24 h, the percentage of radioactivity that penetrated the skin increased gradually with dose, ranging from 19.8 ± 2.9% (lowest dose) to 29.2 ± 3.0% (highest dose). Also, less than 4% of the administered radioactivity was retained in the tissues at 24 h, and evaporation accounted for the remaining percentage. Data from this study were also analyzed in relation to a finite dose diffusion/evaporation model. The results of this analysis indicated that the increase in benzyl alcohol absorption with dose was consistent with a 3-fold increase in diffusivity in the stratum corneum, as the concentration of benzyl alcohol increased from tracer levels to saturation.55

TOXICOLOGICAL STUDIES

An Organization for Economic Cooperation and Development (OECD) Screening Information Data Sets (SIDS) for high volume chemicals initial assessment report on benzoic acid, sodium benzoate, potassium benzoate, and benzyl alcohol was published by the United Nations Environmental Program, Chemicals Branch (UNEP chemicals) in 2001, with the following recommendation relating to human safety: “Taking into account the rapid metabolism and excretion, the non- bioaccumulation, the low toxicity after acute and repeated exposures, the non-reproductive toxicity, the non-genotoxicity and the non-carcinogenicity, the low irritating and non- to very low sensitizing properties of these substances, as well as the 6

CIR Panel Book Page 13 Distributed for Comment Only - Do Not Quote or Cite controlled (industrial settings) and /or regulated (pharma, cosmetics and /or food) uses, these substances will pose a minimal risk to humans (workers and consumers).” Because the database for this safety assessment is similar to that for the 2001 CIR Final Report on Benzyl Alcohol, Benzoic Acid, and Sodium Benzoate, the available data in the OECD SIDS assessment are summarized with minimal details in sections that follow.

Acute and Repeated Dose Inhalation Toxicity

Neither acute nor repeated inhalation exposures to benzyl alcohol or benzoic acid caused death in rats. The results of a repeated dose inhalation toxicity study indicated no test substance-related macroscopic or microscopic findings for either test substance, and the no-observed-effect-level (NOEL) and no-observed-adverse-effect level (NOAEL) were considered to be 1,072 mg/m3 for benzyl alcohol and 12.6 mg/m3 for benzoic acid.

Benzyl Alcohol and Benzoic Acid

According to the OECD SIDS initial assessment report on Benzoates, 4 h of inhalation exposure to benzyl alcohol or benzoic acid at 4 and 12 mg/l aerosol/dust, respectively, did not cause death in rats. Thus, low acute toxicity was associated with these compounds.2

A 4-week inhalation toxicity study of aerosolized benzyl alcohol and benzoic acid was performed using groups of Crl:CD(SD) rats.56 Four groups of rats (10/sex/group) were exposed (nose-only) to aerosolized benzyl alcohol 5 days per week (6 h/day) for 4 weeks. Each animal received a minimum of 20 exposures, and target exposure concentrations were 30, 100, 300, and 1,000 mg/m3 for the four groups. Two additional groups of rats were exposed to aerosolized benzoic acid (2.5 and 12.5 mg/m3) according to the same procedure. The group designated for exposure to benzyl alcohol (30 mg/m3 target) was actually exposed to a mean atmosphere concentration that was 136% of the target concentration. For the remaining groups, all mean atmosphere concentrations were within 96.7% to 107.2% of the respective target concentrations. A concurrent control group was exposed to filtered air only.

There were no test-substance related deaths or effects on any of the following: body weight, food consumption, clinical pathology parameters, and organ weights. Additionally, there were no test substance-related macroscopic or microscopic findings. The (NOEL) and NOAEL were considered to be 1,072 mg/m3 for benzyl alcohol and 12.6 mg/m3 for benzoic acid.56

Acute Oral Toxicity

Some of the LD50 values that have been reported for various species include > 10,000 mg/kg for potassium benzoate in rats, mice, and guinea pigs, and the following values for benzyl benzoate: > 2,000 mg/kg (rats), ≤ 2,002 mg/kg (mice), 1,122 mg/kg (guinea pigs), 1,680 mg/kg (rabbits), 2,240 mg/kg (cats), and 2,244 mg/kg (dogs).

Acute oral toxicity data are summarized in Table 4.

Benzyl Alcohol, Benzoic Acid, Sodium Benzoate, and Potassium Benzoate

According to the OECD SIDS initial assessment report on Benzoates, low acute oral toxicity is associated with benzoic acid and its sodium and potassium salts. The LD50 values were > 2000 mg/kg body weight, except for benzyl alcohol (LD50 = 1610 mg/kg [rats]), which needs to be considered harmful via the oral route.2

Acute Intraperitoneal Toxicity

Benzyl Benzoate

Mice were dosed i.p. with benzyl benzoate (test procedure/number of mice not stated), and an LD50 of > 500 mg/kg body weight was reported.3

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An acute dermal LD50 of 2,000 mg/kg for benzyl alcohol in rabbits and acute dermal LD50s of 4,000 mg/kg (rats) and 4,448 mg/kg (rabbits) for benzyl benzoate have been reported. Death of cats has been observed, following single dermal applications of a benzyl benzoate solution.

Benzyl Alcohol, Benzoic Acid, Sodium Benzoate, and Potassium Benzoate

According to the OECD SIDS initial assessment report on Benzoates, low acute dermal toxicity is associated with benzyl alcohol and benzoic acid and its sodium and potassium salts.2

Benzyl Alcohol and Benzyl Benzoate

The acute dermal toxicity of benzyl benzoate was evaluated using rats (number not stated), and an LD50 of 4,000 mg/kg body weight was reported. Study details were not included. In a study involving rabbits, the acute dermal toxicity of undiluted benzyl benzoate was evaluated according to the Draize procedure (no GLP; number of animals not stated), and an LD50 of 4,448 mg/kg body weight was reported.3 In another study involving rabbits, acute dermal LD50s of 2,000 mg/kg and 4,000 mg/kg have been reported for benzyl alcohol and benzyl benzoate, respectively.57

Single dermal applications of 25% benzyl benzoate (20 ml in isopropanol/water) to the backs of cats resulted in death. Additional study details were not provided.3

Acute Subcutaneous Toxicity

Benzyl Benzoate

Guinea pigs (number not stated) received single s.c. injections of benzyl benzoate, and mortality rates at the administered doses were as follows: 0 (0.5 g/kg), 30% (1 g/kg), 80% (5 g/kg), 100% (10 g/kg), and 100% (20 g/kg).58

Acute Intramuscular Toxicity

Benzyl Benzoate

Following benzyl benzoate injection into cats (number not stated) i.m., doses of 2 and 10 g/kg were described as lethal.58

Repeated Dose Toxicity – Oral Studies

Benzoic acid and its salts and benzyl alcohol caused a low degree of toxicity in repeated dose oral toxicity studies. Toxic effects were not associated with repeated oral doses of benzyl benzoate.

Benzyl Alcohol, Benzoic Acid, Sodium Benzoate, and Potassium Benzoate

According to the OECD SIDS initial assessment report on Benzoates, repeated dose oral toxicity studies yielded an NOAEL of 800 mg/kg/day for benzoic acid and > 1000 mg/kg/day for the salts. At greater doses, increased mortality, reduced weight gain, and liver and kidney effects were observed. Systemic toxic effects of a similar nature (e.g. liver, kidney) were observed after dosing with benzyl alcohol, benzoic acid, sodium benzoate, and potassium benzoate. However, these effects were observed at higher doses of benzoic acid and its salts when compared to dosing with benzyl alcohol. Long-term studies on benzyl alcohol yielded an NOAEL of > 400 mg/kg body weight per day for rats and > 200 mg/kg body weight per day for mice. At higher doses, effects on body weight and lesions in the brain, thymus, skeletal muscle, and kidneys were observed. It was noted that one should take into account that oral administration was by gavage in these studies, whereby saturation of metabolic pathways is likely to occur. It was concluded that benzoic acid and its salts exhibited very low repeated dose toxicity and that benzyl alcohol exhibited low repeated dose toxicity.2

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No effects were observed in a 7-month toxicity study in which mice (no. and strain not stated) were dosed orally with benzyl benzoate (800 mg/kg).52

Repeated Dose Toxicity – Dermal Studies

Systemic/dermal effects induced by benzyl benzoate were observed in rats receiving repeated dermal doses up to 2.0 g/kg and in rabbits receiving repeated dermal doses up to 4.0 ml/kg. Test substance-related toxic signs were not observed in dogs, cows, horses, sheep or pigs that received repeated doses of benzyl benzoate.

Benzyl Benzoate

The following effects were observed in groups of rats (3 males/3 females/group; strain not stated) that received dermal applications amounting to 0.188, 0.488, 0.781, 1.25, or 2.0 g/kg for 30 days: skin effects, systemic effects, hyperplasia of squamous epithelium, degeneration of hair follicles and sebaceous glands, subcutaneous fibrosis, and hyperplasia of the thyroid gland. Seven areas of the shaved back were dosed in rotation. Dosing with 2.0 g/kg also caused 4 deaths, decreased white blood cell numbers, and other blood effects. Body weight changes, decreased white blood cell numbers, and other blood effects were also observed following dosing with 1.25 g/kg. Additional study details were not provided.52

In a study involving rabbits, (non-GLP; number and strain not stated) dermal doses of benzyl benzoate were applied daily for 90 days.3 The range of doses tested was not stated. Deaths occurred at doses of 2.2 g/kg body weight/day and greater. Testicular atrophy was noted at high doses, and, possibly, there was also an increased incidence of focal nephritis and encephalitis. Very mild skin irritation was also observed. Additional details rela(doseting to this study were not provided. Benzyl benzoate was also applied topically to rabbits at doses of 0.5 ml/kg/day daily for 90 days. Slight dermatitis and inanition were observed, and, at higher does, slight to moderate testicular atrophy was apparent. Increased leucocyte counts were reported and results were also suggestive of kidney damage. Benzyl benzoate also caused death, but the number of mortalities and corresponding toxic doses were not reported. Repeated dermal applications of benzyl benzoate (doses not stated) did not cause toxic symptoms in the following species: dogs, horses, cows (young females), sheep, or pigs. Additional study details were not provided.3

In another study, rabbits (no. and strain not stated) received dermal applications of benzyl benzoate at doses of 0.5, 1.0, 2.0, and 4.0 ml/kg daily for 90 days.52 Lethality and effects on the following were reported after dosing: blood effects (at 2.0 ml/kg), kidneys, skin (slight dermatitis and other effects), reproduction, leukocyte (increased 2.5 x normal), testicular damage, and kidney damage. Animal deaths were not preceded by the usual pattern of systemic effects prior to death. Additional details were not included.

Five applications of benzyl benzoate (undiluted) were made to the backs of the following domestic animals: 1 sheep (250 ml dose), 1 pig (200 ml), 1 young cow (500 ml), and 1 horse (1000 ml). The animals were observed for 2 weeks or until death. None of the animals died. In another study, 6 applications of undiluted benzyl benzoate were made to the backs (clipped skin, 4 " x 6" area) of 3 dogs as follows: 1 dog (200 ml volume) and 2 dogs (100 ml). No test substance-related effects were observed.52

Benzyl benzoate was rubbed onto cutaneous lesions on the shaved upper thighs of cats (no. not stated). Three applications per animal were made. Within 24 to 36 h, the test substance was lethal at doses of 8.7 to 12.8 g/kg.52

Repeated Dose Toxicity – Parenteral Studies

Subcutaneous dosing with benzyl benzoate resulted in death of mice (111 mg doses) and rabbits (2.5 ml/kg doses).

Benzyl Benzoate

Groups of adult C57BL/6J mice (5 per group) were injected s.c. with 0.1 ml benzyl benzoate (~111 mg) daily for 4 weeks. An untreated group of sham injected animals served as the untreated concurrent control group. The adrenal glands were

CIR Panel Book Page 16 Distributed for Comment Only - Do Not Quote or Cite prepared for light and electron microscopic examination. The animals died within 3 h of initial injection. Additional study details were not provided.3

Rabbits (1 or 2 animals) were injected s.c. once with benzyl benzoate in olive oil (doses of 1 to 2.5 ml/kg) or received 4 daily 0.25 mg/kg injections. Effects on leukocyte counts were monitored over an 11-day period and the animals were also examined for mortalities and clinical signs. The 2.5 ml/kg dose was lethal and clinical signs were observed. No effects were associated with the 0.25, 1, or 1.5 ml/kg doses.52

Ocular Irritation/Toxicity

Benzyl alcohol (4% aqueous), benzoic acid (undiluted), and its sodium salt (concentration not stated) were irritating to the eyes of rabbits, and it was expected that potassium benzoate would be irritating as well. A vehicle containing benzyl alcohol as a preservative for injected triamcinolone acetonide (TA) was toxic when instilled into the eyes of rabbits. However, it has been noted that retinal toxicity is not observed following injection of 0.1 ml preserved TA in clinical practice. Benzyl benzoate (undiluted) was irritating to the eyes of rabbits and humans.

Benzyl Alcohol

The ocular toxicity of benzyl alcohol was evaluated using 9 New Zealand rabbits. The following test concentrations of benzyl alcohol (in stock solution of carboxymethylcellulose and polysorbate 80) were injected intravitreally, each into 3 eyes (15 eyes total injected): 0.0073%, 0.022%, 0.073%, 0.222%, and 0.733%. Control eyes (2) were injected intravitreally with 0.9% normal saline. Neither clinical nor pathologic evidence of ocular toxicity was observed at the lowest concentration; transient clinical changes (retinal hemorrhage and whitening), but no remarkable pathological changes, were noted in 1 of 3 eyes tested with 0.022% benzyl alcohol. Ocular toxicity was observed at the remaining concentrations; changes in the outer retina included loss and shortening of outer segments and photoreceptors.59

Triamcinolone acetonide (TA) has been increasingly applied intravitreally in therapy for retinal diseases, and the vehicles of TA injections usually contain benzyl alcohol as a preservative. The ocular toxicity of 2 commercial vehicles (A and B) for drugs was evaluated using 3 groups of 12 New Zealand white rabbits (vehicle A, vehicle B, and balanced saline solution [BSS, negative control] groups). The 2 vehicles were similar in that each also contained benzyl alcohol (9.9 mg/ml), sodium carboxymethylcellulose (CBC, 7.5 mg/ml), and polysorbate 80 (0.4 mg/ml). Each group of 12 was divided into 2 groups, each receiving a different intravitreal dose (0.1 or 0.2 ml). Ocular examination (using ophthalmoscope) results were normal for group A and the negative control group; retinal damage was obvious in group B. However, at microscopic examination, retinal toxicity was observed in groups A and B. Vehicle B induced extensive retinal necrosis and atrophy, while vehicle B induced localized retinal changes.60

In a published comment on the preceding publication, the association of the benzyl alcohol vehicle with retinal toxicity was acknowledged. However , it was noted that retinal toxicity is not observed at concentrations achieved following injection of 0.1 ml preserved TA in clinical practice.61

In a similar study, 4 groups of Chinchilla rabbits (pigmented) were injected intravitreally with the following 4 materials, respectively: 0.1 ml BSS, 0.1 ml TA (1.3 mg/ml), 0.1 ml vehicle (0.99% BA and other excipient) alone, and 0.1 ml TA + vehicle. Given the materials tested, it should be noted that commercially prepared TA was described as a suspension containing 40 mg TA plus vehicle containing 0.99% BA, 0.75% excipient sodium carboxymethylcellulose, and 0.04% polysorbate. At microscopic examination, morphologic changes in the ciliary body, lens, and retina were observed only in 2 groups (TA + vehicle and vehicle alone). Thus, it was concluded that benzyl alcohol was toxic to these tissues when injected intravitreally.62

Benzoic Acid, Benzyl Alcohol, Sodium Benzoate, and Potassium Benzoate

According to the OECD SIDS initial assessment report on Benzoates, benzyl alcohol (4% aqueous) and benzoic acid (undiluted) were irritating to the eyes of rabbits and sodium benzoate (concentration not stated) was only slightly irritating. Data on potassium benzoate were not available; however, it was expected that this chemical would induce slight ocular irritation.2 10

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Undiluted benzyl benzoate was instilled (0.1 ml) into the eyes of 6 rabbits (3 males, 3 females), and reactions were scored up to 72 h post-instillation. Ocular irritation was observed in 3 rabbits (only reactions reported) at 1 h post-instillation.52

In an ocular irritation study involving human subjects (test procedure/number of subjects not stated), undiluted benzyl benzoate was classified as an ocular irritant.3

Skin Irritation and Sensitization

Undiluted benzyl alcohol and benzoic acid were slightly irritating and an unspecified concentration of sodium benzoate was non-irritating to the skin of rabbits. In studies involving guinea pigs, benzyl benzoate was non-irritating over the range of test concentrations, 2.5% to 50%. For the lower concentrations evaluated for sensitization in guinea pigs, benzyl benzoate was classified as a non-sensitizer at 5% and a weak sensitizer at 1%. At a concentration of 10%, results were consistent with classifications ranging from non-sensitizer to moderate sensitizer. Benzyl benzoate was classified as a mild sensitizer in guinea pigs when tested at a concentration of 40%. In rabbits, undiluted and 25% benzyl benzoate were classified as non- irritants. Results for healthy human subjects indicate that benzyl benzoate was classified as a non-irritant undiluted and at concentrations down to 2%, except for its classification as a minimal irritant at 50% in a very small percentage of subjects in one of the studies. Benzyl benzoate was classified as non-sensitizer when tested at a concentration of 30% in healthy human subjects. Most of the skin sensitization studies on benzyl benzoate involved groups of patients, and mixed results regarding sensitization potential were reported.

Skin irritation and sensitization data (human and non-human) are included in Table 5.

Benzyl Alcohol, Benzoic Acid, Sodium Benzoate, and Potassium Benzoate

According to the OECD SIDS initial assessment report on Benzoates, benzoic acid and benzyl alcohol were slightly irritating to the skin, whereas, sodium benzoate was non-irritating. Skin irritation data on potassium benzoate were not available; however, it is not expected that this compound would be irritating to the skin.2 In the same report, benzoic did not induce sensitization in animal studies; however, a low incidence of positive reactions was observed in dermatologic patients patch tested. Similar results were reported for sodium benzoate, and it has been suggested that the positive reactions observed were actually non-immunologic contact urticaria. Both positive and negative results were reported for benzyl alcohol in animal skin sensitization studies. A maximum skin sensitization incidence of 1% was reported for benzyl alcohol in human patch tests. Occupational exposure to benzyl alcohol, benzoic acid, or sodium benzoate has not resulted in skin sensitization over a period of decades.2

Case Reports

Case reports included positive patch test reactions to benzoic acid, negative patch test reactions to benzyl benzoate, and positive or negative patch or intradermal/prick test reactions to benzyl alcohol and sodium benzoate.

Case reports involving patch, intradermal, or prick testing of benzyl alcohol, benzoic acid, sodium benzoate, or benzyl benzoate are included in Table 6.

Cytotoxicity

Sodium Benzoate

The potential for sodium benzoate and other preservatives to induce lipid peroxidation in normal and α-linolenic acid -loaded cultured hepatocytes from male Wistar rats was evaluated. Cell injury was measured by the release of lactic acid dehydrogenase into the culture medium after 10 h of incubation. At concentrations up to 10 mM, sodium benzoate was not

CIR Panel Book Page 18 Distributed for Comment Only - Do Not Quote or Cite cytotoxic to either group of hepatocytes. Also, lipid peroxidation was not detected in either group of hepatocytes incubated with sodium benzoate.63

Benzyl Benzoate

In an in vitro assay, benzyl benzoate was toxic to ascites sarcoma BP8 cell cultures. Additional details were not provided.3

Phototoxicity

Benzyl Benzoate

The phototoxicity of benzyl benzoate was evaluated using hairless mice (number and strain not stated). Details relating to the test protocol were not included. Following test substance application, test sites were irradiated for a total of 3 or 4, 24 h exposures. No reactions were observed after 3 exposures; however, a slightly positive reaction was observed after 4 exposures.3

The phototoxicity of benzyl benzoate was evaluated, using 5 female Hartley guinea pigs, at concentrations of 10%, 30%, and 50% in acetone. At 4 h after depilation, the test substance was applied to a circular area (diameter = 1.5 cm) on both sides of the back. Two applications (left and right side) of each concentration were made to the back of each animal. One side of each animal was covered with aluminum foil and the other was irradiated with 5 Toshiba model FL-40 BLB lamps (320 to 400 nm) at a distance of 10 cm for 70 min. Photo-irritation was not observed at either of the 3 test concentrations.52

Photohemolysis

Neither benzyl alcohol (10-3 mol/l or 0.1%) nor sodium benzoate (10-3 mol/l) induced significant photohemolysis of human erythrocytes exposed to UVA/UVB light in vitro. However, in another assay, moderate photohemolysis of human erythrocytes was induced by benzyl alcohol (10-4 mol/l) only in the presence of UVA light, and benzyl benzoate did not induce photohemolysis in the presence of UVA or UVB light.

Benzyl Alcohol, Sodium Benzoate, and Benzyl Benzoate

The photohemolytic activity of benzyl alcohol and sodium benzoate in vitro was evaluated in a test involving human erythrocyte suspensions, incubated with either chemical (10-3 mol/l) for 1 h and exposed to UVA or UVB radiation. The UVA rich light source was a UVASUN 5000 lamp (320 to 460 nm; 42 mW/cm2) and the UVB rich light source was a lamp with TL 20 W/12 light bulbs (between 275 and 365 nm; 1 mW/cm2 [UVB] and 0.4 mW/cm2 [UVA]) Cultures without either chemical served as negative controls. Neither benzyl alcohol nor sodium benzoate induced significant hemolysis.64

A similar in vitro photohemolysis test (human erythrocyte suspensions) was used to evaluate the phototoxicity of benzyl alcohol and benzyl benzoate. The radiation sources described in the preceding study were used. Moderate hemolysis (~6%) was induced by benzyl alcohol (at 10-4 mol/l) only in the presence of UVA, classifying it as a weakly phototoxic compound. Benzyl benzoate was not phototoxic in the presence of UVA or UVB light.65

The photohemolytic activity of benzyl benzoate (0.1%) in vitro was evaluated using human red blood cells. Cultures were exposed to UVA or UVB radiation for up to 3 h and then centrifuged. Release of hemoglobin in the supernatant was determined as cyanmethaemoglobin. No photohemolytic activity was noted for 0.1% benzyl benzoate.52

REPRODUCTIVE AND DEVELOPMENTAL TOXICITY

In an OECD SIDS assessment report, benzoic acid (up to 750 mg/kg/day) was not a reproductive toxicant in rats in a four- generation reproductive toxicity study, and repeated doses of sodium benzoate (0.01g in diet) did not induce reproductive 12

CIR Panel Book Page 19 Distributed for Comment Only - Do Not Quote or Cite effect in rats. Developmental effects were observed in the presence of marked maternal toxicity in rats fed sodium benzoate in the diet ( ≥ 2800 mg/kg/day; NOAEL = 1400 mg/kg/day) in another study; maternal toxicity was not observed in hamsters (doses up to 300 mg/kg/day, also the NOEL), rabbits (doses up to250 mg/kg/day, also the NOEL) ,or CD-1 mice (doses up to 175 mg/kg/day, also the NOEL). Benzyl alcoholwas not a developmental toxicant in CD-1 mice (doses up to 750 mg/kg/day, also the LOAEL) or in mice of an unspecified strain (doses up to550 mg/kg/day, also the NOAEL) dosed by gavage. Benzyl benzoate(up to 1% in the diet) was not found to be teratogenic in rats dosed orally. However, testicular atrophy was observed in rabbits that received repeated dermal doses of benzyl benzoate (up to 1% in diet). There was no evidence of adverse effects on pregnancy outcome in a study on the safety of topical application of benzyl benzoate (25%) lotion in pregnant women. Benzoic acid (up to 1,000 µg/kg, s.c. injection) did not induce any reproducible evidence of estrogenic

activity in uterotrophic assays involving immature female Alpk:APfSD rats and female Alpk:APfCD-1 mice. Benzyl benzoate induced weak estrogenic responses in certain tests using cultured human breast cancer cells in vitro, but not in others.

Benzoic Acid, Sodium Benzoate, Potassium Benzoate, and Benzyl Alcohol

According to the OECD SIDS initial assessment report on benzyl alcohol, benzoic acid, and its sodium and potassium salts, benzoic acid did not induce reproductive effects in a four-generation reproductive toxicity study (NOAEL > 750 mg/kg).2 Groups of rats (20 rats/sex/group) received benzoic acid doses of 375 or 750 mg/kg/day in the diet continuously. Animals of the third generation were killed after 16 weeks. Also, test substance-related effects on reproductive organs (based on gross and microscopic examination) were not observed in subchronic studies (rats and mice) on benzyl alcohol and sodium benzoate.

In groups of rats fed sodium benzoate (doses up to 5600 mg/kg/day) during each day of gestation, developmental effects were observed only in the presence of marked maternal toxicity (reduced food intake and decreased body weight; NOAEL = 1400 mg/kg/day). All developmental effects were observed at doses ≥ 2800 mg/kg/day. On days 6 through 10, 15, or 18 of gestation, dosing with sodium benzoate by gavage in hamsters (NOEL = 300 mg/kg body weight), rabbits (NOEL = 250 mg/kg), and CD-1 mice (NOEL = 175 mg/kg), did not result in maternal toxicity; doses > NOEL were not tested. In mice of an unspecified strain dosed with benzyl alcohol by gavage, an NOAEL of 550 mg/kg (only dose) body weight for developmental toxicity was reported. An LOAEL of 750 mg/kg/day (only dose) for developmental toxicity was reported for CD-1 mice dosed orally (gavage) with benzyl alcohol. In this study, maternal toxicity (increased mortality, reduced body weight, and clinical toxicology) was observed.2

Sodium Benzoate and Benzyl Benzoate

The effect of sodium benzoate on biochemical aspects of pregnant female albino rats and survival of their offspring was evaluated using groups of 10 Sprague-Dawley rats.66 Two groups of weanling female rats were fed (ad libitum) an acceptable dose (0.01 g) and high dose (0.0125 g) of sodium benzoate in the diet daily for 12 weeks. Standard diet was fed to the control group. After mating, the animals were fed ad libitum during pregnancy. When compared to the control group, both doses of sodium benzoate induced a decrease in serum bilirubin and an increase in serum urea.

The high and unacceptable doses of sodium benzoate also induced an increase in serum uric acid (p < 0.01 and p < 0.05, respectively). Serum alanine aminotransferase (ALT) activity was significantly greater (p < 0.01) in the high dose group, but not at the acceptable dose. The high dose did not induce a significant increase in serum creatinine. Statistically significant ( p < 0.01) decreases in food intake, hemoglobin, and hematocrit were also reported. Pregnant rats that received high doses of sodium benzoate sustained a significant decrease (p < 0.05) in mean weight of their pups, and this decrease amounted to 13.6% of the corresponding mean control group weight.66

The teratogenicity of benzyl benzoate was evaluated using 21 rats (strains not stated) per dose; doses administered in the diet were defined as 0.04 or 1.0% (≈ 24 or 595 mg/kg body weight/day).3 The test substance was administered daily from day 0 of gestation to day 21 post-parturition. The results of examinations for external, skeletal, or visceral anomalies indicated that benzyl benzoate did not induce harmful effects in fetuses.

In a reproductive and developmental toxicity study, pregnant rats (no. and strain not stated) were fed diets supplemented with 0.04 or 1.0% benzyl benzoate from day 0 of gestation to day 21 post-parturition.52 Another group of rats was fed a control

CIR Panel Book Page 20 Distributed for Comment Only - Do Not Quote or Cite diet. There were no effects on reproductive or developmental toxicity parameters; no external, skeletal, or visceral anomalies were observed in fetuses from either treatment group. Minor variations were observed. A significantly decreased number of fetuses with incomplete sternebrae was noted in the 1% benzyl benzoate treatment group.

Testicular atrophy was observed in 2 repeated dose dermal toxicity studies (90 days) on benzyl benzoate involving rabbits.3 The exact doses at which this finding occurred were not stated. However, testicular atrophy was observed at daily doses > 0.5 g/kg/day. These studies are summarized in the Repeated Dose Toxicity – Parenteral Studies section earlier in the report text.

Pregnant NMRI mice (34 animals) were injected s.c. with an unspecified dose of benzyl benzoate in castor oil on days 1 and 11 of gestation.52 The animals were killed on gestation day 17. Untreated mice served as controls. Fetal observations relating to the following were made: counts, sex, weight, and malformations. There was no evidence of test substance-related effects on fetuses. Additional details were not included.

A developmental toxicity study was performed using Drosophila melanogaster.52 One group of males and females was raised on food medium with 4% benzyl benzoate in ethanol and the other group was raised on control feed medium. Virgin Canton-S males and females were collected daily for 4 days and then mated. The percentage of eggs hatched was 6.5 times greater in flies on feed containing benzyl benzoate, compared to those on control feed.

The safety of benzyl benzoate lotion (25% benzyl benzoate) as a topical treatment for scabies during pregnancy was assessed using a population of 444 pregnant women and their matched controls (1,776 pregnant women).67 The study population consisted of refugee and migrant women attending antenatal clinics on the Thai-Burmese border between August of 1993 and April of 2006. Most first treatments took place during the second and third trimesters, and the overall median gestation exposure was 24.5 weeks. Treated women (444) received 559 applications of butyl benzoate lotion (79.5%, 15.5%, 4.5%, and 0.l5% receiving 1, 2, 3, and 4 treatments, respectively).

Conditional Poisson regression was used to estimate risk ratios for outcomes of pregnancy (proportion of abortions, congenital abnormalities, neonatal deaths, stillbirths, and premature babies), mean birth weight, and estimated median gestational age for scabies and scabies-free women. Regarding pregnancy outcomes, there were no statistically significant differences between women treated with the lotion and their matched controls. Thus, there was no evidence of adverse effects on pregnancy outcome due to topical application of benzyl benzoate (25%) lotion.67

Benzoic Acid, Benzyl benzoate; Estrogenic effects

The estrogenic activity of benzoic acid was evaluated in the recombinant yeast human estrogen (ERα) receptor assay in vitro and in uterotrophic assays involving immature female Alpk:APfSD rats (21 to 22 days old) and immature female 68 Alpk:APfCD-1 mice. Immature mouse uterotrophic assays involved 3 daily s.c. injections of benzoic acid in corn oil (100 and 1,000 µg/kg doses; 5 ml/kg = dose volume), ending on day 4. The same protocol was used for the rat assays, with the exception that benzoic acid in arachis oil was injected (doses of 10, 100, and 1,000 µg/kg).

-7 -3 Benzoic acid (10 to 10 M) produced negative results in the recombinant yeast human estrogen (ERα) receptor assay in vitro. Overall (rat and mouse assays), benzoic acid produced one statistically significant and 3 statistically nonsignificant increases in average uterine weight and one significant and 6 nonsignificant decreases in average uterine weight. Therefore, in all uterotrophic assays, benzoic acid did not produce any reproducible evidence of estrogenic activity. Results for vehicle controls were negative and the positive control, estradiol, was uterotrophic.68

The estrogenic activity of benzyl benzoate (concentration range: 10-9 to 10-4 M) in vitro was evaluated in the E-screen test using MCF-7 breast cancer cells. Untreated cultures served as controls. In this test, MCF-7 cells proliferate in the presence of estrogen. Cell numbers were assessed by measurement of the total protein content, using the sulforhodamine B assay. Compared to control cultures, benzyl benzoate did not increase the proliferation of MCF-7 cells (p > 0.05) over the range of concentrations tested.69

The estrogenic activity of benzyl benzoate, benzyl salicylate, and butylphenylmethylpropional (Lilial) in the estrogen- responsive MCF7 human breast cancer cell line was evaluated using the following assays: competitive binding assay to 14

CIR Panel Book Page 21 Distributed for Comment Only - Do Not Quote or Cite estrogen receptor (ER) of MCF7 cytosol, competitive binding assay to recombinant ERα and ERβ, and the assay of stably transfected ERE-CAT reporter gene in MCF7 cells.70 In the latter assay, The ERE-CAT vector consisted of the estrogen response element (ERE) of the vitellogenin A2 gene from -331 to -295 bp cloned into the pBLCAT2 vector upstream of the thymidine kinase (tk) promoter. Cell proliferation experiments were also performed.

The following results indicate that all 3 chemicals produced estrogenic responses in cultured human breast cancer cells in vitro. 70 At 3,000,000-fold molar excess, each chemical was able to partially displace [3H]estradiol from recombinant human estrogen receptors ERα and ERβ, and from cytosolic ER of MCF7 cells.

At concentrations in the 5 x 10-5 to 5 x 10-4 M range, benzyl benzoate and the other 2 chemicals were able to increase the expression of a stably integrated estrogen-responsive reporter gene (ERE-CAT) and of the endogenous estrogen-responsive pS2 gene in MCF7 cells; however, these effects were at a lesser extent when compared to 10-8 M 17β-estradiol (5,000 to 50,000 molar excess of benzyl benzoate).

In cell proliferation experiments, each chemical increased the proliferation of estrogen-dependent cells over a 7-day period. Cell proliferation was inhibited by fulvestrant (antiestrogen), suggesting an ER-mediated mechanism. However, over a 35- day period, the extent of proliferation in the presence of 10-4 M benzyl benzoate, benzyl salicylate, or butylphenylmethylpropional increased to the same magnitude as that observed in the presence of 10-8 M 17β-estradiol over a 14-day period (10,000 molar excess of benzyl benzoate).70

GENOTOXICITY

According to the OECD SIDS initial assessment report on benzyl alcohol, benzoic acid, and its sodium and potassium salt, the weight of evidence of the in vitro and in vivo genotoxicity data indicates that these chemicals are not mutagenic or clastogenic. Mixed/equivocal results were apparent in in vitro assays, but negative results were reported in in vivo genotoxicity assays. Other test results appear to be consistent with these findings. Benzoic acid was non-genotoxic in the micronucleus test, but results were positive for sodium benzoate and potassium benzoate in the spore rec-assay to assess DNA damaging activity. Also, sodium benzoate caused a significant increase in sister chromatid exchanges when compared to control cultures. Benzyl benzoate was not mutagenic in the Ames test, but results were positive for benzyl alcohol and benzoic acid in the in vitro comet assay for evaluating DNA-damaging potential.

Benzoic Acid, Sodium Benzoate, Potassium Benzoate, and Benzyl Alcohol

According to the OECD SIDS initial assessment report on benzyl alcohol, benzoic acid, and its sodium and potassium salt, each chemical was not mutagenic in in vitro Ames tests. Various results (negative and positive [chromosomal/chromatid responses]) for sodium benzoate, potassium benzoate, and benzyl alcohol were obtained in other in vitro genotoxicity assays. However, while some mixed and/or equivocal in vitro chromosomal/chromatid responses have been observed, sodium benzoate and benzyl alcohol were not genotoxic in the in vivo cytogenetic assay, the micronucleus test, or in other in vivo assays. The weight of evidence of the in vitro and in vivo genotoxicity data indicates that these chemicals are not mutagenic or clastogenic.2

In the in vitro comet assay, the DNA-damaging potential of benzoic acid and benzyl alcohol was evaluated using human blood cells (lymphocytes).71 This assay is able to detect single and double strand breaks in DNA. Benzoic acid was evaluated at concentrations ranging from 0.05 to 5 mM and, benzyl alcohol, at concentrations ranging from 1 to 50 mM. Benzoic acid was tested at lower concentrations (≤ 5 mM) because this chemical has apoptotic effects at higher concentrations. The tail moment and % tail DNA in chemicals evaluated were compared to results for the solvent control (distilled water). Significantly increased tail moment and % tail DNA were noted only with 5 mM benzoic acid and with 25 and 50 M benzyl alcohol, indicating genotoxicity. A dose response was not observed.

Benzoic acid (in DMSO) was evaluated in the micronucleus test at concentrations ranging from 250 to 1,000 µg/ml (with and without metabolic activation) using L5178Y TK+/- clone 3.7.2C mouse lymphoma cells. Cultures without benzoic acid

CIR Panel Book Page 22 Distributed for Comment Only - Do Not Quote or Cite served as negative controls and mitomycin C served as the positive control. Benzoic acid was non-genotoxic over the range of concentrations tested, both with and without metabolic activation. The positive control was genotoxic.72

The genotoxicity of sodium benzoate was evaluated in the sister chromatid exchanges assay (SCE assay) using human blood lymphocyte cultures incubated for 72 h. Preparations were scored blindly for cells in their first mitosis, second mitosis, and third and subsequent divisions. Forty SCEs second division cells from each culture were scored for SCEs. Compared to negative control cultures, sodium benzoate (0.02, 0.2, 2, 4, and 8 mM) caused statistically significant delays in cell division (p < 0.01), indicative of weak cytostatic activity, but did not induce cytotoxicity. Also, when compared to negative control cultures, 8 mM sodium benzoate induced a statistically significant increase (p < 0.01) in SCEs/cell. Sodium benzoate (2 mM) induced a statistically significant decrease (p < 0.05) in SCEs/cell when compared to the 8 mM sodium benzoate culture.73

The DNA-damaging activity of sodium benzoate and potassium benzoate was evaluated in the spore rec-assay using Bacillus subtilis M45 (rec-) and H17 (rec+) strains with and without metabolic activation. Sodium benzoate (in water) was tested at doses of 16 and 20 mg/disk and potassium benzoate (in mixed solution of water and ethanol [1:1]) was tested at doses of 15 and 20 mg/disk. Results for both chemicals were judged positive in this assay.74

Benzyl Benzoate

In the Ames test, benzyl benzoate was not mutagenic to Salmonella typhimurium strains TA98 or TA 100 at doses up to 5,000 µg/plate (plate incorporation assay) or 5,000 mg/plate (pre-incubation assay). Benzyl Benzoate also was not mutagenic to the following strains at doses up to 3 µmol/plate (pre-incubation assay) with or without metabolic activation: TA98, TA100, TA1525, and TA 1537. In the recombination assay, benzyl benzoate was not mutagenic to Bacillus subtilis strains H17 or M45 at a concentration of 10 mg/disk.3

In another Ames test, the mutagenicity of benzyl benzoate (in ethanol) was evaluated using Salmonella typhimurium strains TA98, TA100, TA1535, and TA1537. Benzyl benzoate was tested at a concentration of 3 µmol/plate with and without metabolic activation and was not found to be mutagenic.52

Photogenotoxicity

The photogenotoxicity of benzoic acid (0.5%) and sodium benzoate (0.5%) has been demonstrated in vitro using bacterial cell suspensions.

Benzoic Acid and Sodium Benzoate

The photogenotoxicity of benzoic acid (0.5%) and sodium benzoate (0.5%) and other food additives was evaluated using Escherichia coli cell suspensions.75 Initially, the cytotoxic and genotoxic potential of each in the absence of sunlight (200 to 3,000 nm; mean intensity = 5.8 x 102 Wm-2) was evaluated by allowing the cells to remain in contact with the additive for 80 min in the dark. Neither benzoic acid nor sodium benzoate affected cell viability (i.e., not cytotoxic) or the number of spontaneous mutations in the absence of sunlight. The number of spontaneous mutations was 2.09 mutations per 108 cells. Also, these chemicals did not induce mutations in the absence of sunlight.

Plates containing the cellular suspension (5 ml) and a food additive (0.5% concentration) were exposed to sunlight for periods ranging from 0 min to 60 min. Exposure to sunlight resulted in cell death in the presence or absence of benzoic acid or sodium benzoate. When cells were exposed to direct sunlight in the absence of benzoic acid or sodium benzoate, the number of induced mutations increased with duration of exposure (5.1 x 102 mutations per 108 cells after 60 min). In the presence of benzoic acid or sodium benzoate, the number of induced mutations increased with the duration of exposure to sunlight. This increase was almost additive over that observed with sunlight exposure alone. After 60 min, there were approximately 1 x 103 mutations per 108 cells.75

Effect on DNA Synthesis

Sodium benzoate caused a significant increase in the DNA content of protozoan nuclei, suggestive of stimulation of mitosis. 16

CIR Panel Book Page 23 Distributed for Comment Only - Do Not Quote or Cite Sodium Benzoate

Aqueous sodium benzoate, at a dose of 7.20 mg/ml, was added to Tetrahymena pyriformis in an experimental series consisting of six 100 ml cultures. Another experimental series, without sodium benzoate, served as the control. The quantitative analysis of DNA content of the protozoan nuclei was performed using an image analysis system. Sodium benzoate caused a statistically significant increase (p < 0.01) in DNA content, suggestive of stimulation of mitosis. The authors speculated that this observation might represent a direct cytotoxic effect due to increased DNA content of nuclei. 76

CARCINOGENICITY

Benzyl alcohol was not carcinogenic when tested dermally on mice at 2.00% in a nonoxidative hair dye. NTP considered benzyl alcohol negative for carcinogenicity following 2 years of oral dosing in rats(doses up to 400 mg/kg) and mice (doses up to 200 mg/kg), but the EPA considered the 3 of 48 incidence of adrenal cortex adenoma to be equivocal evidence of carcinogenic activity rather than negative. Benzoic acid was negative for carcinogenicity when dermally tested on mice at 0.016% in a nonoxidative hair dye. Sodium benzoate was negative for carcinogenicity when administered orally at up to 2% to rats (in feed for up to 2 years) or mice (in a life-time drinking water study).

The following carcinogenicity study summaries are included in the CIR final report on benzyl alcohol, benzoic acid, and sodium benzoate that was published in 2001.1

Benzyl Alcohol and Benzoic Acid

Groups of 100 F344/N rats 50 each sex) were dosed orally with 200 or 400 mg/kg benzyl alcohol in corn oil, 5 days per

week for 103 weeks. Groups of 100 B6C3F1 mice were dosed with 100 or 200 mg/kg benzyl alcohol following the same schedule. During week 80, mice were mistakenly dosed for four days with 375 (low-dose group) and 750 mg/kg (high- dose group) of α-methylbenzyl alcohol. Dose-related negative trends were noted in the incidences of anterior pituitary gland neoplasms in female rats (vehicle control, 29/50; low dose, 17/47; high dose, 9/49) and of Harderian gland adenomas in male mice (8/50; 3/50; 2/50). Epithelial hyperplasia of the nonglandular stomach was noted in 4 of 50 high- dose male rats; it was not found in controls or low-dose male rats. An increased incidence of adenomas of the adrenal cortex noted in high-dose male mice (0/48; 0/44; 3/48) was within historical range and not considered compound-related. The NTP investigators considered the study negative for benzyl alcohol-induced carcinogenicity. However, reviewing the study, the EPA considered the 3 of 48 incidence of adrenal cortex adenoma to be “equivocal evidence of carcinogenic activity rather than negative.” From the Final Report on benzyl alcohol, benzoic acid, and sodium benzoate1

A skin painting study was performed using groups of 120 Eppley Swiss mice (60 per sex). A non-oxidative hair dye containing 2.0% benzyl alcohol and 0.016% benzoic acid was painted onto the skin at a dose of 0.05 ml/application, three times weekly for 20 months. Sites were shaved of hair 24 h before each application and a new bottle of dye was used each week. Two groups of control animals were shaved but not treated. Nine months into the study, 10 mice/sex/group were killed. Body weights and survival differed little between treatment and control groups. Varying degrees of chronic dermal inflammation were noted in all groups, including the controls. A significant (p < .01) increase in malignant lymphomas was noted in treated females (23/60). However, the researchers noted that one concurrent control group had a very low incidence (7/60 or 12%) of that tumor type. The rate was 22% for the other control group and had averaged 33% for three control groups in previous studies. Thus, the findings were not considered treatment- related. The incidence of pulmonary adenomas and hepatic hemangiomas, which are common in this mouse strain, were similar between treated and control groups. No unusual neoplasms were observed.

From the Final Report on benzyl alcohol, benzoic acid, and sodium benzoate1

Sodium Benzoate

For 18 to 24 months, groups of Fischer 344 rats (50 males and 52 females per group) received feed containing 2% or 1% sodium benzoate. The doses corresponded to the maximum tolerated dose (MTD) and ½MTD as determined in 6-week 17

CIR Panel Book Page 24 Distributed for Comment Only - Do Not Quote or Cite toxicity studies. A control group of 25 male and 43 female rats received untreated feed. Average daily sodium benzoate intake was 280 and 202 mg, respectively, for male and female rats of the 2% group, and 141 and 102 mg, respectively, for male and female rats of the 1% group. No clinical signs of toxicity or differences in average body weight or mortality rates were noted in treated rats when compared with controls. Neoplasms that were present in treated rats were similar in type and number to those in controls. No evidence of sodium benzoate-related carcinogenicity was observed. From the Final Report on benzyl alcohol, benzoic acid, and sodium benzoate1

In a life-time drinking water study, 100 Albino Swiss mice (50 of each sex) were supplied with water containing 2% sodium benzoate. A control group of 200 mice was supplied with untreated water. Average daily intake of sodium benzoate was 124.0 and 119.2 mg for males and females, respectively. Sodium benzoate treatment did not affect survival. No carcinogenic effect attributable to treatment was noted at necropsy. From the Final Report on benzyl alcohol, benzoic acid, and sodium benzoate1

SUMMARY

The safety of the following ingredients in cosmetics is reviewed in this safety assessment: Benzyl alcohol, benzoic acid, sodium benzoate, calcium benzoate, magnesium benzoate, potassium benzoate, and benzyl benzoate. Most of these ingredients would function as fragrance ingredients/preservatives in cosmetic products. Together, data reported to the Food and Drug Administration’s Voluntary Cosmetic Registration Program in 2010 and the results of a 2010 Personal Care Products industry survey indicated use of the following ingredients in cosmetics: benzyl alcohol, benzoic acid, sodium benzoate, calcium benzoate, potassium benzoate, and benzyl benzoate. According to this industry survey, ingredient use concentrations have ranged from 0.000001% (sodium benzoate) to 10% (benzyl alcohol).

Among the methods of manufacture identified are the production of benzyl alcohol via the action of sodium or potassium carbonate on benzyl chloride and the production of benzoic acid via the decarboxylation of phthalic anhydride. Hydroxyl radical generated by the metal-catalyzed reduction of O2 and H2O2 by ascorbic acid can attack benzoic acid to produce benzene under conditions that are prevalent in foods and beverages. Additionally, exposure to UV light and elevated temperature over the shelf life of beverages may result in benzene formation in products containing benzoic and ascorbic acids. The U.S. beverage industry voluntarily reformulated beverages that were found to contain benzene levels at or above the 5 ng/g maximum contaminant level for drinking water established by the U.S. Environmental Protection Agency. Limits for heavy metal impurities in food-grade (as Pb, ≤ 10 mg/kg) and pharmaceutical-grade (heavy metals, ≤ 0.001%) potassium benzoate have been established.

Benzyl alcohol is metabolized to benzoic acid via simple oxidation. Benzoic acid and sodium benzoate are rapidly absorbed from the gastrointestinal tract of mammals, conjugated with glycine in the liver, and then excreted as hippuric acid. In in vitro studies, the percutaneous absorption of benzoic acid through guinea pig skin and the percutaneous absorption of benzyl alcohol and benzyl benzoate through human skin was reported. The percutaneous absorption and urinary excretion of benzyl alcohol and benzyl benzoate was observed following dermal application to rhesus monkeys.

Benzoic acid (up to 1,000 µg/kg, s.c. injection) did not induce any reproducible evidence of estrogenic activity in

uterotrophic assays involving immature female Alpk:APfSD rats and female Alpk:APfCD-1 mice. However, benzyl benzoate induced estrogenic responses in cultured human breast cancer cells in vitro. Specifically, at 3,000,000-fold molar excess, benzyl benzoate was able to partially displace [3H]estradiol from recombinant human estrogen receptors ERα and ERβ, and from cytosolic ER of MCF7 cells. The antiparasitic activity of benzyl benzoate has been demonstrated in vitro.

Neither acute (up to 12 mg/l aerosol/dust) nor repeated inhalation (up to 1,000 mg/m3) exposures to benzyl alcohol or benzoic acid caused death in rats. Results of the repeated dose inhalation toxicity study also indicated no test substance-related macroscopic or microscopic findings for either test substance. Overall, the results of acute oral and dermal toxicity studies indicated a low level of toxicity for benzyl alcohol, benzoic acid, and benzyl benzoate. The same was true for repeated dose oral toxicity studies on benzoic acid and its salts and benzyl alcohol. Systemic/dermal effects induced by benzyl benzoate were observed in rats receiving repeated dermal doses up to 2.0 g/kg and in rabbits receiving repeated dermal doses up to 4.0

CIR Panel Book Page 25 Distributed for Comment Only - Do Not Quote or Cite ml/kg. However, test substance-related toxic signs were not observed in other species that received repeated doses of benzyl benzoate.

Benzyl alcohol (4% aqueous), benzoic acid (undiluted), and its sodium salt (concentration not stated) were irritating to the eyes of rabbits, and it was expected that potassium benzoate would be irritating as well. Benzyl benzoate (undiluted) was irritating to the eyes of rabbits and humans. Undiluted benzyl alcohol and benzoic acid were slightly irritating and benzyl benzoate was non-irritating to the skin at concentrations up to 50% in animal studies. Undiluted benzyl benzoate was a non- irritant in human subjects. In animal studies, benzyl benzoate induced sensitization reactions ranging from none to moderate. Moderate sensitization was observed at a concentration of 10% benzyl benzoate, but not at lower concentrations, and there was one report of mild sensitization at a concentration of 40%. Benzyl benzoate was classified as non-sensitizer when tested at a concentration of 30% in healthy human subjects. Most of the skin sensitization studies on benzyl benzoate involved groups of patients, and mixed results regarding sensitization potential were reported.

A slightly positive reaction was observed in a phototoxicity study involving hairless mice tested with an unspecified concentration of benzyl benzoate. Photo-irritation was not observed in guinea pigs tested with benzyl benzoate at concentrations up to 30%. Moderate photohemolytic activity in human erythrocytes was observed in the presence of benzyl benzoate, but significant photohemolytic activity was not associated with sodium benzoate or benzyl alcohol in these cells

Developmental effects were observed in the presence of marked maternal toxicity in rats fed sodium benzoate in the diet ( ≥ 2800 mg/kg/day; NOAEL = 1400 mg/kg/day). Testicular atrophy was observed in rabbits that received repeated dermal doses of benzyl benzoate (up to 1% in diet). However, overall, benzyl alcohol, benzoic acid, and sodium benzoate were not classified as reproductive/developmental toxicants in oral and dermal animal studies. There was no evidence of adverse effects on pregnancy outcome due to topical application of benzyl benzoate (25%) lotion in pregnant women.

Mixed/equivocal results were apparent in in vitro assays, but negative results were reported in in vivo genotoxicity assays on benzyl alcohol, benzyl benzoate, benzoic acid, and its sodium and potassium salt. The photogenotoxicity of benzoic acid (0.5%) and sodium benzoate (0.5%) has been demonstrated in vitro using bacterial cell suspensions. Benzyl alcohol was negative for carcinogenicity when dermally tested on mice at 2.00% in a nonoxidative hair dye. NTP considered it negative for carcinogenicity following 2 years of oral dosing in rats (up to 400 mg/kg) and mice (up to 200 mg/kg), but the EPA considered the 3 of 48 incidence of adrenal cortex adenoma to be equivocal evidence of carcinogenic activity rather than negative. Benzoic acid was negative for carcinogenicity when dermally tested on mice at 0.016% in a nonoxidative hair dye. Sodium benzoate was negative for carcinogenicity when administered orally at up to 2% to rats (in feed for up to 2 years) or mice (in a life-time drinking water study).

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Table 1. Benzyl Alcohol and Benzoic Acid and its Salts and Ester22 Chemical Names/CAS Nos. Definitions* Functions in Cosmetics Benyl Alcohol An aromatic alcohol External analgesics; fragrance Alcohol benzylicus; enzenemethanol; ingredients; oral health care drugs; benzylic alcohol; phenylcarbinol; preservatives; solvents; and viscosity phenylmethanol; phenylmethyl decreasing agents alcohol; and α-toluenol; CAS No. 100- 51-6 Benzoic Acid An aromatic acid Fragrance ingredients; preservatives; Acidum benzoicum; and pH adjusters benzenecarboxylic acid; benzeneformic acid; benzenemethanoic acid; carboxybenzene; dracyclic acid; phenylcarboxylic acid; and phenylformic acid; CAS No. 65-85-0 Sodium Benzoate The sodium salt of benzoic acid Corrosion inhibitors; fragrance Benzoic acid, sodium salt and natrii ingredients; and preservatives benzoas (EP); CAS No. 532-32-1

Calcium Benzoate The calcium salt of benzoic acid Preservatives Benzoic Acid, Calcium Salt and Calcium Dibenzoate; CAS No. 2090- 05-3

Magnesium Benzoate The magnesium salt of benzoic acid Preservatives Benzoic Acid, magnesium salt and magnesium dibenzoate; CAS No. 553- 70-8

Potassium Benzoate The potassium salt of benzoic acid Preservatives Benzoic Acid, Potassium Salt; CAS No. 582-25-2

Benzyl Benzoate The ester of benzyl alcohol and Fragrance ingredients; pesticides; and Benylate; benzoic acid, benzyl ester; benzoic solvents benzoic acid, phenylmethyl ester; benzylis benzoas (EP); and phenylmethyl benzoate; CAS No. 120- 51-4 *See Figure 1 for structures, completing the definition of each chemical.

Table 2. Properties of Benzyl Alcohol and Benzoic Acid and its Salts and Ester77 Benzyl Benzoic Sodium Calcium Magnesium Potassium Benzyl Properties Alcohol Acid Benzoate Benzoate Benzoate Benzoate Benzoate

Form Liquid Monoclinic White White powder White solid2 Leaflets/oily plates granules liquid Molecular weight 108.14 122.12 144.1 121.1278 266.53 160.21 212.24 Density 1.045 1.321 1.118 Melting point -15.19⁰C 122.4⁰C 330.6⁰C2 200⁰ 330.6⁰C2 21⁰C Boiling point 204.7⁰C 249.2⁰C 464.9⁰C2 464.9⁰C2 323-324⁰C Water Practically dissolves 1 g dissolves Soluble in 20 Solubility 2.9 g/l 556 g/l2 insoluble in (1g) in ~ 25 in 1.8 ml parts water ml water water water logP 1.12 1.82 -2.2692 1.5379 -2.2692 3.6880 UV absorption 228.6 nm78 226.8 nm78 256 nm78 max.

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Table 3. Current Frequency and Concentration of Use According to Duration and Type of Exposure Provided in 201123,24 Benzoic Acid Benzyl Alcohol Sodium Benzoate Calcium Benzoate # of # of # of # of Uses Conc. (%) Uses Conc. (%) Uses Conc. (%) Uses Conc. (%) Exposure Type 0.000008 to Eye Area 23 0.00005 to 0.2 109 0.8 58 0.00001 to 0.4 NR NR 0.000006 to Possible Ingestion 64 0.003 to 2 23 0.9 33 0.001 to 0.6 NR NR 0.000003 to Inhalation 9 0.08 431 0.0003 to 3 34 0.0001 to 0.3 NR NR 0.002 to Dermal Contact 550 0.000002 to 5 1694 0.000008 to 2 1075 0.000001 to 1 NR 0.004 Deodorant (underarm) 1 0.03 to 0.5 38 0.03 to 0.5 2 0.5 NR NR Hair - Non-Coloring 97 0.00002 to 1 359 0.0002 to 3 431 0.000005 to 1 NR NR Hair-Coloring 22 0.004 to 0.03 56 0.00007 to 10 18 0.008 to 0.5 NR NR 0.00002 to 0.00001 to Nail 7 0.0005 to 0.2 3 0.5 7 0.001 NR NR Mucous Membrane 134 0.000003 to 2 228 0.000006 to 1 391 0.0004 to 0.6 NR NR 0.000003 to Bath Products 23 0.08 20 0.009 to 0.9 15 0.004 to 1 NR NR Baby Products 3 0.000002 25 0.004 to 0.3 26 0.3 to 0.9 NR NR Duration of Use 0.002 to Leave-On 378 0.000002 to 1 1511 0.000008 to 3 636 0.00001 to 1 NR 0.004 0.000006 to Rinse off 318 0.000003 to 5 649 10 951 0.000001 to 1 NR NR 0.000006 to 0.002 to Totals/Conc. Range 696 0.000002 to 5 2160 10 1587 0.000001 to 1 NR 0.004 Potassium Benzoate Benzyl Benzoate Magnesium Benzoate # of # of # of Uses Conc. (%) Uses Conc. (%) Uses Conc. (%) Exposure Type 0.0006 to Eye Area NR NR 5 0.002 NR NR 0.000005 to Possible Ingestion NR NR 82 0.2 NR NR Inhalation NR NR 368 0.002 to 4 NR NR Dermal Contact NR 0.0002 1402 0.000005 to 4 NR NR Deodorant (underarm) NR NR 47 0.006 to 0.6 NR NR Hair - Non-Coloring 1 NR 259 0.006 to 4 NR NR Hair-Coloring NR NR 21 0.07 to 2 NR NR Nail 1 0.0003 26 0.2 to 0.8 NR NR Mucous Membrane NR 0.0002 115 0.002 to 0.4 NR NR Bath products NR NR 31 0.0005 to 2 NR NR Baby Products NR NR 3 NR NR NR Duration of Use Leave-On 1 0.0003 1403 0.000005 to 4 NR NR Rinse off 1 0.0002 313 0.0001 to 2 NR NR Totals/Conc. Range 2 0.002 to 0.003 1716 0.000005 to 4 NR NR NR = Not Reported; NS = Not Surveyed; Totals = Rinse-off + Leave-on Product Uses. Note: Because each ingredient may be used in cosmetics with multiple exposure types, the sum of all exposure type uses may not equal the sum total uses.

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Table 4. Acute Oral Toxicity Studies on Potassium and Benzyl Benzoate Material Animals Test Procedure Results Potassium Benzoate Rats, mice, and guinea pigs Not stated LD50 > 10,000 mg/kg2 Benzyl Benzoate (undiluted) 10 rats/dose Oral dosing followed by 6-day observation LD50 = 1891 mg/kg3 period; no GLP* Benzyl Benzoate 20 rats Single dose by gavage; 14-day observation LD50 = 2800 mg/kg3 period; no GLP

Benzyl Benzoate Rats (no. not stated) Not stated LD50 = 500 mg/kg3 Benzyl Benzoate (in arachis Rats (no. not stated) OECD guideline 401; 100 to 2000 mg/kg LD50 > 2000 mg/kg3 oil) oral doses; GLP study Benzyl Benzoate (in arachis 10 Sprague-Dawley albino rats Single 5.0 ml/kg dose; 14-day observation LD50 > 2000 mg/kg58 oil) period Benzyl Benzoate Rats (no. not stated) Not stated LD50 = 1700 mg/kg58 Benzyl Benzoate (1% in 5 groups of 10 Sprague-Dawley Doses up to 4000 mg/kg total (i.e., four 250 LD50 = 1.16 g/kg58 gum tragacanth suspension) rats mg/kg doses at 40 ml/kg dose volume/dose); 14-day observation period Benzyl Benzoate (in Tween 6 groups of 10 Sprague-Dawley 6 different dose levels (dose volume = 40 LD50 = 1160 mg/kg58 80 + gum tragacanth rats 5 males, 5 females/group) ml/kg) vehicle) Benzyl Benzoate (in tween Rats (25 males, 25 females) Not stated LD50 = 1550 mg/kg58 80 solution + methylcellulose vehicle) Benzyl Benzoate (in tween Sprague-Dawley rats (5 males, 10 mg/kg total dose (i.e., four 2500 mg/kg LD50 > 10 g/kg58 80 solution + gum 5 femlaes) doses); 14-day observation period tragacanth vehicle) Benzyl Benzoate 10 rats/dose Dosing followed by 6-day observation LD50 = 1.7 ml/kg58 period Benzyl Benzoate 20 rats Dosing followed by 2-week observation LD50 = 2.8 g/kg58 period Benzyl Benzoate (undiluted) 10 mice/dose Dosing followed by 6-day observation LD50 = 1557 mg/kg3 period; no GLP Benzyl Benzoate 10 mice/dose Dosing followed by 6-day observation LD50 = 1.4 ml/kg58 period

Benzyl Benzoate Mice (no. not stated) Not stated LD50 = 1400 mg/kg3 Benzyl Benzoate (undiluted) 10 rabbits/dose Dosing followed by 6-day observation LD50 = 2002 mg/kg3 period; no GLP Benzyl Benzoate 10 rabbits/dose Dosing followed by 6-day observation LD50 = 1.8 ml/kg58 period Benzyl Benzoate 9 rabbits Single dose by gavage; 14-day observation LD50 = 1680 mg/kg3 period; no GLP Benzyl Benzoate 12 rabbits Dosing followed by 2-week observation LD50 = 1.68 g/kg58 period Benzyl Benzoate 11 cats Single dose by gavage; 14-day observation LD50 = 2240 mg/kg3 period; no GLP Benzyl Benzoate 4 dogs Single dose by gavage; 14-day observation LD50 > 2244 mg/kg3 period; no GLP Benzyl Benzoate (undiluted) 10 guinea pigs/dose Oral dosing followed by 6-day observation LD50 = 1112 mg/kg3 period ; no GLP Benzyl Benzoate Guinea pigs (no. not stated) Not stated LD50 = 1000 mg/kg3

Benzyl Benzoate 10 guinea pigs/dose Dosing followed by 6-day observation LD50 = 1 mg/kg58 period *GLP = good laboratory practice

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Table 5. Skin Irritation and Sensitization Studies Doses/Concentrations Test Substance Animals/Subjects Tested Procedure Results Animal Studies Benzyl alcohol Rabbits (no. not undiluted benzyl alcohol dose applied for 24 h Slightly irritating2 (purity not stated) stated) (10 mg) Benzoic Acid (purity 3 albino rabbits undiluted benzoic acid (0.5 dose applied to flank under semi- Slightly irritating2 not stated) g/kg) occlusive dressing for 4 h

Sodium Benzoate Rabbits (no. not undiluted benzyl alcohol dose applied for 24 h Slightly irritating2 stated) (10 mg) Sodium Benzoate Rabbits (no. not Concentration not stated OECD Guide - line 404 protocol Non-irritating2 stated) Benzyl Benzoate Guinea pigs (no. not undiluted benzyl benzoate Dose applied to intact abdominal skin Non-irritating58 stated) (15 g/kg) (duration not stated)

Benzyl Benzoate Guinea pigs (no. not up to 50% Not stated Non-irritant58 (50%) stated) Benzyl Benzoate 10 female Hartley 10% induction 10% w/v in complete adjuvant Non-sensitizer at 5% (40%) albino guinea pigs concentration; challenge (intradermal induction); 10% w/v in and 10% benzyl concentrations up to 40% Freund's complete adjuvant + saline benzoate; mild (topical induction); topical challenge sensitizer at 40% (4 of concentrations up to 40% benzyl 10 animals) 58 benzoate in propylene glycol + acetone

Benzyl Benzoate 4 male guinea pigs up to 25% 24-h occlusive patch test Non-irritant58 (25% in acetone/PEG vehicle)

Benzyl Benzoate Guinea pigs (no. not 10% Maximization test Moderate (10%) stated) sensitization58 Benzyl Benzoate 4 female guinea pigs up to 10% 24-h occlusive patch test; up to 10% Non-irritating at 5 and (10% in ethanol) during induction and challenge with 2.5% 10% concentrations. Barely perceptible erythema (at 24 and 48 h) at 2.5% concentration58

Benzyl Benzoate 10 guinea pigs 10% Modified Fruend's complete adjuvant test Weak sensitization58 (10% in acetone) Benzyl Benzoate (1 Guinea pigs (no. not 1% and 10% (15 or 30 mg) Modified Freund's complete adjuvant test: Weak sensitization at and 10%) stated) Challenge with 1% and 10% benzyl both concentrations58 benzoate (in acetone) Benzyl Benzoate Guinea pigs (no. not Not stated Maximization test (no GLP) Non-sensitizer3 stated) Benzyl Benzoate Guinea pigs (6 to 8 Not stated Open epicutaneous test (24 h application) Non-sensitizer3 per group) Benzyl Benzoate 4 female New Undiluted benzyl benzoate 4 h semiocclusive patch test Non-irritant58 Zealand albino (0.5 ml/2 cm2) rabbits Benzyl Benzoate Rabbits (no. not Not stated 3-week subacute skin toxicity test Very mild gross skin stated) irritation58 Predictive Tests Benzyl Alcohol 31 healthy subjects 0.2 ml benzyl alcohol Hill Top chamber with Webril pad Skin irritation in 1 applied to upper arm for up to 4 h subject81 Benzyl Alcohol 485 subjects Lotion tested as supplied Topical treatment of head lice program Low frequency of lotion 5% (Ulesfia application site TM) irritation (2.3% incidence)44

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Table 5. Skin Irritation and Sensitization Studies Doses/Concentrations Test Substance Animals/Subjects Tested Procedure Results Benzoic Acid 86 healthy subjects 1 M in petrolatum Finn chambers applied to volar arm for Caused contact (mean age = 37.3 20 min urticaria82 years) Benzoic Acid 58 healthy subjects 1 M in petrolatum (50 µl Finn chambers applied to volar arm for Caused contact (mean age = 39 volume) 20 min urticaria83 years) Benzyl Benzoate 18 subjects (18+ 5 to 10 drops of undiluted Closed patch containing benzyl benzoate No skin irritation58 years) benzyl benzoate applied to healthy and locally inflammed skin for up to 48 h Benzyl Benzoate Male and female undiluted benzyl benzoate 24-h closed patch test No skin irritation58 subjects (no. not (1cm-diameter area) stated; 18+ years) Benzyl Benzoate 21 subjects (24 to 70 50% (0.3 ml) 24-h occlusive patch test Minimal erythema in 3 (50% in diethyl years old) subjects; negligible phthalate/ethanol) irritation potential58 Benzyl Benzoate 129 subjects (46 50% (0.3 ml) 24-h RIPT No effects58 (50% in males, 83 females) ethanol/diethyl phthalate)

Benzyl Benzoate 20 healthy males 30% Occlusive patch test; 5 48 h applications No sensitization58 (30% in petrolatum) Benzyl Benzoate 34 healthy male and 20% 48-h closed patch test No skin irritation58 (20% in vaselinum female subjects aldum or unguentum hydrophilicum vehicle)

Benzyl Benzoate (4% 25 healthy male and 4% 48-h closed patch test No skin irritation58 in petrolatum) female subjects Benzyl Benzoate (2% 30 healthy male and 2% 24- to 72-h closed patch test No skin irritation58 in unguentum female subjects (18 + simplex or years) unguentum hydrophilicum vehicle) Provocative Tests/Treatment Benzyl Alcohol (1% 11,373 dermatitis 1% 24 h or 48 h patch test (Finn chambers) Questionable/irritant in petrolatum) patients (males and reactions (62 patients); females) sensitization reactions (46 patients)84

Benzyl Alcohol (5% 102 patients 5% 24 h patch tests (Finn chambers) 8 subjects with positive in petrolatum) reactions85 Benzyl Alcohol (1%) 2,166 dermatitis 1% 24 or 48 h patch tests Sensitization rate of patients 0.3%86 Benzoic Acid (5% in 7 patients (males and 5% 45 min epicutaneous application (AL- Erythema at physiological saline) females) with birch tests on Scanpor) application sites of all pollen allergy patients (contact urticaria)87

Benzoic Acid (5% in 102 patients 5% 24 h patch tests (Finn chambers) 20 subjects with petrolatum) positive reactions85 Benzoic Acid 1,252 patients with Concentration not stated Contact urticaria test; Delayed 366 patients positive oral mucosal disease hypersensitivity test (urticaria test); 124 patients positive (hypersensitivity test)88

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Table 5. Skin Irritation and Sensitization Studies Doses/Concentrations Test Substance Animals/Subjects Tested Procedure Results Benzoic Acid 417 patients Concentration not stated 48 h occlusive patch tests (Finn 5% of patients with chambers; 10 mm diameter) questionable/irritant reactions; 4.3% with allergic reactions89

Benzoic Acid 465 patients Concentration not stated Not stated Positive reactions (allergenicity) in 2.1% of patients90 Sodium Benzoate 465 patients Concentration not stated Not stated Positive reactions (allergenicity) in 1.9% of patients90 Benzyl Benzoate Patients (no. not 30% Maximization test Non-sensitizer3 (30% in petrolatum) stated)

Benzyl Benzoate 1,000 scabies patients 20% Treatment for scabies No dermatitis after (20% emulsion) treatment91 Benzyl Benzoate 175 patients with and 0.1, 2, and 20% Closed patch test (no GLP) Non-sensitizer3 (0.1, 2, and 20% in without dermatoses vaseline ointment or cream base)

Benzyl Benzoate 73 dermatitis patients 10% Patch test (no GLP) No positive reactions (10% in water) (sensitization)3

Benzyl Benzoate 11 male and female 5% 24 h application Sensitization in 3 of 8 (5%) patients (18+ years) patients58 Benzyl Benzoate (5% 20 male and female 5% 48-h patch test (aluminum-backed strips) Sensitization in 1 of 20 in petrolatum) patients (18+ years) patients58

Benzyl Benzoate (5% 45 dermatitis patients 5% 48-h closed patch test No sensitization58 in petrolatum) (18+ years) Benzyl Benzoate (5% 73 eczematous 5% Patch test (no GLP) 1 positive reaction in petrolatum) dermatitis patients (sensitization)3 Benzyl Benzoate (5% Eczema patients (no. 5% Patch test (no GLP) No positive reactions3 in petrolatum) not stated) Benzyl Benzoate (5% 8 dermatitis patients 5% Patch test (no GLP) 1 positive reaction in petrolatum) (sensitization) 3 Benzyl Benzoate (5% 19 dermatitis patients 5% Patch test (no GLP) No positive reactions3 in petrolatum) Benzyl Benzoate (5% 241 patients 5% Patch test (no GLP) No positive reactions3 in yellow soft paraffin) Benzyl Benzoate (5% 103 patients 5% Patch test (no GLP) 12 positive reactions in vaseline) (sensitization) 3

Benzyl Benzoate (5% 115 patients 5% Patch test (no GLP) 14 positive reactions in vaseline) (sensitization)3

Benzyl Benzoate (5% 465 dermatitis 5% Patch test (no GLP) 7 positive reactions3 in vaseline) patients Benzyl Benzoate 443 patients (383 5% Patch test (no GLP) 2 positive reactions (1 tested and 60 cosmetic dermatitis controls) patient; 1 non-cosmetic dermatitis patient) 3

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Table 5. Skin Irritation and Sensitization Studies Doses/Concentrations Test Substance Animals/Subjects Tested Procedure Results Benzyl Benzoate 271 patients (225 5% Patch test (no GLP) 2 positive reactions (1 (5%) tested and 46 cosmetic dermatitis controls) patient; 1 non-cosmetic dermatitis patient)3

Benzyl Benzoate (5% 73 patients (18+ 5% Not stated Sensitization in 1 in petrolatum) years) patient3 Benzyl Benzoate (5% 5 male and female 5% 30 min closed patch test No sensitization58 in petrolatum) urticaria patients (18+ years) Benzyl Benzoate (5% 12 patients (18+ 5% Patch test No sensitization58 in petrolatum) years) Benzyl Benzoate (5% 3 female allergic 5% Patch tests (Finn chambers) No sensitization58 in petrolatum) dermatitis patients Benzyl Benzoate 50 dermatitis patients 5% Patch tests Sensitization in 1 (5%; vehicle not patient58 reported) Benzyl Benzoate (5% 70 dermatitis 5% 48 h patch tests (Finn chambers) 2.9% sensitization rate patients; 19 eyelid (in 70 patients); no dermatitis patients sensitization (19 patients) 58

Benzyl Benzoate (5% 115 patients with 5% 48 h closed patch tests Sensitization in 14 of in petrolatum, positive reactions to 115 patients58 vaseline, or alcohol) Balsam of Peru

Benzyl Benzoate 225 patients (18+ 5% Patch test Sensitization in 2 of (5%; vehicle not years) 225 patients58 stated) Benzyl Benzoate 658 male and female 5% 48 h patch tests (Finn chambers) Sensitization in 1 (5%; vehicle not patients patient58 stated) Benzyl Benzoate (5% 102 contact 5% 24 h patch tests (Finn chambers) Sensitization in 4 in white petrolatum) dermatitis patients patients58 (18+ years) Benzyl Benzoate (5% 317 eczema patients 5% Patch test No sensitization58 in petrolatum) (18 + years) Benzyl Benzoate (5% 73 patients with 5% Patch test 1 positive reaction in petrolatum) eczematous (allergic contact dermatitis dermatitis)91 Benzyl Benzoate (5% 102 patients 5% 24 h patch tests (Finn chambers) 4 subjects with positive in petrolatum) reactions85 Benzyl Benzoate (2 144 patients (14 2 or 5% Closed patch test (no GLP) 1 positive reaction or 5% in vaseline) controls included) per (sensitization in test concentration cosmetic dermatitis patient)3

Benzyl Benzoate (2 64 dermatitis patients 2 or 5% Closed patch test Sensitization in 1 or 5% in petrolatum) (18+ years) patient (5% concentration); no sensitization (2%)58

Benzyl Benzoate Patients (no. not 2% Patch test (no GLP) Negative reaction in (2%) stated) contact dermatitis patient3 Benzyl Benzoate 198 patients 2% Patch test (no GLP) No positive reactions3 (2%) Benzyl Benzoate (2% 335 patients 2% Patch test (no GLP) No positive reactions3 in petrolatum)

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Table 5. Skin Irritation and Sensitization Studies Doses/Concentrations Test Substance Animals/Subjects Tested Procedure Results Benzyl Benzoate (2% 539 patients (18+ 2% 48 h closed patch test Sensitization in 1 of 32 in yellow paraffin) years; 50 with PLE patients58 photosensitivity dermatitis, 32 with polymorphic light eruption (PLE) and 457 with contact dermatitis)

Benzyl Benzoate (2% 284 patients (18+ 2% Patch test 1% sensitization rate in petrolatum) years) (in 284 patients). No sensitization (in 335 patients) 58

Benzyl Benzoate (2% 198 patients (18+ 2% Patch test No sensitization58 in petrolatum) years) Benzyl Benzoate (2% 241 patients (18+ 2% Finn chamber technique Sensitization in 1 male in yellow soft years) patient58 paraffin) Benzyl Benzoate (2% 140 contact 2% Patch test No sensitization58 in white petrolatum) dermatitis patients (30 to 67 years) Benzyl Benzoate (2% 539 patients 2% Patch test (no GLP) 1 positive reaction in yellow soft (sensitization in patient paraffin) with polymorphic light eruption)3

Benzyl Benzoate (1% 28 patients allergic to 1% Not stated No sensitization58 in yellow petrolatum) perfumes (18+ years)

Benzyl Benzoate 2,003 dermatitis 1% 24 or 48 h patch tests No sensitization86 (1%) patients Benzyl Benzoate 111 male and female 0.1% 24-h to 48-h closed patch test No sensitization58 (0.1% in ethanol or dermatitis patients non-irritative cream (18+ years) base)

Benzyl Benzoate Cosmetic dermatitis Concentration not stated Patch test (no GLP) 1 positive reaction patients (no. not (sensitization) 3 stated) Benzyl Benzoate 15 patients Concentration not stated Patch test (no GLP) delayed in reaction in 1 patient sensitized to Peru balsam3 Benzyl Benzoate 111 patients Concentration not stated Patch test (no GLP) No positive reactions3

Benzyl Benzoate 142 patients Concentration not stated Patch test (no GLP) 10 positive reactions (sensitization in patents sensitive to balsam of Peru) 3

Benzyl Benzoate 155 scabies patients Concentration not stated Treatment with benzyl benzoate Dermatitis in 17 (concentration not stated) patients3 Benzyl Benzoate 460 patients Concentration not stated Patch test (no GLP) 3 positive reactions (sensitization in patients sensitive to cosmetics)3

Benzyl Benzoate 25 patients Concentration not stated Maximization test (no GLP) No positive reactions3

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Table 5. Skin Irritation and Sensitization Studies Doses/Concentrations Test Substance Animals/Subjects Tested Procedure Results Benzyl Benzoate 465 patients Concentration not stated Not stated Positive reactions (allergenicity) in 1.5% of patients90

Table 6. Case Reports Test Substance Subjects Procedure/Results Benzyl Alcohol 30-year-old facial dermatitis patient Patch testing (no test concentration) produced macular erythema92

Benzyl Alcohol 38-year old eczema patient 1% aqueous benzyl alcohol: negative prick test results and positive (++) intradermal injection test results. Negative injection test results in 10 healthy controls93 Benzyl Alcohol 39-year-old female with pruritic erythema of foot 5% benzyl alcohol in petrolatum: weak (+) reaction in patch test and strong positive reaction in repeated open application test94

Benzyl Alcohol 67-year-old male with leg dermatitis 1% benzyl alcohol in petrolatum: 2+ occlusive patch test reaction. 0.9% benzyl alcohol in saline: negative prick test reaction at 0.5 h reading, but marked induration and proximal spread over arm at days 3 to 895

Benzyl Alcohol 53-year-old with stasis dermatitis 1% benzyl alcohol in petrolatum: redness and swelling at 1 h after patch application, wheal 1 day later, and mild urticaria at day 596

Benzyl Alcohol 16-year-old female with possible anaphylactic Benzyl alcohol preparation: negative in prick tests, but positive in reactions after i.m. injection with B12 preparation intradermal tests97 containing 0.9% benzyl alcohol Benzyl Alcohol 57-year-old female with pruritic dermatitis Allergic contact dermatitis (+, ++, or +++ reaction) after patch testing with benzyl alcohol98 Benzyl Alcohol 40-year-old female with dermatitis Positive patch test reaction (+++, allergic contact dermatitis) to 9.5% benzyl alcohol in petrolatum99 Benzyl Alcohol 65-year-old female with eyelid dermatitis Macular erythema after patch testing with benzyl alcohol (concentration not stated)100 Benzyl Alcohol 30-year-old female with eyelid dermatitis Positive (1+) patch test reaction to benzyl alcohol (concentration not stated)92 Benzoic Acid 30-year-old facial dermatitis patient Patch testing (no test concentration) produced 1+ reaction92 Benzoic Acid 46-year-old female with history of erythema and 5% benzoic acid in petrolatum: positive reaction101 itching Sodium Benzoate 64-year-old female with erythema and edema of Positive patch test reaction to 5% sodium benzoate on days 2 and 4; finger negative patch test results at interdigital area at 2 months102

Sodium Benzoate 52-year-old female with hand dermatitis Negative patch test reaction to sodium benzoate (concentration not stated)103 Sodium Benzoate 75-year-old female with history of pruritus Relapse of pruritus after 100 mg oral dose of sodium benzoate104 Benzyl Benzoate 25-year-old male with acute bullous eruption after Negative patch test reaction to 1% benzyl benzoate in petrolatum105 application of 30% benzyl benzoate preparation

Benzyl Benzoate 4 scabies patients (ages not stated) Severe skin irritation; pruritic dermatitis within 12 to 20 h after (unspecified concentration treatment (unspecified concentration or 25% emulsion)58 or 25% emulsion in aqueous solution of triethanolamine stearate)

Benzyl Benzoate Scabies patients (no. not included) Treatment with benzyl benzoate solution (concentration not stated) did not cause dermatitis3 Benzyl Benzoate 4 scabies patients Treatment with benzyl benzoate solution caused dermatitis3

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Table 6. Case Reports Test Substance Subjects Procedure/Results Benzyl Benzoate (5% in 1 patient (18 + years) Reaction read at 48 h and 96 h. No sensitization58 petrolatum Benzyl Benzoate 2% in 23-year old male patient Negative patch test results; no sensitization58 petrolatum)

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Figure 1. Chemical Structures

OH O

O- K+ benzyl alcohol potassium benzoate

O OH benzoic acid O

O benzyl benzoate

O- Na+ sodium benzoate

O- Ca2+

-O

O calcium benzoate

O- Mg2+

-O

O magnesium benzoate

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34. Food and Drug Administration (FDA). Substances for use only as components of coatings. Benzyl alcohol and benzyl benzoate. 21 CFR 175.105. 2011.

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62. Zhengyu, S., Fang, W., and Ying, F. Vehicle used for triamcinolone acetonide is toxic to ocular tissues of the pigmented rabbit. Curr.Eye Res. 2009;34:(9):769‐776.

63. Sugihara, N. Shimomichi K. and Furuno K. Cytotoxicity of food preservatives in cultured rat hepatocytes loaded with linoleic acid. Toxicology. 1997;120:(1):29‐36.

CIR Panel Book Page 41 Distributed for Comment Only - Do Not Quote or Cite 64. Placzek, M. Krosta I. Gaube S. Eberlein‐KöNIG B. and Przybilla B. Evaluation of phototoxic properties of antimicrobials used in topical preparations by a photohaemolysis test. Acta Derm.Venereol. 2005;85:(1):13‐16.

65. Placzek, M. Frömel W. Eberlein B. Gilbertz K. P. and Przybilla B. Evaluation of phototoxic properties of fragrances. Acta.Derm.Venereol. 2007;87:(4):312‐316.

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67. Mytton, O. T. McGready R. Lee S. J. Roberts C. H. Ashley E. A. Carrara V. I. Thwai K. L. Jay M. P. Wiangambun T. Singhasivanon P. and Nosten F. Safety of benzyl benzoate lotion and permethrin in pregnancy: a retrospective matched cohort study. BJOG. 2007;114:(5):582‐587.

68. Ashby, J. Lefevre P. A. Odum J. Tinwell H. Kennedy S. J. Beresford N. and Sumpter J. P. Failure to confirm estrogenic activity for benzoic acid and clofibrate: Implications for lists of endocrine‐disrupting agents. Regul.Toxicol.Pharmacol. 1997;26:96‐101.

69. Hashimoto, Y. Kawaguchi M. Miyazaki K. and Nakamura M. Estrogenic activity of tissue cojnditioners in vitro. Dent.Mater. 2003;19:(4):341‐346.

70. Charles, A. K. and Darbre P. D. Oestrogenic activity of benzyl salicylate, benzyl benzoate and butylphenylmethylpropional (Lilial) in MCF7 human breast cancer cells in vitro. J.Appl.Toxicol. 2009;422‐434.

71. Demir, E., lu, S., and Kaya, B. Assessment of genotoxic effects of benzyl derivatives by the comet assay. Food Chem.Toxicol.%2010., May. 48(5):1239‐42.:(5:1239‐42):Food.

72. Nesslany, F. and Marzin D. A micromethod for the in vitro micronucleus assay. Mutagenesis. 1999;14:(4):403‐410.

73. Mpountoukas, P., Vantarakis, A., Sivridis, E., and Lialiaris, T. Cytogenetic study in cultured human lymphocytes treated with three commonly used preservatives. Food Chem.Toxicol.%2008., Jul. 46(7):2390‐3.:(7:2390‐3):Food.

74. Ishizaki, M. and Ueno S. The DNA‐damaging activity of natural and synthetic food additives (V). J.Food Hyg.Soc.Jpn. 1989;30:(5):447‐451.

75. Salih, F. M. Risk assessment of combined photogenotoxic effects of sunlight and food additives. Sci.total Environ. 2006;362:(1‐3):68‐73.

76. Stefanidou, M., Alevizopoulos, G., and Spiliopoulou, C. DNA content of Tetrahymena pyriformis as a biomarker for different toxic agents. Chemosphere.%2008., Dec. 74(1):178‐80.:(1:178‐ 80):Chemosphere.

77. O'Neil, M. J. The Merck Index Online. Whitehouse Station, NJ: Merck & Co., Inc., 2010.

78. Scientific and Technical Information Network. Beilstein record on calcium benzoate. https://stnweb.cas.org. 2010. Date Accessed 12‐7‐2010.

79. National Library of Medicine. ChemIDplus record on magnesium benzoate. http://www.nlm.nih.gov. 2010. Date Accessed 12‐7‐2010.

CIR Panel Book Page 42 Distributed for Comment Only - Do Not Quote or Cite 80. Barratt, M. D. Quantitative structure‐activity relationships for skin irritation and corrosivity of neutral and electrophilic organic chemicals. Toxicology In Vitro. 1996;10:(3):247‐256.

81. Basketter, D. A. Chamberlain M. Griffiths H. A. Rowson M. Whittle E. and York M. The classification of skin irritants by human patch. test Food and Chemical Toxicology. 1997;35:(8):845‐852.

82. Coverly, J. Peters L. Whittle E. and Basketter D. A. Susceptibility of skin stinging, non‐immunologic contact urticaria abd acute skin irritation; is there a relationship? Contact Dermatitis. 1998;38:(2):90‐95.

83. Marriott, M. Holmes J. Peters L. Cooper K. Rowson M. and Basketter D. A. The complex problem of sensitive skin. Contact Dermatitis. 2005;53:(2):93‐99.

84. Schnuch, A. Geier J. Uter W. and Frosch P. J. Patch testing with preservatives, antimicrobials and industrial biocides. Results from a multicentre study. British Journal of Dermatology. 1998;138:(3):467‐476.

85. Hausen, B. M. Contact allergy to Balsam of Peru. II. Patch test results in 102 patients. Am J.Contact Dermat. 2001;12:(2):93‐102.

86. Schnuch, A. Uter W. Geier J. Lessman H. and Frosch P. J. Sensitization to 26 fragrances to be labelled according to current European regulation. Contact Dermatitis. 2007;57:(1):1‐10.

87. Wallengren, J. and Larsson B. Nitric oxide participates in prick test and irritatnt patch test reactions in human skin. Arch.Dermatol.Res. 2001;293:(3):121‐125.

88. Wray, D. Rees S. R. Gibson J. and Forsyth A. The role of allergy in oral mucosal diseases. Q J Med. 2000;93:(8):507‐511.

89. Kanerva, L. Rantanen T. Aalto‐Korte K. et al. A multicenter study of patch test reactions with dental screening series. American Journal of Contact Dermatitis. 2001;12:(2):83‐87.

90. Meynadier, J.‐M. Meynadier J. Colmas A. Castelain P. Y. Ducombs G. Chabeau G. Lacroix M. Martin P. and Ngangu Z. Allergy to preservatives. Ann.Dermatol.Venereol. 1982;109:(12):1017‐1023.

91. Meneghini, C. L. Vena G. A. and Angelini G. Contact dermatitis to scabicides. Contact Dermatitis. 1982;8:(4):285‐286.

92. Jacob, S. E. and Stechschulte, S. Eyelid dermatitis associated with balsam of Peru constituents: benzoic acid and benzyl alcohol. Contact Dermatitis.%2008., Feb. 58(2):111‐2.:(2:111‐2):Contact.

93. Verecken, P. Birringer C. Knitelius A. C. Herbaut D. and Germaux M. A. Sensitization to benzyl alcohol: a possible cause of "corticosteroid allergy". Contact Dermatitis. 1998;38:(2):106‐106.

94. Podda, M. Grundmann‐Kollmann M. Kaufmann R. and Boehncke W. H. Allergic contact dermatitis from benzyl alcohol during topical antimycotic treatment. Contact Dermatitis . 1999;41:(5):302‐303.

95. Shaw, D. W. Allergic contact dermatitis to benzyl alcohol in a hearing aid impression material. Am J.Contact Dermat. 1999;10:(4):228‐232.

96. Guin, J. D. and Goodman J. Contact urticaria from benzyl alcohol presenting as intolerance to saline soaks. Contact Dermatitis. 2001;45:(3):182‐183.

CIR Panel Book Page 43 Distributed for Comment Only - Do Not Quote or Cite 97. Turvey, S. E. Cronin B. Arnold A. D. Twarog F. J. and Dioun A. F. Adverse reactions to vitamin B12 injections dud to benzyl alcohol sensitivity. Allergy. 2004;59:(9):1023‐1024.

98. Curry, E. J. and Warshaw E. M. Benzyl alcohol allergy: Importance of patch testing with personal products. Dermatitis. 2005;16:(4):203‐208.

99. Carrascosa, J.‐M. Domingo H. Soria X. and Ferrándiz C. Allergic contact dermatitis due to benzyl alcohol in a hair dye. Contact Dermatitis. 2006;55:(2):124‐125.

100. Jacob, S. E. Ciclosporin ophthalmic emulsion ‐ a novel therapy for benzyl alcohol‐associated eyelid dermatitis. Contact Dermatitis.%2008., Mar. 58(3):169‐70.:(3:169‐70):Contact.

101. Sestini, S. Mori M. and Francalanci S. Allergic contact dermatitis from benzyl alcohol in multiple medicaments. Contact Dermatitis. 2004;50:(5):316‐317.

102. Vilaplana, J. and Romaguera C. Fixed drug eruptiion from sodium benzoate. Contact Dermatitis. 2003;49:(6):290‐291.

103. Sutton, T. and Nixon R. Allergic contact dermatitis to sodium benzoate chloroacetamide in a sorbolene lotion. Australas.J.Dermatol. 2006;47:(3):209‐210.

104. Asero, R. Sodium benzoate‐induced pruritus. Allergy. 2006;61:(10):1240‐1241.

105. Stransky, L. Vasileva S. and Mateev G. Contact bullous pehphigoid? Contact Dermatitis. 1996;35:(3):182‐ 182.

CIR Panel Book Page 44 Data

Distributed for Comment Only - Do Not Quote or Cite

PersonalCare ‘ProductsCouncil Committedto Safety, Quality& Innovation Memorandum

TO: F. Alan Andersen, Ph.D. Director - COSMETIC iNGREDIENT REVIEW (CW)

FROM: John Bailey, Ph.D. Industry Liaison to the CW Expert Panel

DATE: February 25, 2010

SUBJECT: Inhalation study of Benzyl Alcohol and Benzoic Acid

WIL Research Laboratories, LLC. 2010. A 4-week inhalation toxicity study of aerosolized benzyl alcohol and benzoic acid in Sprague-Dawley rats. WIL-703002.

1101 17th Street, N.W., Suite 300 Washington, D.C. 20036-4702 202.331.1770 202.331.1969 (fax) www.personalcarecouncil.org

CIR Panel Book Page 45 Distributed for Comment Only - Do Not Quote or Cite

FINAL REPORT

Volume 1 of 4 (Text, Figures 1-2, Tables 1-36, and Appendix A - Tables A1-A8)

STUDY TITLE A 4-WEEK INHALATION TOXICITY STUDY OF AEROSOLIZED BENZYL ALCOHOL AND BENZOIC ACID IN SPRAGUE-DAWLEY RATS

STUDY NUMBER WIL-703002

DATA REQUIREMENT OECD Guideline, Section 412

STUDY DIRECTOR Jason M. Roper, PhD

STUDY INITIATION DATE 13 March 2009

STUDY COMPLETION DATE 8 February 2010

PERFORMING LABORATORY WIL Research Laboratories, LLC 1407 George Road Ashland, OH 44805-8946

SPONSOR The Personal Care Products Council 1101 17th Street NW Washington D.C. 20036

Page 1 of 990

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WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

COMPLIANCESTATEMENT

This study, designated WIL-703002, was conducted in compliance with the United States Environmental Protection Agency (EPA) Good Laboratory Practice Standards (40 CFR Part 792), 16 October 1989; the Organisation for Economic Cooperation and Development (OECD) Principles of Good Laboratory Practice [C(97) 186/Final], 26 November 1997; the standard operating procedures of WIL Research Laboratories, LLC; and the protocol as approved by the Sponsor.

The protocol was designed and the study was conducted in general accordance with the following guidelines: Organisation for Economic Cooperation and Development (OECD): Guidelines for Testing Chemicals, Section 4 12.

d F Jasom. Roper, PhD Date Senior Toxicologist, Inhalation Toxicology Study Director

1 Linda Loretz, PhD, &T Date Sponsor Representative

ApplicantBubmitter Date

Page 2 of 990

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WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

TABLE OF CONTENTS

VOLUME 1 Page

Compliance Statement...... 2 Table Of Contents...... 3 Index Of Figures ...... 6 Index Of Tables...... 7 Index Of Appendices...... 10 1. Summary...... 13 1.1. Objective...... 13 1.2. Study Design...... 13 1.3. Results...... 14 1.4. Conclusions...... 14 2. Introduction...... 15 2.1. General Study Information ...... 15 2.2. Key Study Dates ...... 16 2.3. Principal Investigators And Test Sites...... 16 3. Study Design...... 17 4. Experimental Procedures - Materials And Methods...... 18 4.1. Test Substances And Vehicle...... 18 4.1.1. Test Substance 1 Identification...... 18 4.1.2. Test Substance 2 Identification...... 18 4.1.3. Vehicle Identification...... 19 4.2. Inhalation Exposure Methods ...... 19 4.3. Test Atmosphere Generation Methods ...... 20 4.3.1. Benzyl Alcohol ...... 20

4.3.2. Benzoic Acid...... 22 4.4. Exposure Concentrations ...... 23 4.4.1. Actual Exposure Concentrations - Control And Benzoic Acid ...... 23 4.4.2. Analyzed Exposure Concentrations - Benzyl Alcohol ...... 23

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WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

VOLUME 1 (continued) Page

4.4.3. Particle Size Determination ...... 24 4.4.4. Real Time Monitoring - Benzoic Acid ...... 25 4.5. Test Substance Characterization For Stability And Purity ...... 25 4.6. Test System...... 25 4.7. Organization Of Test Groups, Exposure Levels And Treatment Regimen ...... 25 4.8. Animal Receipt And Acclimation/Pretest Period ...... 26 4.9. Animal Housing...... 27 4.10. Diet, Drinking Water And Maintenance...... 27 4.11. Environmental Conditions ...... 28 4.12. Assignment Of Animals To Treatment Groups...... 28 5. Parameters Evaluated ...... 30 5.1. Survival...... 30 5.2. Clinical Observations...... 30 5.3. Body Weights...... 30 5.4. Food Consumption...... 30 5.5. Clinical Pathology...... 31 5.5.1. Hematology And Coagulation ...... 31 5.5.2. Serum Chemistry ...... 32 5.6. Ophthalmic Examinations...... 32 5.7. Anatomic Pathology...... 32 5.7.1. Macroscopic Examination ...... 32 5.7.2. Organ Weights ...... 34 5.7.3. Slide Preparation And Microscopic Examination...... 34 5.8. Statistical Methods...... 34

5.9. Data Retention ...... 35 6. Results ...... 36 6.1. Characterization Of Exposure Atmospheres...... 36 6.1.1. Nominal Exposure Concentrations ...... 36

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WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

VOLUME 1 (continued) Page

6.1.2. Exposure Concentrations ...... 37 6.1.3. Particle Size Determination ...... 37 6.2. Analytical Chemistry ...... 38 6.3. Survival...... 38 6.4. Clinical Observations...... 39 6.5. Body Weights...... 39 6.6. Food Consumption...... 39 6.7. Clinical Pathology...... 40 6.7.1. Hematology And Coagulation ...... 40 6.7.2. Serum Chemistry ...... 40 6.8. Ophthalmic Examinations...... 40 6.9. Anatomic Pathology...... 41 6.9.1. Macroscopic Examination ...... 41 6.9.2. Organ Weights ...... 41 6.9.3. Microscopic Examination ...... 41 7. Conclusions...... 42 8. Key Study Personnel And Report Submission...... 43 9. Quality Assurance Unit Statement...... 44 9.1. Phases Inspected ...... 44 9.2. Approval ...... 47 10. References...... 48 11. Deviations From The Protocol...... 49

Page 5 of 990

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WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

INDEX OF FIGURES

VOLUME 1 (continued) Page

1. Summary Of Body Weights [G] (Males)...... 52

2. Summary Of Body Weights [G] (Females) ...... 53

Page 6 of 990

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WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

INDEX OF TABLES

VOLUME 1 (continued) Page

1. Summary Of Survival And Disposition (Males) ...... 55

2. Summary Of Survival And Disposition (Females)...... 57

3. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (Detailed Physical Examinations/Dispositions - Males)...... 59

4. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (Detailed Physical Examinations/Dispositions - Females)...... 60

5. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (Prior To Exposure - Males)...... 61

6. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (Prior To Exposure - Females) ...... 62

7. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (Mid-Exposure - Males) ...... 63

8. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (Mid-Exposure - Females)...... 64

9. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (1 Hour Post-Exposure - Males)...... 65

10. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (1 Hour Post-Exposure - Females)...... 66

11. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (Daily Observations - Non-Exposure Days - Males)...... 67

12. Summary Of Clinical Findings: Total Occurrence/No. Of Animals (Daily Observations - Non-Exposure Days - Females) ...... 68

13. Summary Of Body Weights [G] (Males)...... 69

14. Summary Of Body Weights [G] (Females) ...... 72

15. Summary Of Body Weight Changes [G] (Males) ...... 75

Page 7 of 990

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WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

VOLUME 1 (continued) Page

16. Summary Of Body Weight Changes [G] (Females)...... 77

17. Summary Of Cumulative Body Weight Changes [G] (Males)...... 79

18. Summary Of Cumulative Body Weight Changes [G] (Females)...... 80

19. Summary Of Food Consumption [G/Animal/Day] (Males)...... 81

20. Summary Of Food Consumption [G/Animal/Day] (Females) ...... 83

21. Summary Of Hematology Values (Males) ...... 85

22. Summary Of Hematology Values (Females)...... 92

23. Summary Of Serum Chemistry Values (Males)...... 99

24. Summary Of Serum Chemistry Values (Females) ...... 104

25. Summary Of Special Chemistry Values (Males)...... 109

26. Summary Of Special Chemistry Values (Females) ...... 110

27. Summary Of Ophthalmic Examination Findings (Week -2 Pretest Examination - Males)...... 111

28. Summary Of Ophthalmic Examination Findings (Week -2 Pretest Examination - Females) ...... 112

29. Summary Of Ophthalmic Examination Findings (Week 3 Examination - Males) ...... 113

30. Summary Of Ophthalmic Examination Findings (Week 3 Examination - Females)...... 114

31. Summary Of Macroscopic Findings (Males)...... 115

32. Summary Of Macroscopic Findings (Females) ...... 117

33. Summary Of Organ Weights And Relative Organ Weights (Males)...... 118

34. Summary Of Organ Weights And Relative Organ Weights (Females) ...... 126

35. Summary Of Microscopic Findings (Males) ...... 134

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WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

VOLUME 1 (continued) Page

36. Summary Of Microscopic Findings (Females)...... 150

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WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

INDEX OF APPENDICES

VOLUME 1 (continued) Page

A. Individual Animal Data...... 163

A1. Individual Survival And Disposition (Males)...... 164

A2. Individual Survival And Disposition (Females)...... 167

A3. Individual Clinical Observations (Detailed Physical Examinations/Dispositions - Males) ...... 170

A4. Individual Clinical Observations (Detailed Physical Examinations/Dispositions - Females)...... 185

A5. Individual Clinical Observations (Prior To Exposure - Males)...... 200

A6. Individual Clinical Observations (Prior To Exposure - Females)...... 251

A7. Individual Clinical Observations (Mid-Exposure - Males)...... 302

A8. Individual Clinical Observations (Mid-Exposure - Females) ...... 303

VOLUME 2

A9. Individual Clinical Observations (1 Hour Post-Exposure - Males) ...... 306

A10. Individual Clinical Observations (1 Hour Post-Exposure - Females)...... 357

A11. Individual Clinical Observations (Daily Observations - Non-Exposure Days - Males) ...... 408

A12. Individual Clinical Observations (Daily Observations - Non-Exposure Days - Females)...... 423

A13. Individual Body Weights [G] (Males)...... 438

A14. Individual Body Weights [G] (Females) ...... 452

A15. Individual Body Weight Changes [G] (Males) ...... 466

A16. Individual Body Weight Changes [G] (Females)...... 473

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VOLUME 2 (continued) Page

A17. Individual Cumulative Body Weight Changes [G] (Males)...... 480

A18. Individual Cumulative Body Weight Changes [G] (Females)...... 487

A19. Individual Food Consumption [G/Animal/Day] (Males)...... 494

A20. Individual Food Consumption [G/Animal/Day] (Females) ...... 501

A21. Individual Hematology Values (Males)...... 508

A22. Individual Hematology Values (Females)...... 536

A23. Individual Serum Chemistry Values (Males)...... 564

A24. Individual Serum Chemistry Values (Females) ...... 578

A25. Individual Special Chemistry Values (Males)...... 592

A26. Individual Special Chemistry Values (Females) ...... 599

A27. Individual Ophthalmic Examination Findings (Week -2 Pretest Examination - Males)...... 606

A28. Individual Ophthalmic Examination Findings (Week -2 Pretest Examination - Females)...... 609

A29. Individual Ophthalmic Examination Findings (Week 3 Examination - Males)...... 612

A30. Individual Ophthalmic Examination Findings (Week 3 Examination - Females) ...... 615

VOLUME 3

A31. Individual Macroscopic And Microscopic Findings (Males) ...... 620

A32. Individual Macroscopic And Microscopic Findings (Females)...... 719

A33. Individual Organ Weights And Final Body Weights [G] (Males)...... 814

A34. Individual Organ Weights And Final Body Weights [G] (Females) ...... 821

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VOLUME 3 (continued) Page

A35. Individual Organ Weights Relative To Final Body Weights [G/100 G] (Males)...... 828

A36. Individual Organ Weights Relative To Final Body Weights [G/100 G] (Females)...... 835

A37. Individual Organ Weights Relative To Brain Weights [G/100 G] (Males)...... 842

A38. Individual Organ Weights Relative To Brain Weights [G/100 G] (Females)...... 849

B. Certificates Of Analysis (Sponsor-Provided Data)...... 856

C. Exposure Atmosphere Generation Report (WIL Research Laboratories, LLC) ...... 862

D. Test Substance Purity And Stability Report (WIL Research Laboratories, LLC) ...... 898

E. Pretest Clinical Observations ...... 921

F. Animal Room Environmental Conditions...... 924

VOLUME 4

G. Clinical Pathology Methods, Procedures, And References ...... 939

H. Pathology Report (WIL Research Laboratories, LLC)...... 945

I. Ophthalmic Examinations (Brian C. Gilger, DVM, MS, DACVO) ...... 957

J. Study Protocol ...... 959

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1. SUMMARY

1.1. OBJECTIVE The objective of this study was to evaluate the potential toxic effects of aerosolized benzyl alcohol when administered to rats by nose-only inhalation on a 5-day per week basis for a period of 4 weeks (minimum of 20 exposures). In addition to a control group exposed to filtered air and treatment groups exposed to 4 concentrations of benzyl alcohol, 2 additional groups were exposed to benzoic acid.

1.2. STUDY DESIGN Aerosolized benzyl alcohol was administered via nose-only inhalation for 6 hours per day on a 5-day/week basis for a period of 4 weeks (a minimum of 20 exposures/animal) to 4 groups (Groups 2-5) of Crl:CD(SD) rats. Target exposure concentrations were 30, 100, 300, and 1000 mg/m3 for Groups 2, 3, 4, and 5, respectively. Aerosolized benzoic acid was administered via nose-only inhalation exposure on a 5-day/week basis for a period of 4 weeks (a minimum of 20 exposures/animal) to 2 groups (Groups 6 and 7) of Crl:CD(SD) rats. Target exposure concentrations were 2.5 and 12.5 mg/m3 for Groups 6 and 7, respectively. A concurrent control group (Group 1) was exposed to filtered air on a comparable regimen. Each group consisted of 10 animals/sex. All animals were euthanized on the day following the last exposure. The first day and week of exposure were designated as study day 0 and study week 0, respectively.

All animals were observed twice daily for mortality and moribundity. Clinical examinations were conducted prior to exposure, at the approximate mid-point of each exposure (beginning on study day 2; third exposure), 0 to 1 hour following exposure, and once daily on non-exposure days; detailed physical examinations were performed at least weekly. Individual body weights and food consumption were recorded approximately weekly. Ophthalmic examinations were conducted once during the pretest period (study week -2) and near the end of the exposure period (study week 3). Blood samples for clinical pathology evaluations (hematology, coagulation, and serum chemistry) were collected from all animals at the scheduled necropsy (study week 3). Complete

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necropsies were conducted on all animals, and selected organs were weighed at the

scheduled necropsy. Selected tissues were examined microscopically from all surviving

animals in the control (Group 1) and high dose benzyl alcohol and benzoic acid groups (Groups 5 and 7); gross lesions were examined from all animals in all groups.

1.3. RESULTS With the exception of the low-concentration benzyl alcohol group (Group 2), which had a mean exposure atmosphere concentration of 136% of the target concentration, all exposure atmosphere mean concentrations were within 96.7% to 107.2% of the respective target concentrations. There were no test substance-related mortalities, clinical or ophthalmic observations, or effects on body weights, food consumption, clinical pathology parameters, and organ weights. There were no test substance-related macroscopic or microscopic findings.

1.4. CONCLUSIONS Based on the results of this study, 6-hour nose-only inhalation exposure to benzyl alcohol at mean exposure concentrations of 41, 102, 290 and 1,072 mg/m3 or exposure to benzoic acid at target exposure concentrations of 2.5 and 12.6 mg/m3 for 4 weeks (minimum of 20 exposures) was well-tolerated with no effects at any exposure level. The no-observed-effect-level (NOEL) and no-observed-adverse-effect level (NOAEL) were considered to be 1,072 mg/m3 for benzyl alcohol and 12.6 mg/m3 for benzoic acid.

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2. INTRODUCTION

The objective of this study was to evaluate the potential toxic effects of aerosolized benzyl alcohol when administered to rats by nose-only inhalation on a 5-day per week basis for a period of 4 weeks (minimum of 20 exposures). In addition to a control group exposed to filtered air and treatment groups exposed to 4 concentrations of benzyl alcohol, 2 additional groups were exposed to benzoic acid.

2.1. GENERAL STUDY INFORMATION This report presents the data from “A 4-Week Inhalation Toxicity Study of Aerosolized Benzyl Alcohol and Benzoic Acid in Sprague-Dawley Rats.” Due to software spacing constraints, the study title appears as “4-Week Inhalation Study of Benzyl Alcohol & Benzoic Acid in Rats” on the report tables. The first day and week of exposure were designated as study day 0 and study week 0, respectively.

The animals were euthanized and necropsied on the day following their last exposure (study day 26, 27, or 28). Although study day 28 was technically the first day of study week 4, all necropsy days are referred to as study week 3 throughout the report text and tables.

The following computer protocols were used for data collection during the study:

Computer Protocol Type of Data Collected WIL-703002M...... Main study data (males) WIL-703002F...... Main study data (females) WIL-703002P...... Pretest data (males) WIL-703002Q ...... Pretest data (females)

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2.2. KEY STUDY DATES

Date(s) Event(s) 31 March 2009...... Experimental starting date (animal receipt) 13 April 2009...... Assignment to study groups 22 April 2009...... Experimental start date (initiation of dose administration; study day 0) 18, 19, 20 May 2009...... Scheduled necropsy (study week 3) 9 July 2009...... Experimental termination (completion) date (last histopathological examination)

2.3. PRINCIPAL INVESTIGATORS AND TEST SITES

Test Site and Principal John Boyce, DVM, PhD, DACVP, DACLAM Investigator for Microscopic Senior Pathologist Evaluation Biotechnics, a WIL Research Laboratories LLC subsidiary 310 Millstone Drive Hillsborough, NC 27278

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3. STUDY DESIGN

Acclimation/pretest period

Assignment of animals to study groups

10 animals/sex/group treated for a minimum of 20 exposures (6 hours per day, 5 days per week for 4 weeks)

Detailed physical examinations performed weekly; clinical observations recorded daily; body weights and food intake recorded approximately weekly

Ophthalmic examinations performed prior to necropsy (study week 3)

Clinical pathology evaluations conducted prior to necropsy (study week 3)

Necropsies performed on all animals; selected organs weighed; selected tissues examined microscopically

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4. EXPERIMENTAL PROCEDURES -MATERIALS AND METHODS

4.1. TEST SUBSTANCES AND VEHICLE 4.1.1. TEST SUBSTANCE 1 IDENTIFICATION Test substance 1, benzyl alcohol, was received from Emerald Kalama Chemical, LLC, Kalama, WA, as follows:

Physical Date of Identification Description Receipt

Benzyl Alcohol Clear colorless liquid 7 October 2008 Lot no. KABCN89307 Exp. date: 27 September 2010 CAS no. 100-51-6 [WIL log no. 8077A]

Benzyl Alcohol Clear colorless liquid 17 April 2009 Lot no. KABCN94301 Exp. date: 1 April 2011 CAS no. 100-51-6 [WIL log no. 8077B]

Documentation regarding the purity and stability of test substance 1 is on file with the Sponsor and WIL Research Laboratories, LLC. Certificates of Analysis were provided by the Sponsor and are presented in Appendix B. The purity was 99.9%. Benzyl alcohol was stored at room temperature and was considered stable under this condition. Reserve samples were collected and stored in the Archives of WIL Research Laboratories, LLC.

4.1.2. TEST SUBSTANCE 2 IDENTIFICATION Test substance 2, benzoic acid, was received from Emerald Kalama Chemical, LLC, Kalama, WA, on 7 October 2008 as follows:

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Physical Identification Description

Benzoic Acid White flakes Lot no. KABZT87212 Re-test date: 29 July 2009 CAS no. 65-85-0 [WIL log no. 8078A]

Documentation regarding the purity and stability of test substance 2 is on file with the Sponsor and WIL Research Laboratories, LLC. A Certificate of Analysis was provided by the Sponsor and is presented in Appendix B. The purity was 99.6%. Benzoic acid was stored at room temperature and was considered stable under this condition. A reserve sample was collected and stored in the Archives of WIL Research Laboratories, LLC.

4.1.3. VEHICLE IDENTIFICATION The vehicle used for exposure of the control group (Group 1) was filtered air.

4.2. INHALATION EXPOSURE METHODS Exposures were conducted using seven 7.9-L conventional nose-only exposure systems (designed and fabricated by WIL Research Laboratories, LLC) with synthetic rubber grommets in exposure ports to engage animal holding tubes. One exposure system was dedicated for each group for the duration of the study.

The following table presents the exposure system used for each group:

Group Number: 1 2A 3A 4A 5A 6B 7B Exposure System: 1 2 3 4 5 6 7 Target Exposure Concentration (mg/m3): 0 30 100 300 1000 2.5 12.5

A = Benzyl Alcohol B = Benzoic Acid

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Air supplied to the nose-only systems was provided from a dry compressed air source.

All test atmosphere exhaust passed through a Solberg filter prior to entering the facility

exhaust system, which consisted of charcoal- and HEPA-filtration.

Animals were housed in normal animal colony rooms during non-exposure hours. For the exposure each day, the animals were placed into nose-only exposure restraint tubes in the colony rooms, transported to the exposure room, placed on the nose-only system, exposed for the requisite duration, and returned to their home cages in the animal colony rooms. Animals were held in restraint tubes for approximately 20-56 minutes prior to animal exposure. Food and water were withheld during the animal exposures. Exposure methods and conditions are presented in detail in Appendix C.

4.3. TEST ATMOSPHERE GENERATION METHODS For Exposure System 1, humidified and dilution air were added to the exposure system. Humidified air was added using a regulator and a rotameter-type flowmeter. Dry compressed air passed through a muffler-type bubbler submerged in a 2-L Erlenmeyer flask filled with deionized water to produce humidified air. Dilution air was added using a Coilhose Pneumatics regulator and a Gilmont rotameter-type flowmeter. Exposure methods and conditions are detailed in Appendix C. The following is a list of parameters used during animal exposures for the control exposure system:

Dilution Airflow Humidified Airflow Total Airflow Exposure System Rate (LPM) Rate (LPM) Rate (LPM) 1 20.1 29.5 49.6

4.3.1. BENZYL ALCOHOL A vapor/aerosol atmosphere was generated using a system which operated as follows. A

syringe pump and appropriate size syringe were used to deliver test substance to an atomizer. The atomizer was comprised of a no. 2850 fluid cap and a no. 64 air cap. Using a Coilhose Pneumatics regulator, compressed air was supplied to the air port of the atomizer at a known, constant pressure to effect the atomization of the test substance.

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The resulting vapor/aerosol atmosphere passed through a liquid trap prior to entering the exposure system. For Exposure Systems 2, 3, and 4, a siphon was placed in-line prior to

the exposure system to reduce the concentration as needed. The siphon was controlled using a rotameter-type flowmeter. The approximate test substance delivery rates and syringe sizes are summarized in the following table:

Exposure Syringe Size Test Substance System (mL) Flow Rate (g/hour) 2 3 0.1 to 1.1 3 5 0.7 to 0.9 4 10 4.2 5 100 19 to 35

On 29 April 2009, the Exposure System 2 siphon rotameter-type flowmeter was replaced with a Dwyer rotameter-type flowmeter. Dilution and humidified air was added to the atmosphere prior to entering the exposure system. Dilution air was added using a Coilhose Pneumatics regulator and metered using a Gilmont rotameter-type flowmeter. Humidified air was added using a Coilhose Pneumatics regulator and metered using a Dwyer rotameter-type flowmeter. Dry compressed air passed through a muffler-type bubbler submerged in a 2-L Erlenmeyer flask filled with DI water to produce humidified air. On 30 April 2009, the siphon rotameter-type flowmeter for Exposure System 2 was moved after the humidified air. Exposure methods and conditions are detailed in Appendix C. The approximate airflows used for exposures are summarized in the following table:

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Siphon Atomizer Dilution Humidified Total Exposure Airflow Rate Airflow Rate Airflow Rate Airflow Rate Airflow Rate System (LPM) (LPM) (LPM) (LPM) (LPM) 2 0 to 9 4 to 19 0.5 to 18 16 to 40 28 to 64 3 0 to 0.1 16 0 to 12 20 to 39 39 to 59 4 1 to 3 18 6 to 20 13 to 26 40 to 57 5 NA 19 8 to 36 0 to 21 37 to 60 NA = Not applicable

4.3.2. BENZOIC ACID A dust aerosol atmosphere of the test substance was generated using a system which operated as follows: The test substance was delivered using a Wright Dust Feeder (WDF) and controller, which fed test substance at a constant rate to a jet mill micronizer operating as a dispersion device. The WDF was equipped with a 5-cm3 stainless steel cup. Prior to packing the test substance into the WDF cups, the test substance was sieved using a no. 25 standard sieve. The sieved test substance was packed in the cup using manual compression. Using a Coilhose Pneumatics regulator, dry compressed air was supplied to the WDF to deliver test substance to the jet mill. The WDF supply air did not add to the total airflow to the nose-only system. Using a Coilhose Pneumatics regulator, dry compressed air was supplied to 2 Ashcroft gauges with needle valves to control the airflow to the micronizing and inlet ports of the jet mill where the test substance was milled to the desired particle size. The resulting aerosol from the jet mill was delivered to the exposure system through ¾-inch ID anti-static tubing. A 28.3-L settling chamber was placed in-line between the jet mill and the exposure system to remove larger particles. A tee was placed between the settling box and exposure system to provide humidified air to the exposure system. Using a Coilhose Pneumatics regulator and a

Dwyer rotameter-type flowmeter, dry compressed air passed through a muffler-type bubbler submerged in a 2-L Erlenmeyer flask filled with DI water to produce humidified air. Exposure methods and conditions are detailed in Appendix C. The following is a list

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of approximate parameters used during animal exposures for the test substance exposure

systems:

Jet Mill Inlet Micronizing Humidified Total WDF WDF Exposure Airflow Airflow Rate Airflow Airflow Rate Setting Airflow System Rate (LPM) (LPM) Rate (LPM) (LPM) (Indicated) (PSI) 6 16.4 11.6 32.3 60.3 0.05-0.13 13 7 15.8 12.6 32.0 60.4 0.20-0.45 14

4.4. EXPOSURE CONCENTRATIONS 4.4.1. ACTUAL EXPOSURE CONCENTRATIONS -CONTROL AND BENZOIC ACID Actual exposure concentrations were determined using standard gravimetric methods for the control and benzoic acid groups. Samples were collected on pre-weighed, 25-mm glass-fiber filters held in an open-faced filter holder positioned in the animal breathing zone of the nose-only exposure system. Following sample collection, the filters were re-weighed and the concentration calculated as the filter weight difference divided by the sample volume. Samples were collected at approximately 90-minute intervals for the test substance exposure systems and 1 sample was collected weekly for the control exposure system. Exposure methods and conditions are detailed in Appendix C. The following table summarizes the approximate sampling conditions:

Exposure Sample Flow Rate Sample Time System (L/minute) (minutes) 1 0.8 85 6 0.9 to 1.0 85 7 1.0 30 to 85

4.4.2. ANALYZED EXPOSURE CONCENTRATIONS -BENZYL ALCOHOL Concentrations of benzyl alcohol in the exposure systems were determined at approximately 90-minute intervals using a gas chromatograph (GC). Additional samples were collected as needed. Samples were collected from the approximate animal-

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breathing zone of the exposure system using a series of 2 impingers containing isopropyl

alcohol (IPA) as a trapping liquid. Test atmosphere samples were pulled through the impinger sampling train. This sampling method was used to collect the test atmosphere vapor, as well as aerosol (if present). Following sample collection, the liquid in the impingers was pooled, mixed using a laboratory vortex, and manually injected into a calibrated, validated GC. Impinger samples for the presence of benzyl alcohol were not collected or analyzed for the control group. Exposure of the control group in a separate room from the benzyl alcohol exposures eliminated the possibility of exposure of the control group to benzyl alcohol. The following table lists the sampling parameters used during animal exposure.

Exposure IPA Volume In Total IPA Sampling Time System Each Impinger (mL) Volume (mL) (Minutes)

2 10 20 10 3 10 20 10 4 10 20 10 to 20 5 20 40 5

4.4.3. PARTICLE SIZE DETERMINATION Aerosol particle size determinations were conducted for Exposure Systems 5, 6, and 7 using a 7-stage cascade impactor. Pre-weighed, 25-mm glass-fiber filters were used as the collection substrates. One sample was collected weekly at approximately 1.8 LPM for 2, 300, and 60 minutes for Exposure Systems 5, 6, and 7, respectively. The filters were re-weighed and the particle size was calculated based on the impactor stage cut-offs. The aerosol size was expressed as the mass median aerodynamic diameter (MMAD) and

the geometric standard deviation (GSD). Exposure methods and conditions are detailed in Appendix C.

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4.4.4. REAL TIME MONITORING -BENZOIC ACID

The benzoic acid atmospheres were monitored in real-time using a HPM-1000 light scattering aerosol photometers. The monitors were connected to an Omega RD6110 Chart Recorder to provide a continuous display of concentration. Real-time aerosol monitoring of the test atmosphere was intended to provide an index of exposure concentrations for system adjustments and was not used to define actual concentrations. Exposure methods and conditions are detailed in Appendix C.

4.5. TEST SUBSTANCE CHARACTERIZATION FOR STABILITY AND PURITY A sample of each test substance was collected prior to the initial use for animal exposures for analysis of concentration (time-zero value). Following the final exposure, a sample of each test substance was collected from the container used for the final exposure for purity determination. The end of use percent concentration was compared to the time zero (pre-use) value for determination of stability. All analyses were conducted by the Analytical Chemistry Department, WIL Research Laboratories, LLC using a validated high performance liquid chromatography method using ultraviolet absorbance detection. Details about the methodology and results of these analyses are presented in Appendix D, and the results are summarized in Section 6.2.

4.6. TEST SYSTEM Crl:CD(SD) rats from Charles River Laboratories, Inc., Raleigh, NC, were used as the test system for this study. This species and strain of animal is generally recognized as appropriate for inhalation studies. The Sprague Dawley rat was selected because it is a widely used strain for which significant historical control data are available. The animals were approximately 9 weeks old at the initiation of dose administration.

4.7. ORGANIZATION OF TEST GROUPS,EXPOSURE LEVELS AND TREATMENT REGIMEN Filtered air (control group), benzyl alcohol or benzoic acid atmospheres were administered as 6-hour, nose-only inhalation exposures on a 5-day per week basis for a

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period of 4 weeks (20-22 exposures for each animal) through the day prior to the

scheduled necropsy. The first day of exposure was defined as study day 0.

The following table presents the study group assignment:

Group Test Target Concentration Level Number of Animals Number Substance (mg/m3) Males Females

1 Filtered Air 0 10 10 2 Benzyl Alcohol 30 10 10 3 Benzyl Alcohol 100 10 10 4 Benzyl Alcohol 300 10 10 5 Benzyl Alcohol 1000 10 10 6 Benzoic Acid 2.5 10 10 7 Benzoic Acid 12.5 10 10

All benzyl alcohol and benzoic acid exposure concentrations were selected by the Sponsor based on known toxicity information. For benzyl alcohol, a mixed aerosol and vapor exposure atmosphere may have been present. Therefore, the exposure concentration was reported as total test substance (i.e., aerosol plus vapor).

The route of administration was inhalation exposure because this is a potential route of human exposure. Nose-only exposure methods were used to reduce the potential for dermal exposure or oral exposure resulting from grooming. The number of animals selected for this study was the minimum needed to yield statistically valid and scientifically meaningful data and to meet the objectives of the study.

4.8. ANIMAL RECEIPT AND ACCLIMATION/PRETEST PERIOD Eighty-four male and 84 female Crl:CD(SD) rats were received in good health on 31 March 2009, from Charles River Laboratories, Inc., Raleigh, NC. The animals were

approximately 41 days old at receipt. Each animal was examined by a qualified technician on the day of receipt and weighed 3 days later. Each animal was uniquely identified with a subcutaneous microchip (BMDS system) in the scapular area. All animals were housed for a 22-day acclimation/pretest period. During this period, each

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animal was observed twice daily for mortality and changes in general appearance or

behavior.

Pretest data collection began on 3 April 2009. Individual body weights and food consumption were recorded and detailed physical examinations were performed periodically during the pretest period. Ophthalmic examination data were also recorded for pretest animals prior to the initiation of exposure. Pretest clinical observations are presented in Appendix E.

To screen animals for poor tolerance of restraint, and to limit potential effects on respiration of the novel environment/conditions of restraint, the animals were acclimated to restraint in nose-only exposure tubes by increasing the restraint time over the acclimation period. On the first, second, third, fourth, and fifth days of restraint acclimation, the animals were acclimated for 1, 2, 3, 4, and 6 hours, respectively. Animals were then acclimated for 6 hours every other day until the initiation of exposure. Following the restraint period, each animal was observed for clinical signs of injury or stress.

4.9. ANIMAL HOUSING Upon arrival, all animals were housed individually in clean, stainless steel, wire-mesh cages suspended above cage-board. Animals were maintained in accordance with the Guide for the Care and Use of Laboratory Animals (National Research Council, 1996). The animal facilities at WIL Research Laboratories, LLC are accredited by the Association for Assessment and Accreditation of Laboratory Animal Care International (AAALAC International). Nylabones® were provided to all animals throughout the study (except during exposure or exposure acclimation periods) for environmental enrichment

and to aid in maintaining the animals’ oral health, and were sanitized weekly.

4.10. DIET,DRINKING WATER AND MAINTENANCE The basal diet used in this study, PMI Nutrition International, LLC, Certified Rodent LabDiet® 5002 (meal), is a certified feed with appropriate analyses performed by the

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manufacturer and provided to WIL Research Laboratories, LLC. Reverse

osmosis-treated (on-site) drinking water, delivered by an automatic watering system, and

the basal diet were provided ad libitum throughout the study, except during exposure or exposure acclimation periods, or any scheduled period of fasting. Municipal water supplying the facility was analyzed for contaminants according to the standard operating procedures. The results of the diet and water analyses are maintained at WIL Research Laboratories, LLC. No contaminants were present in animal feed or water at concentrations sufficient to interfere with the objectives of this study.

4.11. ENVIRONMENTAL CONDITIONS All animals were housed throughout the acclimation period and during the study in environmentally controlled rooms. The room temperature and humidity controls were set to maintain environmental conditions of 71  5F (22  3C) and 50  20% relative humidity. Room temperature and relative humidity were controlled and monitored using the Metasys® DDC Electronic Environmental control system. These data were recorded approximately hourly and are summarized in Appendix F. Actual mean daily temperature ranged from 70.4F to 71.2F (21.3C to 21.8C) and mean daily relative humidity ranged from 37.0% to 52.0% during the study. Fluorescent lighting provided illumination for a 12-hour light (0600 hours to 1800 hours)/12-hour dark photoperiod. Air handling units were set to provide a minimum of 10 fresh air changes per hour.

4.12. ASSIGNMENT OF ANIMALS TO TREATMENT GROUPS On 13 April 2009 (9 days prior to the initiation of exposure), all available rats were weighed and examined in detail for physical abnormalities. Based on the review of all appropriate pretest data, which were collected using the WIL Toxicology Data Management System (WTDMS™), animals judged suitable for assignment to the study

were selected for use in a computerized randomization procedure. A printout containing the animal numbers, corresponding body weights and individual group assignments was generated based on body weight stratification in a block design. The animals were then arranged into groups according to the printout. Individual body weights at randomization

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were within  20% of the mean for each sex. One Group 2 (30 mg/m3 benzyl alcohol)

female was found dead in the restraint tube following the exposure conducted on study day 1 (second exposure). This animal was replaced with a suitable remaining pretest group female prior to exposure on study day 2, and the replacement animal received 20 total exposures.

Each group (Groups 1-7) consisted of 10 males and 10 females. Individual body weights ranged from 255 g to 352 g for males and from 182 g to 234 g for females at the initiation of exposure.

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5. PARAMETERS EVA LUATED

5.1. SURVIVAL All animals were observed twice daily for mortality and moribundity, once in the morning and once in the afternoon, except on the day of scheduled necropsy.

5.2. CLINICAL OBSERVATIONS Clinical examinations were performed prior to exposure, at the approximate mid-point during the exposure period (beginning on study day 2; third exposure), 0 to 1 hour post-exposure (designated as 1 hour post-exposure for report presentation purposes), and once daily on non-exposure days. The absence or presence of findings was recorded for individual animals at the scheduled intervals. Detailed physical examinations were conducted on all animals at least once during the pretest period, at the time of randomization and group assignment, and weekly during the exposure phase (including prior to scheduled necropsy).

5.3. BODY WEIGHTS Individual body weights were recorded approximately weekly during the pretest period, at the time of randomization and group assignment, weekly throughout the study (including the first day of exposure), and a non-fasted body weight was collected on the day before the first scheduled day of necropsy. Mean body weights and mean body weight changes were calculated for the corresponding intervals. Final body weights (fasted) were recorded for all animals on the days of the scheduled necropsies.

5.4. FOOD CONSUMPTION Individual food consumption was recorded weekly beginning at least 1 week prior to exposure. Food intake was calculated as g/animal/day for the corresponding body weight

intervals. When food consumption could not be measured for a given interval (due to spillage, weighing error, obvious erroneous value, etc.), the appropriate interval was footnoted as "NA" (Not Applicable) on the individual tables.

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5.5. CLINICAL PATHOLOGY

Blood samples for clinical pathology evaluations (hematology, coagulation, and serum chemistry) were collected from all animals on the days of the scheduled necropsies (study week 3). The animals were fasted overnight prior to blood collection. Blood was collected for hematology and serum chemistry evaluation via the retro-orbital sinus of animals anesthetized by inhalation of isoflurane. Blood was collected for coagulation parameters at the time of euthanasia via the vena cava of animals anesthetized by inhalation of isoflurane. Blood was collected into tubes containing potassium EDTA (hematology), sodium citrate (coagulation) or no anticoagulant (serum chemistry). Clinical pathology methods, procedures, and references are presented in Appendix G. Interpretation of the clinical pathology data was performed by John Boyce, DVM, PhD, DACVP, DACLAM (Appendix H). The following parameters were evaluated:

5.5.1. HEMATOLOGY AND COAGULATION Total leukocyte count (White Cells) Red cell distribution width Erythrocyte count (Red Cells) (Red CellWidth) Hemoglobin Hemoglobin distribution width Hematocrit (HGB Width) Mean corpuscular volume (MCV) Differential leukocyte count - Mean corpuscular hemoglobin Percent and absolute (MCH) -Neutrophil Mean corpuscular hemoglobin -Lymphocyte concentration (MCHC) -Monocyte Platelet count (Platelet) -Eosinophil Prothrombin time (ProTime) -Basophil Activated partial thromboplastin time -Large unstained cell (APTT) Platelet estimatea Reticulocyte count Red cell morphology Percent (Reticulocyte) (RBC Morphology)a Absolute (Retic Absolute)

( ) - Designates tabular abbreviation a - Presented on individual tables if a manual differential was performed, and the manual data were accepted and reported instead of the automated differential data

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5.5.2. SERUM CHEMISTRY

Albumin Gamma glutamyltransferase Total protein (GlutamylTransfer) Globulin [by calculation] Glucose Albumin/globulin ratio (A/G Ratio) Total cholesterol (Cholesterol) [by calculation] Calcium Total bilirubin (Total Bili) Chloride Urea nitrogen Phosphorus Creatinine Potassium Alkaline phosphatase Sodium (AlkalinePhos’tse) Triglycerides (Triglyceride) Alanine aminotransferase Sorbitol dehydrogenase (Alanine Transfer) (Sorbitol ‘Genase)a Aspartate aminotransferase (AspartatTransfer)

( ) - Designates tabular abbreviation a - Presented on special chemistry tables

5.6. OPHTHALMIC EXAMINATIONS Ocular examinations were conducted on all animals during pretest (9 April 2009; study week -2) and prior to the scheduled necropsy (16 May 2009; study week 3). All ocular examinations were conducted using an indirect ophthalmoscope and slit lamp biomicroscope preceded by pupillary dilation with an appropriate mydriatic agent. Pretest examinations were performed by David A. Wilkie, DVM, MS, DACVO, and study week 3 examinations were performed by Brian C. Gilger, DVM, MS, DACVO (Appendix I).

5.7. ANATOMIC PATHOLOGY 5.7.1. MACROSCOPIC EXAMINATION A complete necropsy was conducted on all animals. Animals were euthanized by

isoflurane inhalation and exsanguination. The necropsies included an examination of the external surface, all orifices, and the cranial, thoracic, abdominal, and pelvic cavities, including viscera. Clinical findings that were confirmed macroscopically were designated CEO (correlates with externally observed) on the individual macroscopic data

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tables. The following tissues and organs were collected and placed in

10% neutral-buffered formalin (except as noted):

Adrenals (2) Lymph nodes Aorta Axillary (2) Bone with marrow Mediastinal and bronchial (if visible) Femur with joint Mesenteric and mandibular Sternum Mammary gland (females only) d Bone marrow smear (from femur) a Nasal cavity with turbinates e Brain Ovaries (2) with oviducts f Cerebrum (2 levels) Pancreas Cerebellum with pons/medulla Peripheral nerve (sciatic) Cervix Peyer’s patches Epididymides (2) b Pharynx Eyes with optic nerves (2) c Pituitary Exorbital lacrimal glands (2) Prostate Gastrointestinal tract Salivary glands [mandibular (2)] Esophagus Seminal vesicles (2) Stomach Skeletal muscle (rectus femoris) Duodenum Skin Jejunum Spinal cord (cervical, thoracic, lumbar) Ileum Spleen Cecum Testes (2) b Colon Thymus Rectum Thyroid gland with parathyroids (2) f Harderian glands (2) Trachea Heart Urinary bladder Kidneys (2) Uterus Larynx Vagina Liver (sections of 2 lobes) All gross lesions Lungs (including bronchi, fixed by constant pressure inflation with fixative)

a -Not placed in formalin; not examined. b - Fixed in Bouin’s solution c - Fixed in Davidson’s solution d - A corresponding section of skin was collected from the same anatomic area for males. e

- Following collection of the appropriate protocol-specified tissues, the entire head was removed and preserved. Following decalcification, 6 cross-sections of the nasal cavities were prepared for microscopic examination in accordance with the method described by Morgan (1991). f -Examined if in plane of section and in all cases when a gross lesion of the organ was present.

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5.7.2. ORGAN WEIGHTS

The following organs were weighed from all animals at the scheduled necropsy:

Adrenals Lungs (prior to inflation with fixative) Brain Ovaries and oviducts Epididymides Spleen Heart Testes Kidneys Thymus Liver Uterus with cervix

Paired organs were weighed together. Organ-to-final-body-weight and organ-to-brain- weight ratios were calculated.

5.7.3. SLIDE PREPARATION AND MICROSCOPIC EXAMINATION After fixation, protocol-specified tissues were trimmed according to standard operating procedures and the protocol. Trimmed tissues were processed into paraffin blocks, sectioned at 4 to 8 microns, mounted on glass microscope slides, and stained with hematoxylin and eosin.

Microscopic examination was performed on all tissues listed in Section 5.7.1. from all animals in the control, 1000 mg/m3 benzyl alcohol, and 12.5 mg/m3 benzoic acid groups (Groups 1, 5, and 7, respectively) at the scheduled necropsy. Gross lesions were examined from all animals. Missing tissues were identified as not found at necropsy, lost at necropsy, lost during processing or other designations as appropriate. Tissues may appear on the report tables as not examined due to the tissue not being in the plane of section, not present at trimming, etc. Microscopic examination was performed by John Boyce, DVM, PhD, DACVP, DACLAM, WIL Research Laboratories, LLC (Appendix H).

5.8. STATISTICAL METHODS All statistical tests were performed using appropriate computing devices or programs. Analyses were conducted using two-tailed tests (except as noted otherwise) for minimum significance levels of 1% and 5%, comparing each test substance-treated group to the

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control group by sex. Each mean was presented with the standard deviation (S.D.),

standard error (S.E.) and the number of animals (N) used to calculate the mean.

Statistical analyses were not conducted if the number of animals was 2 or less. Due to the different rounding conventions inherent in the types of software used, the means and standard deviations on the summary and individual tables may differ by ±1 in the last significant figure.

Body weights, body weight changes, food consumption, clinical pathology values (except gamma glutamyltransferase), and organ weight data were subjected to a parametric one-way analysis of variance (ANOVA) (Snedecor and Cochran, 1980) to determine intergroup differences. If the ANOVA revealed statistically significant (p<0.05) intergroup variance, Dunnett's test (Dunnett, 1964) was used to compare the test substance-treated groups to the control group. Gamma glutamyltransferase values under range were assigned a value of 0.1 (half the lower limit of quantitation) for statistical analysis and reporting. Gamma glutamyltransferase data were subjected to the Kruskal-Wallis nonparametric ANOVA test (Kruskal and Wallis, 1952) to determine intergroup differences. If the ANOVA revealed significance (p<0.05), Dunn’s test (Dunn, 1964) was used to compare the test substance-treated groups to the control group.

5.9. DATA RETENTION The Sponsor has title to all documentation records, raw data, specimens, slides or other work product generated during the performance of the study. All work product generated by WIL Research Laboratories, LLC, including raw paper data and specimens, are retained in the Archives at WIL Research Laboratories, LLC, as specified in the study protocol. Data generated by the Sponsor, will be archived by the Sponsor or the Sponsor's designee.

Reserve samples of the test substances, pertinent electronic storage media, and the original final report are retained in the Archives at WIL Research Laboratories, LLC, in compliance with regulatory requirements.

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6. RESULTS

6.1. CHARACTERIZATION OF EXPOSURE ATMOSPHERES Analyses of Exposure Concentrations: Appendix C

6.1.1. NOMINAL EXPOSURE CONCENTRATIONS The following table summarizes the overall mean nominal concentrations for each test substance exposure system:

Exposure System: 2A 3A 4A 5A 6B 7B Mean Concentration (mg/m3): 94 288 1284 8293 31 145 Standard Deviation: 44.4 75.5 148.1 1290.7 4.9 31.0 N: 22 22 22 22 22 22 A= Benzyl Alcohol, B= Benzoic Acid

Nominal exposure concentrations were calculated by determining the pre-exposure and post-exposure mass of the test substance for a given exposure, and dividing the difference by the total airflow rate through the exposure system. A significant portion of the generated test substance was removed using siphons or liquid traps for the benzyl alcohol groups, and using settling chambers for the benzoic acid groups, prior to the atmosphere reaching the animal exposure chambers. Because a portion of the atmospheres were removed prior to the animal exposure chambers, high mean nominal concentrations were observed compared with the actual measured exposure concentrations. For reporting purposes, the actual measured concentrations at the chamber, as described in Section 6.1.2. are considered to be the test substance concentrations to which the test system was exposed.

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6.1.2. EXPOSURE CONCENTRATIONS

The following table summarizes the overall mean exposure concentrations for each exposure system:

Exposure System: 1 2A 3A 4A 5A 6B 7B

Target Concentration (mg/m3): 0 30 100 300 1000 2.5 12.5 Actual Concentration (mg/m3): 0 41 102 290 1072 2.5 12.6 Percent Difference from Target (%): 0 37 2.0 -3.3 7.2 0.0 0.8 Standard Deviation (mg/m3): 0.0 12.9 16.4 24.6 163.0 0.13 0.91 Concentration (ppm) NA 9.6 23.8 67.4 248.8 NA NA Standard Deviation (ppm): NA 3.00 3.81 5.72 37.86 NA NA N: 4 22 22 22 22 22 22 A= Benzyl Alcohol, B= Benzoic Acid

6.1.3. PARTICLE SIZE DETERMINATION Aerosol particle size was determined for the 1000 mg/m3 benzyl alcohol exposure system (5) and both benzoic acid exposure systems (6 and 7). The aerosol particle size was not determined for the control group (filtered air) or for the 30, 100, or 300 mg/m3 benzyl alcohol exposure systems. In a previous method development and validation study (Kirkpatrick, Draft) it was determined that benzyl alcohol atmospheres at concentrations less than 100 mg/m3 did not contain aerosol particles. Additionally, due to the volatility of benzyl alcohol, and the sampling time required to obtain a measurable amount of test substance on impactor substrates, it was not possible to accurately assess the particle size of the 300 mg/m3 benzyl alcohol exposure atmosphere, although it was determined previously that aerosol particles were present at this concentration

(Kirkpatrick, Draft). It is not anticipated that the MMAD of aerosol particles in the 300 mg/m3 benzyl alcohol exposure atmosphere would have exceeded that observed for the 1000 mg/m3 benzyl alcohol exposure atmosphere. The following table summarizes the overall mean aerosol particle size for Exposure Systems 5, 6, and 7:

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Exposure System: 5A 6B 7B

Mean MMAD (Microns): 3.3 2.1 2.5

Mean GSD: 2.39 3.66 3.03 N: 4 4 4 A= Benzyl Alcohol, B= Benzoic Acid

6.2. ANALYTICAL CHEMISTRY Test Substance Purity And Stability Report: Appendix D

Benzyl alcohol and benzoic acid samples were collected and analyzed prior to use for animal exposures (101% pure and 99.8% pure, respectively) and after use for animal exposures 42 days later (99.5% pure and 101% pure, respectively). Comparison of these purity data indicated that the test substances were stable for the duration of use on the study.

Observed purity values greater than 100% are accounted for by acceptable variability in the analytical method used to assess test substance purity.

6.3. SURVIVAL Summary Data: Table 1, Table 2, Table 3, Table 4 Individual Data: Table A1, Table A2, Table A3, Table A4

There were no test substance-related deaths. One Group 2 female (no. 8307; 30 mg/m3 benzyl alcohol) was found dead in the restraint tube following the exposure conducted on study day 1 (second exposure). Based on the lack of clinical signs of toxicity in any test substance-treated group, this death was considered related to poor tolerance to restraint and was not considered test substance-related.

All other animals survived to the scheduled necropsy.

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6.4. CLINICAL OBSERVATIONS

Summary Data: Table 3, Table 4, Table 5, Table 6, Table 7, Table 8, Table 9, Table 10, Table 11, Table 12 Individual Data: Table A3, Table A4, Table A5, Table A6, Table A7, Table A8, Table A9, Table A10, Table A11, Table A12

There were no test substance-related clinical observations. All clinical findings in the test substance-treated groups were noted with similar incidence in the control group, were limited to one or two animals, were not noted in an exposure concentration-related manner, and/or were common findings for laboratory rats of this age and strain.

6.5. BODY WEIGHTS Summary Data: Table 13, Table 14, Table 15, Table 16, Table 17, Table 18; Figure 1, Figure 2 Individual Data: Table A13, Table A14, Table A15, Table A16, Table A17, Table A18

Body weights were unaffected by test substance administration. There were no statistically significant differences when the control and test substance-treated groups were compared.

6.6. FOOD CONSUMPTION Summary Data: Table 19, Table 20 Individual Data: Table A19, Table A20

Food consumption was unaffected by test substance administration. There were no statistically significant differences when the control and test substance-treated groups were compared.

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6.7. CLINICAL PATHOLOGY

6.7.1. HEMATOLOGY AND COAGULATION Summary Data: Table 21, Table 22 Individual Data: Table A21, Table A22 Pathology Report: Appendix H

Hematology parameters were unaffected by test substance administration. There were no statistically significant differences when the control and test substance-treated groups were compared, and all group mean values were within the WIL historical control reference ranges (version 2.8).

6.7.2. SERUM CHEMISTRY Summary Data: Table 23, Table 24, Table 25, Table 26 Individual Data: Table A23, Table A24, Table A25, Table A26 Pathology Report: Appendix H

Serum chemistry parameters were unaffected by test substance administration. There were no statistically significant differences when the control and test substance-treated groups were compared, and all group mean values were within the WIL historical control reference ranges (version 2.8).

6.8. OPHTHALMIC EXAMINATIONS Summary Data: Table 27, Table 28, Table 29, Table 30 Individual Data: Table A27, Table A28, Table A29, Table A30 Ophthalmic Examination Report: Appendix I

No ophthalmic lesions indicative of toxicity were observed in any of the test

substance-treated groups. All findings observed were typical in prevalence and appearance for laboratory rats of this age and strain.

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6.9. ANATOMIC PATHOLOGY

6.9.1. MACROSCOPIC EXAMINATION Summary Data: Table 31, Table 32 Individual Data: Table A31, Table A32 Pathology Report: Appendix H

There were no test substance-related macroscopic findings at the scheduled necropsy. All macroscopic findings noted were considered to be spontaneous and/or incidental in nature and unrelated to test substance administration.

6.9.2. ORGAN WEIGHTS Summary Data: Table 33, Table 34 Individual Data: Table A33, Table A34, Table A35, Table A36, Table A37, Table A38 Pathology Report: Appendix H

There were no statistically significant test substance-related alterations in final body weight or organ weights. However, non-statistically significant lower mean final body weights in the males exposed to either 300 or 1000 mg/m3 benzyl alcohol resulted in nonadverse, statistically significant, higher mean epididymide weights relative to final body weights but not relative to brain weights. All group mean values were within the WIL historical control reference range (means ± 2 standard deviations; version 2.8).

6.9.3. MICROSCOPIC EXAMINATION Summary Data: Table 35, Table 36 Individual Data: Table A31, Table A32 Pathology Report: Appendix H

There were no test substance-related microscopic findings. All findings observed were consistent with normal background lesions in clinically normal rats of the age and strain used on this study and were considered spontaneous and/or incidental in nature and unrelated to test substance administration.

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7. CONCLUSIONS

Based on the results of this study, 6-hour nose-only inhalation exposure to benzyl alcohol at mean exposure concentrations of 41, 102, 290 and 1,072 mg/m3 or exposure to benzoic acid at target exposure concentrations of 2.5 and 12.6 mg/m3 for 4 weeks (minimum of 20 exposures) was well-tolerated with no effects at any exposure level. The no-observed-effect-level (NOEL) and no-observed-adverse-effect level (NOAEL) were considered to be 1,072 mg/m3 for benzyl alcohol and 12.6 mg/m3 for benzoic acid.

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8. KEYSTUDY PERSONNELAND REPORT SUBMISSION

Report Submitted By:

Jason Id Roper, PhD Date Senior Toxicologist, Inhalation Toxicology Study Director

Report Prepared By: &gd& Jana f,. flrnelrod, BS Study Analyst

Director, Inhala6on Toxicology 3

Jf,] 7/ 1,iI ( [ (( - d~&&~~ $’ c .,< Charlene A. Weygandt, BS ’ 1 Date Lead Analyst and Scientific Advisor, Study Analysis and Reports

Study Personnel:

Susan C. Haley, BS Manager, Clinical Pathology Sally A. Keets, AS Senior Operations Manager, Vivarium Carol A. Kopp, BS, LAT Manager, Gross Pathology and Developmental Toxicology Laboratory Teresa D. Morris, BS Senior Operations Manager, Toxicology Theresa M. Rafeld, CPhT Group Manager, Formulations Laboratory Michael A. Safron, AS, Manager, Histology HT (ASCP)CM Robert A. Wally, BS, RAC Manager, Reporting and Regulatory Technical Services

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9. QUALITY ASSURANCE UNIT STATEMENT

9.1. PHASES INSPECTED Date(s) Date(s) Findings Findings Date(s) of Reported to Reported to Inspection(s) Phase Inspected Study Director Management Auditor(s)

22-Apr-2009 Test Article Exposure 22-Apr-2009 19-May-2009 S.Power

07-May-2009 Post-Dose Observations 08-May-2009 23-Jun-2009 R.Rohr

18-May-2009 Blood Collection and Analysis 18-May-2009 23-Jun-2009 J.Dieterly

18-May-2009 Necropsy 18-May-2009 23-Jun-2009 J.Dieterly

28-May-2009 Trimming of Tissues 28-May-2009 23-Jun-2009 R.Rohr

10-Jul-2009, 14-Jul-2009 Study Records (A-1) 14-Jul-2009 20-Aug-2009 C.Heifner

22-Jul-2009, 23-Jul-2009 Study Records (I-1) 24-Jul-2009 20-Aug-2009 S.Power

23-Jul-2009 Study Records (I-2) 24-Jul-2009 20-Aug-2009 S.Power

24-Jul-2009 Study Records (I-3) 24-Jul-2009 20-Aug-2009 S.Power

24-Jul-2009, 27-Jul-2009 Study Records (C-1) 27-Jul-2009 20-Aug-2009 S.Power

27-Jul-2009 Study Records (N-1) 27-Jul-2009 20-Aug-2009 S.Power

27-Jul-2009 Study Records (N-2) 27-Jul-2009 20-Aug-2009 S.Power

27-Jul-2009, Study Records (H-1) 28-Jul-2009 20-Aug-2009 S.Power 28-Jul-2009

28-Jul-2009 Study Records (P-1) 28-Jul-2009 20-Aug-2009 S.Power

28-Jul-2009, 29-Jul-2009 Study Records (Ex-1) 05-Aug-2009 23-Sep-2009 S.Power

29-Jul-2009, 30-Jul-2009, 03-Aug-2009, 04-Aug-2009 Study Records (Ex-2) 05-Aug-2009 23-Sep-2009 S.Power

31-Jul-2009, 03-Aug-2009, 04-Aug-2009 Study Records (Ex-3) 05-Aug-2009 23-Sep-2009 S.Power

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Date(s) Date(s) Findings Findings Date(s) of Reported to Reported to Inspection(s) Phase Inspected Study Director Management Auditor(s)

05-Aug-2009 Draft Report (Pathology Appendix) 06-Aug-2009 23-Sep-2009 S.Power

06-Aug-2009 Draft Report (Inhalation Appendix) 06-Aug-2009 23-Sep-2009 S.Power

06-Aug-2009, 07-Aug-2009, 10-Aug-2009 Draft Report (without Inhalation and Pathology Appendices) 10-Aug-2009 23-Sep-2009 S.Power

10-Aug-2009 Study Records (Ex-3, Supplemental) 10-Aug-2009 23-Sep-2009 S.Power

18-Aug-2009, 19-Aug-2009 Draft Report (Analytical Chemistry) 19-Aug-2009 23-Sep-2009 C.Heifner

This study was inspected in accordance with the U.S. EPA Good Laboratory Practice Standards (40 CFR Part 792), the OECD Principles of Good Laboratory Practice, the standard operating procedures of WIL Research Laboratories, LLC, and the Sponsor's protocol and protocol amendments, with the following exception. The data located in Appendix B (Certificates Of Analysis) were the responsibility of the Sponsor. Quality Assurance findings, derived from the inspections during the conduct of the study and from the inspections of the raw data and draft report, are documented and have been reported to the study director. Review of the protocol and protocol amendments (if applicable) as well as a yearly internal facility inspection are conducted by the WIL Quality Assurance Unit. A status report is submitted to management monthly.

This report accurately reflects the data generated during the study. The methods and procedures used in the study were those specified in the protocol, its amendments, and the standard operating procedures of WIL Research Laboratories, LLC.

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The raw data, the retention samples, and the final report will be stored in the Archives at

WIL Research Laboratories, LLC, or another location specified by the Sponsor.

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9.2. APPROVAL

This study was inspected according to the criteria discussed in Section 9.1.

Report Audited By: ffi&& : Carrie A. Heifner, BA Associate Compliance Specialist

Steven P. Power Compliance Specialist

Report Released By:

@A-Heather L. Johnson, BS, RQAP-GLP Manager, Quality Assurance

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10. REFERENCES

Dunn, O.J. Multiple comparisons using rank sums. Technometrics 1964, 6(3), 241- 252.

Dunnett, C.W. New tables for multiple comparisons with a control. Biometrics 1964, 20, 482-491.

Kirkpatrick, D.T. An Inhalation Exposure Method Development and Validation Study of Aerosolized Benzyl Alcohol, Benzaldehyde and Benzoic Acid (Study No. WIL-703001). WIL Research Laboratories, LLC, Ashland, OH, Draft.

Kruskal, W.H.; Wallis, W.A. Use of ranks in one-criterion variance analysis. Journal of the American Statistical Association 1952, 47, 583-621.

Morgan, K. T., Approaches to the identification and recording of nasal lesions in toxicology studies, Toxicologic Pathology, 19(4) (Part 1) 1991, 337-351.

National Research Council. Guide for the Care and Use of Laboratory Animals, Institute of Laboratory Animal Resources, Commission on Life Sciences; National Academy Press: Washington, DC, 1996.

Snedecor, G.W.; Cochran, W.G. One-Way Classifications; Analysis of Variance. In Statistical Methods, 7th ed.; The Iowa State University Press: Ames, IA, 1980; pp 215-237.

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11. DEVIATIONS FROM THE PROTOCOL

This study was conducted in accordance with the protocol and protocol amendments, except for the following.

 Protocol Section 2.5 states that the consulting ophthalmologist will be David A. Wilkie, DVM, MS, DACVO. Due to a scheduling conflict, the end-of-study ophthalmic examinations were conducted by Brian C. Gilger, DVM, MS, DACVO.  Protocol Section 7.1 states that animals will be acclimated to restraint in nose-only exposure restraint tubes for 6 hours on the fifth day of restraint acclimation. Females were kept in the restraints for up to 6 minutes outside the allowable time range on the fifth day of acclimation.  Protocol Sections 7.4.1 and 7.4.3 state that animals will be submitted for necropsy on the day following the twentieth exposure. Due to a 3-day stagger start in the necropsy schedule, some animals were euthanized on the days following the twenty-first or twenty-second exposures.  Protocol Section 7.5 states that animal exposure airflow rate, temperature, and humidity for all benzyl alcohol exposure systems will be documented at approximately 60-minute intervals. On 30 April 2009, the above-mentioned parameters were not documented between 2 hours, 30 minutes and 4 hours, 30 minutes into exposure.  Protocol Section 7.5 states that for each day’s exposure, animals will be held in restraint tubes for a minimum of 25-30 minutes prior to the initiation of exposure. On 22 May 2009, animals in the benzyl alcohol-treated groups were acclimated for only 20 minutes prior to the initiation of exposure.  Protocol Section 7.5 states that the average relative humidity of the exposure atmosphere will be 30% to 70%. For Exposure Systems 2 and 3, relative humidities were as low as 28% and 23%, respectively. For Exposure System 5, the relative humidity was as high as 91%.  Protocol Section 7.7.2 states that impinger samples will be collected with the impinger placed in an ice bath. During method development, it was determined that

there was no significant benefit to sampling on ice.  Protocol Section 7.7.2 states that samples for the test substance exposure systems will be collected at approximately 90-minute intervals. For the benzyl alcohol exposure systems, additional samples were collected between the scheduled samples as needed and ranged from a minimum of 57 minutes to a maximum of 136 minutes.

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 Protocol Section 7.7.3 states that aerosol particle size determination will be conducted once per week for all benzyl alcohol groups. Particle size was only determined for the 1000 mg/m3 exposure atmosphere. During a previous method development and validation study (Kirkpatrick, Draft), a calibrated HPM-1000 was used as an aerosol monitoring device and detected no aerosol in the 30 or 100 mg/m3 exposure atmospheres. Aerosol particles were detected in the 300 mg/m3 exposure system at a concentration of ≤ 79 mg/m3. Due to the volatility of benzyl alcohol, and the sampling time required to obtain a measureable amount of test substance on the impactor substrates, it was not possible to measure the particle size of the 300 mg/m3 benzyl alcohol exposure atmosphere.  Protocol Section 8.3 states that a clinical observation will be conducted prior to exposure and 0 to 1 hour following exposure. The following animals did not receive an observation at the appropriate interval: Animal Clinical Observation Number Sex Group Study Day Not Received 8303 Female 3 1 Prior to exposure 8232 Male 5 3 Prior to exposure 8279 Female 7 3 Prior to exposure 8301 Female 6 4 Prior to exposure 8249 Male 5 4 0 to 1 hour post-exposure 8282 Female 4 9 0 to 1 hour post-exposure 8246 Male 3 16 0 to 1 hour post-exposure

 Protocol Section 8.11.2 states that lungs will be weighed. At the time of necropsy on 18 May 2009, the lung weight for female no. 8292 (Group 5) was not recorded.  Protocol Section 8.11.3 states that microscopic examination will be performed on tissues for all animals that die spontaneously. Female no. 8307 (Group 2) was replaced on study day 1 after an apparent accidental death. The tissues for this animal were not processed to slide or examined microscopically. Necropsy data will be maintained in the study records, but not presented in the final report.

These deviations did not negatively impact the quality or integrity of the data nor the outcome of the study.

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The Personal Care Products Council aa

FIGURES 1-2

Page 51 of 990

CIR Panel Book Page 96

PROJECT NO.:WIL-703002 FIGURE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHTS (G) - MALES DATA PRESENTED AS MEAN ± SE

500

450 Distributed 400

350 for

CIR Comment

Panel 300 Page 52 of 990 Book 250 Only - Page Do 200 Not 97 Quote

BODYWEIGHT (GRAMS) 150 or

100 Cite

0 -4 -3 -2 -1 0 1 2 3 4 STUDY WEEK

0 MG/M3 30 MG/M3 BENZYL 100 MG/M3 BENZYL 300 MG/M3 BENZYL 1000MG/M3 BENZYL 2.5MG/M3 BENZOIC 12.5MG/M3BENZOIC

BW v1.00 07/21/2009

PROJECT NO.:WIL-703002 FIGURE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHTS (G) - FEMALES DATA PRESENTED AS MEAN ± SE

300

250 Distributed for

CIR 200 Comment Panel Page 53 of 990 Book 150 Only - Page Do Not 98

100 Quote BODYWEIGHT (GRAMS) or Cite 50

0 -4 -3 -2 -1 0 1 2 3 4 STUDY WEEK

0 MG/M3 30 MG/M3 BENZYL 100 MG/M3 BENZYL 300 MG/M3 BENZYL 1000MG/M3 BENZYL 2.5MG/M3 BENZOIC 12.5MG/M3BENZOIC

BW v1.00 07/21/2009 Distributed for Comment Only - Do Not Quote or Cite

WIL-703002 Benzyl Alcohol and Benzoic Acid The Personal Care Products Council

TABLES 1-36

Page 54 of 990

CIR Panel Book Page 99

TABLE 1 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SURVIVAL AND DISPOSITION

MALES GROUP : 1 2 3 4 ------WEEK LIVE FD EE SE LIVE FD EE SE LIVE FD EE SE LIVE FD EE SE ------0 10 0 0 0 10 0 0 0 10 0 0 0 10 0 0 0 1 10 0 0 0 10 0 0 0 10 0 0 0 10 0 0 0 2 10 0 0 0 10 0 0 0 10 0 0 0 10 0 0 0 Distributed 3-A 0 0 0 10 0 0 0 10 0 0 0 10 0 0 0 10 ------WEEK = WEEK OF STUDY FD = FOUND DEAD EE = EUTHANIZED IN EXTREMIS SE = SCHEDULED EUTHANASIA A = INCLUDES ANIMALS EUTHANIZED ON STUDY DAY 28 (FIRST DAY OF STUDY WEEK 4) for

------CIR

1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL Comment Panel Page 55 of 990 Book Only Page - Do Not 100 Quote or Cite

TABLE 1 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SURVIVAL AND DISPOSITION

MALES GROUP : 5 6 7 ------WEEK LIVE FD EE SE LIVE FD EE SE LIVE FD EE SE ------0 10 0 0 0 10 0 0 0 10 0 0 0 1 10 0 0 0 10 0 0 0 10 0 0 0 2 10 0 0 0 10 0 0 0 10 0 0 0 Distributed 3-A 0 0 0 10 0 0 0 10 0 0 0 10 ------WEEK = WEEK OF STUDY FD = FOUND DEAD EE = EUTHANIZED IN EXTREMIS SE = SCHEDULED EUTHANASIA A = INCLUDES ANIMALS EUTHANIZED ON STUDY DAY 28 (FIRST DAY OF STUDY WEEK 4) for

------CIR

5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC Comment PSURVv4.09 Panel 07/15/2009

Page 56 of 990 R:08/20/2009 Book Only Page - Do Not 101 Quote or Cite

TABLE 2 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SURVIVAL AND DISPOSITION

FEMALES GROUP : 1 2 3 4 ------WEEK LIVE FD EE SE LIVE FD EE SE LIVE FD EE SE LIVE FD EE SE ------0 10 0 0 0 10 0 0 0 10 0 0 0 10 0 0 0 1 10 0 0 0 10 0 0 0 10 0 0 0 10 0 0 0 2 10 0 0 0 10 0 0 0 10 0 0 0 10 0 0 0 Distributed 3-A 0 0 0 10 0 0 0 10 0 0 0 10 0 0 0 10 ------WEEK = WEEK OF STUDY FD = FOUND DEAD EE = EUTHANIZED IN EXTREMIS SE = SCHEDULED EUTHANASIA A = INCLUDES ANIMALS EUTHANIZED ON STUDY DAY 28 (FIRST DAY OF STUDY WEEK 4) for

------CIR

1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL Comment Panel Page 57 of 990 Book Only Page - Do Not 102 Quote or Cite

TABLE 2 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SURVIVAL AND DISPOSITION

FEMALES GROUP : 5 6 7 ------WEEK LIVE FD EE SE LIVE FD EE SE LIVE FD EE SE ------0 10 0 0 0 10 0 0 0 10 0 0 0 1 10 0 0 0 10 0 0 0 10 0 0 0 2 10 0 0 0 10 0 0 0 10 0 0 0 Distributed 3-A 0 0 0 10 0 0 0 10 0 0 0 10 ------WEEK = WEEK OF STUDY FD = FOUND DEAD EE = EUTHANIZED IN EXTREMIS SE = SCHEDULED EUTHANASIA A = INCLUDES ANIMALS EUTHANIZED ON STUDY DAY 28 (FIRST DAY OF STUDY WEEK 4) for

------CIR

5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC Comment PSURVv4.09 Panel 07/15/2009

Page 58 of 990 R:08/20/2009 Book Only Page - Do Not 103 Quote or Cite

TABLE 3 (DETAILED PHYSICAL EXAMINATIONS/DISPOSITIONS - MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- M A L E ------TABLE RANGE: 04-22-09 TO 05-20-09 GROUP: 1 2 3 4 5 6 7 ------

NORMAL -NO SIGNIFICANT CLINICAL OBSERVATIONS 48/10 43/10 48/10 49/10 50/10 48/10 49/10 Distributed

DISPOSITION -PRIMARY NECROPSY (WEEK 3) 10/10 10/10 10/10 10/10 10/10 10/10 10/10

for

BODY/INTEGUMENT CIR

-HAIR LOSS FORELIMB(S) 1/ 1 5/ 3 0/ 0 0/ 0 0/ 0 2/ 1 0/ 0 Comment

Panel EXCRETA

Page 59 of 990 -SOFT FECES 1/ 1 2/ 2 0/ 0 1/ 1 0/ 0 0/ 0 1/ 1

Book ORAL/DENTAL Only -UPPER INCISOR(S) BROKEN 0/ 0 0/ 0 2/ 1 0/ 0 0/ 0 0/ 0 0/ 0 Page - BODY/INTEG III Do Not

104 -WET BROWN MATERIAL ANOGENITAL AREA 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 1/ 1 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC Quote 7-12.5MG/M3BENZOIC PCSUv4.07 07/15/2009 or Cite

TABLE 4 (DETAILED PHYSICAL EXAMINATIONS/DISPOSITIONS - FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- F E M A L E ------TABLE RANGE: 04-22-09 TO 05-20-09 GROUP: 1 2 3 4 5 6 7 ------

NORMAL -NO SIGNIFICANT CLINICAL OBSERVATIONS 47/10 47/10 50/10 50/10 49/10 50/10 50/10 Distributed

DISPOSITION -PRIMARY NECROPSY (WEEK 3) 10/10 10/10 10/10 10/10 10/10 10/10 10/10

for

BODY/INTEGUMENT CIR

-HAIR LOSS FACIAL AREA 0/ 0 0/ 0 0/ 0 0/ 0 1/ 1 0/ 0 0/ 0 Comment -HAIR LOSS DORSAL HEAD 0/ 0 0/ 0 0/ 0 0/ 0 1/ 1 0/ 0 0/ 0 Panel -HAIR LOSS FORELIMB(S) 1/ 1 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0

Page 60 of 990 -HAIR LOSS HINDLIMB(S) 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0

Book EYES/EARS/NOSE Only -DRIED RED MATERIAL AROUND RIGHT EYE 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 Page - ORAL/DENTAL Do Not

105 -UPPER INCISOR(S) BROKEN 0/ 0 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 -LOWER INCISOR(S) BROKEN 0/ 0 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 ------Quote 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC PCSUv4.07 or

07/15/2009 Cite

TABLE 5 (PRIOR TO EXPOSURE - MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- M A L E ------TABLE RANGE: 04-22-09 TO 05-19-09 GROUP: 1 2 3 4 5 6 7 ------

NORMAL -NO SIGNIFICANT CLINICAL OBSERVATIONS 211/10 209/10 210/10 211/10 208/10 210/10 211/10 Distributed ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC PCSUv4.07 for

07/15/2009 CIR

R:07/17/2009 Comment Panel Page 61 of 990 Book Only Page - Do Not 106 Quote or Cite

TABLE 6 (PRIOR TO EXPOSURE - FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- F E M A L E ------TABLE RANGE: 04-22-09 TO 05-19-09 GROUP: 1 2 3 4 5 6 7 ------

NORMAL -NO SIGNIFICANT CLINICAL OBSERVATIONS 209/10 208/10 210/10 209/10 207/10 210/10 208/10 Distributed

EYES/EARS/NOSE -DRIED RED MATERIAL AROUND NOSE 0/ 0 0/ 0 0/ 0 0/ 0 1/ 1 0/ 0 0/ 0

for

ORAL/DENTAL CIR

-DRIED YELLOW MATERIAL DORSAL NECK 0/ 0 0/ 0 0/ 0 0/ 0 2/ 2 0/ 0 0/ 0 Comment -DRIED YELLOW MATERIAL DORSAL HEAD 0/ 0 0/ 0 0/ 0 0/ 0 2/ 2 0/ 0 0/ 0 Panel -DRIED YELLOW MATERIAL FACIAL AREA 0/ 0 0/ 0 0/ 0 0/ 0 2/ 2 0/ 0 0/ 0

Page 62 of 990 ------

Book 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC Only PCSUv4.07 Page 07/15/2009 - R:07/17/2009 Do Not 107 Quote or Cite

TABLE 7 (MID-EXPOSURE - MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- M A L E ------TABLE RANGE: 04-24-09 TO 05-19-09 GROUP: 1 2 3 4 5 6 7 ------NO SIGNIFICANT CLINICAL OBSERVATIONS MID-EXPOSURE ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC Distributed 7-12.5MG/M3BENZOIC PCSUv4.07 08/25/2009 R:08/25/2009 for

CIR Comment Panel Page 63 of 990 Book Only Page - Do Not 108 Quote or Cite

TABLE 8 (MID-EXPOSURE - FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- F E M A L E ------TABLE RANGE: 04-24-09 TO 05-19-09 GROUP: 1 2 3 4 5 6 7 ------NO SIGNIFICANT CLINICAL OBSERVATIONS MID-EXPOSURE ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC Distributed 7-12.5MG/M3BENZOIC PCSUv4.07 08/25/2009 R:08/25/2009 for

CIR Comment Panel Page 64 of 990 Book Only Page - Do Not 109 Quote or Cite

TABLE 9 (1 HOUR POST-EXPOSURE - MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- M A L E ------TABLE RANGE: 04-22-09 TO 05-19-09 GROUP: 1 2 3 4 5 6 7 ------

NORMAL -NO SIGNIFICANT CLINICAL OBSERVATIONS 211/10 207/10 209/10 211/10 208/10 210/10 211/10 Distributed

EXCRETA -SOFT FECES 0/ 0 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0

for

SPECIAL II CIR

-PROLAPSED PENIS 0/ 0 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 Comment ------Panel 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC

Page 65 of 990 7-12.5MG/M3BENZOIC

Book PCSUv4.07 07/15/2009 Only R:07/17/2009 Page - Do Not 110 Quote or Cite

TABLE 10 (1 HOUR POST-EXPOSURE - FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- F E M A L E ------TABLE RANGE: 04-22-09 TO 05-12-09 GROUP: 1 2 3 4 5 6 7 ------

NORMAL -NO SIGNIFICANT CLINICAL OBSERVATIONS 150/10 146/10 150/10 147/10 150/10 150/10 150/10 Distributed

BEHAVIOR/CNS -PARTIAL CLOSURE LEFT EYE 0/ 0 1/ 1 0/ 0 1/ 1 0/ 0 0/ 0 0/ 0 -PARTIAL CLOSURE RIGHT EYE 0/ 0 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 for

CIR

EYES/EARS/NOSE Comment -COMPLETE CLOSURE LEFT EYE 0/ 0 0/ 0 0/ 0 1/ 1 0/ 0 0/ 0 0/ 0 Panel -COMPLETE CLOSURE RIGHT EYE 0/ 0 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0

Page 66 of 990 ------

Book 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC Only PCSUv4.07 Page 07/15/2009 - R:07/17/2009 Do Not 111 Quote or Cite

TABLE 11 (DAILY OBSERVATIONS - NON-EXPOSURE DAYS - MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- M A L E ------TABLE RANGE: 04-27-09 TO 05-12-09 GROUP: 1 2 3 4 5 6 7 ------

NORMAL -NO SIGNIFICANT CLINICAL OBSERVATIONS 60/10 59/10 60/10 60/10 58/10 60/10 60/10 Distributed

EYES/EARS/NOSE -DRIED RED MATERIAL AROUND LEFT EYE 0/ 0 1/ 1 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 -DRIED RED MATERIAL AROUND NOSE 0/ 0 1/ 1 0/ 0 0/ 0 1/ 1 0/ 0 0/ 0 for

CIR

BODY/INTEG III Comment -DRIED YELLOW MATERIAL FORELIMB(S) 0/ 0 0/ 0 0/ 0 0/ 0 1/ 1 0/ 0 0/ 0 Panel ------

Page 67 of 990 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC

Book 7-12.5MG/M3BENZOIC PCSUv4.07 Only 07/15/2009 Page R:07/17/2009 - Do Not 112 Quote or Cite

TABLE 12 (DAILY OBSERVATIONS - NON-EXPOSURE DAYS - FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CLINICAL FINDINGS: TOTAL OCCURRENCE/NO. OF ANIMALS

----- F E M A L E ------TABLE RANGE: 04-27-09 TO 05-12-09 GROUP: 1 2 3 4 5 6 7 ------

NORMAL -NO SIGNIFICANT CLINICAL OBSERVATIONS 60/10 60/10 60/10 60/10 59/10 60/10 59/10 Distributed

EYES/EARS/NOSE -DRIED RED MATERIAL AROUND LEFT EYE 0/ 0 0/ 0 0/ 0 0/ 0 1/ 1 0/ 0 0/ 0 -DRIED RED MATERIAL AROUND NOSE 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 0/ 0 1/ 1 for

------CIR

1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC Comment 7-12.5MG/M3BENZOIC Panel PCSUv4.07

Page 68 of 990 07/15/2009

Book R:07/17/2009 Only Page - Do Not 113 Quote or Cite

TABLE 13 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHTS [G]

------MALES GROUP: 1 2 3 4 5 6 7 ------WEEK -4 MEAN 166. 166. 167. 169. 169. 162. 169. % DIFFERENCE 0.0 0.6 1.8 1.8 -2.4 1.8 S.D. 7.4 7.7 5.4 7.7 5.6 5.0 7.5 Distributed S.E. 2.3 2.4 1.7 2.4 1.8 1.6 2.4 N 10 10 10 10 10 10 10

-3 for

MEAN 203. 202. 203. 204. 203. 199. 203. CIR

% DIFFERENCE -0.5 0.0 0.5 0.0 -2.0 0.0 Comment S.D. 9.6 6.6 8.0 10.0 8.4 8.0 9.9 Panel S.E. 3.0 2.1 2.5 3.2 2.7 2.5 3.1

Page 69 of 990 N 10 10 10 10 10 10 10

Book -2 Only MEAN 257. 258. 257. 258. 257. 258. 258. Page % DIFFERENCE 0.4 0.0 0.4 0.0 0.4 0.4 - S.D. 14.9 13.2 13.8 12.7 14.4 15.1 14.5 Do Not

114 S.E. 4.7 4.2 4.4 4.0 4.6 4.8 4.6 N 10 10 10 10 10 10 10 Quote -1 MEAN 266. 263. 263. 262. 266. 265. 266. % DIFFERENCE -1.1 -1.1 -1.5 0.0 -0.4 0.0 or

S.D. 15.1 14.9 16.1 15.5 16.2 16.3 16.9 Cite S.E. 4.8 4.7 5.1 4.9 5.1 5.1 5.3 N 10 10 10 10 10 10 10

------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC ------None significantly different from control group

TABLE 13 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHTS [G]

------MALES GROUP: 1 2 3 4 5 6 7 ------WEEK 0 MEAN 311. 312. 313. 305. 309. 311. 315. % DIFFERENCE 0.3 0.6 -1.9 -0.6 0.0 1.3 S.D. 24.8 26.8 25.5 20.0 22.0 22.8 23.6 Distributed S.E. 7.9 8.5 8.1 6.3 7.0 7.2 7.5 N 10 10 10 10 10 10 10

1 for

MEAN 344. 343. 349. 338. 336. 344. 353. CIR

% DIFFERENCE -0.3 1.5 -1.7 -2.3 0.0 2.6 Comment S.D. 34.4 36.7 30.3 25.3 26.8 30.7 29.7 Panel S.E. 10.9 11.6 9.6 8.0 8.5 9.7 9.4

Page 70 of 990 N 10 10 10 10 10 10 10

Book 2 Only MEAN 376. 374. 378. 368. 364. 380. 384. Page % DIFFERENCE -0.5 0.5 -2.1 -3.2 1.1 2.1 - S.D. 42.3 41.0 33.0 28.7 32.2 36.9 33.7 Do Not

115 S.E. 13.4 13.0 10.4 9.1 10.2 11.7 10.7 N 10 10 10 10 10 10 10 Quote 3 MEAN 398. 397. 403. 385. 381. 401. 405. % DIFFERENCE -0.3 1.3 -3.3 -4.3 0.8 1.8 or

S.D. 48.3 43.5 38.0 29.2 32.2 39.9 39.8 Cite S.E. 15.3 13.7 12.0 9.2 10.2 12.6 12.6 N 10 10 10 10 10 10 10

------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC ------None significantly different from control group

TABLE 13 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHTS [G]

------MALES GROUP: 1 2 3 4 5 6 7 ------WEEK 3' MEAN 416. 411. 418. 399. 395. 419. 421. % DIFFERENCE -1.2 0.5 -4.1 -5.0 0.7 1.2 S.D. 54.3 44.8 40.6 28.5 36.7 42.7 41.7 Distributed S.E. 17.2 14.2 12.8 9.0 11.6 13.5 13.2 N 10 10 10 10 10 10 10

for

------CIR

1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC Comment 7-12.5MG/M3BENZOIC Panel ------

Page 71 of 990 None significantly different from control group

Book ' = SECOND WEEKLY BODY WEIGHT PBFSTv5.30 Only 08/25/2009 Page R:08/25/2009 - Do Not 116 Quote or Cite

TABLE 14 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHTS [G]

------FEMALES GROUP: 1 2 3 4 5 6 7 ------WEEK -4 MEAN 149. 145. 146. 147. 146. 146. 150. % DIFFERENCE -2.7 -2.0 -1.3 -2.0 -2.0 0.7 S.D. 7.2 9.4 6.4 5.1 6.3 8.7 8.9 Distributed S.E. 2.3 3.0 2.0 1.6 2.0 2.7 2.8 N 10 10 10 10 10 10 10

-3 for

MEAN 163. 163. 161. 161. 163. 161. 164. CIR

% DIFFERENCE 0.0 -1.2 -1.2 0.0 -1.2 0.6 Comment S.D. 8.0 8.7 8.8 7.2 7.0 9.4 8.2 Panel S.E. 2.5 2.8 2.8 2.3 2.2 3.0 2.6

Page 72 of 990 N 10 10 10 10 10 10 10

Book -2 Only MEAN 185. 184. 184. 183. 184. 184. 184. Page % DIFFERENCE -0.5 -0.5 -1.1 -0.5 -0.5 -0.5 - S.D. 10.7 11.0 10.3 8.4 9.0 9.7 10.0 Do Not

117 S.E. 3.4 3.5 3.3 2.6 2.8 3.1 3.2 N 10 10 10 10 10 10 10 Quote -1 MEAN 186. 184. 185. 182. 185. 184. 184. % DIFFERENCE -1.1 -0.5 -2.2 -0.5 -1.1 -1.1 or

S.D. 9.8 9.3 10.3 9.9 9.0 9.3 11.1 Cite S.E. 3.1 3.0 3.3 3.1 2.8 2.9 3.5 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 14 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHTS [G]

------FEMALES GROUP: 1 2 3 4 5 6 7 ------WEEK 0 MEAN 206. 201. 203. 201. 206. 204. 200. % DIFFERENCE -2.4 -1.5 -2.4 0.0 -1.0 -2.9 S.D. 11.6 13.0 14.2 12.3 10.8 11.5 13.1 Distributed S.E. 3.7 4.1 4.5 3.9 3.4 3.6 4.2 N 10 10 10 10 10 10 10

1 for

MEAN 223. 218. 219. 214. 217. 219. 216. CIR

% DIFFERENCE -2.2 -1.8 -4.0 -2.7 -1.8 -3.1 Comment S.D. 13.5 13.2 14.4 12.0 13.2 13.2 14.0 Panel S.E. 4.3 4.2 4.5 3.8 4.2 4.2 4.4

Page 73 of 990 N 10 10 10 10 10 10 10

Book 2 Only MEAN 233. 226. 229. 222. 229. 231. 226. Page % DIFFERENCE -3.0 -1.7 -4.7 -1.7 -0.9 -3.0 - S.D. 12.9 19.1 16.9 16.6 15.1 15.9 14.0 Do Not

118 S.E. 4.1 6.0 5.4 5.2 4.8 5.0 4.4 N 10 10 10 10 10 10 10 Quote 3 MEAN 243. 234. 238. 233. 235. 237. 235. % DIFFERENCE -3.7 -2.1 -4.1 -3.3 -2.5 -3.3 or

S.D. 14.8 17.9 16.1 16.9 14.4 13.2 13.2 Cite S.E. 4.7 5.7 5.1 5.3 4.5 4.2 4.2 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 14 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHTS [G]

------FEMALES GROUP: 1 2 3 4 5 6 7 ------WEEK 3' MEAN 249. 239. 241. 236. 239. 242. 240. % DIFFERENCE -4.0 -3.2 -5.2 -4.0 -2.8 -3.6 S.D. 14.7 17.7 16.3 18.7 16.1 13.2 14.9 Distributed S.E. 4.6 5.6 5.2 5.9 5.1 4.2 4.7 N 10 10 10 10 10 10 10

------for

None significantly different from control group CIR

' = SECOND WEEKLY BODY WEIGHT Comment ------Panel 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC

Page 74 of 990 7-12.5MG/M3BENZOIC

Book PBFSTv5.30 Only 07/15/2009 Page R:07/28/2009 - Do Not 119 Quote or Cite

TABLE 15 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHT CHANGES [G]

------MALES GROUP: 1 2 3 4 5 6 ------WEEK -4 TO -3 MEAN 37. 36. 36. 35. 34. 37. 35. S.D. 3.8 4.1 4.8 3.8 4.0 4.7 5.1 S.E. 1.2 1.3 1.5 1.2 1.3 1.5 1.6 Distributed N 10 10 10 10 10 10 10

-3 TO -2 MEAN 54. 56. 54. 54. 54. 59. 55. for

S.D. 7.7 10.3 8.6 4.8 9.5 7.7 8.8 CIR

S.E. 2.4 3.3 2.7 1.5 3.0 2.4 2.8 Comment N 10 10 10 10 10 10 10 Panel

Page 75 of 990 -2 TO -1

Book MEAN 9. 6. 6. 4. 9. 7. 8. S.D. 2.5 4.0 5.8 4.9 4.1 4.5 3.6 Only S.E. 0.8 1.3 1.8 1.5 1.3 1.4 1.1 Page N 10 10 10 10 10 10 10 - Do Not

120 -1 TO 0 MEAN 46. 49. 49. 43. 43. 46. 49. S.D. 11.6 14.2 11.1 5.4 10.5 10.0 8.8 Quote S.E. 3.7 4.5 3.5 1.7 3.3 3.1 2.8 N 10 10 10 10 10 10 10 or

0 TO 1 Cite MEAN 32. 31. 36. 33. 28. 34. 39. S.D. 10.2 10.9 8.1 7.2 9.2 12.0 9.0 S.E. 3.2 3.4 2.6 2.3 2.9 3.8 2.8 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 15 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHT CHANGES [G]

------MALES GROUP: 1 2 3 4 5 6 ------WEEK 1 TO 2 MEAN 32. 31. 30. 30. 28. 36. 31. S.D. 8.7 8.4 7.0 3.7 8.6 12.2 5.7 S.E. 2.7 2.7 2.2 1.2 2.7 3.8 1.8 Distributed N 10 10 10 10 10 10 10

2 TO 3 MEAN 22. 22. 25. 17. 17. 21. 21. for

S.D. 8.6 7.1 6.9 4.3 6.1 6.1 7.3 CIR

S.E. 2.7 2.3 2.2 1.4 1.9 1.9 2.3 Comment N 10 10 10 10 10 10 10 Panel

Page 76 of 990 3 TO 3'

Book MEAN 19. 15. 15. 14. 15. 18. 16. S.D. 8.7 4.5 7.8 4.8 8.1 6.8 6.5 Only S.E. 2.7 1.4 2.5 1.5 2.6 2.2 2.1 Page N 10 10 10 10 10 10 10 - Do Not

121 ------None significantly different from control group ' = SECOND WEEKLY BODY WEIGHT Quote ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC or

PBFSTv5.30 Cite 07/15/2009 R:07/28/2009

TABLE 16 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHT CHANGES [G]

------FEMALES GROUP: 1 2 3 4 5 6 ------WEEK -4 TO -3 MEAN 14. 18. 15. 14. 17. 15. 14. S.D. 2.4 6.5 4.1 2.9 3.1 1.5 1.9 S.E. 0.8 2.1 1.3 0.9 1.0 0.5 0.6 Distributed N 10 10 10 10 10 10 10

-3 TO -2 MEAN 22. 21. 22. 22. 21. 23. 19. for

S.D. 5.5 5.0 4.9 3.0 6.6 4.2 3.9 CIR

S.E. 1.7 1.6 1.6 0.9 2.1 1.3 1.2 Comment N 10 10 10 10 10 10 10 Panel

Page 77 of 990 -2 TO -1

Book MEAN 1. 0. 1. -2. 1. 0. 1. S.D. 4.2 5.8 2.8 3.3 7.9 3.9 3.2 Only S.E. 1.3 1.8 0.9 1.1 2.5 1.2 1.0 Page N 10 10 10 10 10 10 10 - Do Not

122 -1 TO 0 MEAN 20. 17. 18. 20. 22. 20. 16. S.D. 4.9 5.2 7.3 5.1 6.1 6.3 4.0 Quote S.E. 1.6 1.7 2.3 1.6 1.9 2.0 1.3 N 10 10 10 10 10 10 10 or

0 TO 1 Cite MEAN 17. 17. 16. 13. 11. 15. 16. S.D. 6.9 6.1 6.9 3.2 4.8 4.2 5.8 S.E. 2.2 1.9 2.2 1.0 1.5 1.3 1.8 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 16 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF BODY WEIGHT CHANGES [G]

------FEMALES GROUP: 1 2 3 4 5 6 ------WEEK 1 TO 2 MEAN 10. 8. 10. 8. 11. 12. 10. S.D. 6.2 7.7 3.4 6.8 6.5 4.3 4.0 S.E. 2.0 2.4 1.1 2.1 2.0 1.3 1.3 Distributed N 10 10 10 10 10 10 10

2 TO 3 MEAN 10. 8. 9. 11. 6. 6. 9. for

S.D. 6.0 8.5 3.9 3.0 6.5 4.8 6.8 CIR

S.E. 1.9 2.7 1.2 1.0 2.0 1.5 2.2 Comment N 10 10 10 10 10 10 10 Panel

Page 78 of 990 3 TO 3'

Book MEAN 6. 5. 3. 3. 4. 5. 5. S.D. 3.4 3.4 3.5 3.3 3.1 3.0 3.0 Only S.E. 1.1 1.1 1.1 1.0 1.0 0.9 1.0 Page N 10 10 10 10 10 10 10 - Do Not

123 ------None significantly different from control group ' = SECOND WEEKLY BODY WEIGHT Quote ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC or

PBFSTv5.30 Cite 07/15/2009 R:07/28/2009

TABLE 17 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CUMULATIVE BODY WEIGHT CHANGES [G]

------MALES GROUP: 1 2 3 4 5 6 ------WEEK 0 TO 1 MEAN 32. 31. 36. 33. 28. 34. 39. S.D. 10.2 10.9 8.1 7.2 9.2 12.0 9.0 S.E. 3.2 3.4 2.6 2.3 2.9 3.8 2.8 Distributed N 10 10 10 10 10 10 10

0 TO 2 MEAN 64. 62. 66. 63. 55. 70. 69. for

S.D. 18.0 15.1 13.1 10.6 13.9 16.0 12.3 CIR

S.E. 5.7 4.8 4.1 3.3 4.4 5.0 3.9 Comment N 10 10 10 10 10 10 10 Panel

Page 79 of 990 0 TO 3

Book MEAN 86. 84. 90. 80. 72. 91. 91. S.D. 24.4 18.7 17.1 11.4 15.3 19.2 18.7 Only S.E. 7.7 5.9 5.4 3.6 4.8 6.1 5.9 Page N 10 10 10 10 10 10 10 - Do Not

124 0 TO 3' MEAN 105. 99. 105. 94. 86. 109. 107. S.D. 31.0 20.9 22.3 10.9 20.9 21.3 20.8 Quote S.E. 9.8 6.6 7.0 3.5 6.6 6.7 6.6 N 10 10 10 10 10 10 10 or

------Cite None significantly different from control group ' = SECOND WEEKLY BODY WEIGHT ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC PBFSTv5.30 07/15/2009 R:07/28/2009

TABLE 18 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF CUMULATIVE BODY WEIGHT CHANGES [G]

------FEMALES GROUP: 1 2 3 4 5 6 7 ------WEEK 0 TO 1 MEAN 17. 17. 16. 13. 11. 15. 16. S.D. 6.9 6.1 6.9 3.2 4.8 4.2 5.8 S.E. 2.2 1.9 2.2 1.0 1.5 1.3 1.8 Distributed N 10 10 10 10 10 10 10

0 TO 2 MEAN 27. 25. 26. 21. 23. 27. 26. for

S.D. 9.1 8.2 7.8 7.6 5.0 7.2 7.6 CIR

S.E. 2.9 2.6 2.5 2.4 1.6 2.3 2.4 Comment N 10 10 10 10 10 10 10 Panel

Page 80 of 990 0 TO 3

Book MEAN 37. 33. 35. 32. 29. 33. 35. S.D. 8.5 9.2 6.1 8.3 6.2 4.7 6.8 Only S.E. 2.7 2.9 1.9 2.6 2.0 1.5 2.2 Page N 10 10 10 10 10 10 10 - Do Not

125 0 TO 3' MEAN 43. 38. 38. 35. 33. 38. 40. S.D. 8.9 10.2 7.5 9.4 7.6 4.9 6.2 Quote S.E. 2.8 3.2 2.4 3.0 2.4 1.5 2.0 N 10 10 10 10 10 10 10 or

TABLE 19 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF FOOD CONSUMPTION [G/ANIMAL/DAY]

------MALES GROUP: 1 2 3 4 5 6 7 ------WEEK -3 TO -2 MEAN 26. 25. 26. 26. 25. 27. 25. S.D. 1.7 1.9 1.6 2.2 2.1 1.7 1.9 S.E. 0.5 0.6 0.5 0.7 0.7 0.5 0.6 Distributed N 10 10 10 10 10 10 10

-1 TO 0 MEAN 26. 26. 26. 25. 26. 27. 26. for

S.D. 2.4 2.8 2.8 1.9 1.7 2.8 1.8 CIR

S.E. 0.7 0.9 0.9 0.6 0.6 0.9 0.6 Comment N 10 10 10 10 9 10 10 Panel

Page 81 of 990 0 TO 1

Book MEAN 26. 25. 26. 25. 24. 25. 26. S.D. 2.8 3.8 2.6 2.1 2.0 3.1 2.8 Only S.E. 0.9 1.2 0.8 0.7 0.6 1.0 0.9 Page N 10 10 10 10 10 9 10 - Do Not

126 1 TO 2 MEAN 27. 27. 27. 26. 25. 27. 27. S.D. 3.1 2.8 1.9 3.8 2.7 3.3 2.6 Quote S.E. 1.0 0.9 0.6 1.2 0.9 1.0 0.9 N 10 10 10 10 9 10 9 or

2 TO 3 Cite MEAN 28. 27. 29. 27. 26. 27. 28. S.D. 3.3 2.9 3.0 2.3 2.8 3.1 2.8 S.E. 1.0 0.9 1.0 0.7 0.9 1.0 0.9 N 10 10 10 10 9 10 9

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 19 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF FOOD CONSUMPTION [G/ANIMAL/DAY]

------MALES GROUP: 1 2 3 4 5 6 7 ------WEEK 3 TO 3' MEAN 28. 26. 26. 25. 24. 26. 26. S.D. 3.6 3.3 2.9 3.7 3.3 3.8 3.7 S.E. 1.1 1.0 0.9 1.2 1.1 1.2 1.2 Distributed N 10 10 10 10 10 10 10

------None significantly different from control group for

3' = FOOD OUT MEASURED AT END OF WEEK 3 CIR

------Comment 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC Panel 7-12.5MG/M3BENZOIC

Page 82 of 990 PBFSTv5.30

Book 07/21/2009 R:07/28/2009 Only Page - Do Not 127 Quote or Cite

TABLE 20 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF FOOD CONSUMPTION [G/ANIMAL/DAY]

------FEMALES GROUP: 1 2 3 4 5 6 ------WEEK -3 TO -2 MEAN 18. 17. 19. 18. 18. 17. 17. S.D. 3.7 1.5 2.1 0.9 0.9 1.3 1.2 S.E. 1.2 0.5 0.7 0.3 0.3 0.4 0.4 Distributed N 9 10 10 10 10 10 10

-1 TO 0 MEAN 18. 18. 18. 17. 18. 18. 18. for

S.D. 0.8 1.6 1.2 0.9 1.2 1.4 1.2 CIR

S.E. 0.3 0.5 0.4 0.3 0.4 0.4 0.4 Comment N 10 10 10 10 9 10 10 Panel

Page 83 of 990 0 TO 1

Book MEAN 19. 19. 19. 19. 18. 18. 19. S.D. 1.4 1.0 1.5 1.6 1.2 1.3 1.5 Only S.E. 0.4 0.3 0.5 0.5 0.4 0.4 0.5 Page N 10 10 10 10 10 10 10 - Do Not

128 1 TO 2 MEAN 19. 20. 22. 19. 19. 19. 19. S.D. 1.2 2.4 6.5 2.0 1.4 1.6 1.2 Quote S.E. 0.4 0.8 2.1 0.6 0.4 0.5 0.4 N 10 10 10 10 10 10 10 or

2 TO 3 Cite MEAN 20. 18. 19. 18. 18. 18. 18. S.D. 3.9 1.2 1.6 2.4 1.3 1.7 1.1 S.E. 1.2 0.4 0.5 0.7 0.4 0.5 0.4 N 10 8 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 20 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF FOOD CONSUMPTION [G/ANIMAL/DAY]

------FEMALES GROUP: 1 2 3 4 5 6 ------WEEK 3 TO 3' MEAN 19. 18. 19. 18. 19. 18. 18. S.D. 0.9 1.1 1.7 1.3 1.1 1.0 1.4 S.E. 0.3 0.3 0.5 0.4 0.3 0.3 0.4 Distributed N 10 10 10 10 10 10 10

------None significantly different from control group for

3' = FOOD OUT MEASURED AT END OF WEEK 3 CIR

------Comment 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC Panel 7-12.5MG/M3BENZOIC

Page 84 of 990 PBFSTv5.30

Book 07/21/2009 R:07/28/2009 Only Page - Do Not 129 Quote or Cite

TABLE 21 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------WHITE CELL (thous/uL) WEEK 3 MEAN 12.88 11.97 12.79 12.72 11.70 12.68 13.18 % DIFFERENCE -7.1 -0.7 -1.2 -9.2 -1.6 2.3 S.D. 3.535 2.951 3.153 4.126 3.249 3.396 3.307 Distributed S.E. 1.118 0.933 0.997 1.459 1.027 1.074 1.046 N 10 10 10 8 10 10 10

RED CELL (mil/uL) for

WEEK 3 MEAN 8.93 8.79 8.89 9.09 8.92 8.78 8.86 CIR

% DIFFERENCE -1.6 -0.4 1.8 -0.1 -1.7 -0.8 Comment S.D. 0.309 0.389 0.441 0.269 0.171 0.454 0.366 Panel S.E. 0.098 0.123 0.139 0.095 0.054 0.144 0.116

Page 85 of 990 N 10 10 10 8 10 10 10

Book HEMOGLOBIN (g/dL) Only WEEK 3 MEAN 16.4 16.4 16.4 16.6 16.5 16.4 16.2 Page % DIFFERENCE 0.0 0.0 1.2 0.6 0.0 -1.2 - S.D. 0.42 0.59 0.48 0.34 0.35 0.64 0.82 Do Not

130 S.E. 0.13 0.19 0.15 0.12 0.11 0.20 0.26 N 10 10 10 8 10 10 10 Quote HEMATOCRIT (%) WEEK 3 MEAN 48.3 48.5 48.4 48.9 48.3 48.4 47.7 % DIFFERENCE 0.4 0.2 1.2 0.0 0.2 -1.2 or

S.D. 1.84 2.38 1.70 1.34 1.04 1.97 2.32 Cite S.E. 0.58 0.75 0.54 0.47 0.33 0.62 0.73 N 10 10 10 8 10 10 10

------thous/uL = THOUSANDS/MICROLITER, mil/uL = MILLIONS/MICROLITER, fL = FEMTOLITERS, pg = PICOGRAMS, g/dL = GRAMS/DECILITER ------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 21 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------MCV (fL) WEEK 3 MEAN 54.1 55.2 54.5 53.8 54.1 55.2 53.8 % DIFFERENCE 2.0 0.7 -0.6 0.0 2.0 -0.6 S.D. 1.61 1.68 1.16 1.66 1.22 1.81 1.57 Distributed S.E. 0.51 0.53 0.37 0.59 0.39 0.57 0.50 N 10 10 10 8 10 10 10

MCH (pg) for

WEEK 3 MEAN 18.4 18.6 18.5 18.2 18.5 18.7 18.3 CIR

% DIFFERENCE 1.1 0.5 -1.1 0.5 1.6 -0.5 Comment S.D. 0.62 0.33 0.52 0.53 0.43 0.61 0.51 Panel S.E. 0.20 0.10 0.17 0.19 0.13 0.19 0.16

Page 86 of 990 N 10 10 10 8 10 10 10

Book MCHC (g/dL) Only WEEK 3 MEAN 34.0 33.8 33.9 33.9 34.1 33.8 34.1 Page % DIFFERENCE -0.6 -0.3 -0.3 0.3 -0.6 0.3 - S.D. 0.84 0.68 0.43 0.39 0.61 0.43 0.59 Do Not

131 S.E. 0.27 0.21 0.14 0.14 0.19 0.14 0.19 N 10 10 10 8 10 10 10 Quote PLATELET (thous/uL) WEEK 3 MEAN 1279. 1381. 1378. 1376. 1349. 1409. 1350. % DIFFERENCE 8.0 7.7 7.6 5.5 10.2 5.6 or

S.D. 175.7 174.7 137.3 213.3 142.4 182.7 215.3 Cite S.E. 55.6 55.2 43.4 75.4 45.0 57.8 68.1 N 10 10 10 8 10 10 10

TABLE 21 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------PROTIME (seconds) WEEK 3 MEAN 17.8 17.8 18.0 17.8 18.0 17.7 17.7 % DIFFERENCE 0.0 1.1 0.0 1.1 -0.6 -0.6 S.D. 0.81 0.53 0.69 0.72 0.46 0.57 0.65 Distributed S.E. 0.27 0.17 0.22 0.24 0.15 0.18 0.21 N 9 10 10 9 10 10 10

APTT (seconds) for

WEEK 3 MEAN 20.5 20.8 20.9 21.6 21.4 20.1 20.2 CIR

% DIFFERENCE 1.5 2.0 5.4 4.4 -2.0 -1.5 Comment S.D. 1.26 1.72 0.76 1.49 1.72 2.73 2.45 Panel S.E. 0.42 0.54 0.24 0.50 0.55 0.86 0.78

Page 87 of 990 N 9 10 10 9 10 10 10

Book RETICULOCYTE (%) Only WEEK 3 MEAN 2.0 1.9 1.8 1.7 1.7 1.9 2.2 Page % DIFFERENCE -5.0 -10.0 -15.0 -15.0 -5.0 10.0 - S.D. 0.36 0.37 0.25 0.19 0.28 0.31 0.38 Do Not

132 S.E. 0.11 0.12 0.08 0.07 0.09 0.10 0.12 N 10 10 10 8 10 10 10 Quote RETIC ABSOLUTE (thous/uL) WEEK 3 MEAN 177.7 165.7 155.9 155.5 149.2 168.6 192.9 % DIFFERENCE -6.8 -12.3 -12.5 -16.0 -5.1 8.6 or

S.D. 31.00 34.00 24.57 20.07 26.21 25.67 32.40 Cite S.E. 9.80 10.75 7.77 7.10 8.29 8.12 10.25 N 10 10 10 8 10 10 10

TABLE 21 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 4 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------NEUTROPHIL (%) WEEK 3 MEAN 15.1 17.2 11.9 15.4 13.0 19.1 14.1 % DIFFERENCE 13.9 -21.2 2.0 -13.9 26.5 -6.6 S.D. 3.45 14.21 2.67 5.89 2.44 17.90 4.24 Distributed S.E. 1.09 4.49 0.84 2.08 0.77 5.66 1.34 N 10 10 10 8 10 10 10

LYMPHOCYTE (%) for

WEEK 3 MEAN 81.2 79.3 84.5 81.2 84.0 77.5 82.5 CIR

% DIFFERENCE -2.3 4.1 0.0 3.4 -4.6 1.6 Comment S.D. 3.64 14.79 3.39 6.51 3.06 18.40 4.16 Panel S.E. 1.15 4.68 1.07 2.30 0.97 5.82 1.31

Page 88 of 990 N 10 10 10 8 10 10 10

Book MONOCYTE (%) Only WEEK 3 MEAN 1.6 1.8 1.7 1.5 1.3 1.7 1.6 Page % DIFFERENCE 12.5 6.3 -6.3 -18.8 6.3 0.0 - S.D. 0.42 0.95 0.77 0.47 0.49 0.63 0.47 Do Not

133 S.E. 0.13 0.30 0.24 0.17 0.15 0.20 0.15 N 10 10 10 8 10 10 10 Quote EOSINOPHIL (%) WEEK 3 MEAN 1.1 0.8 0.9 1.1 0.8 0.8 0.8 % DIFFERENCE -27.3 -18.2 0.0 -27.3 -27.3 -27.3 or

S.D. 0.39 0.36 0.23 0.50 0.28 0.27 0.32 Cite S.E. 0.12 0.11 0.07 0.18 0.09 0.09 0.10 N 10 10 10 8 10 10 10

TABLE 21 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 5 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------BASOPHIL (%) WEEK 3 MEAN 0.3 0.2 0.2 0.2 0.2 0.2 0.2 % DIFFERENCE -33.3 -33.3 -33.3 -33.3 -33.3 -33.3 S.D. 0.08 0.07 0.05 0.07 0.07 0.09 0.05 Distributed S.E. 0.03 0.02 0.01 0.03 0.02 0.03 0.01 N 10 10 10 8 10 10 10

LG UNSTAIN CELL (%) for

WEEK 3 MEAN 0.7 0.6 0.8 0.6 0.7 0.8 0.7 CIR

% DIFFERENCE -14.3 14.3 -14.3 0.0 14.3 0.0 Comment S.D. 0.33 0.16 0.28 0.23 0.26 0.30 0.30 Panel S.E. 0.10 0.05 0.09 0.08 0.08 0.10 0.10

Page 89 of 990 N 10 10 10 8 10 10 10

Book NEU ABSOLUTE (thous/uL) Only WEEK 3 MEAN 1.91 2.35 1.52 1.88 1.53 2.66 1.87 Page % DIFFERENCE 23.0 -20.4 -1.6 -19.9 39.3 -2.1 - S.D. 0.621 2.947 0.456 0.806 0.608 3.376 0.811 Do Not

134 S.E. 0.196 0.932 0.144 0.285 0.192 1.068 0.256 N 10 10 10 8 10 10 10 Quote LYMPH ABSOLUTE (thous/uL) WEEK 3 MEAN 10.49 9.20 10.81 10.41 9.81 9.58 10.86 % DIFFERENCE -12.3 3.1 -0.8 -6.5 -8.7 3.5 or

S.D. 2.939 1.656 2.739 3.664 2.657 3.128 2.730 Cite S.E. 0.929 0.524 0.866 1.295 0.840 0.989 0.863 N 10 10 10 8 10 10 10

TABLE 21 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 6 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------MONO ABSOLUTE (thous/uL) WEEK 3 MEAN 0.20 0.23 0.22 0.19 0.16 0.21 0.22 % DIFFERENCE 15.0 10.0 -5.0 -20.0 5.0 10.0 S.D. 0.083 0.191 0.105 0.082 0.078 0.103 0.114 Distributed S.E. 0.026 0.060 0.033 0.029 0.025 0.033 0.036 N 10 10 10 8 10 10 10

EOS ABSOLUTE (thous/uL) for

WEEK 3 MEAN 0.15 0.09 0.11 0.13 0.10 0.09 0.10 CIR

% DIFFERENCE -40.0 -26.7 -13.3 -33.3 -40.0 -33.3 Comment S.D. 0.058 0.032 0.034 0.063 0.036 0.040 0.031 Panel S.E. 0.018 0.010 0.011 0.022 0.011 0.013 0.010

Page 90 of 990 N 10 10 10 8 10 10 10

Book BASO ABSOLUTE (thous/uL) Only WEEK 3 MEAN 0.04 0.03 0.03 0.03 0.03 0.03 0.03 Page % DIFFERENCE -25.0 -25.0 -25.0 -25.0 -25.0 -25.0 - S.D. 0.018 0.010 0.011 0.019 0.012 0.012 0.014 Do Not

135 S.E. 0.006 0.003 0.003 0.007 0.004 0.004 0.004 N 10 10 10 8 10 10 10 Quote LUC ABSOLUTE (thous/uL) WEEK 3 MEAN 0.10 0.08 0.10 0.07 0.08 0.11 0.10 % DIFFERENCE -20.0 0.0 -30.0 -20.0 10.0 0.0 or

S.D. 0.057 0.028 0.041 0.029 0.049 0.068 0.061 Cite S.E. 0.018 0.009 0.013 0.010 0.016 0.021 0.019 N 10 10 10 8 10 10 10

TABLE 21 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 7 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------RED CELLWIDTH (%) WEEK 3 MEAN 12.2 12.1 12.1 12.0 11.9 12.2 12.4 % DIFFERENCE -0.8 -0.8 -1.6 -2.5 0.0 1.6 S.D. 0.27 0.73 0.24 0.37 0.34 0.33 0.55 Distributed S.E. 0.09 0.23 0.08 0.13 0.11 0.10 0.17 N 10 10 10 8 10 10 10

HGB WIDTH (g/dL) for

WEEK 3 MEAN 2.50 2.51 2.40 2.51 2.41 2.42 2.53 CIR

% DIFFERENCE 0.4 -4.0 0.4 -3.6 -3.2 1.2 Comment S.D. 0.116 0.186 0.105 0.140 0.161 0.099 0.192 Panel S.E. 0.037 0.059 0.033 0.049 0.051 0.031 0.061

Page 91 of 990 N 10 10 10 8 10 10 10

Book ------Only thous/uL = THOUSANDS/MICROLITER, mil/uL = MILLIONS/MICROLITER, fL = FEMTOLITERS, pg = PICOGRAMS, g/dL = GRAMS/DECILITER Page ------None significantly different from control group Do Not

136 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC Quote PCPSv5.27 07/15/2009 R:07/28/2009 or Cite

TABLE 22 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------WHITE CELL (thous/uL) WEEK 3 MEAN 10.23 9.13 9.83 10.15 9.44 10.33 9.77 % DIFFERENCE -10.8 -3.9 -0.8 -7.7 1.0 -4.5 S.D. 2.914 3.402 2.453 3.428 2.172 2.185 3.699 Distributed S.E. 0.971 1.076 0.776 1.143 0.687 0.728 1.170 N 9 10 10 9 10 9 10

RED CELL (mil/uL) for

WEEK 3 MEAN 8.52 8.55 8.31 8.63 8.46 8.40 8.50 CIR

% DIFFERENCE 0.4 -2.5 1.3 -0.7 -1.4 -0.2 Comment S.D. 0.319 0.240 0.300 0.531 0.407 0.410 0.465 Panel S.E. 0.106 0.076 0.095 0.177 0.129 0.137 0.147

Page 92 of 990 N 9 10 10 9 10 9 10

Book HEMOGLOBIN (g/dL) Only WEEK 3 MEAN 15.7 15.8 15.4 15.7 15.6 15.7 15.6 Page % DIFFERENCE 0.6 -1.9 0.0 -0.6 0.0 -0.6 - S.D. 0.59 0.42 0.55 0.62 0.87 0.72 0.59 Do Not

137 S.E. 0.20 0.13 0.17 0.21 0.27 0.24 0.19 N 9 10 10 9 10 9 10 Quote HEMATOCRIT (%) WEEK 3 MEAN 45.0 45.8 44.0 45.3 45.4 45.2 45.2 % DIFFERENCE 1.8 -2.2 0.7 0.9 0.4 0.4 or

S.D. 1.90 1.58 1.75 2.07 2.58 2.30 2.23 Cite S.E. 0.63 0.50 0.55 0.69 0.82 0.77 0.71 N 9 10 10 9 10 9 10

TABLE 22 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------MCV (fL) WEEK 3 MEAN 52.9 53.6 53.0 52.6 53.6 53.8 53.2 % DIFFERENCE 1.3 0.2 -0.6 1.3 1.7 0.6 S.D. 1.11 1.39 1.61 1.68 1.72 0.91 1.94 Distributed S.E. 0.37 0.44 0.51 0.56 0.54 0.30 0.61 N 9 10 10 9 10 9 10

MCH (pg) for

WEEK 3 MEAN 18.4 18.5 18.5 18.3 18.5 18.7 18.4 CIR

% DIFFERENCE 0.5 0.5 -0.5 0.5 1.6 0.0 Comment S.D. 0.44 0.41 0.51 0.61 0.45 0.47 0.47 Panel S.E. 0.15 0.13 0.16 0.20 0.14 0.16 0.15

Page 93 of 990 N 9 10 10 9 10 9 10

Book MCHC (g/dL) Only WEEK 3 MEAN 34.7 34.6 34.9 34.7 34.4 34.7 34.6 Page % DIFFERENCE -0.3 0.6 0.0 -0.9 0.0 -0.3 - S.D. 0.51 0.44 0.42 0.43 0.39 0.69 0.72 Do Not

138 S.E. 0.17 0.14 0.13 0.14 0.12 0.23 0.23 N 9 10 10 9 10 9 10 Quote PLATELET (thous/uL) WEEK 3 MEAN 1464. 1466. 1482. 1473. 1523. 1683. 1488. % DIFFERENCE 0.1 1.2 0.6 4.0 15.0 1.6 or

S.D. 151.8 185.9 197.7 196.5 149.9 161.3 141.2 Cite S.E. 50.6 58.8 62.5 65.5 47.4 53.8 44.7 N 9 10 10 9 10 9 10

TABLE 22 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------PROTIME (seconds) WEEK 3 MEAN 18.2 18.1 18.2 18.3 18.0 18.0 18.1 % DIFFERENCE -0.5 0.0 0.5 -1.1 -1.1 -0.5 S.D. 0.98 0.96 0.70 0.91 0.62 1.03 1.22 Distributed S.E. 0.31 0.32 0.22 0.29 0.20 0.33 0.38 N 10 9 10 10 10 10 10

APTT (seconds) for

WEEK 3 MEAN 17.4 17.5 17.9 17.7 16.8 17.5 18.7 CIR

% DIFFERENCE 0.6 2.9 1.7 -3.4 0.6 7.5 Comment S.D. 1.77 2.57 1.95 1.49 2.30 1.99 4.84 Panel S.E. 0.56 0.86 0.62 0.47 0.73 0.63 1.53

Page 94 of 990 N 10 9 10 10 10 10 10

Book RETICULOCYTE (%) Only WEEK 3 MEAN 1.9 1.9 1.9 1.8 2.2 1.9 2.1 Page % DIFFERENCE 0.0 0.0 -5.3 15.8 0.0 10.5 - S.D. 0.34 0.32 0.35 0.27 0.68 0.39 0.49 Do Not

139 S.E. 0.11 0.10 0.11 0.09 0.22 0.13 0.15 N 9 10 10 9 10 9 10 Quote RETIC ABSOLUTE (thous/uL) WEEK 3 MEAN 162.5 164.9 161.7 157.9 185.3 158.8 175.1 % DIFFERENCE 1.5 -0.5 -2.8 14.0 -2.3 7.8 or

S.D. 31.69 29.41 25.00 19.27 50.98 32.82 40.28 Cite S.E. 10.56 9.30 7.91 6.42 16.12 10.94 12.74 N 9 10 10 9 10 9 10

TABLE 22 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 4 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------NEUTROPHIL (%) WEEK 3 MEAN 10.3 11.7 9.7 11.7 9.8 11.4 10.0 % DIFFERENCE 13.6 -5.8 13.6 -4.9 10.7 -2.9 S.D. 2.46 3.18 3.09 3.46 2.76 3.31 3.03 Distributed S.E. 0.82 1.01 0.98 1.15 0.87 1.10 0.96 N 9 10 10 9 10 9 10

LYMPHOCYTE (%) for

WEEK 3 MEAN 86.0 84.7 86.8 84.6 87.0 85.1 86.1 CIR

% DIFFERENCE -1.5 0.9 -1.6 1.2 -1.0 0.1 Comment S.D. 2.35 3.68 3.44 3.99 3.01 3.76 3.55 Panel S.E. 0.78 1.16 1.09 1.33 0.95 1.25 1.12

Page 95 of 990 N 9 10 10 9 10 9 10

Book MONOCYTE (%) Only WEEK 3 MEAN 1.4 1.5 1.3 1.5 1.2 1.4 1.5 Page % DIFFERENCE 7.1 -7.1 7.1 -14.3 0.0 7.1 - S.D. 0.36 0.58 0.28 0.55 0.29 0.41 0.84 Do Not

140 S.E. 0.12 0.18 0.09 0.18 0.09 0.14 0.27 N 9 10 10 9 10 9 10 Quote EOSINOPHIL (%) WEEK 3 MEAN 1.1 1.1 1.2 1.2 1.0 1.1 1.2 % DIFFERENCE 0.0 9.1 9.1 -9.1 0.0 9.1 or

S.D. 0.26 0.46 0.38 0.42 0.32 0.47 0.36 Cite S.E. 0.09 0.15 0.12 0.14 0.10 0.16 0.11 N 9 10 10 9 10 9 10

TABLE 22 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 5 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------BASOPHIL (%) WEEK 3 MEAN 0.2 0.2 0.2 0.2 0.2 0.2 0.2 % DIFFERENCE 0.0 0.0 0.0 0.0 0.0 0.0 S.D. 0.06 0.05 0.07 0.06 0.05 0.05 0.07 Distributed S.E. 0.02 0.02 0.02 0.02 0.01 0.02 0.02 N 9 10 10 9 10 9 10

LG UNSTAIN CELL (%) for

WEEK 3 MEAN 1.0 0.9 0.9 0.9 0.8 0.8 1.1 CIR

% DIFFERENCE -10.0 -10.0 -10.0 -20.0 -20.0 10.0 Comment S.D. 0.25 0.29 0.21 0.35 0.20 0.26 0.38 Panel S.E. 0.08 0.09 0.07 0.12 0.06 0.09 0.12

Page 96 of 990 N 9 10 10 9 10 9 10

Book NEU ABSOLUTE (thous/uL) Only WEEK 3 MEAN 1.05 0.99 0.96 1.14 0.93 1.16 0.99 Page % DIFFERENCE -5.7 -8.6 8.6 -11.4 10.5 -5.7 - S.D. 0.411 0.211 0.434 0.384 0.365 0.411 0.501 Do Not

141 S.E. 0.137 0.067 0.137 0.128 0.115 0.137 0.158 N 9 10 10 9 10 9 10 Quote LYMPH ABSOLUTE (thous/uL) WEEK 3 MEAN 8.80 7.83 8.52 8.64 8.20 8.82 8.40 % DIFFERENCE -11.0 -3.2 -1.8 -6.8 0.2 -4.5 or

S.D. 2.469 3.307 2.104 3.274 1.898 2.112 3.178 Cite S.E. 0.823 1.046 0.665 1.091 0.600 0.704 1.005 N 9 10 10 9 10 9 10

TABLE 22 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 6 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------MONO ABSOLUTE (thous/uL) WEEK 3 MEAN 0.14 0.13 0.13 0.14 0.11 0.14 0.14 % DIFFERENCE -7.1 -7.1 0.0 -21.4 0.0 0.0 S.D. 0.050 0.049 0.033 0.049 0.037 0.043 0.070 Distributed S.E. 0.017 0.016 0.010 0.016 0.012 0.014 0.022 N 9 10 10 9 10 9 10

EOS ABSOLUTE (thous/uL) for

WEEK 3 MEAN 0.11 0.09 0.11 0.11 0.09 0.11 0.11 CIR

% DIFFERENCE -18.2 0.0 0.0 -18.2 0.0 0.0 Comment S.D. 0.044 0.028 0.047 0.038 0.031 0.036 0.041 Panel S.E. 0.015 0.009 0.015 0.013 0.010 0.012 0.013

Page 97 of 990 N 9 10 10 9 10 9 10

Book BASO ABSOLUTE (thous/uL) Only WEEK 3 MEAN 0.02 0.02 0.02 0.02 0.02 0.02 0.02 Page % DIFFERENCE 0.0 0.0 0.0 0.0 0.0 0.0 - S.D. 0.011 0.011 0.008 0.012 0.006 0.008 0.013 Do Not

142 S.E. 0.004 0.003 0.003 0.004 0.002 0.003 0.004 N 9 10 10 9 10 9 10 Quote LUC ABSOLUTE (thous/uL) WEEK 3 MEAN 0.10 0.08 0.09 0.09 0.08 0.08 0.11 % DIFFERENCE -20.0 -10.0 -10.0 -20.0 -20.0 10.0 or

S.D. 0.040 0.040 0.030 0.072 0.028 0.025 0.052 Cite S.E. 0.013 0.013 0.010 0.024 0.009 0.008 0.017 N 9 10 10 9 10 9 10

TABLE 22 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 7 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF HEMATOLOGY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------RED CELLWIDTH (%) WEEK 3 MEAN 11.7 11.5 11.7 11.7 11.8 11.6 11.7 % DIFFERENCE -1.7 0.0 0.0 0.9 -0.9 0.0 S.D. 0.63 0.32 0.37 0.41 0.94 0.45 0.30 Distributed S.E. 0.21 0.10 0.12 0.14 0.30 0.15 0.09 N 9 10 10 9 10 9 10

HGB WIDTH (g/dL) for

WEEK 3 MEAN 2.39 2.31 2.40 2.38 2.36 2.41 2.37 CIR

% DIFFERENCE -3.3 0.4 -0.4 -1.3 0.8 -0.8 Comment S.D. 0.117 0.168 0.121 0.217 0.129 0.117 0.104 Panel S.E. 0.039 0.053 0.038 0.072 0.041 0.039 0.033

Page 98 of 990 N 9 10 10 9 10 9 10

143 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC Quote PCPSv5.27 07/15/2009 R:07/28/2009 or Cite

TABLE 23 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------ALBUMIN (g/dL) WEEK 3 MEAN 4.4 4.3 4.3 4.4 4.4 4.3 4.3 % DIFFERENCE -2.3 -2.3 0.0 0.0 -2.3 -2.3 S.D. 0.18 0.23 0.20 0.19 0.15 0.25 0.27 Distributed S.E. 0.06 0.07 0.06 0.06 0.05 0.08 0.08 N 10 10 10 10 10 10 10

TOTAL PROTEIN (g/dL) for

WEEK 3 MEAN 6.6 6.5 6.4 6.4 6.5 6.5 6.5 CIR

% DIFFERENCE -1.5 -3.0 -3.0 -1.5 -1.5 -1.5 Comment S.D. 0.25 0.24 0.38 0.29 0.19 0.27 0.26 Panel S.E. 0.08 0.07 0.12 0.09 0.06 0.09 0.08

Page 99 of 990 N 10 10 10 10 10 10 10

Book GLOBULIN (g/dL) Only WEEK 3 MEAN 2.2 2.2 2.1 2.0 2.1 2.2 2.3 Page % DIFFERENCE 0.0 -4.5 -9.1 -4.5 0.0 4.5 - S.D. 0.18 0.16 0.25 0.17 0.16 0.18 0.23 Do Not

144 S.E. 0.06 0.05 0.08 0.05 0.05 0.06 0.07 N 10 10 10 10 10 10 10 Quote A/G RATIO WEEK 3 MEAN 2.04 2.01 2.14 2.19 2.15 2.01 1.93 % DIFFERENCE -1.5 4.9 7.4 5.4 -1.5 -5.4 or

S.D. 0.197 0.215 0.218 0.194 0.186 0.205 0.238 Cite S.E. 0.062 0.068 0.069 0.061 0.059 0.065 0.075 N 10 10 10 10 10 10 10

------mg/dL = MILLIGRAMS/DECILITER, U/L = INTERNATIONAL UNIT/LITER, g/dL = GRAMS/DECILITER, mEq/L = MILLIEQUIVALENTS/LITER ------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 23 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------TOTAL BILI (mg/dL) WEEK 3 MEAN 0.09 0.10 0.08 0.08 0.07 0.09 0.09 % DIFFERENCE 11.1 -11.1 -11.1 -22.2 0.0 0.0 S.D. 0.039 0.035 0.016 0.042 0.017 0.023 0.031 Distributed S.E. 0.012 0.011 0.005 0.013 0.005 0.007 0.010 N 10 10 10 10 10 10 10

UREA NITROGEN (mg/dL) for

WEEK 3 MEAN 15.2 15.4 15.5 15.7 13.4 13.5 14.8 CIR

% DIFFERENCE 1.3 2.0 3.3 -11.8 -11.2 -2.6 Comment S.D. 2.89 3.20 1.77 1.83 0.61 2.43 3.57 Panel S.E. 0.91 1.01 0.56 0.58 0.19 0.77 1.13 Page 100 of 990 N 10 10 10 10 10 10 10

Book CREATININE (mg/dL) Only WEEK 3 MEAN 0.2 0.2 0.2 0.3 0.2 0.3 0.2 Page % DIFFERENCE 0.0 0.0 50.0 0.0 50.0 0.0 - S.D. 0.05 0.07 0.10 0.05 0.08 0.05 0.04 Do Not

145 S.E. 0.02 0.02 0.03 0.02 0.03 0.02 0.01 N 10 10 10 10 10 10 10 Quote ALKALINEPHOS'TSE (U/L) WEEK 3 MEAN 181. 175. 167. 151. 147. 154. 149. % DIFFERENCE -3.3 -7.7 -16.6 -18.8 -14.9 -17.7 or

S.D. 34.1 44.0 30.3 31.2 26.1 21.6 25.9 Cite S.E. 10.8 13.9 9.6 9.9 8.3 6.8 8.2 N 10 10 10 10 10 10 10

TABLE 23 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------ALANINE TRANSFER (U/L) WEEK 3 MEAN 39. 43. 41. 42. 44. 38. 34. % DIFFERENCE 10.3 5.1 7.7 12.8 -2.6 -12.8 S.D. 6.2 5.8 5.8 4.2 8.6 6.7 4.7 Distributed S.E. 2.0 1.8 1.8 1.3 2.7 2.1 1.5 N 10 10 10 10 10 10 10

ASPARTATTRANSFER (U/L) for

WEEK 3 MEAN 103. 101. 97. 106. 102. 96. 87. CIR

% DIFFERENCE -1.9 -5.8 2.9 -1.0 -6.8 -15.5 Comment S.D. 21.3 16.6 12.2 34.6 16.0 15.5 9.9 Panel S.E. 6.7 5.2 3.9 11.0 5.1 4.9 3.1 Page 101 of 990 N 10 10 10 10 10 10 10

Book GLUTAMYLTRANSFER (U/L) Only WEEK 3 MEAN 0.2 0.2 0.1 0.3 0.2 0.1 0.2 Page % DIFFERENCE 0.0 -50.0 50.0 0.0 -50.0 0.0 - S.D. 0.19 0.26 0.00 0.39 0.28 0.13 0.25 Do Not

146 S.E. 0.06 0.08 0.00 0.12 0.09 0.04 0.08 N 10 10 10 10 10 10 10 Quote GLUCOSE (mg/dL) WEEK 3 MEAN 115. 102. 112. 115. 108. 104. 110. % DIFFERENCE -11.3 -2.6 0.0 -6.1 -9.6 -4.3 or

S.D. 15.7 11.6 11.9 17.8 13.0 10.7 7.2 Cite S.E. 5.0 3.7 3.7 5.6 4.1 3.4 2.3 N 10 10 10 10 10 10 10

TABLE 23 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 4 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------CHOLESTEROL (mg/dL) WEEK 3 MEAN 63. 51. 59. 58. 50. 65. 56. % DIFFERENCE -19.0 -6.3 -7.9 -20.6 3.2 -11.1 S.D. 13.2 5.8 10.3 11.4 8.7 12.5 11.9 Distributed S.E. 4.2 1.8 3.3 3.6 2.7 3.9 3.8 N 10 10 10 10 10 10 10

CALCIUM (mg/dL) for

WEEK 3 MEAN 10.7 10.6 10.7 10.7 10.7 10.6 10.8 CIR

% DIFFERENCE -0.9 0.0 0.0 0.0 -0.9 0.9 Comment S.D. 0.24 0.29 0.28 0.34 0.30 0.35 0.30 Panel S.E. 0.08 0.09 0.09 0.11 0.09 0.11 0.09 Page 102 of 990 N 10 10 10 10 10 10 10

Book CHLORIDE (mEq/L) Only WEEK 3 MEAN 102. 102. 102. 102. 102. 103. 102. Page % DIFFERENCE 0.0 0.0 0.0 0.0 1.0 0.0 - S.D. 2.1 2.0 1.1 1.9 1.6 1.1 1.3 Do Not

147 S.E. 0.7 0.6 0.3 0.6 0.5 0.3 0.4 N 10 10 10 10 10 10 10 Quote PHOSPHORUS (mg/dL) WEEK 3 MEAN 8.0 7.9 7.9 8.0 7.6 8.0 8.0 % DIFFERENCE -1.3 -1.3 0.0 -5.0 0.0 0.0 or

S.D. 0.52 1.23 0.72 0.65 0.54 0.72 0.34 Cite S.E. 0.16 0.39 0.23 0.21 0.17 0.23 0.11 N 10 10 10 10 10 10 10

TABLE 23 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 5 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------POTASSIUM (mEq/L) WEEK 3 MEAN 4.95 4.72 4.83 4.79 4.71 4.88 4.65 % DIFFERENCE -4.6 -2.4 -3.2 -4.8 -1.4 -6.1 S.D. 0.503 0.545 0.437 0.502 0.473 0.473 0.311 Distributed S.E. 0.159 0.172 0.138 0.159 0.149 0.150 0.098 N 10 10 10 10 10 10 10

SODIUM (mEq/L) for

WEEK 3 MEAN 143. 144. 144. 144. 144. 144. 144. CIR

% DIFFERENCE 0.7 0.7 0.7 0.7 0.7 0.7 Comment S.D. 1.1 1.9 1.4 1.3 1.2 1.2 1.3 Panel S.E. 0.3 0.6 0.5 0.4 0.4 0.4 0.4 Page 103 of 990 N 10 10 10 10 10 10 10

Book TRIGLYCERIDE (mg/dL) Only WEEK 3 MEAN 57. 55. 55. 47. 46. 51. 57. Page % DIFFERENCE -3.5 -3.5 -17.5 -19.3 -10.5 0.0 - S.D. 30.8 40.9 20.8 16.7 12.1 20.7 13.8 Do Not

148 S.E. 9.8 12.9 6.6 5.3 3.8 6.5 4.4 N 10 10 10 10 10 10 10 Quote ------mg/dL = MILLIGRAMS/DECILITER, U/L = INTERNATIONAL UNIT/LITER, g/dL = GRAMS/DECILITER, mEq/L = MILLIEQUIVALENTS/LITER ------or

None significantly different from control group Cite ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC PCPSv5.27 07/15/2009 R:07/28/2009

TABLE 24 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------ALBUMIN (g/dL) WEEK 3 MEAN 4.9 4.9 4.8 4.9 4.9 4.9 4.9 % DIFFERENCE 0.0 -2.0 0.0 0.0 0.0 0.0 S.D. 0.22 0.14 0.18 0.29 0.26 0.26 0.19 Distributed S.E. 0.07 0.04 0.06 0.09 0.08 0.08 0.06 N 10 10 10 10 10 10 10

TOTAL PROTEIN (g/dL) for

WEEK 3 MEAN 6.9 6.8 6.8 6.8 6.8 6.8 6.9 CIR

% DIFFERENCE -1.4 -1.4 -1.4 -1.4 -1.4 0.0 Comment S.D. 0.24 0.24 0.25 0.42 0.37 0.27 0.21 Panel S.E. 0.08 0.08 0.08 0.13 0.12 0.09 0.07 Page 104 of 990 N 10 10 10 10 10 10 10

Book GLOBULIN (g/dL) Only WEEK 3 MEAN 2.0 1.9 2.0 1.9 1.9 1.9 2.0 Page % DIFFERENCE -5.0 0.0 -5.0 -5.0 -5.0 0.0 - S.D. 0.17 0.18 0.13 0.20 0.24 0.14 0.09 Do Not

149 S.E. 0.05 0.06 0.04 0.06 0.08 0.05 0.03 N 10 10 10 10 10 10 10 Quote A/G RATIO WEEK 3 MEAN 2.49 2.65 2.47 2.62 2.63 2.61 2.41 % DIFFERENCE 6.4 -0.8 5.2 5.6 4.8 -3.2 or

S.D. 0.256 0.234 0.142 0.247 0.352 0.276 0.145 Cite S.E. 0.081 0.074 0.045 0.078 0.111 0.087 0.046 N 10 10 10 10 10 10 10

TABLE 24 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------TOTAL BILI (mg/dL) WEEK 3 MEAN 0.12 0.13 0.14 0.12 0.14 0.12 0.12 % DIFFERENCE 8.3 16.7 0.0 16.7 0.0 0.0 S.D. 0.019 0.043 0.028 0.039 0.024 0.038 0.043 Distributed S.E. 0.006 0.013 0.009 0.012 0.007 0.012 0.014 N 10 10 10 10 10 10 10

UREA NITROGEN (mg/dL) for

WEEK 3 MEAN 19.0 18.1 18.6 17.3 18.8 17.2 17.9 CIR

% DIFFERENCE -4.7 -2.1 -8.9 -1.1 -9.5 -5.8 Comment S.D. 2.44 1.53 1.61 1.48 2.80 2.73 1.40 Panel S.E. 0.77 0.48 0.51 0.47 0.89 0.86 0.44 Page 105 of 990 N 10 10 10 10 10 10 10

Book CREATININE (mg/dL) Only WEEK 3 MEAN 0.3 0.3 0.3 0.3 0.3 0.3 0.3 Page % DIFFERENCE 0.0 0.0 0.0 0.0 0.0 0.0 - S.D. 0.04 0.03 0.08 0.03 0.06 0.03 0.07 Do Not

150 S.E. 0.01 0.01 0.03 0.01 0.02 0.01 0.02 N 10 10 10 10 10 10 10 Quote ALKALINEPHOS'TSE (U/L) WEEK 3 MEAN 116. 102. 97. 100. 110. 98. 97. % DIFFERENCE -12.1 -16.4 -13.8 -5.2 -15.5 -16.4 or

S.D. 22.0 19.0 23.9 20.5 21.2 25.6 21.3 Cite S.E. 7.0 6.0 7.6 6.5 6.7 8.1 6.7 N 10 10 10 10 10 10 10

TABLE 24 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------ALANINE TRANSFER (U/L) WEEK 3 MEAN 36. 47. 46. 41. 40. 40. 40. % DIFFERENCE 30.6 27.8 13.9 11.1 11.1 11.1 S.D. 6.1 12.8 23.2 10.7 7.9 10.7 11.3 Distributed S.E. 1.9 4.0 7.3 3.4 2.5 3.4 3.6 N 10 10 10 10 10 10 10

ASPARTATTRANSFER (U/L) for

WEEK 3 MEAN 104. 110. 103. 101. 102. 104. 111. CIR

% DIFFERENCE 5.8 -1.0 -2.9 -1.9 0.0 6.7 Comment S.D. 16.8 14.7 21.9 15.5 17.7 19.2 18.0 Panel S.E. 5.3 4.6 6.9 4.9 5.6 6.1 5.7 Page 106 of 990 N 10 10 10 10 10 10 10

Book GLUTAMYLTRANSFER (U/L) Only WEEK 3 MEAN 0.2 0.2 0.4 0.2 0.2 0.2 0.4 Page % DIFFERENCE 0.0 100.0 0.0 0.0 0.0 100.0 - S.D. 0.13 0.30 0.54 0.19 0.21 0.19 0.39 Do Not

151 S.E. 0.04 0.09 0.17 0.06 0.07 0.06 0.12 N 10 10 10 10 10 10 10 Quote GLUCOSE (mg/dL) WEEK 3 MEAN 106. 108. 101. 109. 97. 103. 101. % DIFFERENCE 1.9 -4.7 2.8 -8.5 -2.8 -4.7 or

S.D. 9.1 13.1 10.6 10.4 6.8 10.4 4.6 Cite S.E. 2.9 4.1 3.4 3.3 2.2 3.3 1.5 N 10 10 10 10 10 10 10

TABLE 24 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 4 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------CHOLESTEROL (mg/dL) WEEK 3 MEAN 69. 60. 60. 60. 60. 68. 67. % DIFFERENCE -13.0 -13.0 -13.0 -13.0 -1.4 -2.9 S.D. 12.7 15.4 11.3 11.4 14.4 13.0 10.5 Distributed S.E. 4.0 4.9 3.6 3.6 4.6 4.1 3.3 N 10 10 10 10 10 10 10

CALCIUM (mg/dL) for

WEEK 3 MEAN 10.7 10.5 10.6 10.7 10.7 10.8 10.5 CIR

% DIFFERENCE -1.9 -0.9 0.0 0.0 0.9 -1.9 Comment S.D. 0.29 0.22 0.33 0.38 0.42 0.22 0.21 Panel S.E. 0.09 0.07 0.10 0.12 0.13 0.07 0.07 Page 107 of 990 N 10 10 10 10 10 10 10

Book CHLORIDE (mEq/L) Only WEEK 3 MEAN 102. 102. 103. 102. 103. 104. 103. Page % DIFFERENCE 0.0 1.0 0.0 1.0 2.0 1.0 - S.D. 1.8 2.1 1.6 2.3 1.1 1.8 1.6 Do Not

152 S.E. 0.6 0.7 0.5 0.7 0.3 0.6 0.5 N 10 10 10 10 10 10 10 Quote PHOSPHORUS (mg/dL) WEEK 3 MEAN 7.3 7.4 7.1 7.1 7.4 7.4 7.1 % DIFFERENCE 1.4 -2.7 -2.7 1.4 1.4 -2.7 or

S.D. 0.72 0.47 0.67 0.92 0.76 0.75 0.63 Cite S.E. 0.23 0.15 0.21 0.29 0.24 0.24 0.20 N 10 10 10 10 10 10 10

TABLE 24 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 5 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SERUM CHEMISTRY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------POTASSIUM (mEq/L) WEEK 3 MEAN 4.46 4.79 4.45 4.37 4.54 4.61 4.47 % DIFFERENCE 7.4 -0.2 -2.0 1.8 3.4 0.2 S.D. 0.415 0.274 0.362 0.444 0.329 0.353 0.426 Distributed S.E. 0.131 0.087 0.115 0.140 0.104 0.111 0.135 N 10 10 10 10 10 10 10

SODIUM (mEq/L) for

WEEK 3 MEAN 144. 143. 143. 143. 144. 145. 144. CIR

% DIFFERENCE -0.7 -0.7 -0.7 0.0 0.7 0.0 Comment S.D. 2.3 1.5 1.9 2.4 2.3 1.8 1.4 Panel S.E. 0.7 0.5 0.6 0.7 0.7 0.6 0.5 Page 108 of 990 N 10 10 10 10 10 10 10

Book TRIGLYCERIDE (mg/dL) Only WEEK 3 MEAN 41. 41. 42. 37. 42. 45. 42. Page % DIFFERENCE 0.0 2.4 -9.8 2.4 9.8 2.4 - S.D. 11.5 10.4 6.3 6.0 8.1 13.0 7.7 Do Not

153 S.E. 3.6 3.3 2.0 1.9 2.5 4.1 2.4 N 10 10 10 10 10 10 10 Quote ------mg/dL = MILLIGRAMS/DECILITER, U/L = INTERNATIONAL UNIT/LITER, g/dL = GRAMS/DECILITER, mEq/L = MILLIEQUIVALENTS/LITER ------or

TABLE 25 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SPECIAL CHEMISTRY VALUES

------MALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------SORBITOL'GENASE (U/L) WEEK 3 MEAN 13. 13. 15. 12. 13. 16. 13. % DIFFERENCE 0.0 15.4 -7.7 0.0 23.1 0.0 S.D. 3.7 6.2 5.2 3.9 4.7 4.8 5.1 Distributed S.E. 1.2 2.0 1.6 1.2 1.5 1.5 1.6 N 10 10 10 10 10 10 10

------for

U/L = INTERNATIONAL UNIT/LITER CIR

------Comment None significantly different from control group Panel ------Page 109 of 990 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC

Book 7-12.5MG/M3BENZOIC PCPSv5.27 Only 07/15/2009 Page R:07/28/2009 - Do Not 154 Quote or Cite

TABLE 26 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF SPECIAL CHEMISTRY VALUES

------FEMALES ANALYSIS GROUP: 1 2 3 4 5 6 7 ------SORBITOL'GENASE (U/L) WEEK 3 MEAN 15. 18. 14. 16. 15. 13. 13. % DIFFERENCE 20.0 -6.7 6.7 0.0 -13.3 -13.3 S.D. 6.3 5.5 3.0 4.7 6.6 5.8 5.8 Distributed S.E. 2.0 1.8 0.9 1.5 2.1 1.8 1.8 N 10 10 10 10 10 10 10

------for

U/L = INTERNATIONAL UNIT/LITER CIR

------Comment None significantly different from control group Panel ------Page 110 of 990 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC

Book 7-12.5MG/M3BENZOIC Only PCPSv5.27 Page 07/15/2009 - R:07/28/2009 Do Not 155 Quote or Cite

TABLE 27 (WEEK -2 PRETEST EXAMINATION - MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF OPHTHALMIC EXAMINATION FINDINGS

------M A L E ----- GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED EXAM 1(WEEK -2) 10 10 10 10 10 10 10

CORNEAL CRYSTALS - BILATERAL 1 3 0 2 1 4 1 Distributed NO OCULAR LESIONS - BILATERAL 9 7 10 8 9 6 9 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC for

POPSI2v4.02 CIR

07/15/2009 Comment R:08/24/2009 Panel Page 111 of 990 Book Only Page - Do Not 156 Quote or Cite

TABLE 28 (WEEK -2 PRETEST EXAMINATION - FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF OPHTHALMIC EXAMINATION FINDINGS

------F E M A L E ----- GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED EXAM 1(WEEK -2) 10 10 10 10 10 10 10

NO OCULAR LESIONS - BILATERAL 10 10 10 10 10 10 10 Distributed ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC POPSI2v4.02 for

07/15/2009 CIR

R:08/24/2009 Comment Panel Page 112 of 990 Book Only Page - Do Not 157 Quote or Cite

TABLE 29 (WEEK 3 EXAMINATION - MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF OPHTHALMIC EXAMINATION FINDINGS

------M A L E ----- GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED EXAM 2(WEEK 3) 10 10 10 10 10 10 10

CORNEAL CRYSTALS - BILATERAL 3 2 1 1 3 3 2 Distributed NO OCULAR LESIONS - BILATERAL 7 8 9 9 7 7 8 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC for

POPSI2v4.02 CIR

07/15/2009 Comment Panel Page 113 of 990 Book Only Page - Do Not 158 Quote or Cite

TABLE 30 (WEEK 3 EXAMINATION - FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF OPHTHALMIC EXAMINATION FINDINGS

------F E M A L E ----- GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED EXAM 2(WEEK 3) 10 10 10 10 10 10 10

NO OCULAR LESIONS - BILATERAL 10 10 10 9 10 10 10 Distributed CORNEAL CRYSTALS - BILATERAL 0 0 0 1 0 0 0 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC for

POPSI2v4.02 CIR

07/15/2009 Comment Panel Page 114 of 990 Book Only Page - Do Not 159 Quote or Cite

TABLE 31 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MACROSCOPIC FINDINGS

SCHEDULED NECROPSY ------M A L E ----- GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 10 10 10 10 10 10 Distributed

EPIDIDYMIDES -SMALL 1 0 0 0 0 0 0

for

HARDERIAN GLANDS CIR

-PALE 0 1 0 0 0 0 0 Comment

Panel ILEUM Page 115 of 990 -DIVERTICULUM 1 0 0 0 0 0 0

Book JEJUNUM Only -DIVERTICULUM 0 0 0 0 0 0 1 Page - KIDNEYS Do Not

160 -AREA(S), DEPRESSED 0 1 0 0 0 1 0 -DILATED PELVIS 1 1 0 0 0 0 1 -AREA(S), WHITE 0 0 0 0 0 0 1 Quote

LIVER -AREA(S), WHITE 1 0 0 0 0 0 0 or

Cite LN, ILIAC -ENLARGED 0 1 0 0 0 0 0

PROSTATE -MASS 0 0 0 0 0 1 1 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 31 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MACROSCOPIC FINDINGS

SCHEDULED NECROPSY ------M A L E ----- GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 10 10 10 10 10 10 Distributed

PROSTATE -- CONTINUED -SMALL 0 1 0 0 0 0 0

for

SPLEEN CIR

-ACCESSORY 0 0 0 0 0 0 1 Comment

Panel TEETH Page 116 of 990 -FRACTURED 0 0 1 0 0 0 0

Book TESTES Only -SOFT 1 0 0 0 0 0 0 Page - THYMUS Do Not

161 -AREA(S), DARK RED 2 0 0 2 2 3 0 -DISCOLORATION, DARK RED 0 0 1 0 0 0 0 Quote URETERS -DISTENDED 0 1 0 0 0 0 1 -CALCULUS(I) 0 1 0 0 0 0 0 or

Cite URINARY BLADDER -THICKENED 0 1 0 0 0 0 1 -DISTENDED 0 0 0 0 0 0 1 -CALCULUS(I) 0 1 0 0 0 0 1

NO SIGNIFICANT CHANGES OBSERVED - ALL EXAMINED TISSUES 5 8 8 8 8 6 6 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC PGRSI2v4.07 07/15/2009

TABLE 32 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MACROSCOPIC FINDINGS

SCHEDULED NECROPSY ------F E M A L E ----- GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 10 10 10 10 10 10 Distributed

ADRENAL GLANDS -PALE 0 0 0 0 1 0 1

for

KIDNEYS CIR

-DILATED PELVIS 1 0 0 0 0 0 0 Comment -AREA(S), DEPRESSED 0 1 0 0 0 0 0 Panel

Page 117 of 990 LIVER

Book -AREA(S), WHITE 0 0 1 1 0 1 1 Only LN, AXILLARY Page -AREA(S), DARK RED 0 0 0 0 0 0 1 - Do Not

162 LN, MEDIASTINAL -DISCOLORATION, DARK RED 0 0 0 0 0 0 1 Quote OVIDUCTS -CYST(S) 0 0 0 1 0 0 0 or

THYMUS Cite -AREA(S), DARK RED 0 0 0 3 0 1 1 -DISCOLORATION, DARK RED 0 1 0 0 0 0 0

UTERUS -CONTENTS, CLEAR FLUID 1 0 1 5 1 1 2

NO SIGNIFICANT CHANGES OBSERVED - ALL EXAMINED TISSUES 8 8 8 4 8 8 5 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC PGRSI2v4.07 07/15/2009

TABLE 33 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------MALES GROUP: 1 2 3 4 5 6 7 ------FINAL BODY WT (G) MEAN 383. 376. 383. 363. 358. 383. 386. % DIFFERENCE -1.8 0.0 -5.2 -6.5 0.0 0.8 S.D. 49.9 43.5 38.7 26.9 34.7 41.6 38.7 Distributed S.E. 15.8 13.8 12.2 8.5 11.0 13.2 12.2 N 10 10 10 10 10 10 10

ADRENAL GLANDS (G) for

MEAN 0.0684 0.0733 0.0652 0.0718 0.0632 0.0674 0.0694 CIR

% DIFFERENCE 7.2 -4.7 5.0 -7.6 -1.5 1.5 Comment S.D. 0.01290 0.01162 0.01483 0.01439 0.00754 0.00824 0.01571 Panel S.E. 0.00408 0.00367 0.00469 0.00455 0.00238 0.00260 0.00497 Page 118 of 990 N 10 10 10 10 10 10 10

Book ADRENAL GLANDS (G/100 G FINAL BODY WEIGHT) Only MEAN 0.018 0.020 0.017 0.020 0.018 0.018 0.018 Page % DIFFERENCE 11.1 -5.6 11.1 0.0 0.0 0.0 - S.D. 0.0028 0.0029 0.0034 0.0041 0.0030 0.0020 0.0043 Do Not

163 S.E. 0.0009 0.0009 0.0011 0.0013 0.0010 0.0006 0.0014 N 10 10 10 10 10 10 10 Quote ADRENAL GLANDS (G/100 G BRAIN) MEAN 3.469 3.648 3.195 3.551 3.105 3.302 3.504 % DIFFERENCE 5.2 -7.9 2.4 -10.5 -4.8 1.0 or

S.D. 0.6081 0.5526 0.6750 0.7297 0.3873 0.4346 0.7598 Cite S.E. 0.1923 0.1747 0.2135 0.2308 0.1225 0.1374 0.2403 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 33 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------MALES GROUP: 1 2 3 4 5 6 7 ------BRAIN (G) MEAN 1.97 2.01 2.04 2.03 2.04 2.04 1.98 % DIFFERENCE 2.0 3.6 3.0 3.6 3.6 0.5 S.D. 0.059 0.122 0.071 0.113 0.081 0.101 0.083 Distributed S.E. 0.019 0.038 0.022 0.036 0.026 0.032 0.026 N 10 10 10 10 10 10 10

BRAIN (G/100 G FINAL BODY WEIGHT) for

MEAN 0.521 0.541 0.538 0.559 0.572 0.539 0.516 CIR

% DIFFERENCE 3.8 3.3 7.3 9.8 3.5 -1.0 Comment S.D. 0.0600 0.0634 0.0622 0.0314 0.0447 0.0513 0.0369 Panel S.E. 0.0190 0.0200 0.0197 0.0099 0.0141 0.0162 0.0117 Page 119 of 990 N 10 10 10 10 10 10 10

Book EPIDIDYMIDES (G) Only MEAN 1.05 1.15 1.16 1.14 1.15 1.13 1.12 Page % DIFFERENCE 9.5 10.5 8.6 9.5 7.6 6.7 - S.D. 0.084 0.099 0.085 0.062 0.093 0.099 0.082 Do Not

164 S.E. 0.027 0.031 0.027 0.020 0.029 0.031 0.026 N 10 10 10 10 10 10 10 Quote EPIDIDYMIDES (G/100 G FINAL BODY WEIGHT) MEAN 0.279 0.307 0.304 0.314* 0.322** 0.297 0.292 % DIFFERENCE 10.0 9.0 12.5 15.4 6.5 4.7 or

S.D. 0.0377 0.0173 0.0259 0.0257 0.0314 0.0232 0.0271 Cite S.E. 0.0119 0.0055 0.0082 0.0081 0.0099 0.0073 0.0086 N 10 10 10 10 10 10 10

------* = Significantly different from the control group at 0.05 using Dunnett's test ** = Significantly different from the control group at 0.01 using Dunnett's test ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 33 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------MALES GROUP: 1 2 3 4 5 6 7 ------EPIDIDYMIDES (G/100 G BRAIN) MEAN 53.594 57.281 56.797 56.165 56.360 55.403 56.667 % DIFFERENCE 6.9 6.0 4.8 5.2 3.4 5.7 S.D. 3.7822 5.3986 4.8325 3.4156 4.6654 4.3194 3.6127 Distributed S.E. 1.1960 1.7072 1.5282 1.0801 1.4753 1.3659 1.1424 N 10 10 10 10 10 10 10

HEART (G) for

MEAN 1.45 1.37 1.38 1.31 1.32 1.47 1.43 CIR

% DIFFERENCE -5.5 -4.8 -9.7 -9.0 1.4 -1.4 Comment S.D. 0.236 0.138 0.152 0.128 0.139 0.303 0.161 Panel S.E. 0.075 0.044 0.048 0.040 0.044 0.096 0.051 Page 120 of 990 N 10 10 10 10 10 10 10

Book HEART (G/100 G FINAL BODY WEIGHT) Only MEAN 0.380 0.365 0.362 0.361 0.369 0.382 0.371 Page % DIFFERENCE -3.9 -4.7 -5.0 -2.9 0.5 -2.4 - S.D. 0.0364 0.0322 0.0375 0.0263 0.0303 0.0565 0.0312 Do Not

165 S.E. 0.0115 0.0102 0.0119 0.0083 0.0096 0.0179 0.0099 N 10 10 10 10 10 10 10 Quote HEART (G/100 G BRAIN) MEAN 73.768 67.902 67.966 64.855 64.787 71.507 72.166 % DIFFERENCE -8.0 -7.9 -12.1 -12.2 -3.1 -2.2 or

S.D. 10.8674 4.8695 9.0860 6.9948 6.7518 12.2927 7.2888 Cite S.E. 3.4366 1.5399 2.8733 2.2120 2.1351 3.8873 2.3049 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 33 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 4 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------MALES GROUP: 1 2 3 4 5 6 7 ------KIDNEYS (G) MEAN 2.76 2.85 2.68 2.52 2.63 2.79 2.78 % DIFFERENCE 3.3 -2.9 -8.7 -4.7 1.1 0.7 S.D. 0.339 0.472 0.377 0.250 0.296 0.361 0.354 Distributed S.E. 0.107 0.149 0.119 0.079 0.094 0.114 0.112 N 10 10 10 10 10 10 10

KIDNEYS (G/100 G FINAL BODY WEIGHT) for

MEAN 0.725 0.759 0.699 0.693 0.735 0.729 0.722 CIR

% DIFFERENCE 4.7 -3.6 -4.4 1.4 0.6 -0.4 Comment S.D. 0.0485 0.0904 0.0569 0.0523 0.0684 0.0424 0.0744 Panel S.E. 0.0153 0.0286 0.0180 0.0166 0.0216 0.0134 0.0235 Page 121 of 990 N 10 10 10 10 10 10 10

Book KIDNEYS (G/100 G BRAIN) Only MEAN 140.338 141.762 131.494 124.188 129.084 136.322 140.262 Page % DIFFERENCE 1.0 -6.3 -11.5 -8.0 -2.9 -0.1 - S.D. 14.3176 20.4330 19.6029 10.9906 14.8848 14.9838 15.6268 Do Not

166 S.E. 4.5276 6.4615 6.1990 3.4755 4.7070 4.7383 4.9416 N 10 10 10 10 10 10 10 Quote LIVER (G) MEAN 10.45 10.21 10.12 9.68 9.29 10.14 10.56 % DIFFERENCE -2.3 -3.2 -7.4 -11.1 -3.0 1.1 or

S.D. 1.604 1.553 1.544 1.052 1.189 1.404 1.653 Cite S.E. 0.507 0.491 0.488 0.333 0.376 0.444 0.523 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 33 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 5 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------MALES GROUP: 1 2 3 4 5 6 7 ------LIVER (G/100 G FINAL BODY WEIGHT) MEAN 2.727 2.710 2.636 2.661 2.588 2.645 2.733 % DIFFERENCE -0.6 -3.3 -2.4 -5.1 -3.0 0.2 S.D. 0.1119 0.1881 0.1739 0.1714 0.1514 0.1613 0.2229 Distributed S.E. 0.0354 0.0595 0.0550 0.0542 0.0479 0.0510 0.0705 N 10 10 10 10 10 10 10

LIVER (G/100 G BRAIN) for

MEAN 530.425 506.886 497.022 478.265 455.230 495.277 533.081 CIR

% DIFFERENCE -4.4 -6.3 -9.8 -14.2 -6.6 0.5 Comment S.D. 71.5465 64.6129 77.0886 50.4584 48.7902 57.8948 69.0935 Panel S.E. 22.6250 20.4324 24.3776 15.9564 15.4288 18.3080 21.8493 Page 122 of 990 N 10 10 10 10 10 10 10

Book LUNGS (G) Only MEAN 1.43 1.45 1.43 1.38 1.40 1.45 1.45 Page % DIFFERENCE 1.4 0.0 -3.5 -2.1 1.4 1.4 - S.D. 0.144 0.146 0.129 0.117 0.141 0.157 0.100 Do Not

167 S.E. 0.046 0.046 0.041 0.037 0.045 0.050 0.032 N 10 10 10 10 10 10 10 Quote LUNGS (G/100 G FINAL BODY WEIGHT) MEAN 0.377 0.388 0.375 0.380 0.391 0.379 0.378 % DIFFERENCE 2.9 -0.5 0.8 3.7 0.5 0.3 or

S.D. 0.0441 0.0342 0.0386 0.0326 0.0379 0.0271 0.0266 Cite S.E. 0.0139 0.0108 0.0122 0.0103 0.0120 0.0086 0.0084 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 33 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 6 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------MALES GROUP: 1 2 3 4 5 6 7 ------LUNGS (G/100 G BRAIN) MEAN 72.567 72.243 70.100 68.034 68.540 70.687 73.347 % DIFFERENCE -0.4 -3.4 -6.2 -5.5 -2.6 1.1 S.D. 6.2849 6.4201 6.8684 4.7131 6.9406 6.6843 4.0497 Distributed S.E. 1.9875 2.0302 2.1720 1.4904 2.1948 2.1137 1.2806 N 10 10 10 10 10 10 10

SPLEEN (G) for

MEAN 0.73 0.70 0.66 0.66 0.64 0.73 0.76 CIR

% DIFFERENCE -4.1 -9.6 -9.6 -12.3 0.0 4.1 Comment S.D. 0.106 0.127 0.093 0.048 0.127 0.080 0.229 Panel S.E. 0.034 0.040 0.029 0.015 0.040 0.025 0.073 Page 123 of 990 N 10 10 10 10 10 10 10

Book SPLEEN (G/100 G FINAL BODY WEIGHT) Only MEAN 0.191 0.186 0.174 0.182 0.177 0.193 0.198 Page % DIFFERENCE -2.6 -8.9 -4.7 -7.3 1.0 3.7 - S.D. 0.0184 0.0275 0.0276 0.0190 0.0322 0.0318 0.0536 Do Not

168 S.E. 0.0058 0.0087 0.0087 0.0060 0.0102 0.0101 0.0170 N 10 10 10 10 10 10 10 Quote SPLEEN (G/100 G BRAIN) MEAN 37.011 34.578 32.257 32.499 31.159 35.694 38.434 % DIFFERENCE -6.6 -12.8 -12.2 -15.8 -3.6 3.8 or

S.D. 4.6940 5.5354 4.0413 2.8134 6.0162 4.2012 10.6219 Cite S.E. 1.4844 1.7504 1.2780 0.8897 1.9025 1.3285 3.3589 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 33 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 7 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------MALES GROUP: 1 2 3 4 5 6 7 ------TESTES (G) MEAN 3.30 3.31 3.51 3.41 3.37 3.29 3.41 % DIFFERENCE 0.3 6.4 3.3 2.1 -0.3 3.3 S.D. 0.345 0.377 0.257 0.230 0.386 0.282 0.260 Distributed S.E. 0.109 0.119 0.081 0.073 0.122 0.089 0.082 N 10 10 10 10 10 10 10

TESTES (G/100 G FINAL BODY WEIGHT) for

MEAN 0.875 0.881 0.924 0.941 0.943 0.866 0.893 CIR

% DIFFERENCE 0.7 5.6 7.5 7.8 -1.0 2.1 Comment S.D. 0.1453 0.0567 0.0919 0.0919 0.1025 0.0843 0.1026 Panel S.E. 0.0460 0.0179 0.0291 0.0291 0.0324 0.0267 0.0324 Page 124 of 990 N 10 10 10 10 10 10 10

Book TESTES (G/100 G BRAIN) Only MEAN 167.849 164.508 172.400 168.368 165.515 161.131 172.739 Page % DIFFERENCE -2.0 2.7 0.3 -1.4 -4.0 2.9 - S.D. 16.9538 17.4562 14.3738 12.5753 19.9978 11.7078 12.4991 Do Not

169 S.E. 5.3613 5.5201 4.5454 3.9767 6.3239 3.7023 3.9526 N 10 10 10 10 10 10 10 Quote THYMUS (G) MEAN 0.4251 0.4786 0.3985 0.4072 0.4051 0.5207 0.4771 % DIFFERENCE 12.6 -6.3 -4.2 -4.7 22.5 12.2 or

S.D. 0.06368 0.10392 0.10207 0.08381 0.07873 0.09595 0.08408 Cite S.E. 0.02014 0.03286 0.03228 0.02650 0.02490 0.03034 0.02659 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 33 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 8 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------MALES GROUP: 1 2 3 4 5 6 7 ------THYMUS (G/100 G FINAL BODY WEIGHT) MEAN 0.112 0.127 0.105 0.113 0.113 0.136 0.123 % DIFFERENCE 13.4 -6.3 0.9 0.9 21.4 9.8 S.D. 0.0167 0.0207 0.0301 0.0266 0.0165 0.0230 0.0164 Distributed S.E. 0.0053 0.0066 0.0095 0.0084 0.0052 0.0073 0.0052 N 10 10 10 10 10 10 10

THYMUS (G/100 G BRAIN) for

MEAN 21.604 23.875 19.483 20.087 19.855 25.541 24.109 CIR

% DIFFERENCE 10.5 -9.8 -7.0 -8.1 18.2 11.6 Comment S.D. 2.9908 5.2640 4.6346 3.9516 3.5978 5.0648 4.0670 Panel S.E. 0.9458 1.6646 1.4656 1.2496 1.1377 1.6016 1.2861 Page 125 of 990 N 10 10 10 10 10 10 10

Book ------Only None significantly different from control group Page ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC Do Not

170 7-12.5MG/M3BENZOIC POFBSTv5.15 07/15/2009 Quote R:07/28/2009 or Cite

TABLE 34 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------FEMALES GROUP: 1 2 3 4 5 6 7 ------FINAL BODY WT (G) MEAN 225. 217. 221. 213. 218. 223. 217. % DIFFERENCE -3.6 -1.8 -5.3 -3.1 -0.9 -3.6 S.D. 13.7 18.1 15.2 15.5 15.8 13.8 12.6 Distributed S.E. 4.3 5.7 4.8 4.9 5.0 4.4 4.0 N 10 10 10 10 10 10 10

ADRENAL GLANDS (G) for

MEAN 0.0697 0.0663 0.0691 0.0739 0.0679 0.0633 0.0698 CIR

% DIFFERENCE -4.9 -0.9 6.0 -2.6 -9.2 0.1 Comment S.D. 0.00771 0.00723 0.00667 0.01677 0.00773 0.00567 0.00761 Panel S.E. 0.00244 0.00229 0.00211 0.00530 0.00244 0.00179 0.00241 Page 126 of 990 N 10 10 10 10 10 10 10

Book ADRENAL GLANDS (G/100 G FINAL BODY WEIGHT) Only MEAN 0.031 0.031 0.031 0.035 0.031 0.028 0.032 Page % DIFFERENCE 0.0 0.0 12.9 0.0 -9.7 3.2 - S.D. 0.0040 0.0048 0.0038 0.0056 0.0046 0.0015 0.0036 Do Not

171 S.E. 0.0013 0.0015 0.0012 0.0018 0.0014 0.0005 0.0011 N 10 10 10 10 10 10 10 Quote ADRENAL GLANDS (G/100 G BRAIN) MEAN 3.665 3.613 3.718 3.934 3.672 3.384 3.773 % DIFFERENCE -1.4 1.4 7.3 0.2 -7.7 2.9 or

S.D. 0.3502 0.4323 0.3529 0.8186 0.4900 0.3481 0.4339 Cite S.E. 0.1108 0.1367 0.1116 0.2589 0.1549 0.1101 0.1372 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 34 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------FEMALES GROUP: 1 2 3 4 5 6 7 ------BRAIN (G) MEAN 1.90 1.84 1.86 1.87 1.86 1.87 1.85 % DIFFERENCE -3.2 -2.1 -1.6 -2.1 -1.6 -2.6 S.D. 0.050 0.038 0.103 0.066 0.104 0.082 0.092 Distributed S.E. 0.016 0.012 0.032 0.021 0.033 0.026 0.029 N 10 10 10 10 10 10 10

BRAIN (G/100 G FINAL BODY WEIGHT) for

MEAN 0.848 0.852 0.845 0.883 0.856 0.841 0.857 CIR

% DIFFERENCE 0.5 -0.4 4.1 0.9 -0.8 1.1 Comment S.D. 0.0549 0.0627 0.0499 0.0546 0.0758 0.0495 0.0641 Panel S.E. 0.0174 0.0198 0.0158 0.0173 0.0240 0.0157 0.0203 Page 127 of 990 N 10 10 10 10 10 10 10

Book HEART (G) Only MEAN 0.90 0.85 0.88 0.88 0.88 0.86 0.89 Page % DIFFERENCE -5.6 -2.2 -2.2 -2.2 -4.4 -1.1 - S.D. 0.067 0.091 0.095 0.166 0.065 0.098 0.198 Do Not

172 S.E. 0.021 0.029 0.030 0.052 0.020 0.031 0.063 N 10 10 10 10 10 10 10 Quote HEART (G/100 G FINAL BODY WEIGHT) MEAN 0.401 0.393 0.401 0.413 0.404 0.387 0.408 % DIFFERENCE -2.0 0.0 3.0 0.7 -3.5 1.7 or

S.D. 0.0232 0.0356 0.0409 0.0543 0.0315 0.0392 0.0789 Cite S.E. 0.0073 0.0113 0.0129 0.0172 0.0100 0.0124 0.0249 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 34 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------FEMALES GROUP: 1 2 3 4 5 6 7 ------HEART (G/100 G BRAIN) MEAN 47.438 46.286 47.495 47.052 47.407 46.056 48.117 % DIFFERENCE -2.4 0.1 -0.8 -0.1 -2.9 1.4 S.D. 3.1262 4.9547 5.1811 8.1981 4.2620 4.8519 11.6432 Distributed S.E. 0.9886 1.5668 1.6384 2.5925 1.3478 1.5343 3.6819 N 10 10 10 10 10 10 10

KIDNEYS (G) for

MEAN 1.68 1.61 1.62 1.58 1.65 1.67 1.59 CIR

% DIFFERENCE -4.2 -3.6 -6.0 -1.8 -0.6 -5.4 Comment S.D. 0.205 0.072 0.139 0.134 0.121 0.155 0.144 Panel S.E. 0.065 0.023 0.044 0.042 0.038 0.049 0.046 Page 128 of 990 N 10 10 10 10 10 10 10

Book KIDNEYS (G/100 G FINAL BODY WEIGHT) Only MEAN 0.746 0.748 0.737 0.742 0.759 0.745 0.732 Page % DIFFERENCE 0.3 -1.2 -0.5 1.7 -0.1 -1.9 - S.D. 0.0641 0.0598 0.0581 0.0472 0.0557 0.0456 0.0625 Do Not

173 S.E. 0.0203 0.0189 0.0184 0.0149 0.0176 0.0144 0.0198 N 10 10 10 10 10 10 10 Quote KIDNEYS (G/100 G BRAIN) MEAN 88.360 87.761 87.349 84.219 89.198 88.882 85.812 % DIFFERENCE -0.7 -1.1 -4.7 0.9 0.6 -2.9 or

S.D. 10.3789 3.7459 7.0069 5.8392 8.2042 7.7009 8.5277 Cite S.E. 3.2821 1.1846 2.2158 1.8465 2.5944 2.4352 2.6967 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 34 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 4 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------FEMALES GROUP: 1 2 3 4 5 6 7 ------LIVER (G) MEAN 6.29 5.93 6.20 5.87 5.97 6.28 6.06 % DIFFERENCE -5.7 -1.4 -6.7 -5.1 -0.2 -3.7 S.D. 0.589 0.842 0.704 0.631 0.569 0.431 0.414 Distributed S.E. 0.186 0.266 0.223 0.199 0.180 0.136 0.131 N 10 10 10 10 10 10 10

LIVER (G/100 G FINAL BODY WEIGHT) for

MEAN 2.797 2.728 2.812 2.756 2.742 2.814 2.795 CIR

% DIFFERENCE -2.5 0.5 -1.5 -2.0 0.6 -0.1 Comment S.D. 0.1941 0.2184 0.2303 0.2158 0.2110 0.1102 0.1581 Panel S.E. 0.0614 0.0690 0.0728 0.0682 0.0667 0.0348 0.0500 Page 129 of 990 N 10 10 10 10 10 10 10

Book LIVER (G/100 G BRAIN) Only MEAN 331.217 322.649 333.498 312.896 322.701 335.396 327.826 Page % DIFFERENCE -2.6 0.7 -5.5 -2.6 1.3 -1.0 - S.D. 30.5532 45.0645 32.4395 27.8556 36.1780 22.0077 28.5486 Do Not

174 S.E. 9.6618 14.2507 10.2583 8.8087 11.4405 6.9595 9.0278 N 10 10 10 10 10 10 10 Quote LUNGS (G) MEAN 1.05 1.06 1.10 1.06 1.01 1.06 1.02 % DIFFERENCE 1.0 4.8 1.0 -3.8 1.0 -2.9 or

S.D. 0.119 0.091 0.126 0.107 0.076 0.101 0.099 Cite S.E. 0.038 0.029 0.040 0.034 0.025 0.032 0.031 N 10 10 10 10 9 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 34 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 5 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------FEMALES GROUP: 1 2 3 4 5 6 7 ------LUNGS (G/100 G FINAL BODY WEIGHT) MEAN 0.469 0.492 0.500 0.500 0.471 0.474 0.471 % DIFFERENCE 4.9 6.6 6.6 0.4 1.1 0.4 S.D. 0.0530 0.0393 0.0529 0.0527 0.0370 0.0303 0.0315 Distributed S.E. 0.0168 0.0124 0.0167 0.0167 0.0123 0.0096 0.0100 N 10 10 10 10 9 10 10

LUNGS (G/100 G BRAIN) for

MEAN 55.270 57.869 59.179 56.624 55.543 56.630 55.292 CIR

% DIFFERENCE 4.7 7.1 2.4 0.5 2.5 0.0 Comment S.D. 5.2131 3.9945 5.3301 5.3759 5.1288 5.4748 5.9721 Panel S.E. 1.6485 1.2632 1.6855 1.7000 1.7096 1.7313 1.8885 Page 130 of 990 N 10 10 10 10 9 10 10

Book OVARIES/OVIDUCTS (G) Only MEAN 0.1291 0.1179 0.1241 0.1194 0.1240 0.1333 0.1161 Page % DIFFERENCE -8.7 -3.9 -7.5 -4.0 3.3 -10.1 - S.D. 0.01791 0.01248 0.01183 0.02026 0.02282 0.01811 0.01091 Do Not

175 S.E. 0.00566 0.00395 0.00374 0.00641 0.00722 0.00573 0.00345 N 10 10 10 10 10 10 10 Quote OVARIES/OVIDUCTS (G/100 G FINAL BODY WEIGHT) MEAN 0.058 0.055 0.056 0.056 0.057 0.060 0.053 % DIFFERENCE -5.2 -3.4 -3.4 -1.7 3.4 -8.6 or

S.D. 0.0079 0.0065 0.0046 0.0089 0.0118 0.0075 0.0026 Cite S.E. 0.0025 0.0020 0.0015 0.0028 0.0037 0.0024 0.0008 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 34 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 6 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------FEMALES GROUP: 1 2 3 4 5 6 7 ------OVARIES/OVIDUCTS (G/100 G BRAIN) MEAN 6.804 6.415 6.685 6.361 6.721 7.133 6.273 % DIFFERENCE -5.7 -1.7 -6.5 -1.2 4.8 -7.8 S.D. 0.9627 0.6685 0.6775 0.9963 1.3918 1.0664 0.6224 Distributed S.E. 0.3044 0.2114 0.2142 0.3151 0.4401 0.3372 0.1968 N 10 10 10 10 10 10 10

SPLEEN (G) for

MEAN 0.47 0.42 0.45 0.49 0.46 0.48 0.46 CIR

% DIFFERENCE -10.6 -4.3 4.3 -2.1 2.1 -2.1 Comment S.D. 0.056 0.087 0.079 0.113 0.070 0.062 0.061 Panel S.E. 0.018 0.027 0.025 0.036 0.022 0.020 0.019 Page 131 of 990 N 10 10 10 10 10 10 10

Book SPLEEN (G/100 G FINAL BODY WEIGHT) Only MEAN 0.209 0.193 0.203 0.230 0.210 0.216 0.209 Page % DIFFERENCE -7.7 -2.9 10.0 0.5 3.3 0.0 - S.D. 0.0174 0.0353 0.0311 0.0414 0.0300 0.0219 0.0196 Do Not

176 S.E. 0.0055 0.0112 0.0098 0.0131 0.0095 0.0069 0.0062 N 10 10 10 10 10 10 10 Quote SPLEEN (G/100 G BRAIN) MEAN 24.742 22.734 24.147 26.173 24.688 25.772 24.593 % DIFFERENCE -8.1 -2.4 5.8 -0.2 4.2 -0.6 or

S.D. 2.7514 4.5478 4.1612 5.5162 3.9200 3.1400 3.4949 Cite S.E. 0.8701 1.4381 1.3159 1.7444 1.2396 0.9929 1.1052 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 34 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 7 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------FEMALES GROUP: 1 2 3 4 5 6 7 ------THYMUS (G) MEAN 0.3909 0.3881 0.4211 0.3651 0.3766 0.4079 0.3651 % DIFFERENCE -0.7 7.7 -6.6 -3.7 4.3 -6.6 S.D. 0.06098 0.12642 0.09606 0.10811 0.06731 0.08272 0.07424 Distributed S.E. 0.01928 0.03998 0.03038 0.03419 0.02129 0.02616 0.02348 N 10 10 10 10 10 10 10

THYMUS (G/100 G FINAL BODY WEIGHT) for

MEAN 0.174 0.177 0.190 0.170 0.172 0.183 0.168 CIR

% DIFFERENCE 1.7 9.2 -2.3 -1.1 5.2 -3.4 Comment S.D. 0.0256 0.0426 0.0360 0.0420 0.0239 0.0384 0.0331 Panel S.E. 0.0081 0.0135 0.0114 0.0133 0.0075 0.0121 0.0105 Page 132 of 990 N 10 10 10 10 10 10 10

Book THYMUS (G/100 G BRAIN) Only MEAN 20.602 21.073 22.522 19.413 20.345 21.850 19.739 Page % DIFFERENCE 2.3 9.3 -5.8 -1.2 6.1 -4.2 - S.D. 3.2861 6.5598 4.4649 5.4665 3.7763 4.9356 4.1692 Do Not

177 S.E. 1.0392 2.0744 1.4119 1.7287 1.1942 1.5608 1.3184 N 10 10 10 10 10 10 10 Quote UTERUS/CX (G) MEAN 0.66 0.57 0.56 0.65 0.51 0.61 0.55 % DIFFERENCE -13.6 -15.2 -1.5 -22.7 -7.6 -16.7 or

S.D. 0.219 0.216 0.260 0.201 0.201 0.259 0.255 Cite S.E. 0.069 0.068 0.082 0.063 0.064 0.082 0.081 N 10 10 10 10 10 10 10

------None significantly different from control group ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC

TABLE 34 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 8 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF ORGAN WEIGHTS AND RELATIVE ORGAN WEIGHTS

------FEMALES GROUP: 1 2 3 4 5 6 7 ------UTERUS/CX (G/100 G FINAL BODY WEIGHT) MEAN 0.296 0.266 0.254 0.302 0.232 0.274 0.256 % DIFFERENCE -10.1 -14.2 2.0 -21.6 -7.4 -13.5 S.D. 0.1067 0.1023 0.1158 0.0874 0.0839 0.1143 0.1223 Distributed S.E. 0.0337 0.0324 0.0366 0.0276 0.0265 0.0361 0.0387 N 10 10 10 10 10 10 10

UTERUS/CX (G/100 G BRAIN) for

MEAN 34.685 31.232 29.917 34.517 27.533 33.069 29.627 CIR

% DIFFERENCE -10.0 -13.7 -0.5 -20.6 -4.7 -14.6 Comment S.D. 11.5223 11.7106 13.0586 10.7059 11.6316 15.1362 12.7579 Panel S.E. 3.6437 3.7032 4.1295 3.3855 3.6782 4.7865 4.0344 Page 133 of 990 N 10 10 10 10 10 10 10

Book ------Only None significantly different from control group Page ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC Do Not

178 7-12.5MG/M3BENZOIC POFBSTv5.15 07/15/2009 Quote R:07/28/2009 or Cite

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

Distributed ADRENAL CORTEX TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 9 NA NA NA 9 NA 9 -HYPERTROPHY 1 NA NA NA 1 NA 1 for

MINIMAL 1 NA NA NA 1 NA 1 CIR

Comment ADRENAL MEDULLA Panel TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Page 134 of 990 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

Book AORTA Only TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Page EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 - Do Not

179 BRAIN TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Quote

CECUM TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 or

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Cite ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

COLON Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

DUODENUM for

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Comment

Panel EPIDIDYMIDES Page 135 of 990 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10

Book EXAMINED, UNREMARKABLE 7 NA NA NA 7 NA 10 -ASPERMIA 1 NA NA NA 0 NA 0 Only PRESENT 1 NA NA NA NONE NA NONE Page -INFILTRATE, MONONUCLEAR 1 NA NA NA 3 NA 0 - MINIMAL 1 NA NA NA 3 NA NONE Do Not

180 -INFLAMMATION, ACUTE 1 NA NA NA 0 NA 0 MINIMAL 1 NA NA NA NONE NA NONE Quote ESOPHAGUS TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 or

------Cite 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

EYES Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 9 NA NA NA 10 NA 9 -DYSPLASIA, RETINAL 1 NA NA NA 0 NA 1 MINIMAL 1 NA NA NA NONE NA 1 for

CIR

FEMUR Comment TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Panel EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Page 136 of 990

Book HARDERIAN GLANDS TOTAL NUMBER EXAMINED 10 1 NA NA 10 NA 10 Only EXAMINED, UNREMARKABLE 10 1 NA NA 10 NA 10 Page - HEART Do Not

181 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 6 NA NA NA 10 NA 7 -CARDIOMYOPATHY 4 NA NA NA 0 NA 2 Quote MINIMAL 4 NA NA NA NONE NA 2 -INFILTRATE, MONONUCLEAR 0 NA NA NA 0 NA 1 MINIMAL NONE NA NA NA NONE NA 1 or

------Cite 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 4 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

ILEUM Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 9 NA NA NA 10 NA 10 -DIVERTICULUM 1 NA NA NA 0 NA 0 PRESENT 1 NA NA NA NONE NA NONE for

CIR

JEJUNUM Comment TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Panel EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 9 Page 137 of 990 -DIVERTICULUM 0 NA NA NA 0 NA 1

Book PRESENT NONE NA NA NA NONE NA 1 Only JOINT Page TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 - EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Do Not

182 KIDNEYS TOTAL NUMBER EXAMINED 10 2 NA NA 10 1 10 Quote EXAMINED, UNREMARKABLE 4 0 NA NA 3 1 4 -CYST, TUBULAR 1 0 NA NA 0 0 0 PRESENT 1 NA NA NA NONE NA NONE or

-DILATATION, PELVIS 1 1 NA NA 0 0 1 Cite MINIMAL NONE 1 NA NA NONE NA 1 MILD 1 NA NA NA NONE NA NONE ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 5 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

KIDNEYS - CONTINUED Distributed -HYPERPLASIA, UROTHELIAL 0 0 NA NA 0 0 1 MILD NONE NA NA NA NONE NA 1 -HYPOPLASIA, TUBULAR 0 1 NA NA 0 0 0 MINIMAL NONE 1 NA NA NONE NA NONE for

-INFILTRATE, MONONUCLEAR 0 0 NA NA 1 0 1 CIR

MINIMAL NONE NA NA NA 1 NA 1 Comment -INFLAMMATION, SUBACUTE 0 1 NA NA 0 0 1 Panel MILD NONE NA NA NA NONE NA 1 Page 138 of 990 MODERATE NONE 1 NA NA NONE NA NONE

Book -NECROSIS, PAPILLARY 0 1 NA NA 0 0 0 MODERATE NONE 1 NA NA NONE NA NONE Only -NEPHROPATHY, CHRONIC PROGRESSIVE 5 1 NA NA 7 0 5 Page MINIMAL 5 1 NA NA 6 NA 5 - MILD NONE NA NA NA 1 NA NONE Do Not

183 LAC. GLAND EXOR TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Quote EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

LARYNX or

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Cite EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 9 -INFILTRATE, MONONUCLEAR 0 NA NA NA 0 NA 1 MILD NONE NA NA NA NONE NA 1 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 6 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

LIVER Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 3 NA NA NA 3 NA 2 -INFILTRATE, MONONUCLEAR 7 NA NA NA 7 NA 8 MINIMAL 6 NA NA NA 6 NA 7 for

MILD 1 NA NA NA 1 NA 1 CIR

-LIPIDOSIS, TENSION 1 NA NA NA 0 NA 0 Comment PRESENT 1 NA NA NA NONE NA NONE Panel -NECROSIS 0 NA NA NA 1 NA 0 Page 139 of 990 MINIMAL NONE NA NA NA 1 NA NONE

Book LN, AXILLARY Only TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Page EXAMINED, UNREMARKABLE 9 NA NA NA 10 NA 9 - -HYPERPLASIA, LYMPHOID 1 NA NA NA 0 NA 1 Do Not

184 MINIMAL 1 NA NA NA NONE NA 1

LN, BRONCHIAL Quote TOTAL NUMBER EXAMINED 7 NA NA NA 9 NA 9 EXAMINED, UNREMARKABLE 7 NA NA NA 9 NA 9 NOT PRESENT FOR EXAMINATION 3 NA NA NA 1 NA 1 or

------Cite 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 7 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

LN, ILIAC Distributed TOTAL NUMBER EXAMINED NA 1 NA NA NA NA NA EXAMINED, UNREMARKABLE NA 0 NA NA NA NA NA -HYPERPLASIA, LYMPHOID NA 1 NA NA NA NA NA MINIMAL NA 1 NA NA NA NA NA for

CIR

LN, MANDIBULAR Comment TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Panel EXAMINED, UNREMARKABLE 10 NA NA NA 8 NA 8 Page 140 of 990 -HEMORRHAGE 0 NA NA NA 1 NA 0

Book MILD NONE NA NA NA 1 NA NONE -HYPERPLASIA, LYMPHOID 0 NA NA NA 1 NA 2 Only MINIMAL NONE NA NA NA 1 NA 2 Page - LN, MEDIASTINAL Do Not

185 TOTAL NUMBER EXAMINED 10 NA NA NA 9 NA 10 EXAMINED, UNREMARKABLE 9 NA NA NA 7 NA 9 NOT PRESENT FOR EXAMINATION 0 NA NA NA 1 NA 0 Quote -HEMORRHAGE 1 NA NA NA 2 NA 1 MINIMAL 1 NA NA NA 2 NA 1 or

LN, MESENTERIC Cite TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 8 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

LUNGS Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 8 NA NA NA 5 NA 8 -HYPERPLASIA, TYPE II PNEUMOCYTE 0 NA NA NA 1 NA 0 MINIMAL NONE NA NA NA 1 NA NONE for

-HYPERTROPHY, BRONCHIOLAR 0 NA NA NA 1 NA 1 CIR

MINIMAL NONE NA NA NA NONE NA 1 Comment MILD NONE NA NA NA 1 NA NONE Panel -INFILTRATE, EOSINOPHIL 2 NA NA NA 0 NA 1 Page 141 of 990 MINIMAL 2 NA NA NA NONE NA 1

Book -INFILTRATE, MONONUCLEAR 0 NA NA NA 5 NA 2 MINIMAL NONE NA NA NA 5 NA 2 Only

Page MARROW, FEMUR - TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Do Not

186 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

MARROW, STERN Quote TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 or

NASAL LEVEL I Cite TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 7 NA NA NA 8 NA 8 -HYPERPLASIA, SQUAMOUS 1 NA NA NA 2 NA 0 MINIMAL 1 NA NA NA 2 NA NONE ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 9 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

NASAL LEVEL I - CONTINUED Distributed -INFLAMMATION, SUBACUTE 3 NA NA NA 1 NA 2 MINIMAL 2 NA NA NA 1 NA 2 MILD 1 NA NA NA NONE NA NONE

for

NASAL LEVEL II CIR

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Comment EXAMINED, UNREMARKABLE 10 NA NA NA 8 NA 9 Panel -INFLAMMATION, ACUTE 0 NA NA NA 1 NA 0 Page 142 of 990 MINIMAL NONE NA NA NA 1 NA NONE

Book -INFLAMMATION, SUBACUTE 0 NA NA NA 1 NA 0 MINIMAL NONE NA NA NA 1 NA NONE Only -MINERALIZATION 0 NA NA NA 0 NA 1 Page MINIMAL NONE NA NA NA NONE NA 1 - Do Not

187 NASAL LEVEL III TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 9 NA NA NA 10 NA 8 Quote -INFILTRATE, MONONUCLEAR, NASOLACRIMAL DUCT 0 NA NA NA 0 NA 1 MINIMAL NONE NA NA NA NONE NA 1 -INFLAMMATION, SUBACUTE 1 NA NA NA 0 NA 1 or

MINIMAL 1 NA NA NA NONE NA NONE Cite MILD NONE NA NA NA NONE NA 1 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 10 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

NASAL LEVEL IV Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 9 -HYPERPLASIA, LYMPHOID 0 NA NA NA 0 NA 1 MINIMAL NONE NA NA NA NONE NA 1 for

CIR

NASAL LEVEL V Comment TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Panel EXAMINED, UNREMARKABLE 9 NA NA NA 10 NA 10 Page 143 of 990 -INFLAMMATION, NASOLACRIMAL DUCT 1 NA NA NA 0 NA 0

Book MILD 1 NA NA NA NONE NA NONE Only NASAL LEVEL VI Page TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 - EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Do Not

188 NERVE, SCIATIC TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Quote EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

NERVES, OPTIC or

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Cite EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 11 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

PANCREAS Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

PARATHYROIDS for

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Comment

Panel PEYER'S PATCHES Page 144 of 990 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10

Book EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Only PHARYNX Page TOTAL NUMBER EXAMINED 10 NA NA NA 9 NA 10 - EXAMINED, UNREMARKABLE 10 NA NA NA 9 NA 9 Do Not

189 NOT PRESENT FOR EXAMINATION 0 NA NA NA 1 NA 0 -INFILTRATE, MONONUCLEAR 0 NA NA NA 0 NA 1 MINIMAL NONE NA NA NA NONE NA 1 Quote

PITUITARY TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 or

EXAMINED, UNREMARKABLE 10 NA NA NA 9 NA 10 Cite -MALFORMATION, RATHKE'S POUCH 0 NA NA NA 1 NA 0 PRESENT NONE NA NA NA 1 NA NONE ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 12 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

PROSTATE Distributed TOTAL NUMBER EXAMINED 10 1 NA NA 10 1 10 EXAMINED, UNREMARKABLE 8 0 NA NA 7 0 7 -INFILTRATE, MONONUCLEAR 2 0 NA NA 3 0 0 MINIMAL 1 NA NA NA 3 NA NONE for

MILD 1 NA NA NA NONE NA NONE CIR

-INFLAMMATION, SUBACUTE 0 1 NA NA 0 0 2 Comment MINIMAL NONE NA NA NA NONE NA 1 Panel MODERATE NONE 1 NA NA NONE NA 1 Page 145 of 990 -INFLAMMATION, SUPPURATIVE 0 0 NA NA 0 1 1

Book MODERATE NONE NA NA NA NONE NA 1 SEVERE NONE NA NA NA NONE 1 NONE Only

Page RECTUM - TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Do Not

190 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

SAL. GLAND MAND Quote TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 or

SEMINAL VESICLES Cite TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 13 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

SKELETAL MUSCLE Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 9 -NECROSIS 0 NA NA NA 0 NA 1 MINIMAL NONE NA NA NA NONE NA 1 for

CIR

SKIN Comment TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Panel EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Page 146 of 990

Book SPINAL CORD TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Only EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Page - SPLEEN Do Not

191 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Quote STERNEBRAE TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 or

------Cite 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 14 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

STOMACH, GLAN Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 9 NA NA NA 10 NA 10 -INFLAMMATION, ACUTE 1 NA NA NA 0 NA 0 MINIMAL 1 NA NA NA NONE NA NONE for

CIR

STOMACH, NON Comment TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Panel EXAMINED, UNREMARKABLE 9 NA NA NA 10 NA 10 Page 147 of 990 -INFLAMMATION, ACUTE 1 NA NA NA 0 NA 0

Book MINIMAL 1 NA NA NA NONE NA NONE Only TESTES Page TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 - EXAMINED, UNREMARKABLE 9 NA NA NA 10 NA 9 Do Not

192 -APLASIA 1 NA NA NA 0 NA 0 PRESENT 1 NA NA NA NONE NA NONE -MINERALIZATION 0 NA NA NA 0 NA 1 Quote MINIMAL NONE NA NA NA NONE NA 1

THYMUS or

TOTAL NUMBER EXAMINED 10 NA 1 2 10 3 10 Cite EXAMINED, UNREMARKABLE 9 NA 1 0 8 0 10 -HEMORRHAGE 1 NA 0 2 2 3 0 MINIMAL 1 NA NA 2 2 3 NONE ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 15 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

THYROID GLANDS Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

TRACHEA for

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Comment

Panel URETERS Page 148 of 990 TOTAL NUMBER EXAMINED NA 1 NA NA NA NA 1

Book EXAMINED, UNREMARKABLE NA 0 NA NA NA NA 0 -DILATATION NA 1 NA NA NA NA 0 Only MINIMAL NA 1 NA NA NA NA NA Page -HYPERPLASIA, UROTHELIAL NA 0 NA NA NA NA 1 - MINIMAL NA NA NA NA NA NA 1 Do Not

193 -INFILTRATE, MONONUCLEAR NA 0 NA NA NA NA 1 MINIMAL NA NA NA NA NA NA 1 Quote URINARY BLADDER TOTAL NUMBER EXAMINED 10 1 NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 0 NA NA 10 NA 9 or

-CRYSTALS 0 1 NA NA 0 NA 1 Cite PRESENT NONE 1 NA NA NONE NA 1 -HYPERPLASIA, UROTHELIAL 0 1 NA NA 0 NA 1 SEVERE NONE 1 NA NA NONE NA 1 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 35 (MALES) PROJECT NO.:WIL-703002M 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 16 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- MALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 2 1 2 10 4 10

URINARY BLADDER - CONTINUED Distributed -INFLAMMATION, SUBACUTE 0 0 NA NA 0 NA 1 MILD NONE NA NA NA NONE NA 1 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC for

7-12.5MG/M3BENZOIC CIR

NA = NOT APPLICABLE Comment PHSI2v4.30 Panel 07/15/2009 Page 149 of 990 Book Only Page - Do Not 194 Quote or Cite

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 1 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

Distributed ADRENAL CORTEX TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

for

ADRENAL MEDULLA CIR

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Comment EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Panel

Page 150 of 990 AORTA

Book TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Only

Page BRAIN - TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Do Not

195 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

CECUM Quote TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 or

CERVIX Cite TOTAL NUMBER EXAMINED 10 NA 1 5 10 1 10 EXAMINED, UNREMARKABLE 10 NA 0 0 9 0 10 -ESTROUS CYCLE: ESTRUS 0 NA 0 1 0 0 0 PRESENT NONE NA NA 1 NONE NA NONE ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 2 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

CERVIX - CONTINUED Distributed -ESTROUS CYCLE: PROESTRUS 0 NA 1 4 0 1 0 PRESENT NONE NA 1 4 NONE 1 NONE -MUCIFICATION 0 NA 0 0 1 0 0 MODERATE NONE NA NA NA 1 NA NONE for

CIR

COLON Comment TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Panel EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Page 151 of 990

Book DUODENUM TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Only EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Page - ESOPHAGUS Do Not

196 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Quote EYES TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 8 NA NA NA 9 NA 10 or

-DYSPLASIA, RETINAL 2 NA NA NA 1 NA 0 Cite MINIMAL 2 NA NA NA NONE NA NONE SEVERE NONE NA NA NA 1 NA NONE ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 3 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

FEMUR Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

HARDERIAN GLANDS for

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Comment

Panel HEART Page 152 of 990 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10

Book EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 9 -CARDIOMYOPATHY 0 NA NA NA 0 NA 1 Only MINIMAL NONE NA NA NA NONE NA 1 Page - ILEUM Do Not

197 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Quote JEJUNUM TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 or

------Cite 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 4 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

JOINT Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

KIDNEYS for

TOTAL NUMBER EXAMINED 10 1 NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 7 0 NA NA 7 NA 7 Comment -CYST, TUBULAR 1 0 NA NA 1 NA 0 Panel PRESENT 1 NA NA NA 1 NA NONE Page 153 of 990 -INFILTRATE, MONONUCLEAR 0 1 NA NA 0 NA 0

Book MILD NONE 1 NA NA NONE NA NONE -NEPHROPATHY, CHRONIC PROGRESSIVE 3 1 NA NA 2 NA 3 Only MINIMAL 3 NA NA NA 1 NA 2 Page MILD NONE 1 NA NA 1 NA 1 - Do Not

198 LAC. GLAND EXOR TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Quote

LARYNX TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 or

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Cite ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 5 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

LIVER Distributed TOTAL NUMBER EXAMINED 10 NA 1 1 10 1 10 EXAMINED, UNREMARKABLE 5 NA 1 0 6 0 6 -INFILTRATE, MONONUCLEAR 5 NA 0 0 4 0 4 MINIMAL 5 NA NA NA 4 NA 4 for

-LIPIDOSIS, TENSION 0 NA 0 1 0 1 1 CIR

MINIMAL NONE NA NA NA NONE 1 1 Comment MILD NONE NA NA 1 NONE NA NONE Panel

Page 154 of 990 LN, AXILLARY

Book TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 8 Only -HEMORRHAGE 0 NA NA NA 0 NA 2 Page MINIMAL NONE NA NA NA NONE NA 2 - Do Not

199 LN, BRONCHIAL TOTAL NUMBER EXAMINED 9 NA NA NA 6 NA 9 EXAMINED, UNREMARKABLE 9 NA NA NA 6 NA 9 Quote NOT PRESENT FOR EXAMINATION 1 NA NA NA 4 NA 1

LN, MANDIBULAR or

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Cite EXAMINED, UNREMARKABLE 10 NA NA NA 7 NA 9 -HYPERPLASIA, LYMPHOID 0 NA NA NA 3 NA 1 MINIMAL NONE NA NA NA 3 NA 1 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 6 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

LN, MEDIASTINAL Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 9 -HEMORRHAGE 0 NA NA NA 0 NA 1 MINIMAL NONE NA NA NA NONE NA 1 for

CIR

LN, MESENTERIC Comment TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Panel EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Page 155 of 990

Book LUNGS TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Only EXAMINED, UNREMARKABLE 10 NA NA NA 9 NA 8 Page -INFILTRATE, EOSINOPHIL 0 NA NA NA 1 NA 2 - MINIMAL NONE NA NA NA NONE NA 2 Do Not

200 MILD NONE NA NA NA 1 NA NONE -INFILTRATE, MONONUCLEAR 0 NA NA NA 1 NA 2 MINIMAL NONE NA NA NA NONE NA 2 Quote MILD NONE NA NA NA 1 NA NONE

MAMMARY GLAND or

TOTAL NUMBER EXAMINED 9 NA NA NA 9 NA 10 Cite EXAMINED, UNREMARKABLE 9 NA NA NA 9 NA 10 NOT PRESENT FOR EXAMINATION 1 NA NA NA 1 NA 0 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 7 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

MARROW, FEMUR Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

MARROW, STERN for

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Comment

Panel NASAL LEVEL I Page 156 of 990 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10

Book EXAMINED, UNREMARKABLE 9 NA NA NA 6 NA 10 -HYPERPLASIA, SQUAMOUS 0 NA NA NA 2 NA 0 Only MINIMAL NONE NA NA NA 2 NA NONE Page -INFILTRATE, MONONUCLEAR 1 NA NA NA 2 NA 0 - MINIMAL 1 NA NA NA 2 NA NONE Do Not

201 -INFLAMMATION, SUBACUTE 0 NA NA NA 2 NA 0 MINIMAL NONE NA NA NA 2 NA NONE Quote NASAL LEVEL II TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 9 NA 10 or

-INFILTRATE, MONONUCLEAR 0 NA NA NA 1 NA 0 Cite MINIMAL NONE NA NA NA 1 NA NONE ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 8 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

NASAL LEVEL III Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

NASAL LEVEL IV for

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Comment

Panel NASAL LEVEL V Page 157 of 990 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10

Book EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Only NASAL LEVEL VI Page TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 - EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Do Not

202 NERVE, SCIATIC TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Quote EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

NERVES, OPTIC or

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 9 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

OVARIES Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

OVIDUCTS for

TOTAL NUMBER EXAMINED 10 NA NA 1 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA 1 10 NA 10 Comment

Panel PANCREAS Page 158 of 990 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10

Book EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Only PARATHYROIDS Page TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 9 - EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 9 Do Not

203 NOT PRESENT FOR EXAMINATION 0 NA NA NA 0 NA 1

PEYER'S PATCHES Quote TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 or

PHARYNX Cite TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 10 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

PITUITARY Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

RECTUM for

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Comment

Panel SAL. GLAND MAND Page 159 of 990 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10

Book EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Only SKELETAL MUSCLE Page TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 - EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Do Not

204 SKIN TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 Quote EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

SPINAL CORD or

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 11 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

SPLEEN Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

STERNEBRAE for

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Comment

Panel STOMACH, GLAN Page 160 of 990 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10

Book EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Only STOMACH, NON Page TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 - EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Do Not

205 THYMUS TOTAL NUMBER EXAMINED 10 NA NA 3 10 1 10 Quote EXAMINED, UNREMARKABLE 10 NA NA 1 10 0 9 -HEMORRHAGE 0 NA NA 2 0 1 1 MINIMAL NONE NA NA 2 NONE 1 1 or

------Cite 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 12 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

THYROID GLANDS Distributed TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10

TRACHEA for

TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 CIR

EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Comment

Panel URINARY BLADDER Page 161 of 990 TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10

Book EXAMINED, UNREMARKABLE 10 NA NA NA 10 NA 10 Only UTERUS Page TOTAL NUMBER EXAMINED 10 NA 1 5 10 1 10 - EXAMINED, UNREMARKABLE 9 NA 0 0 9 0 8 Do Not

206 -DILATATION, LUMEN 1 NA 1 5 1 1 2 MINIMAL NONE NA NA NA NONE NA 1 MILD 1 NA 1 5 1 1 1 Quote

VAGINA TOTAL NUMBER EXAMINED 10 NA NA NA 10 NA 10 or

EXAMINED, UNREMARKABLE 0 NA NA NA 0 NA 0 Cite -ESTROUS CYCLE: DIESTRUS 4 NA NA NA 7 NA 3 PRESENT 4 NA NA NA 7 NA 3 ------1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE

TABLE 36 (FEMALES) PROJECT NO.:WIL-703002F 4-WEEK INHALATION STUDY OF BENZYL ALCOHOL & BENZOIC ACID IN RATS PAGE 13 SPONSOR:PERSONAL CARE PRODUCTS SUMMARY OF MICROSCOPIC FINDINGS

----- FEMALE ------GROUP: 1 2 3 4 5 6 7 ------NUMBER OF ANIMALS IN DOSE GROUP 10 10 10 10 10 10 10 NUMBER OF ANIMALS EXAMINED WEEK 3 10 1 2 6 10 2 10

VAGINA - CONTINUED Distributed -ESTROUS CYCLE: ESTRUS 2 NA NA NA 1 NA 4 PRESENT 2 NA NA NA 1 NA 4 -ESTROUS CYCLE: PROESTRUS 4 NA NA NA 1 NA 3 MODERATE NONE NA NA NA 1 NA NONE for

PRESENT 4 NA NA NA NONE NA 3 CIR

-MUCIFICATION 0 NA NA NA 1 NA 0 Comment MODERATE NONE NA NA NA 1 NA NONE Panel ------Page 162 of 990 1- 0 MG/M3 2-30 MG/M3 BENZYL 3-100 MG/M3 BENZYL 4-300 MG/M3 BENZYL 5-1000MG/M3 BENZYL 6-2.5MG/M3 BENZOIC

Book 7-12.5MG/M3BENZOIC NA = NOT APPLICABLE Only PHSI2v4.30 Page 07/15/2009 - Do Not 207 Quote or Cite Distributed for Comment Only - Do Not Quote or Cite

PersonalCare ProductsCounci Committedto Safety, Quality& Innovation

Memorandum

TO: F. Alan Andersen, Ph.D. Director - COSMETIC INGREDIENT REV]EW (CIR)

FROM: John Bailey, Ph.D. Industry Liaison to the CIR Expert Panel

DATE: October 28, 2010

SUBJECT: RIFM Synopsis on Benzyl Benzoate

Research Institute for Fragrance Materials. 2010. Benzyl Benzoate Synopsis.

1101 17th N.W., Suite Washington, D.C. 20036-4702 (fax) www.personalcarecouncil.org Street, 300 202.331.1770 202.331.1969 CIR Panel Book Page 208 Benzyl benzoate Distributed for Comment Only - Do Not Quote or Cite Page 1 of 67

DATA IN THIS SYNOPSIS HAVE NOT BEEN PEER REVIEWED BY A SCIENTIFIC PANEL Benzyl benzoate

Synonyms Benylate Benzoic acid, benzyl ester Benzoic acid, phenylmethyl ester CAS 120-51-4 Benzyl benzoate Principal EINECS FMA RIFM ‘!J Benzyl phenylformate Phenylmethyl benzoate

CAS Number RIFM ID FEMA EINECS Registration

120-51-4 108 2138 204-402-9 EINECS DSL TSCA

RIFM Monograph: 108 (Published 1973: FCT,vll,p1015 (Binder, p129))

Fragrance Structure-Activity Group: Esters/Aryl Alkyl Alcohol Aryl Alkyl Acid/Benzoic Acid Derivatives (Benzoates) Formula 02CH1412 Structure -C0-CH Molecular Weight 212.25 SMILES Notation O=C(OCc(cccc 1)cCH-C1)c(cccc2)c2 Generic Class Aromatic Esters 652 Description A clear, coloreslss to very pale yellow liquid having a slight aromatic odor.

Physical Data

Acid Value (XV.B) 1.0 Max FMA 5.0 g. Boiling Point 323 LIC FMA Boiling Point (calculated) 317.89 DC EPI Suite Congealing Point (I.E) 18DC FMA Flash Point >212E1F;CC FMA Henrys Law (calculated) 2.802e-006 /molPaEm EPI Suite Log 0K (calculated) 3.54 3 EPI Suite Melting Point (calculated) 70.75 DC EPI Suite as Purity (X.B.2.b) 99.0 Mm. FMA

Refractive Index @ 2OLIC 1.568 to 1.570 FMA (1.B) Specific Gravity 2OLIC(I.A) 1.118 - 1.122 FMA Specific Gravity 25L1C(I.A) 1.116 - 1.120 FMA

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Vapor Pressure <0.00 1 mm Hg 20C FMA 0.000555 mm Hg @ Vapor Pressure (calculated) EPI Suite 25 DC Water Solubility (Flask 15.3 mg/L at 20 +1-0.5 HaarmannReimer, 1992u Method) C Water Solubility (calculated) 15.39 mgIL EPI Suite

PreparationBy the interaction of sodium benzoate and benzyl chloride (Bedoukian,1967) Natural Occurrence l[Benzyl benzoate is reported to occur in nature. Use LevelsilIn public use since the 1900s.

Flavor Consumption (in kg)

[1995 IIEUROPE 11131051 1995 USA 11318271 1987 USA 12701 I 1982 USA 28801

1976 USA 01 1975 USA ( 11112001 1970 USA fl78901

Uses (in ppm)

Average Average Mean Daily Product . . Updated Usual Maximum Consumption (gms) FAlcoholic Beverage 29.6711 98.91 32.51121-Jul-881 IBakedGoods 45.8411 99.41 137.21121-Jul-881 iChewing Gum 6.4811 6.4811 0.21121-Jul-881 IFrozenDairy 22.77 69.6711 25.61121-Jul-881 IGelatinPudding 31.48 72.611 20.41121-Jul-881

Gravies 10 2011 8.31121-Jul-88] IHardCandy 3.1 49.6111 0.61121-Jul-88] [t Products l.3 2.5511 78.41121-Jul-881 INon-alcoholic Beverage 11.3811 33.0411 104.01121-Jul-881 Candy ISoft II 39.981189.7311 5.81121-Jul-881 PAD! 10.0810.0810.0810.0810.0810.0810.0810.0810.0810.08

Status

Benzyl benzoate was included by the Council of Europe in the list of substances granted A - may be used in foodstuffs (COE No. 262) , was approved by the FDA

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as a flavor ( 21 CFR 172.5 15) , Has an IFRA Standard See.

Flavor and Extract Manufacturers’ Association states: Generally Recognized as Safe as a flavor ingredient - GRAS 3. (2138) Hall,1965

Indicative Non-Exhaustive List states: Listed

The Industrial Safety and Health Law (Japan) states: ISHL Number (4-(7)-200)

Joint Expert Committee on Food Additives states: Group ADI 0-5 mg/kg for the benzyllbenzoic moiety, representing total benzoate for all food-additive sources. (2004)

Joint Expert Committee on Food Additives states: The Joint FAQ/WHO Expert Committee on Food Additives (JECFA) concluded that the substance does not present a safety concern at current levels of intake when used as a flavouring agent. (24)

Joint Expert Committee on Food Additives states: The Benzyl Derivatives Safety Evaluation is available here.

Joint Expert Committee on Food Additives states: The JECFA Evaluation Summary is available here.

United Nations Transport Classification Codes states: 9; III; 3082

FFIDS Volume VI Updated 1-Nov-85

European Hazard Classification Labeling I R22 Harmful if swallowed. I R51/53 “Toxic to aquatic organisms, may cause long-term adverse effects in the aquatic environment.I S(2) Keep out of the reach of children. (When material is sold to the general public) S25 Avoid contact with eyes. S46 If swallowed, seek medical advice immediately and show this container or label. S61 Avoid release to the environment. Refer to special instructions/Safety data sheets. Harmful Dangerous for the environment US OSHA Health Hazard Statements Acute Oral May be harmful if swallowed. 1-Jan-00 Target Organ 28 Prolonged or repeated dermal exposure may cause damage to the 1-Jan-00 Effects skin and kidneys.

Global Harmonized System Hazard Statements I Hazard CategorySignal WordlICodell Hazard Statement I

I I I II I

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EH C2 1411 IToxic to aquatic life with long lasting effects EH A2 to I aquatic I 1H401llToxic life ATO 4 Warning 1H302IHarmful if swallowed

Precautionary Precautionary Statement Code

P264 Wash .. (hands/face) thoroughly after handling P270 Do not eat, drink or smoke when using this product P273 Avoid release to the environment IF SWALLOWED: call a POISON CENTER or doctor/physician if you feel P301/312 unwell P330 Rinse mouth P391 Collect spillage Dispose of contents/container to ... (in accordance with P501 local/regional/national/international regulation).

Human Health Data

Acute toxicity Route: gavage. Species: rat. A group of 10 Sprague-Dawley albino rats (5 male and 5 female) were used. Animals were fasted overnight prior to dosing and for approximately 2 hours after dosing. Initial body weights were 119-148 grams for males and 118-148 grams for females. All animals received one dose of 5.0 mi/kg test material in arachis oil B.P. Animals were observed for mortality and/or systemic effects at 30 minutes, 1, 2, 3, 4 and 5 hours and then once daily for 14 days. In addition, individual bodyweights were recorded on days 0, 7 and 14. Surviving animals were sacrificed on Day 14. Gross necropsies were conducted on all animals. Summary The acute oral LD5O for combined sexes > 2000 mg/kg body weight based on no mortality observed at this dose level. 2000 mg/kg clinical signs, 0/10 deaths. Dose was mg/kg bodyweight. Clinical signs observed were hunched posture, lethargy, piloerection, ptosis and decreased respiratory rate in 10/10 animals; occasional tremors were observed in 4/10 animals (3 males and 1 female). Recovery of all animals was complete by Day 4. Necropsy revealed congestion of the lungs in 1/10 female animals and a pale area on the glandular region of the stomach in 1/10 male animals. Necropsy was normal for the remaining 8/10 animals. (HaarmannReimer, 1985) Route: oral. Species: rat. In an acute oral toxicity study, the LD5O of the test material in rats was analyzed by using various parameters of the test material, and was reported. No further details were provided. Article in Japanese. LD5O 1700 MG/KG 1700 mg/kg calculated LD5O. (Nishimura,1994a)

Route: gavage. Species: rat. An acute oral toxicity study was conducted using five groups of ten Sprague Dawley rats (5 Male and 5 Female) dosed orally at 1000, 1600, 2000, 3200 and 4000 mg/kg (four doses separated by one-half hour intervals; example, 1000 mg/kg - 4 doses 250 mg/kg, 40 mi/kg volume). Animals were fasted 18 hours prior to dosing. Initial weights ranged from 180 - 280 grams after fasting. Animals were housed individually in stainless steel 1/2” wire mesh cages. Animals received Wayne Lab Blox and water ad libitum. Light cycle was 12 hours lightll2 hours dark; temperature was maintained at 22 +/- 3 C with a humidity of 30 to 70%. Acclimation period was 5 days. The rats were observed immediately, and at one, four and 24 hrs after dosing and twice daily for

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14 days for pharmacotoxic, CNS effects and mortality. Body weights were recorded on the 14th day. Surviving rats were sacrificed by C02 inhalation and a gross necropsy performed. Additional Acute Oral Toxicity Data were reported at the end of this study on: a) 50% Benzyl Benzoate in polyethylene granules (0.5-1.0 mm diameter) as a suspension in 1% gum Tragacenth. The LD5O> 10 mg/kg body weight. b) Polyethylene granules in a 1% Gum Tragacanth suspension. The LD5O> 10 mg/kg body weight. c) Benzyl Benzoate (neat) in 1% Gum Tragacanth suspension. The LD5Owas 1.16 g/kg body weight. The incorporation of fragrance material Benzyl Benzoate in plastic polyethylene beads provides at least a 10 fold saftey factor in No-lethal effect levels (NLEL) (5.0 g/kg vs. 0.5 g/kg) when compared to reported data on the same material tested at 100%. Subjects: 25 Male 25 Female Study Length: 14 days (Litchfield and Wilcoxon.) The vehicle was 0.25% methylcellulose plus one drop of 5% Tween 80 solution for each 5 ml of vehicle.LD5O 1550 mg/kg with 95% confidence limits of 1260 to 1907 mg/kg. Summary Based upon the results of the Acute Oral Toxicity Study in Rats, the calculated acute oral LD5Owas determined to be 1550 mg/kg with 95% confidence limits of 1260 and 1907 mg/kg. The calculated acute oral LD5Ofor females was determined to be 1230 mg/kg with 95% confidence limits of 932 to 1624 mg/kg. The LD5Ofor males was not calculated due to Litchfield and Wilcoxon method restrictions. 2000 mg/kg clinical signs, lethal, 10/10 deaths. 4/5 deaths occurred in male and female rats on day 1, and 1/5 deaths occurred in male and female rats on day 2. Clinical signs included piloerection, arched back, decreased activity, salivation, chromaturia, diarrhea, decreased body tone, clonic-tonic convulsions, hopping convulsions, tremors, body drop, prostration, chromodacryorrhea, clonic convulsions, semiprostration, polyuria, abnormal gait, flaccid body tone, exophthalmus, hypersensitivity and poor grooming. Necropsy of the animals dying on study revealed distention of the gastrointestinal tract, hemorrhages in the lobes of the lungs and in the thymus, small erosions or ulcers in the glandular mucosa of the stomach, perforation in the pyloric section of the stomach, ascitic fluid, congested lungs with adhesions, discoloration of the genital-anal and oral region and discoloration of the thymus, intestines and pancreas. Terminal necropsy revealed congested lungs and slight discoloration of the thymus. 1000 mg/kg clinical signs, lethal, 1/10 deaths. One death occurred in female rats on day 2. Clinical signs included piloerection, arched back, decreased activity, salivation, chromaturia, diarrhea, decreased body tone, clonic-tonic convulsions, hopping convulsions, tremors, body drop, prostration, chromodacryorrhea, clonic convulsions, semiprostration, polyuria, abnormal gait, flaccid body tone, exophthalmus, hypersensitivity and poor grooming. Necropsy of the animals dying on study revealed distention of the gastrointestinal tract, hemorrhages in the lobes of the lungs and in the thymus, small erosions or ulcers in the glandular mucosa of the stomach, perforation in the pyloric section of the stomach, ascitic fluid, congested lungs with adhesions, discoloration of the genital-anal and oral region and discoloration of the thymus, intestines and pancreas. Terminal necropsy revealed congested lungs and slight discoloration of the thymus. 1600 mg/kg clinical signs, lethal, 5/10 deaths. 3/5 deaths occurred in female rats on day 1, and 1/5 deaths on day 2. 1/5 deaths occurred in male rats on day 1. Clinical signs included piloerection, arched back, decreased activity, salivation, chromaturia, diarrhea, decreased body tone, clonic-tonic convulsions, hopping convulsions, tremors, body drop, prostration, chromodacryorrhea, clonic convulsions, semiprostration, polyuria, abnonnal gait, flaccid body tone, exophthalmus, hypersensitivity and poor grooming. Necropsy of the animals dying on study revealed distention of the gastrointestinal tract, hemorrhages in the lobes of the lungs and in the thymus, small erosions or ulcers in the glandular mucosa of the stomach, perforation in the pyloric section of the stomach, ascitic fluid, congested lungs with adhesions, discoloration of the genital-anal and oral region and discoloration of the thymus, intestines and pancreas. Terminal necropsy revealed congested lungs and slight discoloration of the thymus. 3200 mg/kg clinical signs, lethal, 10/10 deaths. 4/5 deaths occurred in male and female rats on day 1, and 1/5 deaths occurred in male and female rats on day 2. Clinical signs included piloerection, arched back, decreased activity, salivation, chromaturia, diarrhea, decreased body tone, clonic-tonic convulsions, hopping convulsions, tremors, body drop, prostration, chromodacryorrhea, clonic convulsions, semiprostration, polyuria, abnormal gait, flaccid body tone, exophthalmus, hypersensitivity and poor grooming. Necropsy of the animals dying on study revealed distention of the gastrointestinal tract, hemorrhages in the lobes of the

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lungs and in the thymus, small erosions or ulcers in the glandular mucosa of the stomach, perforation in the pyloric section of the stomach, ascitic fluid, congested lungs with adhesions, discoloration of the genital-anal and oral region and discoloration of the thymus, intestines and pancreas. Terminal necropsy revealed congested lungs and slight discoloration of the thymus. 4000 mg/kg clinical signs, lethal, 10/10 deaths. 5/5 deaths occurred in female rats on day 1, 4/5 deaths occurred in male rats on day 1, and 1/5 deaths occurred in male rats on day 2. Clinical signs included piloerection, arched back, decreased activity, salivation, chromaturia, diarrhea, decreased body tone, clonic-tonic convulsions, hopping convulsions, tremors, body drop, prostration, chromodacryorrhea, clonic convulsions, semiprostration, polyuria, abnormal gait, flaccid body tone, exophthalmus, hypersensitivity and poor grooming. Necropsy of the animals dying on study revealed distention of the gastrointestinal tract, hemorrhages in the lobes of the lungs and in the thymus, small erosions or ulcers in the glandular mucosa of the stomach, perforation in the pyloric section of the stomach, ascitic fluid, congested lungs with adhesions, discoloration of the genital-anal and oral region and discoloration of the thymus, intestines and pancreas. Terminal necropsy revealed congested lungs and slight discoloration of the thymus. 1230 mg/kg calculated LD5O,calculated acute oral LD5Ofor females was determined to be 1230 mg/kg with 95% confidence limits of 932 to 1624 mg/kg. 1550 mg/kg calculated LD5O, calculated acute oral LD5Ofor male and female rats was determined to be 1550 mg/kg with 95% confidence limits of 1260 and 1907 mg/kg. (IFF,1982a)

Route: oral. Species: rat. An acute oral toxicity study was conducted using 1 group of 5 male and 5 female Sprague Dawley rats dosed orally at 10 gm/kg in four doses of 2500 mg/kg separated by 1/2 hour intervals. Animals were fasted 18 hours prior to dosing. Initial weights ranged from 180 - 280 grams after fasting. Animals were housed individually in stainless steel 1/2” wire mesh cages. Animals received Wayne Lab Blox and water ad libitum. Light cycle was 12 hours light/12 hours dark; temperature was maintained at 22 +/- 3 C with a humidity of 30 tO70%. Acclimation period was 5 days. Animals were observed immediately and at one, four, 24 hours after dosing twice daily for fourteen days for pharmacotoxic, CNS effects and mortality. Surviving rats were sacrificed by C02 inhalation and a gross necropsy performed. Dose levels were based on the results of an associated dose range-finding study. Subjects: 5 Male 5 Female Study Length: 14 days (Acute Oral Toxicity Study in Rats 14 day) Gum Tragacanth (1%) was vehicle plus one drop of 5% Tween 80 solution for each 5 ml of vehicle.Summary Based upon the results from the Acute Oral Toxicity Study in Rats, the estimated acute oral LD5Ofor the test substance was determined to be greater than 10 gmfkg. 10 g/kg clinical signs, Signs observed were: decreased activity, piloerection, abnormal gait, diarrhea, decreased body tone, exophthalmus, hypersensitivity, poor grooming, and chromodacryorrhea. 0/10 rats died at the 10 gmfkg dose level. Terminal necropsy revealed congestion, abscesses and hemorrhages in the lungs and hemorrhages in the thymus. No visible lesions were observed in the remaining animals. (IFF,1982b)

Route: oral. Species: rat. Preliminary: Four animals (2 Male and 2 Female) were administered the test substance at 10 gm/kg. To achieve this, four 2500 mg/kg doses were given at 30 minute intervals. Results: No signs of mortality were observed during the dose-range-finding study. Acute Oral Study: The test substance to ten mice (5 Male & 5 Female) by oral gavage at a total dose of 10 gmfkg. Four doses of 2500 mg/kg were given every 30 minutes. Body weights were measured initially and on day 14 of the study. Observations were made immediately and at 1,4 and 24 hrs. following dosing and then twice daily for the remainder of the study. 1% Gum Tragacanth was used as a vehicle, plus 1 drop of 5% Tween 80 for each 5 ml. of vehicle. A discussion on the variation in LD5O ‘srecorded, using various test substance vehicles, is given at the end of this report. Subjects: 5 Male 5 Female Study Length: 14 days LC5O >10 gmfkgSummary Based upon the results from the Acute Oral Toxicity study in 10 rats, the estimated acute oral LD5Ofor the test substance was determined to be greater than 10 gm/kg. 10 g/kg calculated LD5O,Signs observed during the study were decreased activity, piloerection, abnormal gait, exophthalmus, decreased body tone, abnormal stance, tremors, hypersensitivity, body drop, poor grooming, and diarrhea. 0/10 animals died during the study. Terminal necropsy revealed hemorhages in the lungs and thymus. No visible lesions were observed in the

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remaining animals. (IFF,1982c)

Route: oral. Species: rat. The acute oral toxicity of the test material was evaluated in male and female Sprague-Dawley rats (weighing 180-280 g). The animals were acclimated for at least 5 days prior to the start of the study. They were individually housed in suspended stainless steel wire-mesh cages with food and water availiable ad libitum. The animal room was maintained at a temperature of 22 +1-3 degrees C, a humidity of 30-70% and with a 12 hour lightldark cycle. Based on the results of a preliminary range finding study, a volume of 40 mI/kg test material was administered to groups of ten animals at one-fourth of the given dose in one-half hour intervals (total number of doses = 4). Gum Tragacanth was used as the vehicle plus one drop of 5% Tween 80 solution for each 5 ml vehicle. Six groups of ten rats (5 males/S females) were fasted for 18 hours and treated with neat test material in gum tragacanth at 6 different does levelsby oral gavage. The rats were observed immediately and at 1, 4 and 24 hours after dosing and twice daily for 14 days for pharmacotoxic, CNS effects and mortality. On the fourteenth day body weights were recorded. Rats were sacrificed and gross necropsy conducted. Necrospy included the examination of the following organs and tissues: Brainlpituitary, thyroid/parathyroid, esophagus/trachea, thymus, heart, lungs, liver, spleen, stomach, intestines, pancreas, kidneys, adrenals, urinary bladder, ovaries, uterus and testes. The LD5Owas calculated based on the method of Litchfield and Wilcoxin, 1949. Subjects: 30 Male 30 Female Study Length: 14 days Vehicle was 1% Gum Tragacenth.LD5O 1160 mg/kgSummary Under the conditions of this study, the calculated acute oral LD5Oin male and female rats was 1160 mg/kg (95% C.I. 1009-1334 mg/kg). 1160 mg/kg calculated LD5O,The calculated acute oral LD5O in male and female rats was 1160 mg/kg (95% C.I. 1009-1334 mg/kg). 5000 mg/kg clinical signs, lethal, 10/10 animals died at a dose level of 5000 mg/kg test material. Signs observed during the study were body drop, prostration, salivation, chromaturia, excess urination, diarrhea, piloerection, decreased activity, abnormal gait, tremors, arched back, clonic convulsions, salivation, flaccid body tone, straub tail, poor grooming cyanosis, hypersensitivity, ptosis and hunched back. Necropsy of those dying during the study revealed distention of the gastointestinal tract, hemorrhages in the superior and left lobes of lung and in the thymus, small erosions or ulcers in the glandular mucosa of stomach, diarrhea, congested lungs, yellow idscoloration of intestines, hemorrhagic areas in pyloric portion of stomach, red exudate around nasal cavity, discolored adrenals, intestines and lungs, and hemorrhages in small intestines. 3200 mg/kg clinical signs, lethal, 10/10 animals died at a dose level of 3200 mg/kg test material. Signs observed during the study were body drop, prostration, salivation, chromaturia, excess urination, diarrhea, piloerection, decreased activity, abnormal gait, tremors, arched back, clonic convulsions, salivation, flaccid body tone, straub tail, poor grooming cyanosis, hypersensitivity, ptosis and hunched back. Necropsy of those dying during the study revealed distention of the gastointestinal tract, hemorrhages in the superior and left lobes of lung and in the thymus, small erosions or ulcers in the glandular mucosa of stomach, diarrhea, congested lungs, yellow idscoloration of intestines, hemorrhagic areas in pyloric portion of stomach, red exudate around nasal cavity, discolored adrenals, intestines and lungs, and hemorrhages in small intestines. 2500 mg/kg clinical signs, lethal, 10/10 animals died at a dose level of 2500 mg/kg test material. Signs observed during the study were body drop, prostration, salivation, chromaturia, excess urination, diarrhea, piloerection, decreased activity, abnormal gait, tremors, arched back, clonic convulsions, salivation, flaccid body tone, straub tail, poor grooming cyanosis, hypersensitivity, ptosis and hunched back. Necropsy of those dying during the study revealed distention of the gastointestinal tract, hemorrhages in the superior and left lobes of lung and in the thymus, small erosions or ulcers in the glandular mucosa of stomach, diarrhea, congested lungs, yellow idscoloration of intestines, hemorrhagic areas in pyloric portion of stomach, red exudate around nasal cavity, discolored adrenals, intestines and lungs, and hemorrhages in small intestines. 1600 mg/kg clinical signs, lethal, 9/10 animals died at a dose level of 1600 mg/kg test material. Signs observed during the study were body drop, prostration, salivation, chromaturia, excess urination, diarrhea, piloerection, decreased activity, abnormal gait, tremors, arched back, clonic convulsions, salivation, flaccid body tone, straub tail, poor grooming cyanosis, hypersensitivity, ptosis and hunched back. Necropsy of those dying during the

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study revealed distention of the gastointestinal tract, hemorrhages in the superior and left lobes of lung and in the thymus, small erosions or ulcers in the glandular mucosa of stomach, diarrhea, congested lungs, yellow idscoloration of intestines, hemorrhagic areas in pyloric portion of stomach, red exudate around nasal cavity, discolored adrenals, intestines and lungs, and hemorrhages in small intestines. 1250 mg/kg clinical signs, lethal, 7/10 animals died at a dose of 1250 mg/kg test material. Signs observed during the study were body drop, prostration, salivation, chromaturia, excess urination, diarrhea, piloerection, decreased activity, abnormal gait, tremors, arched back, clonic convulsions, salivation, flaccid body tone, straub tail, poor grooming cyanosis, hypersensitivity, ptosis and hunched back. Necropsy of those dying during the study revealed distention of the gastointestinal tract, hemorrhages in the superior and left lobes of lung and in the thymus, small erosions or ulcers in the glandular mucosa of stomach, diarrhea, congested lungs, yellow idscoloration of intestines, hemorrhagic areas in pyloric portion of stomach, red exudate around nasal cavity, discolored adrenals, intestines and lungs, and hemorrhages in small intestines. 1000 mg/kg clinical signs, lethal, 2/10 animals died at a dose of 1000 mg/kg test material. Signs observed during the study were body drop, prostration, salivation, chromaturia, excess urination, diarrhea, piloerection, decreased activity, abnormal gait, tremors, arched back, clonic convulsions, salivation, flaccid body tone, straub tail, poor grooming cyanosis, hypersensitivity, ptosis and hunched back. Necropsy of those dying during the study revealed distention of the gastointestinal tract, hemorrhages in the superior and left lobes of lung and in the thymus, small erosions or ulcers in the glandular mucosa of stomach, diarrhea, congested lungs, yellow idscoloration of intestines, hemorrhagic areas in pyloric portion of stomach, red exudate around nasal cavity, discolored adrenals, intestines and lungs, and hemorrhages in small intestines. 1190 mg/kg calculated LD5O, The calculated acute oral LD5O for male rats was 1190 (95% C.I. 960- 1476 mg/kg). 1120 mg/kg calculated LD5O, The calculated acute oral LD5O for female rats was 1120 (95% C.I. 966-1299 mg/kg). (IFF,1982d) Route: oral. Species: guinea pig. 10 guinea pigs per dose. 18 hour predose fast. 6 day observation period. LD5O estimated from dosage-mortality curve. Material used undiluted. (DRAIZE,1944)LD50 1 MG/KG 1 mg/kg calculated LD5O. (Draize,1948) Route: oral. Species: mouse. 10 mice per dose. 6 day observation period. LD5O estimated from dosage-mortality curve. Material used undiluted. (DRAIZE,1944)LD5O 1.4 ML/KG 1.4 ml/kg calculated LD5O. (Draize,1948) Route: oral. Species: rat. 10 rats per dose. 18 hour predose fast. 6 day observation period. LD5O estimated from dosage-mortality curve. Material used undiluted. (DRAIZE,1944)LD5O 1.7 ML/KG 1.7 ml/kg calculated LD5O. (Draize,1948) Route: oral. Species: rabbit. 10 rabbits per dose. 6 day observation period. LD5O estimated from dosage-mortality curve. Material used undiluted. (DRAIZE,1944)LD5O 1.8 ML/KG 1.8 mlfkg calculated LD5O. (Draize,1948) Route: skin. Species: rabbit. (DRAIZE,1944)LD5O 4 ML/KG 4 mlfkg calculated LD5O. (Draize,1948) Route: oral. Species: rat. The LD5O, orally by stomach tube was determined. Animals observed for 2 wk or until death. Strain not specified. Method for determining LD5O not specified. LD5O 2.8 G/KG 2.8 g/kg calculated LD5O, using 20 rats. (Graham,1945) Route: oral. Species: rabbit. The LD5O, orally by stomach tube was determined. Animals observed for 2 wk or until death. Strain not specified. Method for determining LD5O not specified. LD5O 1.68 G/KG 1.68 g/kg calculated LD5O, using 12 rabbits. (Graham,1945) Route: oral. Species: cat. The LD5O, orally by stomach tube was determined. Animals observed for 2 wk or until death. Method for determining LD5O not specified. LD5O 2.24 G/KG 2.24 g/kg calculated LD5O, using 11 cats. (Graham,1945) Route: oral. Species: dog. The LD5O, orally by stomach tube was determined. Animals observed for 2 wk or until death. Method for determining LD5O not specified. LD5O > 22.44 g/kg using 4 dogs. (Graham, 1945)

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Route: skin. Species: cat. Benzyl compounds neat or in various vehicles was applied one or two times to the clipped backs (4 x 6 in) by cotton balls held firmly with hemostatic forceps. The area was massaged during application to facilitate absorption. Animals observed for 2 wk or until death. 100%. 20 ml lethal, 2 cats used; died within 22 hr. 60% H20/33% benzyl benzoate: 2 cats used; died within 69 hr. 7.5 ml lethal, 2 cats used; died within 48hr. (Graham,1945) Route: skin. Species: dog. Benzyl benzoate was applied 6 times to the clipped backs (4 x 6 in) by cotton balls held firmly with hemostatic forceps. The area was massaged during application to facilitate absorption. Animals observed for 2 wk or until death. 100%. 200 ml no effects, used 1 dog. 100 ml no effects, used 2 dogs. (Graham,1945) Route: skin. Species: domestic animals. Benzyl benzoate was applied 5 times to the backs of a horse, a heifer, a sheep (clipped) & a pig using a 4 inch pig bristle paint brush. The area was massaged during application to facilitate absorption. Animals observed for 2 wk or until death. 100%. 250 ml no effects, in 1 sheep. 200 ml no effects, in 1 pig. 500 ml no effects, in 1 heifer. 1000 ml no effects, in 1 horse. (Graham, 1945) Route: oral. Species: rat. A preliminary range finding study was performed to select dose levels for the LD5O acute toxicity study. Four fasted Sprague-Dawley outbred albino rats (2 males and 2 females) with initial body weights of 180-19 1 grams were administered test material at 10 grnlkg. Due to the physical characteristics of the test material and large dosing volume the group of 4 animals were administered 2500 mg/kg 4 times at 1/2 hour intervals. They were individually housed in suspended stainless steel wire-mesh cages with food and water availiable ad libitum. Acclimation period was 5 days. Gum Tragacanth (1%) was vehicle plus one drop of 5% Tween 80 solution for each 5 ml of vehicle. No effects, no signs of mortality observed during dose range-finding study. (IFF,1982b) Route: gavage. Species: rat. A preliminary dose-range finding study was performed to select dose levels for the LD5O phase of the experiment. Three groups of four Sprague Dawley rats two male and two female) were dosed orally at 500, 1600 and 5000 mg/kg body weight. Animals were fasted 18 hours prior to dosing. Initial weights ranged from 180 - 280 grams after fasting. Animals were housed individually in stainless steel 1/2’ wire mesh cages. Animals received Wayne Lab Blox and water ad libitum. Light cycle was 12 hours lightll2 hours dark; temperature was maintained at 22 +1-3 C with a humidity of 30 to 70%. Acclimation period was 5 days. Rats were observed immediately after dosing and at 1 and 4 hours and daily thereafter for seventy-two hours for overt pharmacotoxic, CNS effects and mortality. Body weights were recorded on the third day and surviving rats were sacrificed by C02 inhalation. Subjects: 6 Male 6 Female Study Length: 72 hours (Litchfield and Wilcoxon.) The vehicle was 0.25% methylcellulose plus one drop of 5% Tween 80 solution for each 5 ml of vehicle. 500 mg/kg clinical signs, no deaths in male or female rats. Observations included slightly decreased body tone. 1600 mg/kg clinical signs, lethal, 1/4 deaths. 1/2 deaths occurred in male rats 48 hours following dosing. Observations included decreased body tone, abnormal gait, arched back, chromodacryorrhea, hypersensitivity, poor grooming and decreased activity. 5000 mg/kg clinical signs, lethal, 4/4 deaths. 1/2 deaths occurred in female rats 4 hours following dosing, and 3/3 deaths in male and female rats 24 hours following dosing. Observations included decreased body tone, abnormal gait, and decreased activity. (1FF,1982a) Route: skin. Species: cow. (Graham, 1945) Route: skin. Species: domestic animals. (Graham,1945) Route: skin. Species: pig. (Graham,1945) Route: oral. Species: rat. A preliminary range finding study was performed to select dose levels for the LD5O acute toxicity study. Four fasted Sprague-Dawley rats (2/sex) were treated orally by gavage with three doses of a prepared test material in a vehicle of 0.25% methyl cellulose. Individual body weights were recorded prior to dosing. No additional detail provided on preliminary testing. Subjects: 2 Male 2 Female Test material in vehicle of 0.25% methyl cellulose (-01).Summary Based on the results of this dose range-finding study, test material-Ol (neat in 0.25% methyl cellulose) was recoded as test

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material-04 (neat in 1% Gum Tragacanth), and given to rats at six dose levels (one-fourth of total dose every half-hour for a total of 4 doses) of 5000, 3200, 2500, 1600, 1250 and 1000 mg/kg. 1600 mg/kg clinical signs, lethal, 1/10 animals died at a dose level of 1600 mg/kg. Signs observed included decreased body tone, abnormal gate, decreased activity, body drop, arched back, chromodacryorrhea, poor grooming and hypersensitivity. 500 mg/kg clinical signs, None (0/10) of the animals died at a dose level of 500 mg/kg. Signs observed included decreased body tone, abnormal gate, decreased activity, body drop, arched back, chromodacryorrhea, poor grooming and hypersensitivity. 5000 mg/kg lethal, 10/10 anumals died at a dose level of 5000 mg/kg. Signs observed included decreased body tone, abnormal gate, decreased activity, body drop, arched back, chromodacryorrhea, poor grooming and hypersensitivity. (1FF,1982d) Route: gavage. Species: rat. A range finding study was conducted to establish a dosing regimen for the main oral LD5O study. Groups of 2 male and 2 female, Sprague-Dawley albino rats were used. Animals were fasted overnight prior to dosing and for approximately 2 hours after dosing. Initial body weights were 119-148 grams for males and 118-148 grams for females. All animals received one dose of 5.0 mi/kg test material in arachis oil B.P. Animals were observed for mortality and/or systemic effects at 30 minutes, 1 and 4 hours and then once daily for 7 days or until all evidence of toxicity had subsided. 1500 mg/kg clinical signs, 0/2 deaths. Dose was mg/kg bodyweight. Clinical signs observed were abnormal body carriage (hunched posture), lethargy, pilo-erection, a decreased respiratory rate and ptosis. Recovery of all animals was complete by Day 2. Based on no deaths at any of the dose levels, 2000 mg/kg was selected for the main oral LD5O study. 1000 mg/kg clinical signs, 0/2 deaths. Dose was mg/kg bodyweight. Clinical signs observed were abnormal body carriage (hunched posture), lethargy, pilo-erection, and a decreased respiratory rate. Recovery of all animals was complete by Day 2. Based on no deaths at any of the dose levels, 2000 mg/kg was selected for the main oral LD5O study. 100 mg/kg clinical signs, 0/2 deaths. Dose was mg/kg bodyweight. Clinical signs observed were abnormal body carriage (hunched posture), lethargy, pilo-erection, a decreased respiratory rate and ptosis. Recovery of all animals was complete by Day 2. Based on no deaths at any of the dose levels, 2000 mg/kg was selected for the main oral LD5O study. 500 mg/kg clinical signs, 0/2 deaths. Dose was mg/kg bodyweight. Clinical signs observed were abnormal body carriage (hunched posture), lethargy, pilo-erection, a decreased respiratory rate and ptosis. Recovery of all animals was complete by Day 2. Based on no deaths at any of the dose levels, 2000 mg/kg was selected for the main oral LD5O study. 2000 mg/kg clinical signs, 0/2 deaths. Dose was mg/kg bodyweight. Clinical signs observed were abnormal body carriage (hunched posture), lethargy, pilo-erection, a decreased respiratory rate and ptosis. All symptoms persisted up to and including Day 3 but had resolved completely by Day 4. Based on no deaths at any of the dose levels, 2000 mg/kg was selected for the main oral LD5O study. (HaarmannReimer, 1985) Route: in vitro. Species: yeast. The yeast test was conducted on Saccharomyces cerevisiae, using a thermostatized incubation mixture with nutrients and trace elements. The test substance was added to the mixture by increasing concentration, and the effect upon the growth rate of the yeast cells was monitored at 30, 90, 150, and 210 minutes after beginning the experiment by counting the cell number. Every experiment was repeated several times under identical conditions. A simple counting chamber under the microscope or an electronic coulter counter was used to count cell number and calculate percent growth of cells in relation to the untreated control. The mean inhibitory concentration or 1C50 value of a given compound was calculated and correlated with the known LD5O values found in animals with the same substance. A blank was carried along with every experiment as control. 94 mg/l, 1C50. (Koch,1993) Route: skin. Species: cat. Cutaneous lesions on the shaven upper thighs were rubbed three times with test material. Survival and clinical signs were monitored. Lethal, within 24 to 36 hours at doses of 8.7 to 12.8 g/kg. (Postel,1943) Route: intramuscular. Species: cat. A single injection was administered. Clinical signs and survival were monitored. 2 g/kg lethal. 10 g/kg lethal. (Postel,1943)

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Route: subcutaneous. Species: guinea pig. A single injection of neat material was given under the abdominal skin. Clinical signs and survival were monitored. 5 g/kg clinical signs, lethal, 80% mortality. 0.5 glkg no effects. 1 glkg lethal, 30% mortality. 20 g/kg clinical signs, lethal, 100% mortality. 10 g/kg clinical signs, lethal, 100% mortality. (Postel,1943) Route: skin. Species: rat. A liniment containing the test material was applied to the skin of a rat. The liniment contained black soap and 90% alcohol. Clinical signs and survival were monitored. Clinical signs, the application of 7 to 8 gm of the liniment to the skin of rats led to agitation. No effect was seen after the application of 5 gm. (Jaulmes,1946) Route: gavage. Species: rat. The acute-toxic-class test was conducted using Wistar rats. The tests were conducted at 6 laboratories as part of a validation study of the method. The doses were 25, 200 or 2000 mg/kg depending upon the results generated. A step wise procedure was used with 3 fasted rats/group. Observation was 14 d ATC = acute-toxic-class. Nonspecific effects, ATC = unclassified at 2 of the laboratories and harmful at 4 laboratories. (Schlede,1992) Irritation Route: skin. Species: human 18+ yrs. A closed 12, 24 or 48 hour patch test was conducted on healthy and locally inflamed skin. Neat material was applied. 100 % no effects, 5 to 10 drops were applied to the patch. (Postel,1943) Route: skin. Species: guinea pig. Neat test material was applied to shaven, undamaged abdominal skin. Local and absorptive disruptions were monitored Duration not given. 15 g/kg no effects. (Postel,1943) Route: skin. Species: guinea pig. An acute dermal irritation study was performed as part of the accompanying phototoxicity test. 30 % no effects. 50 % no effects, Benzyl benzoate was classified as ‘non-irritant primary skin irritant” up to 50% concentration. 10 % no effects. (Takasago,1981)

Route: surface of eye. Species: rabbit. The undiluted test substance was instilled in the right eye(0. 1 ml) of six (6) rabbits. The test substance was placed in the right eye of each animal by gently pulling the lower lid away from the eyeball to form a cup. The upper and lower lids were then gently held together for one second to prevent loss of material. The eyes were examined at 1, 24, 48 and 72 hrs. and 7 days after treatment. Subjects: 3 Male 3 Female Study Length: 7 days (Acute Eye Irritation Test in Rabbits)Summary The test substance was not considered to be an eye irritant, based upon the observations made in this eye irritation test in rabbits. 100 % eye effects, Postive ocular responses (conjunctiva & Iris) were observed in 3 Albino New Zealand white rabbits (3 Male & 3 Female) at (1) hour after treatment. No other positive ocular responses were observed in any of the test animals. (1FF,1982) Route: skin. Species: rabbit. A four hour semi-occlusive patch test conducted on four female New Zealand white albino rabbits. A 0.5 ml aliquot of undiluted test material was applied to a 2.5 cm square of surgical lint B.P. which was then placed on an area of clipped, intact dorsal skin. The lint patches were held in place by encircling the trunk of the animal with a length of “Elastoplast” elastic adhesive bandage 10 cm wide. After four hours, the adhesive tapes were removed and the treated sites were cleansed by gentle swabbing with cotton wool soaked in warm water. One hour after removal of the patches and excess test material, the treated sites were assessed for reactions. Similar examinations were made at 24, 48, 72 and 168 hours after patch removal. Irritation was assessed and allocated a numerical value based on the following criteria: Erythema and Eschar Formation: 0 = No erythema; 1 = Very slight erythema (barely perceptible); 2 = Well-defined erythema; 3 = Moderate to severe erythema; 4 = Severe erythema (beet redness) to slight eschar formation (injuries in depth); Oedema Formation: 0 = No oedema; 1 = Very slight oedema (barely perceptible); 2 = Slight oedema (edges of area well defined by definite raising); 3 = Moderate oedema (raised approximately 1mm); 4 = Severe oedema (raised more than 1mm and extending beyond area of exposure). This test was conducted according to Annex V of EEC Directive 79/831. The EEC classification system indicates that when the mean value obtained for either erythema or edema equals or exceeds 2, the material is considered an irritant. 100 % no effects, erythema=1 .2, edema=0.4 Average scores calculated from the numerical

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values given to irritation observed at 24, 48, and 72 hrs. (RIFM,1985) [Haynes,1985] Route: skin. Species: rat. (RIFM, 1980) [Moreno,1980] Route: skin. Species: rabbit. (Lehman,1955) Route: skin. Species: guinea pig. Occluded patch preliminary irritation testing was conducted on 4 female guinea pigs using test material in a vehicle of ethanol. Three concentrations of the test material were applied. Readings were made at 24 and 48 hours after patch removal. Scoring was according to the following scale: 0 = no reaction, +I. = barely perceptible erythema, + = scattered, mild erythema (faint pink), ++ = moderate and diffuse erythema (pale pink), +++ = intense erythema (deep pink) and oedema. A challenge concentration for follow-up study was selected based on preliminary patch test results. Subjects: 4 Female Study Length: 24 hours vehicle was ethanol.Summary 2% test material in ethanol was selected for the challenge application concentration for the main study as it was suitably non-irritant. 10 % irritant effects, 1/4 animals exhibited barely perceptible erythema at the 24 hour reading only. 5 % no effects. 2.5 % irritant effects, 1/4 animals exhibited barely perceptible erythema at the 24 and 48 hour reading. (Quest,1977) Route: skin. Species: guinea pig. Occluded patch preliminary irritation testing was conducted on 4 male guinea pigs using test material in a vehicle of acetone/PEG. Three concentrations of the test material were applied. Readings were made at 24 and 48 hours after patch removal. Scoring was according to the following scale: 0 no reaction, 0.5 = very faint erythema (usually non-confluent), 1 faint erythema (usually confluent), 2 = moderate erythema, 3 = marked erythema (with or without oedema), n = necrosis, sp = small spots of erythema. Subjects: 4 Male Study Length: 24 hours vehicle was acetone/PEG.Summary No observed effects in any of the test animals at the concentrations tested (5%, 10%, 25%). 5 % no effects. 10 % no effects. 25 % no effects. (Quest,1981)

Route: skin. Species: human 18+ yrs. The purpose of this study was to evaluate the potential of the test material to induce a primary skin irritation in humans. Twenty-two subjects, 24-70 years old, were empaneled for the study: 3 male and 19 female. Twenty-one subjects, 3 male and 18 female, completed the test. One subject discontinued due to personal reasons not due to test material reaction. 0.3 ml test material in 3:1 diethyl phthalate:ethanol was applied to a 25 mm Hill Top Chamber patch occlusively. Test material was allowed to volatilize for at least 15 minutes but no longer than 40 minutes prior to application. The test material was applied to the patch at room temperature. The test sites were wiped clean with a dry Kimwipe prior to patching. Patches were applied on the back of each subject. Each subject was patched on Monday and instructed that the patches were to remain in place and dry. The subjects returned to the laboratory approximately 24 hours post-patching (Tuesday) and the patches were removed. The subjects returned to the laboratory on Wednesday and each test site was scored (reading 1) and re-patched. The subjects were again instructed to keep the patches in place and dry. The subjects returned to the laboratory on Thursday and the patches were removed. The subjects returned to the laboratory on Friday and each test site was scored (reading 2). If the Friday readings showed signs of significant irritancy, these subjects would be requested to return to the laboratory for additional readings. 3:1 diethyl phthalate:ethanol and saline were tested individually at the same time as the test material and used for control/comparision. The test sites were scored using the modified scoring scale of the International Contact Dermatitis Research Group System (Fisher, A., Contact Dermatitis, 1986) as follows: 0 = no visible reaction, +/- = faint, minimal erythema, 1 = erythema, 2 = intense erythema, 3 = intense erythema, induration, vesicles, 4 = severe reaction with erythema, induration, vesicles, pustules (may be weeping), E = edema, TR = tape reaction, A = hyperpigmentation, DR = dryness, X = discontinued. Subjects: 3 Male 18 Female Study Length: 96 hours 102-1 vehicle was 3:1 diethyl phthalate:ethanol. 50 % no effects, 3/21 subjects exhibited faint, minimal erythema at Reading 2 (96 hours) only. Authors stated that the test material elicited negligible irritation potential. The vehicle 3:1 diethyl phthalate:ethanol elicited 1/21 erythema reactions and 2/21 faint, minimal erythema reactions at Reading 1 (48 hours) and 8/21 faint, minimal erythema reactions at Reading 2 (96 hours). Authors stated that 3:1 diethyl phthalate:ethanol elicited minimal irritation potential. Saline elicited 0/21

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reactions at Reading 1 (48 hours) and 1/21 faint, minimal erythema reactions at Reading 2 (96 hours). Authors stated that saline elicited negligible ilTitation potential. (RIFM,2003) [Harrison,2003] Route: skin. Species: human 18+ yrs. Irritation was assessed during an associated repeat insult patch test. 129 subjects, 46 males and 83 females completed the study. During the induction phase a webril/adhesive patch (25 nmi Hill Top Chamber System) was used occiusively. 0.3 ml of test material (99.9% purity) was applied to each designated patch. The test sites were wiped with a dry Kimwipe prior to each patching. The left side of the back was used for the test area during the induction phase. Patches remained in place and were kept dry for approximately 24 hours, after which time they were removed. A 24 hour period, during which no test materials were applied, followed the removal of the Monday and Wednesday patch applications; a 48 hour period followed the Friday patch applications. On Mondays, Wednesdays and Fridays, the test sites were observed and any reactions scored and recorded. The identical test site was then repatched until 9 induction patchings were completed over a period of approximately 3 weeks. Reactions were scored based on the following scale: No visible reaction 0; faint, minimal reaction +1-;erythema- 1; intense erythema,induration-2; intense erythema, induration, vesicles-3; severe reaction with erythema, induration, vesicles, pustules-4. Test conducted according to HRL Protocol #100RIFM and HRL Standard Operating Procedures. Subjects: 46 Male 83 Female Study Length: 3 weeks Vehicle was 1:3 EtOH:DEP. 50 % no effects, Three subjects exhibited low-level transient reactions to the test material during the induction phase. (RIFM,2004) [Harrison,2004] Route: skin. Species: rabbit. acute and 3-week subacute skin toxicity testing. (DRAIZE,1944) Irritant effects, very mild gross skin irritation. Primary irritation index = 7.3. (Draize,1948) Route: skin. Species: human 18+ yrs. In a pre-test for a human maximization study, a 48 hour closed patch test was conducted on the forearms or backs of 25 healthy, male and female volunteers. Vehicle was petrolatum. 77-4-ROB. 4 % no effects, Costus and Benzyl Benzoate. (RIFM,1978) [Kligman,1978] Route: skin. Species: rabbit. A 4 hour, semi-occlusive patch test was conducted on three New Zealand white albino rabbits using a 0.5 ml or 0.5 g aliquot of undiluted test material which was placed evenly over a 2.5 cm square of surgical lint B.P. The lint square was then placed onto a 6 cm square area of clipped, intact dorsal skin. The lint patches were held in place by encircling the trunk of the animals with a length of ‘Elastoplast’ elastic adhesive bandage 10 cm wide. After a period of 4 hours the adhesive tapes were removed and the treated sites cleansed by gentle swabbing with cotton wool soaked in warm water. Reactions were assessed at 1, 24, 48, 72 and 168 hours after patch removal. Irritation was assessed and allocated a numerical value based on the following: Erythema and Eschar Formation - No erythema = 0; Very slight erythema (barely perceptible) = 1; Well-defined erythema = 2; Moderate to severe erythema = 3; Severe erythema (beet redness) to slight eschar formation (injuries in depth) = 4. Oedema Formation - No oedema = 0; Very slight oedema (barely perceptible) 1; Slight oedema (edges of area well-defined by definite raising) = 2; Moderate oedema (raised approximately 1 mm) = 3; Severe oedema (raised more than 1 mm and extending beyond area of exposure) = 4. This test was conducted according to Annex V of EEC Directive 79/831. The EEC classification system indicates that when the mean value obtained for erythema or edema equals or exceeds 2, the material is considered an irritant. No vehicle mentioned. 100 % no effects, avg. scores: erythema=0, edema=0. Very slight erythema in all 3 rabbits at 1 hour only. No other effects observed. (RIFM,1984) [Haynes,1984]

Route: skin. Species: human 18+ yrs. A 24 - 72 hour closed patch test was conducted on 1,830 healthy male and female subjects. The test material was applied to the upper inside of the subject’s arms, in a vehicle of Unguentum Simplex or Unguentum Hydrophilicum. Reactions were recorded according to the following criteria: (-)= No reaction [negative reaction]; (+1-) = Slight Erythema [suspected positive reaction]; (+) = Erythema [positive reaction]; (++) = Erythema and Dropsical swelling [positive reaction]; (+++) = Erythema, Papules and Vesicles [positive reaction]. The article is in Japanese. Information obtained from the English translated summary and tables. 2 % no effects, 0/30 subjects.

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(Fujii, 1972) Route: skin. Species: human 18+ yrs. A 48 hour closed patch test was conducted on 1,763 healthy male and female subjects. The test material was applied to the small of the back of the subjects, in a vehicle of Vaselinum Aldum or Unguentum Hydrophilicum. Reactions were recorded according to the following criteria: (-)= No reaction [negative reaction]; (+1-)= Slight Erythema [suspected positive reactioni; (+) = Erythema [positive reaction]; (++) = Erythema and Dropsical swelling [positive reaction]; (+++) = Erythema, Papules and Vesicles [positive reaction]. The article is in Japanese. Infonnation was obtained from the English translated summary and tables. 20 % no effects, 0/34 subjects. (Fujii,1972) Route: skin. Species: human 18+ yrs. A 24 hour closed patch test was conducted on male and female volunteers. Test material was applied undiluted to an area on the inner arm of the subjects measuring 1 cm in diameter. Crystalline substances were applied in the same manner except as saturated alcoholic solutions. Immediately following application, the area was covered with an adhesive bandage for 24 hours. Reactions were read daily for 5 days. Applications which produced visible inflammation, irritation, formation of a popular rash, or any abnormal condition of the area, were considered positive reactions. All positive reactions were confirmed by repetition of the test on the same subject after complete disappearance of positive systems. Results listed are number of positive reactions observed in the number of tests conducted. No effects, no positive reactions were observed in 13 tests. (Katz,1946) Route: intramuscular. Species: chicken. 0.5 ml of neat test material was injected 1/2 inch deep in the right and left pectoral muscles of 6 male Hubbard Crossbred broilers. 2 chickens were sacrificed days 1, 3 and 7 after injection for analysis of gross lesions at the injection sites. 0.5 ml no effects. (Hem,1975) Sensitization Route: skin. Species: guinea pig. A guinea pig maximization test was conducted, using the same concentration of the test material for induction and challenge. The vehicle was not given. No further details were available. Data taken from English table and abstract. (Magnusson,1969) 10 % sensitization effects, Rate III or moderate; Score 0.4. (Ishthara,1986) Route: skin. Species: guinea pig. An open epicutaneous test was conducted in guinea pigs. Induction consisted of 21 daily open applications to the shaved flank of 6-8 guinea pigs per group. One to six experimental and one control group was used. Open challenge applications were made on days 21 and 35. Reactions were read at 24, 48 and 72 hours. No further details were provided. Vehicles were not specified by material. (Klecak,1977) Greater than or equal to 98% purity. 30 % no effects. (Klecak, 1979)

Route: in vitro. Species: unknown. In order to determine if reactivity correlates with sensitization potential, chemicals representing allergens of different potencies (weak to extreme) and nonsensitizers were evaluated for their ability to react with glutathione (GSH). GSH reactivity of a test material was evaluated in a 15 ml plastic Corning conical tube by transferring 50 ul of the GSH stock solution to 400 ul of sodium phosphate buffer, 100 mM (pH 7.4) followed by the addition of 50 ul of the test chemical stock solution. The final reaction contained 0.2 mM GSH and 20 mlvi of the test chemical. The reaction was incubated in a 25 C shaking water bath for 15 minutes. Additional samples containing GSH (0.05 to 200 mM) or glutathione disulfide (GSSG) (0.025 to 100 mM) were prepared without test material and these samples were used to define the calibration curve for each analysis. Immediately following incubation, GSH and GSSG were derivatized with 2,4-dinitrofluorobenzene (DNFB) according to the method of Farriss and Reed (1987). To each sample, 50 ul of bathophenanthrolinedisulfonic acid (BPDS), 100 ul of concentrated perchioric acid, 50 ul of 100 mM iodoacetic acid/0.1 mM m-cresol purple and 480 ul of the working solution of 2 M potassium hydroxide/2.4 M potassium bicarbonate were added. The sample was mixed on a Vortex and placed in the dar at room temperature for 10 minutes. Untreated GSH was derivatized by adding 1 ml of 1% DNFB. The sample was mixed on a Vortex and placed in the dark at room temperature for 60 minutes. The sample was spun on a

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centrifuge at 2000 rpm for 10 minutes and supernatant was analyzed by HPLC. Derivatized GSH and GSSG were separated and quantitated by reversed-phase (RP) HPLC according to the method of Farriss and Read (1987) on a Waters Alliance 2695 system using a Waters 2487 dual channel UV detector (365 nm) and a Waters Spherisorb analytical column. All test materials were prepared and analyzed in duplicate on two different days. The mass of GSH and GSSG on column was calculated with the respective calibration curves. GSH depletion was determined by dividing the GSH in a sample containing a test material by the GSH in a sample with no test chemical and multiplying by 100. GSSG was measured to determine if GSH depletion was a result of GSH conjugation to the test material or due to an oxido-reduction reaction with the test material Authors chose a peptide depletion of greater than 10% to indicate that the chemical had reacted with the starting peptide. 20 millimolar , reactivity of 20 mM test material to 0.2 mM glutathione caused 0.7% (5.5% SD) depletion of glutathione. (Gerberick,2004) Route: in vitro. Species: unknown. In order to determine if reactivity correlates with sensitization potential, chemicals representing allergens of different potencies (weak to extreme) and nonsensitizers were evaluated for their ability to react with three synthetic peptides containing either cysteine, lysine, or histidine. Peptide stock solutions were prepared to a final concentration of 1.25 mM in either 100 mM ammonium acetate buffer, pH 10.2 (lysine peptide) or 100 mM phosphate buffer, pH 7.5 (histidine and cysteine peptides). Test chemical solutions at a concentration of 100 mM were prepared in acetonitrile or solubilized in dimethylsulfoxide and then diluted with an equal part acetonitrile. Triplicate reactivity samples were prepared containing 0.5 mM peptide and either 5 mM or 25 mM test chemical for a peptide test chemical ratio of 1:10 or 1:50. A Biomek 2000 automated workstation (Beckman Coulter) was used to make additions of the peptide stock solution (400 ul), the appropriate buffer (350 ul) and the test chemical solutions (50 or 250 ul) into autosampler vials. In the case of the 1:10 ratio reactivity samples, 200 ul of acetonitrile was added to each vial. Samples without the test chemicals were also prepared in triplicate to function as controls. The autosampler vials were capped, gently vortexed and incubated for 24 hours at room temperature in the autosampler (dark) prior to HPLC analysis. Calibration standards were prepared manually from the peptide stock solution, diluted into the appropriate buffer for the peptide and contained either 5 or 25% acetonitrile. The peptide concentrations were 0.0156, 0.03 13, 0.0625, 0.125, 0.25, 0.50, 1.0 mM. A Waters Alliance 2695 and 996 PDA detector comprised the chromatographic system. A 10 ul injection of the reactivity samples was made onto the column. The peptides were separated from the test chemicals and products on a Zorbax SB-C18 (2.1 x 100 mm) stationary phase (Agilent Technologies) which was preceded by a SecurityGuard cartridge guard system (Phenomenex) containing a C18 cartridge (2.0 x 4.0 mm). The column temperature was 30 C. The mobile phase consisted of 0.1% trifluoroacetic acid (TFA) in water and 0.085% TFA in acetonitrile. A gradient of 90% to 60% over 25 minutes at a flow rate of 0.3 mI/mm was used for the separation. The diode array detector scanned the wavelengths 210-400 nm. Chromatograms were extracted at 220 nm. Quantitation was performed using either Millenium32 or Empower software packages. Peptide reactivity with the test chemicals was reported as percent peptide depletion, which was determined as the reduction of the peptide concentration in the samples relative to the average concentration of the controls. Sensitivity, specificity, positive predictivity, negative predictivity and accuracy were calculated for each peptide based on an arbitrary depletion cut-off value used to determine whether the chemical had reacted with the peptide. For the sensitizers, LLNA EC3 values an the peptide depletion percentages were compared using a Spearman correlation analysis (Hollander and Wolfe, 1973). Correlation analyses and graphs were generated using S-Plus 6.2. A level of significance (alpha) of 0.05 was used for the correlation analyses Authors chose a peptide depletion of greater than 10% to indicate that the chemical had reacted with the starting peptide. Reactivity of 25 mM test material to 0.5 mM lysine caused 3.0% (5.3% SD) depletion of lysine. Reactivity of 5 mM test material to 0.5 mM cysteine caused 0.2% (1.1% SD) depletion of cysteine. Reactivity of 25 mM test material to 0.5 mM histidine caused -2.5% (0.8% SD) depletion of histidine. (Gerberick,2004)

Route: in vitro. Species: human 18+ yrs. This study evaluated a peptide reactivity assay to predict

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fragrance allergens in vitro. Three novel peptide sequences were derived from proteins with specific CYS residues that were highly reactive according to Dennehy et al. (2006). Of the 125 Cys residues identified that react with both target electrophiles, peptides around Cys residues with very high frequency of adduct formation were selected for the study. Two additional peptides were derived from a region of the human protein Keap 1 with known reactive Cys residues (Eggler et al., 2005; Wakabayashi et al., 2004). The reactives residues in Keap 1 were selected to yield a peptide containing an aromatic aminoacid for ease of HPLC detection. The peptide reactivity assay in HPLC vials was conducted as described by Gerberick et al. (2004a) with modifications as follows. The test peptide was used at a concentration of 0.25 mM. The concentration of the phosphate buffer was 20mM. These changes resulted in less observed peptide precipitation. Samples were incubated at 30 C for 24 h with shaking at 140 rpm prior to HPLC analysis with a DAD dector. 5 ul samples were injected onto a Waters polarity column (5-micron). Chromatograms were extracted at 214 nm. Peptide depletion was expressed as the concentration of peptide compared to control samples with the peptide in solvent only. To measure dose response curves, reactivity assays were performed under equal conditions in microtiter plates with a final volume of 100 ul. Unreacted cysteine in the parent peptide was determined by adding 50 ul of a solution of monobromobimane (0.75 mM in 100 mM NaCO3, pH 8.8) and reading the fluorescence at 385/480 nm. GSH depletion assays were performed under equal incubation conditions as for other peptides, but unreacted GSH was detected with the enzymatic test described by Tietze (1969). To detect adducts of the peptides with the chemicals, LC-MS analysis was performed on a Finnigan LCQ classic Mass spectrometer (Thermo Finnigan, USA). Mass spectra were recorded from 200 to 2000 amu. Peptides Corl-C420 and Cys-react were run on specialized columns. LLNA data for comparison were determined under standard conditions as defined in the OECD guideline 429. LLNA data are either from literature as summarized by Gerberick et a!. (2004a, b) or from the Research Institute on Fragrance Materials. CysReact sequence: Ac-RFAACAA; Keap-257 sequence: Ac KYDCEQR; Keap-297 sequence: Ac-DSRCKDY; RP17-34 sequence: Ac-KRVCEEF; Corl-C420 sequence: Ac-NKKCDLF; Cofill-138 sequence: Ac-QANCYEE Additional references: Tietze 1969;Gerberick et al., 2004b. Study Length: 24 hours (Dennehy et a!., 2006;Eggler et al., 2005;Wakabayashi et al., 2004;Gerberick et al., 2004a) Test material was tested undiluted.Summary The current peptide reactivity assay was found to have high reproducibility and was a good positive predictor of moderate and strong allergens, but it is not currently optimized for weak sensitizers. 100 % , The test article was used undiluted. The test article caused -2.1 +- 0.1% GSH depletion and 0.2 +- 0.8% CysReact depletion. In comparison, the reference EC3 value for the LLNA test was >50, and the test article was in the sensitization class very weak/none. (Natsch,2007) Route: skin. Species: guinea pig. Ten test and three control animals, female Heartley strain albino guinea pig, microbial grade, clean purchased from Japan SLC, Inc., weighing 370-420 g (c.a. 7 wks old) at the start of experiment, were used for the main study. Hair on the back of the animal was cut with an electric hair clipper and an electric shaver. Four hours after depilation, based on the results of sighting tests, the concentrations of test material for the induction and the challenge phases were selected. For the purpose of this study the test material was freshly prepared as follows: [Intradermal induction]: 10% w/v in Complete Adjuvant, 10% w/v in a mixture of Freund’s Complete Adjuvant plus physiological saline (1:1); [Topical Induction]: 10% w/v in Complete Adjuvant; [Topical Challengel: 40, 20, 10, 5% w/v in mixture of propylene glycol plus acetone (1:1). The concentration, homogeneity and stability of the test material formulations were not determined. The intensity of skin reaction was graded from 0 to 8 scale. The criteria and scoring system for assessment of dermal reaction are those of Draize J.H. (1959). The scores for erythema and edema at 24-hour reading were totaled for the five test ginea pigs (5 values) and this total was divided by 10 to give the sensitisation index of the text material. 20 % positive effects, 4/10 animals exhibited a response. Response classified as “mild” sensitization. 10 % no effects, The test material was classified as non-sensitizer up to 10% to guinea pig skin. 40 % positive effects, 4/10 animals exhibited a response. Response classified as “mild” sensitization. 5 % no effects. (Takasago,1981)

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Route: skin. Species: guinea pig. A guinea pig open epidutaneous test (OET) was conducted on groups of 6 - 8 male and female guinea pigs weighting 300 - 450 grams. Daily applications were made for 3 weeks to a clipped 8-cm2 area on the flank of each guinea pig. The test sites were not covered and the reactions were read 24 hours after each application. A total of 21 applications of 0.1 ml test material in an unspecified vehicle were made for 21 days. The 10 controls were either left untreated or treated with 0.1 ml of the vehicle for 21 days. At the challenge phase, both the test and control animals were treated on days 21 and 35 on the contralateral flank with the test material at the minimal irritating concentration and some lower primary non-irritating concentrations. The results were not reported for the irritation pre-screen. It was not specified if the concentration reported in the results was the minimal irritating concentration or 1 of the lower primary nonirritating concentration. 30 % no effects, No sensitization (-)was produced. (Klecak,1985) Route: skin. Species: guinea pig. A modified Freund’s complete adjuvant test was conducted. For induction with crude material or with fractions, 30 mg was used. In the case of pure isolated, synthesized or purchased compounds, 15 mg was applied. For challenge, vehicle was acetone. The challenge dose appears in the results. The mean response was computed as the quotient of the sum of all reactions obtained divided by the total number of treated animals. A mean response of 0 to 1 was considered as weak, 1 to 2 as moderate, and greater than 2 as strong No further details were provided. (Hausen,1992b) 10 % sensitization effects, weak. 1 % sensitization effects, weak. (Hausen,1995) Route: skin. Species: guinea pig. The modified FCA method was conducted. Vehicle was acetone. Groups of 10 guinea pigs were used No further details provided. (flausen,1988) 10 % sensitization effects, weak. (Hausen,1992) Local Lymph Node Assay [Note that this report does not contain the original data, please see Location Number 496381 A project was undertaken to classify a recently published mouse local lymph node assay (LLNA) dataset for more than 200 test materials into various chemical reaction mechanistic domains known to be associated with sensitization. The reactive domains used for classification were Michael acceptors, S (N)2-electrophiles, S(N)Ar-electrophiles, Schiff base electrophiles, acyl transfer electrophiles, and non reactive and non-pro-reactive. The main goals of this work were to 1) characterize the population of each of the major domains by test materials in the dataset and to identify any major domains that may be under-represented; 2) to determine the extent to which the major domains covered the range of test materials in the dataset and to consider what could be learned from test materials outside these domains; and 3) to consider the potential of using the domain-classified dataset for predicting sensitization potential of new test materials by employing the strategy of mechanistic read-across or the use of quantitative mechanistic models (QMM). Specific rules were applied in order to try to classify the test materials into the six major reaction mechanistic domains. These rules were specified in detail in the paper. There was also a category for special cases which included: test materials that could be assigned to one of the six domains, although sometimes provisionally, until further comment was made; and test materials which did not fit into any of the stated domains. The authors theorized about how the classified data set could be used for mechanism-based read-across, stating that if two compounds in the same mechanistic domain are similar in their toxicity-determining parameters, then they should be similar in their toxicity, irrespective of whether or not they are similar in structure. The feasibility to develop QMMs was discussed because skin sensitization correlations were found to exist between potency and a combination of reactivity and hydrophobicity parameters within a reaction mechanism domain. The authors also illustrated the application of QMM and read-across for two test materials that are not in the LLNA database. The understanding gained from this work was intended as a step towards developing alternative non-animal methods in the predictive identification of chemicals with the intrinsic ability to cause skin sensitization. (Gerberick,2005;Basketter,2001)EC3 17%Summary The test material was classified as belonging to the S(N)2 domain. The potency category was weak. The test material was classified as belonging to the S(N)2 domain. The potency category was weak. (Roberts,2007)

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Route: skin. Species: mouse. The purpose of this study was to assess the skin sensitization potential of the test material using the Local Lymph Node Assay and to calculate the estimated concentration giving rise to a 3 fold increase in lymphocyte proliferation (EC3). A total of 24 female CBA/CaJOlaJHsd mice were divided into 6 groups consisting of 4 animals in each group. Groups included 1 control (vehicle) group and 5 test groups. Animals were 8-12 weeks of age. Animals were housed 4 per cage in an environmentally controlled animal room with a room temperature of 22+/-3 LI C, relative humidity of 30-70%, a minimum of 15 air changes per hour, and a 12 hour lightldark cycle. Food and water were available ad libitum. Acclimation period was for a least 5 days prior to dosing. The control group was treated with the vehicle only. Animals were checked at least once daily for signs of systemic toxicity and body weights were recorded prior to treatment and prior to injection with methyl thymidine. A volume of approximately 25 ul of 5 different concentrations of test material (% v/v) in a vehicle of 1:3 ethanol:diethyl phthalate was applied topically to the dorsal surface of each ear on three consecutive days. Three days after the third topical application all mice were injected intravenously into a tail vein with 250 ul of phosphate buffered saline (PBS) containing approximately 20 uCi of a 2.OCiJmmol specific activity [3]H-methyl thymidine. Approximately 5 hours after the intravenous injection, the animals were sacrificed, the draining auricular lymph nodes excised and pooled per group (8 nodes per group). Single cell suspensions of lymph node cells were prepared by mechanical disintegration of pooled lymph nodes by passing through 200-mesh stainless steel gauze. Cell suspensions were then washed three times by centrifugation with approximately 10 ml of PBS. Approximately 3 ml of 5% w/v trichioroacetic acid (TCA) was added, precipitated overnight at 4L1 C, then cells were pelleted by centrifugation and resuspended in TCA. The proliferative capacity of pooled lymph node cells was determined by the incorporation of [3]H-methyl thymidine measured in a B- scintillation counter as disintegrations per minute (DPM). DPM per node for test material relative to controls was defined as the stimulation index (SI). Test material was regarded as a sensitizer in the LLNA if one or more concentrations of the test material elicited a 3-fold or greater increase in isotope incorporation relative to the vehicle control group (S1>1=3.0). The estimated concentration of the test material required to produce a 3-fold increase in the draining lymph node cell proliferative activity (EC3) was calculated by interpolating between two SI points using the following equation; EC3 = [(3 LI d)/(b LId)J x (a LIc) + c, where a = concentration with SI immediately above 3, b = STof a, c = concentration with SI immediately below 3, and d = SI of c. EC3 values less than 0.1% were considered as the strongest sensitizers, the weaker sensitizers have a range of EC3 values of 1-10%, and non-sensitizers have EC3 values greater than 100% (Basketter, 2000; Gerberick, 2001). A separate positive control study with a different test material diluted in acetone: olive oil (4:1) was performed to validate the study. Subjects: 24 Female Study Length: 3 days Regulatory Compliance OECD guideline reference 429(2002):Skin sensitization: Local Lymph Node Assay; US Environmental Protection Agency, Health Effects Test Guidelines, OPPTS 870.2600(2003):Skin sensitization. GLP Yes (Kimber and Basketter, 1992, Kimber et al 1994) Y13056/001.Vehicle EtOH:DEP [1:3jSummary the test material in 1:3 ethanol:diethyl phthalate is unlikely to be a skin sensitizer under the conditions of the study. The EC3 value could not be calculated but is estimated to be greater than 50% w/v (greater than 12500 ug/cm2). 5.0 % DPM per Lymph Node 208Stimulation Index 0.7no effects, Test material was not a sensitizer at 5.0% with a stimulation index of 0.7. 2.5 % DPM per Lymph Node l92Stimulation Index 0.7no effects, Test material was not a sensitizer at 2.5% with a stimulation index of 0.7. 10 % DPM per Lymph Node 230Stimulation Index 0.8no effects, Test material was not a sensitizer at 10% with a stimulation index of 0.8. 25 % DPM per Lymph Node 475Stimulation Index 1.7no effects, Test material was not a sensitizer at 25% with a stimulation index of 1.7. 50 % DPM per Lymph Node 753Stimulation Index 2.7no effects, Test material was not a sensitizer at 50% with a stimulation index of 2.7. The test material in 1:3 ethanol:diethyl phthalate is unlikely to be a skin sensitier under the conditions of the study. The EC3 value could not be calculated but is estimated to be greater than 50% w/v (greater than 12500 ug/cm2). (RIFM,2005) [Betts,2005] Sensitization

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Route: skin. Species: human 18+ yrs. Tests were conducted on 11 male and female patients who had previously reacted to propolis. Test material was applied to the back for 1 day and reactions were read after 1 and 3 days. 5 % positive effects, in 3/8 patients. (Hausen,1992) Route: skin. Species: human 18+ yrs. 20 perfume-sensitive male and female patients were given standard 48-hour patch tests using aluminum backed strips. Reactions were read at removal or 24 hours after removal of patches. The vehicle was petrolatum. No further details were provided. All fragrance materials were tested on 50 control patients with negative results. To avoid the “angry back” phenomenon, patients were tested during a period of several months. 5 % sensitization effects, 1 reaction. (Larsen, 1977) Route: skin. Species: human 18+ yrs. A 48 hour closed patch test was conducted on the backs of 155 patients with cosmetic dermatitis Test material in petrolatum was applied on the cloth disks of Toni’s adhesive plaster for patch tests. The plaster was applied to the upper back of the patient for 48 hours. Readings were made at 1 and 24 hours and 1 week and 2 weeks after patch removal. Same patients as those in Itoh,1982 sub-reference 3. 5 % no effects, 0/45. (Itoh,1982) Route: skin. Species: human 18+ yrs. 50 subjects with photosensitivity dermatitis with actinic reticuloid syndrome (PD/AR), 32 subjects with polymorphic light eruption (PLE) and 457 with contact dermatitis (CD) were tested. A 48 hour closed patch was conducted on the upper back of each subject using Al-test patches and Scanpor. Vehicle was yellow paraffin. Patches were removed at 48 hours and any reactions read at that time, and at 72 hours after the application. The following grading system was used: 0 = no response; 1/2 = doubtful; 1 = faint but definite erythema; 2 = definite erythema; 3 erythema + oedema; 4 = erythema + oedema + vesicles. 2 % sensitization effects, reaction in 1/32 PLE subject. (Addo,1982) Route: skin. Species: human 18+ yrs. This report presents two case studies of contact dermatitis with Brazilian propolis. Propolis, called Ebee glue, E is used by bees to construct and maintain their hives. It is a mixed product with material that bees collect from pollen and tree barks, and a secretion of bees. In Japan, propolis, as an alcoholic extract, is widely used for its properties as an antiseptic, antimycotic, bacteriostatic, spasmolytic, antiinflammatory and anesthetic. Case 1 was a 26-year old housewife with a history of atopic dermatitis. She developed a severe dermatitis on her hands and feet after using an undiluted solution of Brazilian propolis in June of 1996. The propolis solution and its 70% ethanol extract (balsam) were analyzed by gas chromatography-mass spectrometry. Patch tests were performed on Case 1 using propolis solution at 100, 50, 10 and 1% (water vehicle); and rosin, petrolatum, fragrance mix, 48 standard perfume allergens, 10 plant allergens, and several test materials, all with petrolatum as vehicle. Propolis solution caused positive (++) reactions at 48 and 72 hours at all concentrations tested. Case 2 was a 3-year old boy whose mother had applied a propolis cream (supplied by a beekeeper) to the boy LIs face injury. After 2 days, the boy developed dermatitis on the area where the cream was applied and the mother immediately stopped using the cream. His mother then applied Oronine H ointment and Biscosal ointment (contains fluocinonide), but the boy Es condition became worse. Patch tests were performed on Case 2 using propolis cream, Oronine H ointment and Biscosal ointment, all at 100% and abietic acid and several test materials, all with petrolatum as vehicle. Propolis cream at 100% caused positive (+) reactions at 48 and 72 hours. The closed patch testing was performed with Finn Chambers attached to the patient LIsupper back with Scanpor Tape for 48 hours. Skin reactions were read at one hour and 24 hours after removal of the patch in accord with ICDRG recommendations. Vehicle was petrolatum. no effects, No skin reactions were observed after patch testing of Case 2 with test material. (Kato,1999) Route: skin. Species: human 18+ yrs. Patients reacting to propolis (bee glue) participated in further testing with the breakdown constituents of the bee glue. Twenty-seven patients agreed to be tested. Reactions were scored at 72 hours No further details were provided. Subjects: 27 Unspecified Sex Summary 5% test material produced no positive reactions in any of the subjects tested. 5.0 % no effects, 5% test material produced no positive reactions in any of the subjects tested. (Hausen,2005)

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Route: skin. Species: human 18+ yrs. A series of patch tests were conducted on various groups of patients from January 2003 - December 2004, according to recommendations of the International Contact Dermatitis Research Group (ICDRG) and the German Contact Dermatitis Research Group (DKG). The patch test materials (Hermal/Trolab) were applied to the back and outer side of upper arms using a well-fitting nonporous tape. The patches were applied for 24 or 48 hours. Readings were conducted until a minimum of 72 hours according to international grading standards. No further information was provided. (Fregert,1981) 2003 patients were tested. 1 % allergic reaction, Allergic reactions were reported in 1/2003 patients. (Schnuch,2007) Route: skin. Species: human 18+ yrs. Twenty-eight patients allergic to perfumes and sweet-smelling constituents were patch tested with a standard series recommended by the International Contact Dermatitis Research Group (ICDRG), and also with various components of essential oils. No further details were available. Article in French. Information obtained from English summary. in yellow petrolatum. 1 % no effects. (Meynadier,1986) Route: skin. Species: human 18+ yrs. Of 5539 male and female patients patch tested, 460 were considered to have positive tests related to cosmetics. Of these, 80 were positive to a fragrance mix. These 80 were patch tested with fragrance chemicals No further information provided. Concentrations not provided. Sensitization effects, 3/80 positive reactions. (Romaguera,1983) Route: skin. Species: human 18+ yrs. (FREGERT,1969A) Vehicle was likely petrolatum 73 patients were tested. 5 % positive effects, in 1 patient. (Angelini,1985) Route: skin. Species: human 18+ yrs. A 24 - 48 hour closed patch test on 7,703 male and female subjects with dermatoses was conducted. The test material was applied to the upper inside arm of the subjects, in a vehicle of 99% Ethanol or non-irritative cream base. Reactions were recorded according to the following criteria: (-)= No reaction [negative reaction]; (+/-) = Slight Erythema [suspected positive reaction]; (+) = Erythema [positive reaction]; (++) = Erythema and Dropsical swelling [positive reaction]; (+++) = Erythema, Papules and Vesicles [positive reactionJ. The article is in Japanese. Information obtained from the English translated summary and tables. 0.1 % no effects, 0/111 subjects. (Fujii,1972) Route: skin. Species: human 18+ yrs. Tests were conducted on 6 patients who had allergic contact dermatitis from the use of a proprietary antiseptic ointment containing oil of cinnamon. Vehicle was petrolatum. Reactions were read at 48 and/or 96 hours. Tested on 1 patient only. 5 % no effects. (Calnan, 1976) Route: skin. Species: human 18+ yrs. A closed patch test was applied for 30 minutes to the upper part of the back of adult male and female patients with chronic urticaria of more than 3 months duration and with varying dermatoses. Test material was applied in petrolatum. The result was read immediately after removal and after every hour until the reaction disappeared A conventional 48 hour closed patch test was performed in the 25 patients that reacted in the first test. The result was read 24 hour after removal of the test. 5 % no effects, in 5 patients tested. (Forsbeck,1977) Route: skin. Species: human 18+ yrs. 12 Patients who were previously patch tested and found to be sensitive to Peru balsam and 1% propolis (a resin used by bees) were further patch tested with 4 components identified in Peru balsam and propolis. The vehicle was petrolatum. No further details were provided. 5 % no effects. (Rudzki,1983) Route: skin. Species: man. A 23-year-old bee-keeper with recurrent episodes of eczema was patch tested with a series of allergens suggested by the case history and clinical examination. The vehicle was petrolatum. 2 % no effects. (Melli,1983) Route: skin. Species: woman. Tests using Finn chambers were conducted on 3 subjects who developed allergic dermatitis from contact preparations containing estradiol benzoate. Reactions were assessed at 72 hours. Vehicle was petrolatum. 5 % no effects. (Ljunggren,1981)

Route: skin. Species: human 18+ yrs. Results of standard patch tests on eczema patients conducted by

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the North American Contact Dermatitis Research Group (NACDG) from July 1977 to 1978 are reported All test substances have since been deleted from the routine series. Vehicle was petrolatum. 2 % positive effects, in 1% of 284 patients tested in 1979/80. No effect in 335 patients tested in 1978/79. (Also tested as part of a mixture but dose not given.). (Mitchell,1982) Route: skin. Species: human 18+ yrs. The principal patch test results of the North American Contact Dermatitis Group (NACDG) for July 1, 1975 to June 30, 1976 are summarized. The Al-Test and Dermicel tape were used. Reactions were evaluated at 48 and 96 hours. A total of 900-2000 patients were tested. Not all patients received the complete “battery’ of tests Vehicles not provided. Some results in this paper were previously published by the author in the 1975 methodology reference, LC 4005. (RUDNER,1975) 2 % no effects, in 198 patients. (Rudner,1977) Route: skin. Species: human 18+ yrs. The principal patch test results of the North American Contact Dermatitis Group (NACDRG) for the period July, 1, 1975 to June 30, 1976 were reported by Rudner (1977). A total of 900-2000 patients were tested. The Al-Test and Dermicel tape were used. Reactions were read at 48 and 96 hours. Not all patients received the complete “battery” of patch tests. Vehicles used were not reported. (Rudner et al,1975) 2 % no effects, no reactions in 198 patients tested. (Rudner,1978) Route: skin. Species: human 18+ yrs. Human patch tests were conducted on patients with and without cosmetic dermatitis and in patients with facial melanosis from the period 1978-1980. Frequency of sensitization as well as cross-sensitization to various test compounds was assessed. Vehicle was not reported. [Article in Japanese, no English abstract or summary]. 99.4 % purity. 5 % positive effects, 2.0% (1/50), subject with a prior history of eczema and dermatitis responded positively. (Ishihara,1981) Route: skin. Species: human 18+ yrs. The North American Contact Dermatitis Group conducted a multicenter study in North America from May 15, 1977 to September 15, 1980. 487 cases of cosmetic related contact dermatitis were identified among 8093 patients tested for contact dermatitis. Of the 487 patients, 385 were female and 102 were male. One hundred and forty nine of the cosmetic contact dermatitis patients were patch tested with cosmetic related products. Patch tests were applied to the upper back with Al-Test strips or Finn Chambers for 48 hours. Reactions were read at 48 and/or 72 hours. In most centers, delayed readings at 96 or 120 hours were also made Percent or amount of test material applied was not provided. Sensitization effects, 1/149 positive reaction. (Eiermann,1982) Route: skin. Species: human 18+ yrs. 142 patients who had positive patch reactions to 25% Balsam of Peru in petrolatum were patch tested with components of Balsam of Peru at 8 worldwide centers. Percent and amount of test material applied was not provided. Sensitization effects, 10/142 reactions. (Mitchell, 1975) Route: skin. Species: human 18+ yrs. Incidence of sensitivity to fragrances in a perfume screening series in 241 consecutive patients (180 female, 61 male) from October 1981-June 1983. The Finn chamber technique was used. Vehicle was PMF (yellow soft paraffin) No further information was provided. 2 % sensitization effects, 0.4% incidence of sensitivity (1/241 male). (Ferguson,1984) Route: skin. Species: human 18+ yrs. Patch tests were carried out on 34 patients with contact allergy to hair cosmetics, 52 with allergic contact dermatitis of the eye lids, 144 with positive reactions to fragrance mix and 54 with contact cheilitis. Subjects ages ranged from 30 to 67 years old and approximately 64% were over 40. Vehicle was white petrolatum. Materials with “no effect” in the results were not necessarily tested on all subjects. 2 % no effects. (Angelini,l985a) Route: skin. Species: human 18+ yrs. 79 male and female patients with eyelid dermatitis and 1012 male and female patients with dermatitis at other sites were assessed and given patch tests in the Contact Dermatitis Clinic of St. Michael’s Hospital, Toronto, Canada, between January, 1980 and May, 1987. The first 630 were tested with a modified series of the European Contact Dermatitis Research Group, 450 were tested with a modified series of the, North American Contact Dermatitis Group, and 11 were tested with other chemicals. Seventy of the total were tested with a perfume series. Test materials were applied to the upper back with Al-Test strips or Finn Chambers and Scanpor tape. Patch

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tests were applied on Mondays, Wednesdays, or Fridays and were first read 48 hours after the test materials were applied. When the chambers or strips were removed at the first reading, sites were scored 30 to 60 minutes after the appliance had been removed to allow adequate time for the physical effects of the pressure of the appliance on the skin to resolve. The second reading was scored at 48 hours or 72 hours after the first reading. Reactions were scored according to the International Contact Dermatitis Research Group (ICDRG). in petrolatum. 5 % positive effects, 2.9% positive reactions in all cases (70 patients); no reactions in eyelid cases (19 patients). (Nethercott,1989) Route: skin. Species: human 18+ yrs. Closed 48 hour patch tests were conducted on patients who had previously exhibited positive reactions to Balsam of Peru. The test used a Lysaplast Special patch which consists of 35 mm wide adhesive tape with a 17 mm circular cellophane disc attached to the center of the tape. Test material (in petrolatum, vaseline or alcohol) was applied to the linen that was then placed on the cellophane disc. The patch was then applied to the anterior and lateral thigh surfaces, then covered with adhesive tape. Reactions were read 10 - 20 minutes after patch removal, and at 72 or 96 hours after patch removal. After 7 - 8 days, reactions were again read in 50% of the tested patients. No further information was provided. (HJORTH,1961B) Vehicle was vaseline. 5 % positive effects, positive reactions in 14/115 patients. (Hjorth,1961a) Route: skin. Species: human 18+ yrs. Repeat tests were conducted on 17 patients after an interval of 7 months to 4 years from the original testing. The re-examination involved the repetition of 2-15 patch tests. Each patient had exhibited a positive reaction to balsam of Peru prior to the original testing See original testing in this series for details on vehicles and doses. This paper includes several detailed case reports. (HJORTH,1961B) Positive effects, new positive reactions were seen in 1 case. (Hjorth,1961b) Route: skin. Species: human 18+ yrs. Patch tests were conducted on patients with cosmetic dermatitis, facial melanosis or non-cosmetic dermatitis or eczema. Article in Japanese. Data from English tables only. Vehicle not given. 5 % positive effects, in 2/225 patients. No effect in 46 controls. (Nishimura, 1984) Route: skin. Species: human 18+ yrs. A total of 658 patients (61.4% women and 38.6% men in the working age group) with hand eczema, that were referred to 1 of 3 participating clinics (Gentofte, Malmo and Odense), were tested with the European standard series and a selection of 14 fragrances found in household products from March 2001 to September 2002. Of the 658 patients tested, 67 (10.2%) reacted to the fragrance allergens used. Patch tests were applied to the upper back for 2 days using Finn Chambers on Scanpor tape. ICDRG recommendations were followed. Readings were taken on day 2 and/or day 3-4 and on day 7. A positive patch test is defined as a +, ++ or +++ reaction. A + reaction was defined as homogeneous erythema and infiltration, however only erythema was not. In addition, on the first day of patch-testing, all subjects were asked to fill out a short questionnaire. The present study does not contain patch-test data from patients without hand eczema. Subjects: 254 Male 404 Female Vehicle was white petrolatum. 5 % positive effects, One patient (0.2%) had a positive patch-test reaction to the test material. (Heydorn,2003a) Route: skin. Species: human 18+ yrs. From 1995 to 1998, 2273 patients with contact dermatitis were tested with a standard series, which included Peru balsam (BP). Of the 445 BP positive patients, those having at least a 1+ or greater reaction to BP were asked to participate in an additional test with the BP series, which consisted of 20 single BP constituents. 102 patients agreed to be tested with the BP series. The patients were 70 females and 32 males, with ages ranging from 18 - 88 years old. The 102 patients were tested 1 or 2 weeks after the results of the standard series had been obtained. The patch tests were applied to the backs of the patients for 24 hours, using Finn Chamber on Scanpor and self-adhesive tape. The vehicle was white petrolatum. Readings were conducted after 24 and 72 hours, according to the International Contact Dermatitis Research Group (ICDRG) rules. Only the 72-hour readings were used for the evaluation. 5 % sensitization effects, 4 patients tested positive, exhibiting 3 + and 1 ++ reactions. (Hausen,2001)

Route: skin. Species: human 18+ yrs. Consecutive hand eczema patients (n=315) were patch tested

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with test compounds in petrolatum. Pure petrolatum was used as the control substance. [no further experimental detail provided], in petrolatum. 5 % no effects. (Heydorn,2002) Route: skin. Species: woman. A case of pigmented contact dermatitis attributed to musk moskene in a cheek rouge is presented. Closed 48 hour patch tests with, Finn chamber and Scanpor tape, were conducted on a 42 year old female patient after the occurence of itchy face hyperpigmentation. The vehicle was petrolatum. Reactions were assessed 1 and 24 hours after patch removal No further information provided. Subjects: 1 Female No effects, (Dose not given). (Hayakawa,1991) Route: skin. Species: human 18+ yrs. Tests were conducted at 8 worldwide centers on 142 patients who had previously reacted to 25% balsam Peru in petrolatum The dose and vehicle were not given. Positive effects, in 10 patients. (Mitchell,1976) Route: skin. Species: human 18+ yrs. Tests were conducted on a 25-year-old man after the occurence of an acute bullous eruption from treatment with a 30% benzyl benzoate preparation. The vehicle was petrolatum. Immunotoxicology, direct immunofluorescence on perilesional skin showed linear deposits of IgG and complement C3 along the basement membrane zone. 1 % No effects. (Stransky,1996) Route: skin. Species: woman. Tests were conducted with 30 vulval vestibulitis patients. All patients were patch tested with European standard series of contact allergens and a special series relefant to perianal and vulval disorders. Sanitary wear, condoms and other potential allergens identified by the patients as causing aggravation were also included. Reactions were graded on Days 2 and 4 The doses and vehicles were not given. No further details were provided. Subjects: 30 Female No effects. (Nunns, 1997) Route: skin. Species: human 18+ yrs. 500 mg of bark samples of the patch-tested twigs were extracted w/ 10 ml of 0.1 M borax in water. After extraction, 2 ml were made acidic w/ 0.1 M HC1 to pH 3-4. The acidic sol’n was extracted by diethyl ether and the extract was analyzed for aromatic acids. Another 2 ml of borax extract was acetylated w/ acetic anhydride (in pyridine) and the phenolic compounds were simultaneously extracted into the hexane phase. Extracts of the bark from the patch-tested twigs were analyzed by gas chromatography and mass spectrometry. 4 chemicals were tentatively identified, i.e. benzyl benzoate, benzoic acid, salicylaldehyde and salicyl alcohol by comparing their mass spectra to the spectra given in the NB library. All chemicals were patch tested on subject described in sub-ref 01. in petrolatum. 5 % no effects. (Jolanki,1997) Route: skin. Species: human 18+ yrs. Closed patch tests in female patients with and without cosmetic dermatitis and facial melanosis using fragrance materials, essential oils, preservatives and lanolin. A control group was also used. Vehicle was vaseline. Article in Japanese. Information derived from English abstract and Tables. 99.4% pure; in vaseline. 2 % no effects. 5 % positive effects, cosmetic dermatitis in 1/64 (1.6%). (Ishihara,1979) Route: skin. Species: human 18+ yrs. The North American Contact Dermatitis Group conducted a multicenter study in North America during a 64-month period from 1977-1983. 713 patients (561 female and 152 male) with cosmetic dermatitis were identified among 13,216 patients with contact dermatitis. Ingredient patch test were perfomed on 56% of the cosmetic dermatitis patients using the Al-Test strips or Finn Chambers to the upper back for 48 hours. Reactions were read at patch removal and 24 hours after patch removal. In most centers, additional readings at 48 or 72 hours after the patch removal were also made. When photoallergy was suspected because of morphologic features and lesion distribution, photopatch tests were done with standard techniques See method reference for details. Results from the first 40 months were previously published. Dose and vehicle not provided. (NACDG & FDA,1977-1980) Positive effects, in 1 patient. (Adams,1985) Route: skin. Species: man. A 22-year-old white male was patch tested after the occunence of dermatitis in the axillary area after use of a solid roll-on antiperspirant Vehicle and other test details not given. 2 % no effects. (Larsen,1983) Route: skin. Species: human 18+ yrs. Summary of the frequency of moderately positive reactions in patch tests using cosmetic ingredients and fragrance materials on eczema and dermatitis patients

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between 1978 and 1986. Article in Japanese. Information derived from English Tables. Vehicle not reported. Results are for total (eczema and dermatitis) patients. Same study as Location 5585. 99.8% pure. 5 % positive effects, 0.4% (2/459). In 1 case, inflammation persisted for over 2 weeks. Control results for Benzyl benzoate were 0% (0/67). (Itoh,1988) Route: skin. Species: human 18+ yrs. Closed patch tests were conducted on contact dermatitis patients with the test material in white petrolatum. No further details were provided. Article in Japanese; data obtained from English abstract and tables only. 5 % no effects, in 69 patients. (Ishihara,1977) Route: skin. Species: human 18+ yrs. Patch test were conducted from 1978-1985 with cosmetic ingredients, on patients with eczema or dermatitis. Article in Japanese. Information obtained from English abstract and tables. No further details provided. Same study as Location 7115. 5 % positive effects, in 2/383 eczema or dermatitis patients and in 0/60 control patients. (Itoh,1986) Route: skin. Species: human 18+ yrs. Seventy-three patients with widespread eczematous dermatitis following treatment of scabies with topical scabicides for not less than 10 days were patch tested with the European standard series, with mesuiphen (5% pet.) & with benzyl benzoate (5% pet.). Testing & result reading techniques were those described by Fregert et al. (1969). 5% pet. Positive effects, in one female patient. Simultaneous skin test with balsam of Peru (which contains benzyl benzoate) was negative in this patient. (Meneghini,1982) Route: skin. Species: human 18+ yrs. 465 patients with dermatitis were patch tested with 29 preservatives. 7 Dermatology clinics were involved. Patients ages varied between 2 yrs old & 80 yrs old. ARTICLE IN FRENCH; INFORMATION TAKEN FROM ENGLISH ABSTRACT AND TABLES. Positive effects, 7 patients reacted. (Meynadier,1982) Route: skin. Species: human 18+ yrs. Patch tests were conducted with ingredients of Balsam of Peru on 53 - 103 subjects. No further information provided. Vehicle was vaseline. 5 % positive effects, in 12/103 subjects. (Hjorth,1961) Route: skin. Species: human 18+ yrs. A 24 to 48 hour occluded patch test was conducted on 30-200 patients. The test material concentration was 0.05 - 0.5% in a vehicle which was either a base cream or 99% ethanol (type of vehicle per test material was not specifically identified). Patch tests were always carried out with both perfumed and non-perfumed cream base at the same time. Patches consisted of a piece of 1 cm2 lint with a 2 cm2 cellophane disc placed on the lint and then covered with 4 cm2 plaster. The patches were applied to the back, the forearm and the inside of the upper arm for 24 to 48 hours. Reactions were evaluated 30 minutes after removal of patch. The skin reaction was graded as follows: - No visible reaction; +- Slight erythema; + Erythema; ++ Erythema and swelling or marked erythema. The period of study extended over 4 yrs and 3 months and patch testing was performed 1-3 times per month; September to May (summer months were excluded). The total number of subjects was 4737 (2341 Japanese men and 2396 Japanese women aged 16-45, mostly 17-30, living in Tokyo and Osaka). Volume dose not provided. 99% purity. No effects, 0/111. (Takenaka,1986)

Route: skin. Species: human 18+ yrs. A repeat insult patch test was conducted to determine if the test material would induce dermal sensitization in human subjects. 108 subjects, 36 males and 72 females completed the study. During the induction phase a webril/adhesive patch (25 mm Hill Top Chamber System) was used occlusively. 0.3 ml of test material was applied to each designated patch. The test sites were wiped with a dry Kimwipe prior to each patching. The left side of the back was used for the test area during the induction phase. Patches remained in place and were kept dry for approximately 24 hours, after which time they were removed. A 24 hour period, during which no test materials were applied, followed the removal of the Monday and Wednesday patch applications; a 48 hour period followed the Friday patch applications. On Mondays, Wednesdays and Fridays, the test sites were observed and any reactions scored and recorded. The identical test site was then repatched until 9 induction patchings were completed over a period of approximately 3 weeks. A rest period of approximately 2 weeks followed the last induction patching. No test materials were applied during the rest period. At the challenge phase, the original induction test sites were observed and each subject

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queried as to whether any reaction was experienced during the rest period. The right side of the back was used for the virgin test site for the challenge phase. Patches were applied as in the induction phase and kept in place for 24 hours after which time they were removed and the challenge site scored. The original test sites were also observed. Scoring of the test sites was done at 48, 72, and 96 hours post- patching using the modified scoring scale of the International Contact Dermatitis Research Group System, 1986. Reactions were scored based on the following scale: No visible reaction 0; faint, minimal reaction +1-;erythema-1; intense erythema,induration-2; intense erythema, induration, vesicles-3; severe reaction with erythema, induration, vesicles, pustules-4. In addition, edema (E) was recorded if present. A rechallenge was conducted approximately 12 weeks later on one female subject. During rechallenge, the patient was patch tested with the original test material in a vehicle of 1:3 EtOH:DEP, DEP and EtOH alone. The subject was also tested with each vehicle individually (no test material). All test materials were applied occluded and semi-occluded. Following the rechallenge the test materials were sent for analysis to confirm their purity by GCMS. Test conducted according to HRL Protocol #100RIFM and HRL Standard Operating Procedures. Subjects: 36 Male 72 Female Study Length: 6 weeks 50 % no effects, Under the conditions of this study, repeat insult patch testing with the test material in a vehicle of 1:3 EtOH:DEP and the vehicle alone induced a 3-level irritation plus edema reaction in one subject. At re-challenge, the female subject had the same reaction when tested with 50% test material in EtOH:DEP and when tested with 1:3 EtOH:DEP alone. However, 50% test material in DEP or 50% test material in EtOH did not produce any irritation or edema. Preliminary results indicated that test materials were free of contamination. After a review of the data by the REXPAN, it was concluded that benzyl benzoate was not a sensitizer. (RIFM,2004) [Harrison,2004] Route: skin. Species: human 18+ yrs. A rechallenge was conducted on one subject who exhibited a 3- level plus edema reaction during an original repeat insult patch test (see RIFM# 47159) using 25 mm Hill Top Chambers. During the rechallenge the patient was patch tested with the original test material in a vehicle of 1:3 EtOH:DEP, DEP and EtOH alone. The subject was also tested with each vehicle individually (no test material) using 25 mm Hill Top Chambers and semi-occlusive patches. Methodology and scoring system not reported. Subjects: 1 Female 50 % no effects, Subject again reacted to both benzyl benzoate in DEP:EtOH (3:1) and to DEP:EtOH (3:1) when tested alone. The subject did not react to saline or 50% test material in DEP, 50% test material in EtOH, EtOH alone, or DEP alone. It was concluded that benzyl benzoate had not produced sensitization. Upon a review of this data, the REXPAN agreed that benzyl benzoate was not a sensitizer. (RIFM,2004) [Harrison,2004a] Route: skin. Species: man. 20 healthy adult males completed the study. Vehicle was petrolatum. Test material was applied to the volar forearm sites under occlusion for a total of five 48 hour periods, always to the same site. The 4th and 5th applications were preceded by 24 hour occlusive applications of 5% aqueous sodium lauryl sulfate. Following a 10 day rest period, challenge patches of test material plus a petrolatum control were applied under occlusion to the backs of each subject following 1 hour pretreatment with 10% aqueous sodium lauryl sulfate. Challenge patches were read at 48 and 72 hours No pre-irritation patch study was conducted. HKO. 30 % no effects. (RIFM,1970) [Kligman,19701 Route: skin. Species: human 18+ yrs. A maximization test was carried out with test material in petrolatum on 26 healthy Japanese-Americans. Application was under occlusion to the same site on the forearms of all subjects for five alternate-day 48 hour periods. Patch sites were pretreated for 24 hours wtth 5% aqueous sodium lauryl sulfate (SLS) under occlusion for the initial patch only. Following a ten to fourteen day rest period, challenge patches were applied under occlusion to fresh sites for 48 hours. Challenge applications were preceded by 30 minute applications of 5% aqueous SLS under occlusion on the left side whereas the test material was applied without SLS treatment on the right side. Additional SLS controls were placed on the left and petrolatum on the right. (Kligman,1966) 79-20- 1OJ.20 % sensitization effects, 1/26 allergic reaction was observed. (RIFM,1979) [Epstein,1979] Phototoxicity

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Route: skin. Species: guinea pig. The animals used were female Heartley strain albino guinea pigs, microbial grade, clean purchased from Japan SLC, Inc., weighing 335-365 g at starting of experiment. Hair on the back of the animal was cut with an electric hair clipper and an electric shaver. Four hours after depilation, the test agent dissolved in acetone was applied on a circle of 1.5 cm in diameter in the depilated area on both sides of the animal. A total of 8 such applications were made, that is, one spot each of a 50% , 30% and 10% solution on the right and also on the left side. Immediately after application, one side was covered with aluminum foil. The other side was irradiated with five Toshiba model FL-40 BLB lamps (40w x 6, emission: 320-400 nrn) that had been equipped with window glass filter to eliminate radiation below 320 nm. The distance from the light source to the skin was 10 cm. Irradiation was continued for 70 mm. The test sites were observed for reaction 24 and 48 hr after irradiation. The intensity of skin reaction was graded from 0 to 8 scale. The criteria and scoring system for assessment of dermal reaction are those of Draize, J.H. (1959). The scores for erythema and edema at 24-hour reading were totaled for the five test guinea pigs (5 values) and this total was divided by 5 to give the primary irritation index of the test material. 50 % no effects, Benyl benzoate was classified as “non-photoirritant’ up to 50% concentration. 30 % no effects. 10 % no effects. (Takasago,1981) Route: in vitro. Species: yeast. Candida utilis was exposed to test material on duplicate plates. The material was applied to filter paper plates and placed on culture media that had been seeded with yeast. One set was exposed to UV-A from a battery of four fluorescent tubes (Thorn: Ultraviolet non filter 20 W; irradiance approximately 1.2 mW/cm2) for 48 hours and the other kept in the dark. If no effect, a test using UV-B was conducted. This consisted of a 15-minute exposure (approximately 1350 mJ/cm2) to the emission from a battery of four Westinghouse FS2O sunlamp fluorescent tubes (irradiance approximately 1.5 mW/cm2) which did not kill the yeast, but by which a phototoxic effect of this wavelength region could be demonstrated. Vehicle was paraffin. 10 % no effects. (Addo,1982) Route: in vitro. Species: human 18+ yrs. Photohaemolysis was studied using washed human red blood cells suspended in barbitone buffered saline, pH 7.4, at a dilution of 1:500. 1 ml of test material at various dilutions in ethyl alcohol were added to 99 ml of the red blood cell suspension. Aliquots of 5 ml, forming cell monolayers in petri dishes, were exposed to UV-A or UV-B. UV-A exposure was from batteries of four fluorescent tubes (Thorn: Ultraviolet non filter 20 W; irradiance approximately 1.2 mW/cm2). UV-B exposure consisted of approximately 1350 mJ/cm2 from a battery of four Westinghouse FS2O sunlamp fluorescent tubes (irradiance approximately 1.5 mW/cm2) which did not kill the yeast, but by which a phototoxic effect of this wavelength region could be demonstrated. Exposures for UV-A or UV-B were for limited times up to 3 hours. After the exposure the dishes were kept in the dark for 30 minutes and then the suspensions were centrifuged, the release haemoglobin in the supernatant being determined as cyanmethaemoglobin. Controls were run using ethyl alcohol alone and the test material without exposure to radiation. 0.1 % no effects. (Addo, 1982) Route: in vitro. Photosensitized oxidation of histidine was measured. Test materials in ethyl alcohol were added to 0.61 mM L-histidine monochloride monohydrate in 0.01 M sodium phosphate buffer,, pH 7.4. The solutions were exposed to UV-A (from two parallel FR 74-T12 PUVA fluorescent tubes); and irradiance approximately 3 mW/cm2 of UV-B. After exposure of 2-3 hours, remaining histidine was determined by a modified Pauly reaction (Sluyterman, 1960). The final solution was read at 530 nm against a prepared blank solution and the amount of histidine present determined by reference to the absorbance obtained with appropriate standard solutions Concentrations of test materials not provided. No effects. (Addo,1982) Route: in vitro. Species: human 18+ yrs. A study was conducted to evaluate the phototoxic potential of a series of test materials invitro with a photohemolysis test using suspensions of human erythrocytes exposed to radiation sources rich in UV-A (320-460 nm) and UV-B (275-365 nm) in the presence of the test materials. Red blood cells obtained from healthy human volunteers were washed and suspended at a dilution of 1:200 in TCM buffer containing 0.03% human albumin. A 0.4 ml volume of this suspension and a correspondingly prepared erythrocyte-free sample (blank) were incubated with 0.1 ml of test material preparations for 1 hour at 37 C. Both the test material sample nd the blank were

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exposed to 0, 5, 25, or 50 J/cm2 UVA or to 0, 500, 1000 or 2000 mJ/cm2 UVB from the TL 20 W/12 light bulbs. During irradiation, samples were kept in a shaking bath at 37 C. 100% heamolysis was obtained by exposure of the erythrocytes to distilled water. After an incubation period of 30 minutes in the dark supernatants were recovered by centrifugation. The released heamologlobin in the supernatants was determined after incubating the samples for 15 minutes in Drabkin’s solution. Hernolysis was measured by reading the absorbance values at 550 nm with a MR 700 microplate reader, and photoheamolysis was calculated as a percentage of total heamolysis. Values of >5% were regarded as significant photohemolysis. Summary Under the conditions of this study, test material did not induce photohemolysis. No effects, Test material did not induce photohemolysis. (Placzek,2007) Skin absorption Route: in vitro. Species: human 18+ yrs. Lower abdominal skin was excised from a human cadaver during autopsy, kept at t20 degree celsius and thawed prior to usage. Subcutaneous tissue was removed. Using a modification of Baumbergerzs method, the subcutaneous tissue was removed and the epidermis was separated from the dermis. The upper surface of the epidermis was fixed to a glass tube which was then placed inside one arm of a U-shaped glass chamber. Approximately 5 ml of saline was added to the chamber and was in complete contact with the bottom of the epidermis. A 0.2 ml aliquot of test material was applied to the top of the epidermis. To avoid evaporation, parafilm was placed over the mouth of the glass tube. The chamber was kept at 21 C and 55% relative humidity for 72 hours. The glass tube was removed from the glass chamber at 72 hours and the saline was poured into a test tube. The U-shaped chamber and the bottom of the epidermis attached to the glass tube were both washed 3X with saline and poured into the same test tube. The final volume of the the tube of original saline and saline used for washing was approximately 10 ml. Saturated salt water and ether were added to the flask and mixed vigorously. The compound was extracted in ether, dehydrated filtered and condensed. A 2 ul aliquot of the condensed sample was injected into a Shimazu GC-6A gas chromatograph. Expt. was repeated 6X. The amount of compound (ug) which penetrated human epidermis was calculated by the following formula: ug (Peak area test sample/peak area standard sample) x concentration of compound in standard sample (ug/ml) x volume test sample (ml). Percent penetration calculated using formula: %penetration = (amount penetration (ug)/(200 ul x specific gravity x 103) x 100 Because the experiment was repeated six times, the percent penetration +/- standard error calculated. Solubility of test material in water and octanol was also conducted. The results suggest that the degree of penetration through human epidermis is not related to sensitization or allergenic potency. Skin absorption, Penetration = 0.018 +1- 0.002%. Solubility in water = <1 ppm. Solubility in octanol = freely soluble. (Jimbo,1983) Route: non-specific parenteral. Species: monkey. Urinary excretion of radioactivity was measured after a single dose to 4 rhesus monkeys. This test was conducted to provide a correction factor for the associated skin absorption studies. [7-14C)benzyl benzoate. Urinary excretion was 65.3 +f 13.4%. (Bronaugh, 1990) Route: skin. Species: monkey. Urinary excretion of radioactivity was measured for 5 d after a single occluded application of test material to 4 female rhesus monkeys. Vehicle was acetone. Material was applied at 4 ug/cm2 to a 1 cm2 area of lightly clipped abdominal skin. The application site was washed & re-occluded after 24 h. [7-14C]benzyl benzoate. Skin absorption, 71.2 +1-4.4% of the applied dose was absorbed with plastic wrap used for occlusion. 64.7 +/- 10.2% of the dose was absorbed with a glass chamber used for occlusion. (Bronaugh,1990) Route: skin. Species: monkey. Urinary excretion of radioactivity was measured for 5 d after a single unoccluded application of test material to 4 female rhesus monkeys. Vehicle was acetone. Material was applied at 4 ug/cm2 to a 1 cm2 area of lightly clipped abdominal skin. [7-l4Cjbenzyl benzoate. Skin absorption, 57.0 +1- 10.4% of the applied dose was absorbed. (Bronaugh,1990)

Route: skin. Species: monkey. Urinary excretion of radioactivity was measured for 5 d after a single unoccluded application of test material to 4 female rhesus monkeys. Vehicle was a moisturizing lotion.

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Material was applied at 4 ug/cm2 to a 1 cm2 area of lightly clipped abdominal skin. [7-l4Clbenzyl benzoate. Skin absorption, was significantly greater then that noted with acetone as the vehicle. The moisturizing lotion was used at 10 mg lotion per cm2 skin. (Bronaugh,1990) Route: in vitro. Species: guinea pig. Test compounds were tested as to their penetrating agent properties by assessing depth to which various ‘active principles” such as malachite greeen, Rhodamine B., orcein penetrated skin of animals in the presence of a test compound. Histological findings in epithelium (epi), hair follicles (hi), corium (c), and subcutis (Sc)were evaluated by following criterion: + (strong coloring/fluorescence), (+) (noticeable coloring/fluorescence), (slight coloring/fluorescence), and - (no coloring/fluorescence). Genuine penetration defined as active principle detected in corium or subcutis. t duration of action. Vehicle usually 50% ethyl glycol. 50 % no effects, rhodamine, t=2 hr, epi,hf,c,sc: =,=,-,-, resp. (Meyer,1965) Pharmacokinetics and metabolism studies Route: in vitro. Species: human 18+ yrs. The ability of human plasma derived arylesterase to hydrolyse the test material was evaluated. Most test materials were solubilized using Tween 80. The Warburg manometric technique was used for esterase determination at pH 7.4, during which the initial substrate concentration was 8 mM. Not hydrolysed. (Augustinsson,1962) Route: gavage. Species: dog. A single dose of test materia1was given to fasted female dogs. Urine was collected for 5 to 6 days. 2 ml/kg , The oral administration of 2 ml/kg benzyl benzoate to dogs led to creatinuria and the urinary excretion of glycuronate conjugates. Urinary acidity, total nitrogen and phosphate increased. 1 ml/kg , The oral administration of 0.5 mI/kg benzyl benzoate to dogs led to creatinuria and the urinary excretion of glycuronate conjugates. Urinary acidity, total nitrogen and phosphate increased. 0.5 mI/kg , The oral administration of 0.5 mi/kg benzyl benzoate to dogs led to creatinuria and the urinary excretion of glycuronate conjugates. (Pack, 1922) Route: oral. Species: human 18+ yrs. Urinary hippuric acid levels were measured after a single dose of test material. Urine was collected for 6 hours. 2.02 gm , Urinary hippuric acid accounted for about 90% of a single oral dose of 2.02 gm benzyl benzoate to a human subject. 1 gm, Urinary hippuric acid elimination during the first 6 hours after administration accounted for 80% of a single oral dose of 1 gm benzyl benzoate to a human subject. In a second subject, urinary hippuric acid accounted for 71% of the dose during the first 6 hours and an additional 14% of the dose during the second six hours after dosing. The urinary elimination of hippiric acid was somewhat slower in a third patient who became sick with influenza. (Snapper,1925) Route: in vitro. Species: human 18+ yrs. The rate of hydrolysis of the test material by 80% human plasma was studied. Vehicle was acetonitrile. The tl/2 for the in vitro hydrolysis of benzyl benzoate to benzoic acid by human plasma was 19 minutes. (Nielsen,1987) Route: in vitro. One ml of essential oil constituents, eugenol (1.065 g), benzyl acetate (1.057 g) & benzyl benzoate (1.118 g), in 10 ml isopropyl alcohol solution were irradiated with a low pressure mercury lamp (253.7 nm) or exposed to sunlight in the presence of atmospheric 02 with & without photosensitizer, (10 mg) acetophenone or benzophenone. The GC retention index & the MS fragmentation pattern of each component of the samples were compared to those of the authentic compound to identify products in the reaction solutions. Major photochemical products benzoic acid, benzyl alcohol & 2-methyl-1-phenyl-1,2-propanediol proposed to form from benzoyloxyl & benzyi radicals. Sunlight exposure resulted in numerous additional products. (Schunk,1986a) Pharmacology Species: rat. Benzyl benzoate, tested as the solvent, was found to produce a rapid, significant and lasting increase in the blood flow of the testis Data from English abstract only. Further details not given. Reproductive effects. (Joffre, 1973)

Route: oral. Species: dog. The effects on blood pressure & pulse of orally administered test materials (by stomach tube) washed down with 20 - 50 ml H20 were examined by the indirect & direct methods

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in unanesthetized & anesthetized dogs, respectively, pure form & in 20% alcoholic solutions. No effects, on blood pressure or pulse rate. Moderate doses of 20% in alcohol caused fall in blood pressure probably due to alcohol. (Gruber,1924) Route: oral. Species: dog. The effects of benzyl esters on the alimentary canal of unanesthetized dogs were noted. Drugs given by stomach tube followed by 20 - 50 ml H20. Animals then turned loose in runways & observed, pure form & in 20% alcoholic solutions. Gastrointestinal tract, emetic & cathartic action in some animals due to local irritation. (Gruber,1924)

Route: subcutaneous. Species: rabbit. One or two rabbits per dose were given 1 injection of test material at 1 to 2.5 ml/kg or 4 daily injections at 0.25 mi/kg. Effects on leucocyte counts were measured for up to 11 days. Mortality and clinical signs were monitored. The vehicle was olive oil. 2 mi/kg no effects. .5 ml/kg clinical signs, lethal. 1 mi/kg no effects. 1.5 mlfkg no effects. 0.25 ml/kg no effects. (Heller,1920) Route: gavage. Species: rabbit. The effects of test material on the tone of the vesical sphincter were studied using anesthetized rabbits. The vehicle was alcohol. A single dose was given to normal or morphine treated rabbits. 20 % positive effects, sphincter tone was reduced in normal and morphine treated rabbits. (Stater,1922) Route: oral. Species: human 18+ yrs. The use of benzyl benzoate as a treatment for dysentery in 9 patients was reported. A 20% alcoholic solution was given in a small amount of cold water 3x/day. The dose was 20 to 30 drops of the alcoholic solution. Additional treatments were used along with the benzyl benzoate. Positive effects, a definite and favorable influence on the course of endamebic dysentery was seen without any undesirable side effects. (Haughwout,1919) Route: in vitro. Species: rat. The depressor or relaxing effect of the test material on active sections of intestines was studied. The vehicle was alcohol and Locke’s solution The dose was 50% in alcohol for liquid esters and 20% in alcohol for solid esters. Positive effects. (Lynn,1926) Route: intravenous. Species: dog. The effects of test material on intestinal muscles was studied. Effects on blood pressure and respiration were also measured. Solid test materials were diluted with sweet almond oil (1:5) and injected at a temperature slightly above that of the body at 39-40 C. Liquids were administered neat No further details provided. 5 % positive effects, decreased blood pressure and respiration with intestinal muscle relaxation. (Nielsen,1921) Route: oral. Species: human 18+ yrs. Plethysmograms were taken after the injestion of XXX, L, XC or CXX drops of a 25% alcoholic solution of test material diluted in water. Measurements were taken for 60 minutes. No effects. (Simici,1926) Route: intramuscular. Species: human 18+ yrs. Blood pressure and plethysmographic readings were taken after a single injection of test material. The vehicle was oil of vaseline. Normal and hypertensive subjects were used. 5 ml blood pressure, 5 ml of a 20% solution produced a pronounced vasodilation and a decreased blood pressure. 3 ml positive effects, 3 ml of a 50% solution produced a moderate vasodilation. 8 ml blood pressure, 8 ml of a 20% solution produced a very strong vasodilation followed by a vasoconstriction. Blood pressure was reduced throughout.blood pressure, vasodilation and decreased blood pressure were seen in a patient with hypertension but without arteriosclerosis. No effects were seen in patients with arteriosclerosis. 2 ml positive effects, 2 ml of a 20% solution produced a pronounced vasodilation but no change of blood pressure. (Simici,1926) Route: oral. Species: human 18+ yrs. The effects of the test material on the blood pressure of 16 patients with arterial hypertension were studied. A 20% acoholic solution of the test material was given in 25- and 30-drop doses four times a day for 4 to 18 days. 20 % no effects. (Gruber,1924b)

Route: skin. Species: human 18+ yrs. The efficacy of the test material against scabies was evaluated, on 21 patients with clinically diagnosed scabies. Scabies was defined as the association of pruritus and at least one classical burrow. The patients were instructed to administer the test material onto their entire body except the head, and to keep the body unwashed for a 12-hour period. Following the initial

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12-hour period, the patients were to shower and then reapply the test material in the previous manner. Assessment of adverse reactions was conducted on day I and 2, and cutaneous examination on day 7, 14 and 30. The criterion for recovery was defined as a complete disappearance of initial skin lesions and pruritus. 10 % positive effects, No patients were recovered on day 7, but 3/21 and 10/21 patients were recovered on days 14 and 30, respectively. A mild and transient increase in pruritus on day 2 was reported by 5 patients. (Glaziou,1993) Route: skin. Species: human 18+ yrs. The efficacy of the test material against scabies was evaluated, on 29 patients with clinically diagnosed scabies. Scabies was defined as the association of pruritus and at least one classical burrow or with multiple pustular eruptions, vesicles or nodules distributed over both flexor or extensor surfaces of the extremities as well as the abdomen, back, perineum and buttocks. Skin scrapings were taken from the affected areas and microscopically examined for the presence of eggs, larvae or adult mites. The patients were instructed to administer an emulsion of the test material on their entire body except the head for 72 hours. The patients were re-examined every week for 4 weeks after treatment, at which time skin scrapings were taken again. The criterion for recovery was defined as a complete disappearance of initial skin lesions and pruritus. 25 % positive effects, On days 7, 14 and 30, recovery was observed in 3/29, 10/29 and 14/29 patients, respectively. A mild to moderate increase in irritation and pruritus by day 2 was reported by 7 patients. (Nnoruka,2001) Route: skin. Species: human 18+ yrs. The efficacy of the test material against scabies was evaluated, on 31 patients with diagnosed scabies. The test material was topically administered over the whole body surface at bed-time for 3 consecutive nights, and following the treatment period, the patients were bathed with soap and water. The patients were examined after the treatment period, as well as after 1 and 3 weeks, to asses the tolerance and efficacy of the test materials. 20 % positive effects, By day 22, 100% (31/31) of the patients were cured. Burning in 6/31, redness in 6/31 and exsiccation in 2/21 patients was observed. 10 % positive effects, By day 22, 100% (31/31) of the patients were cured. Burning in 3/31 and redness in 3/31 patients was observed. (Haustein,1989) Route: skin. Species: human 18+ yrs. A 30-year old female patient was presented with a 3-month history of scaly plaques on her scalp and posterior neck. Slight pruritis, extensive hyperkeratinization with crusting, and crusting, erythematous scaling plaques on the scalp were also present. A biopsy specimen showed multiple subcorneal burrows containing mites and scabies was confirmed by the presence of Sarcoptes scabiei in skin scrapings and preparations made from the skin lesions. The test material was applied to the entire skin surface once a day for 2 days, then reapplied 1 week later. Skin scrapings were conducted after 2 weeks. 25 % positive effects, At the 2-week skin scraping, no mites were found. Within 1 month, a complete disappearance of the cutaneous lesion of crusted scabies was noted. (Chutimunt, 1996) Route: intravenous. Species: dog. A single injection of emulsified test material was given to a dog. Effects on blood pressure, heart rate and respiration were studied. 20 mg blood pressure, decreased blood pressure was seen. (Macht,1920) Route: oral. Species: human 18+ yrs. A beneficial effect was seen with the use of benzyl benzoate as a treatment for hypertension. The most effective form of adminstration was as a 20% alcoholic solution in water or milk. The dose was 20 to 30 drops of the solution, 3 or 4 times per day. Blood pressure, no adverse side-effects were seen even with up to 2 years of treatment. (Macht,1920) Route: intravenous. Species: dog. Respiration, right carotid pressure & pylorus contraction were measured in a dog weighing 13 kilos after inj. of test cpd soln’s into the femoral vein. [No further experimental detail provided]. 10% solution. 2 ml no effects. 4 ml positive effects, depression of respiration, fall in blood pressure & a very slight inhibitory effect on pyloric contractions noted. An inj. of 1 ml adrenaline produced a very obvious inhibition of the pyloric contractions. 6 ml lethal, complete relaxation of the pylorus noted, as well as death of the animal apparently from central respiratory paralysis. (Mason, 1920)

Route: intravenous. Species: dog. In order to measure effect of test cpd on intestinal wall, a small

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rubber balloon was filled w/ water & placed w/in the intestinal lumen & connected by rubber tubing wI a burette which carried a stopper in the upper end. From the stopper a rubber tube led to the recording tambourine. A small dose Of barium chloride was inj. to stir up marked contractions of the intestine followed by an inj. of test cpd. After an interval of about a mm, there occurred a relaxation of the

intestine and @ this pt a further inj. in the same amt . was given. 2 ml positive effects, depressed respiration noted & slight lowering of blood pressure but not immediate change in the tone of the intestine observed after 1st dose. After second dose, a slight increase in the tonus of the intestine noted. (Mason, 1920) Route: intravenous. Species: dog. One animal receiving artificial respiration was inj. w/ test cpd and blood pressure & heart acton were monitored. [No further experimental detail provided]. 3 ml cardiovascular effects, a fall in blood pressure noted, the heart @ the same time became weak and, after an interval almost stopped in diastole. After a prolonged series of feeble, irregular beats, however, heart again recovered. (Mason,1920) Route: intravenous. Species: dog. The action of test cpd on respiration, spleen volume & blood pressure of an animal weighing 6.5 kilos was evaluated. One ml of barium chloride was inj. to produce tonus in the spleen. 4 ml positive effects, marked contractions, profound fall in blood pressure & asphyxia noted. (Mason,1920) Route: intravenous. Species: dog. Respiration, kidney volume (oncometer) and blood pressure were monitored after test cpd sol’n was inj into one animal. [No further experimental detail provided]. 7 ml positive effects, same effects as those noted @ 4 ml but more pronounced. 4 ml positive effects, a small shrinkage in kidney volume (due to fall in blood presure noted.). (Mason,1920) Route: intravenous. Species: dog. Test cpd was inj. into one animal & effects of respiration, blood pressure & uterine contraction were monitored. [No further detail provided]. 3 ml positive effects, obvious results on respiration & blood pressure noted (similar to those in other experiments). Effects on the uterus were very slight. 5 ml lethal, positive effects, uterine contractons were stopped, but not until animal died. (Mason,1920) Route: intravenous. Species: dog. Two animals were inj. w/ test cpd & effect on bronchioles as recorded by a special aspiration method was assesed. [No further experimental detail provided], Respiratory tract, a shortening of the amplitude of the lung indicated contraction of the bronchioles. (Mason, 1920) Route: intravenous. Species: dog. One animal was inj. iv w/ ergamine or codeine followed by inj. w/test cpd. Effect on bronchiolar contraction was evaluated. [No futher experimental detail provided.]. 30 ml no effects, no relaxation fo bronchiolar contractoins induced by ergamine noted. 10 ml no effects, no relaxation of bronchiolar contractions induced by ergamine noted. 3 ml no effects, [20% benzyl benzoate made up in 75% alcohol.]. (Mason,1920) Route: intravenous. Species: dog. The action of test cpd on the pulmonary blood presure ws investigated by the foliwoing method. A cannula was inserted into the left pulmonary artery connectd w/ a water manometer, the distal end of which wasjoined to the lower end of a burette. The manometer & tubes to the artery (and the cannula) Were filled w/ normal salt soin. The upper end of the burette was connected w/ a recording tambourin which wrote on a drum. The tambour, uppr part of the burette & the connecting tubes were filled w/ air. Test cpd was inj. near the beginning of the tracing & towards the end. 3 ml positive effects, Near the beginning of the tracing, a typical prolonged fall in carotid pressure was obtained. A temporary rise in pulmonary pressure was succeeded by a few gasping movements. After a 2nd inj., typical fall produced. (Mason,1920) Route: oral. Species: rabbit. Animals (strain, sex, no. unspecified) received test cpd in sterile olive oil and hematological parameters were measured. [No further experimental detail provided]. 3 gm blood effects, diphasic leucocytosis ultimately ending in a late leucopenia noted. 0.05 gm blood effects, a decided leucocytosis appeared a few days after the daily adm. of the drug. Peak or climax is reached rapidly but is not maintained, and there is an even but slow drop which end in mild leucopenia at end of

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one week. 0.1 gm blood effects, a decided leucocytosis was produced. Time of onset much more rapid than that noted w/ lower doses. (Emge,1921) Route: oral. Species: rabbit. In order to determine the effect of a single large dose of test cpd, a large amt. was adm. by stomach tube in olive oil or dilute alcohol. [No further experimental details provided]. 3 gm blood effects, diphasic leucocytosis ultimately ending in a late leucopenia noted. 1.5 gm blood effects, clinical signs, a sharp increase in lymphocytic element which ultimately develops and into a lymphocytosis was noted. At the end of one week animal began to suffer from ‘snuffles”. (Emge,1921) Route: oral. Species: rabbit. In order to determine the effect of a single large dose of test cpd, a large amt. was adm. by stomach tube in olive oil or dilute alcohol. Subsequent doses followed. [No further experimental detail provided]. Blood effects, clinical signs, the animal suffered again and more severely from “snuffles” as w/ an initial dose. Also a marked rise in total leucocyte count noted. [Doses were 1.5 gm (twice) or 3.0 gm (once).]. (Emge,1921) Route: subcutaneous. Species: rabbit. One test animal (strain & sex unspecified) suffering mildy from “snuffles” but otherwise fat, lively, and of smooth hair, was adm. test cpd daily, beginning w/ 100 mg, and adding an equal amt. each day for approx. 22 days. 1000 mg blood effects, after 10th inj, “snuffles” became very acute, leucocytosis broke and remained so for two days, in spite of increased dosage, and then suddenly entered into an extreme phase again. 100 mg blood effects, a diphasic leukocytosis occurred w/ the small mononuclear element predominating. “Snuffles” recurred after 3 days when the animal was given a rest. With ea. subsequent dose, the animal snuffled more severely. (Emge,1921) Route: intravenous. Species: dog. Thirty-two animals (strain unspecified) were anesthetized either wI ether or w/ paraldehyde. Some of the animals were decerebrated under anesthesia and the ether then withdrawn. After anesthesia, the animals were tracheotomized. Respirations were recorded upon a kymograph by placing a pneumograph about the thorax. Blood pressure was taken from a cannula in the left carotid artery. Blood pressure in the superior vena cava also was recorded by passing a cannula down the right jugular vein. The organs of the abdomen were exposed through a mid-line incision for plethysmography. Cardiovascular effects, respiratory tract, acceleration of the rate & decrease in the depth of respiration, congested lungs @ necropsy, permanent increase in pressure of the superior vena cava and lowered arterial pressure were noted. (Gruber,1924a) Route: intravenous. Species: rabbit. Six animals (strain unspecified) were anesthetized w/ urethane. After anesthesia, animals were trachetomized. Respirations were recorded w/a kymograph by placing a pneumograph about the thorax. Blood pressure was taken from a cannula in the left carotid artery. Blood pressure in the superior vena cava also was recorded by passing a cannula down the right jugular vein. The organs of the abdomen were exposed through a mid-line incision for plethysmography. Cardiovascular effects, respiratory tract, acceleration of the rate & decrease in the depth of respiration, congested lungs @ necropsy, permanent increase in pressure of the superior vena cava and lowered arterial pressure were noted. (Gruber, 1924a) Route: intravenous. Species: cat. 22 animals (strain unspecified) were anesthetized w/ urethane. After anesthesia, animals were tracheotomized. Respirations were recorded w/a kymograph by placing a pneumograph about the thorax. Blood pressure was taken from a cannula in the left carotid artery. Blood pressure in the superior vena cava also was recorded by passing a cannula down the right jugular vein. The organs of the abdomen were exposed through a mid-line incision for plethysmography. Cardiovascular effects, respiratory tract, acceleration of the rate & decrease in the depth of respiration, congested lungs @ necropsy, permanent increase in pressure of the superior vena cava and lowered arterial pressure were noted. (Gruber,1924a)

Route: intraperitoneal. Species: dog. Animals (sex and strain unspecified) were anesthetized either w/ paraldehyde by mouth or w/ ether by inhalation. Blood pressure was measured from left carotid artery. Cannulas were placed in ea. ureter & drops of urine secreted were recorded on the kymography by receiving & recording tambours. Kidney, increased urine secretion (171%), and length of latent period

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between inj & beginning of increased secretion was noted. (Gruber, 1925) Route: intraperitoneal. Species: rabbit. Animals (sex and strain unspecified) were anesthetized either w/ urethane by mouth or w/ ether by inhalation. Blood pressure was measured from left carotid artery. Cannulas were placed in ea. ureter & drops of urine secreted were recorded on the kymography by receiving & recording tambours. Kidney, increased urine secretion (171%), and length of latent period between inj & beginning of increased secretion was noted. (Gruber,1925) Route: intraperitoneal. Species: dog. See complete study: Microfilm location #9653. (Gruber,1924c) Route: oral. Species: rabbit. Animals w/ and w/o hyperpyrexia weighing 1200-1800 kg were adm. test substance w/ 50 ml of water. Temperature was measured by reaction w/ a clinical thermometer. Temperature was read one minute before and after treatment. Hyperpyrexia in animals was produced by a number of different methods. 1 ml positive effects, a distinct lowering of temperature in all cases in which temp was abnormally high was observed. (Dose range 1-5 ml). Due to greater dissipation of heat through dilation of blood vessels. (Macht,1929) Route: unreported. Species: rat. Adult albino animals were trained to find their way from the periphery of the maze to the center where food was placed as an incentive. After preliminary training, animals found their way to the center of the maze w/ no errors. The running time & number of errors & other peculiarities of a neuro muscular nature were noted. Animals were then inj w/ test substances and behavior before & after drug adm. was noted. Vehicle was olive oil. 0.1 ml positive effects, average running time = 13 sec (controls = 14.6 sec). Error 0 in both treated & control trials when dosing soln 0.1%. Slowing of movement thought to be due to direct depressant action of test subst. on skel. muscle. 0.4 ml positive effects, average running time = 6.6 sec(controls = 8.0 sec). Error = 0 in both treated & control trials when dosing soln 0.1%. Slowing of movement thought to be due to the direct depressant action of test subst. on skel. muscle. (Macht,1929) Route: in vitro. Species: frog. An animal’s lower extremities wre perfused w/ Ringer’s solution before & after adding a small quantity of test subst. to the sol’n. 0.1 % positive effects, a greater outflow of the perfusing fluid occurred indicating vasodilation. (Macht,1929) Route: in vitro. Species: rabbit. One animal’s ear was perfused according to the method of Krafkoff. The effect of test substances on the blood vessels in the ear were assessed. 0.5 % positive effects, distinct vasodilation was noted. (Macht,1929) Interaction Route: intramuscular. Species: chicken. 0.5 ml of a 1:1 mixture of sesame oil and benzyl benzoate was injected 1/2 inch deep in the right and left pectoral muscles of 6 male Hubbard Crossbred broilers. 2 chickens were sacrificed days 1, 3 and 7 after injection for analysis of gross lesions at the injection sites. No effects. (Hem,1975) Subchronic toxicity Route: skin. Species: rabbit. Test material was applied at 0.5, 1.0, 2.0 & 4.0 ml/kg, daily for 90 days. Animals observed for irritation & sensitization. Hemoglobin values & morphological changes of blood cells were followed in most of the rabbits. All rabbits living more than 2 days examined grossly with selected histopathology See Draize,1948 for original LD5Odata on these materials. The 90 day studies have been entered here since this paper contains more details. 2 ml/kg blood effects, kidney, lethal, reproductive effects, skin effects, rabbits died without showing usual pattern of systemic effects prior to death; gradual increase in leukocytes to 2-1/2x’s normal with increased heterophyle count; slight dermatitis; atrophy of testes; some kidney damage. (Lehman,1955)

Route: subcutaneous. Species: mouse. Groups of 5 adult C57BL/6J mice received 0.1 ml castor oil, 0.1 ml benzyl benzoate or 0.1 ml castor oil + benzyl benzoate (4:1), daily for 4 wks. Mice then systemically perfused & fixed with 3% glutaraldehyde. Adrenal glands also fixed in situ with glutaraldehyde, excised & examined by light & EM. Combination castor oil + benzyl benzoate caused dramatic morphological alterations including changes in mitochondrial shape, size & membrane matrix

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ratio. Also, accumulation of globular structures within large vacuoles in the macrophages. Lethal, mice died within 3 hr of initial injections. (Migally,1979) Route: skin. Species: rabbit. See Lehman, 1955 for publication of full details. (Draize,1948) Route: skin. Species: rat. 3 males & 3 females/dose group. 7 areas on the shaved back were dosed in rotation. Summary 30 day 0.488 g/kg micropathology in organs other than list, skin effects, systemic effects, histopath showed minor grades of hyperplasia of squamous epithelium, degeneration of hair follicles & sebaceous glands, subcut fibrosis; hyperplasia of thyroid. 1.25 g/kg body weight changes, blood effects, micropathology in organs other than list, skin effects, systemic effects, decreased WBC; gained less wt; histopath showed hyperplasia of squamous epithelium, degen hair follicles & sebaceous glands, subcut fibrosis; hyperplasia thyroid. 0.301 g/kg micropathology in organs other than list, skin effects, systemic effects, histopath showed minor grades of hyperplasia of squamous epithelium, degeneration of hair follicles & sebaceous glands, subcut fibrosis; hyperplasia of thyroid. 0.188 g/kg micropathology in organs other than list, skin effects, systemic effects, histopath showed minor grades of hyperplasia of squamous epithelium, degeneration of hair follicles & sebaceous glands, subcut fibrosis; hyperplasia of thyroid. 2 g/kg blood effects, lethal, micropathology in organs other than list, skin effects, systemic effects, 4/6 deaths; decreased WBC; histopath showed hyperplasia of squamous epithelium, degen of hair follicles & sebaceous glands, subcut fibrosis; hyperplasia thyroid. 0.781 g/kg micropathology in organs other than list, skin effects, systemic effects, histopath showed minor grades of hyperplasia of squamous epithelium, degeneration of hair follicles & sebaceous glands, subcut fibrosis; hyperplasia of thyroid. (RIFM,1980) [Moreno,1980] Chronic toxicity Route: oral. Species: mouse. A 7 month study was conducted Data from CA abstract only. Text in French. 800 mg/kg no effects, (No antitumer effect was noted following injection or grafting of guerin tumors into mice although body weight was increased. A slight cytostatic activity was noted). (Ornellas,1965) Carcinogenesis and mutagenesis studies Route: in vitro. Species: Salmonella typhimurium. (Schunk, 1986a) Route: in vitro. Species: Salmonella lyphimurium. Ames test with and without S-9 using strains TA98, TA100, TA1535 and TA1537. Each material was tested at 3 umol/plate, unless otherwise mentioned. Vehicle was ethanol. Materials giving uncertain results in spot test were tested quantitatively. The positive controls were N-methyl-N’-nitro-N-nitrosoguanidin (without activation) and 2- aminoanthracene (with activiation). No further details provided. No effects. (Florin,1980) Route: in vitro. Species: Salmonella typhimurium. One ml of essential oil constituents, eugenol (1.065 g), benzyl acetate (1.057 g) & benzyl benzoate (1.118 g), in 10 ml isopropyl alcohol solution were irradiated with a low pressure mercury lamp (253.7 nm) or exposed to sunlight in the presence of atmospheric 02 with & without photosensitizer, (10 mg) acetophenone or benzophenone. These samples in DMSO were tested in both the plate incorporation & the pre-incubation assays using strains TA98 & TA100, with & without polychlorinated biphenyl-induced rat liver S-9. No effects, no mutagenic activity in either type assay with either strain, with or without S-9. (Schunk,1986a) Developmental and reproduction studies Route: food. Species: Drosophila melanogaster. Food medium for Drosophila melanogaster was prepared with & without benzyl benzoate (4% in ETOH). Virgin Canton-S males & females from stock cultures raised on control medium were collected daily for 4 days & then mated. One grp laid eggs on control food & another on food with benzyl benzoate. Percent egg hatch determined & compared. 4% (v/v) in ethanol. 4 % Developmental effects, the percentage of unhatched eggs was approximately 6.5 times greater than controls. Suggest not using benzyl benzoate in food medium of Drosophila. (Gonzalez, 1971)

Route: food. Species: rat. Pregnant Wistar rats were given diets supplemented with 0.04 or 1.0%

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benzyl benzoate, or 0.02 or 0.5% 2-(morpholinothio)-benzothiazole from day 0 of gestation to day 21 postparturition. Compared reproductive & developmental parameters to control-fed rats. Article in Japanese. Summary & tables in English. 1 % no effects, on reproductive or developmental parameters. No external, skeletal or visceral anomalies in fetuses. Minor variations observed. Significantly decreased number of fetuses with incomplete sternebrae at 1.0% level. 0.04 % no effects, on reproductive or developmental parameters. No external, skeletal or visceral anomalies in fetuses. Minor variations observed. (Morita, 1980) Route: subcutaneous. Species: mouse. Thirty four Pregnant NMRI mice received a single injection of test material between gestation days 1 and 11. The vehicle was castor oil. Sacrifice was gestation day 17. The observations were fetal sex, weight, count and malformations. This is the solvent control group for a study of sex hormone effects. The solvent was benzyl benzoate in castor oil (concentration not given). Untreated controls were included. No effects. (Eibs,1979) Endocrine Assays Route: in vitro. Species: human 18+ yrs. The estrogenic activities of plasticizers and commercial tissue conditioners were assessed by the E-screen assay. Human breast (MCF-7) cells were trypsinized, plated in 24-well plates at initial concentrations of 2 x 10 (4) cells/mi in 5% fetal bovine serum (FBS), and allowed to attach for 24 hours. The medium was replaced with DMEM with 5% FBS which had been treated to remove endogenous steroids, and contained different concentrations of the test material dissolved in dimethyl sulfoxide (DMSO). The negative control was cell culture medium containing 0.1% DMSO, and the negative control monolayers were cultured for the same periods. After 144 hours, the assay was completed by removing the medium from the wells. The cells were stained with sulforhodamine-B (SRB) for the assessment of cell proliferation by the SRB assay, in which absorbance was measured with a microplate reader equipped with a 540 nm filter. Three independent wells were used for each experiment, and each 3 independent experiments were conducted. No effects, Compared to the controls, no cell proliferation was produced at 10(-8) and 10 (-9) M. .000001 Molar No effects, Compared to the controls, no cell proliferation was produced. .00001 Molar No effects, Compared to the controls, no cell proliferation was produced. .0000001 Molar No effects, Compared to the controls, no cell proliferation was produced. .0001 Molar No effects, Compared to the controls, no cell proliferation was produced. (Hashimoto,2003) Route: in vitro. Species: human 18+ yrs. The purpose of this study was to determine the estrogenic activity of these compounds using the MCF7 human breast cancer cell line as a validated assay system. The ability of benzyl salicylate, benzyl benzoate and butylphenymethylpropional (lilial) to bind to estrogen receptors, stimulate estrogen-responsive gene expression and to increase proliferation of these cells was investigated. The stock MCF7 cells were grown as a monolayer in Dulbecco Es modified EagleLls medium which was supplemented with 5% fetal calf serum, 10 Dg/ml insulin and lOe-8 M 17 El-estradiol in a humidified 10% C02 environment at 37 DC. The cell stocks were sub-cultured on a weekly basis by suspension with 0.06% trypsin-0.02% EDTA at pH 7.3. COMPETITIVE BINDING ASSAY TO ER OF MCF7 CYTOSOL: For this assay, the MCF7 cells were grown as a monolayer in phenol red-free RPMI 1640 medium for at least 3 days to deplete steroid hormone levels in the cells. The cells were then harvested, pelleted and homogenized in 4 volumes of buffer at 4 DC. The homogenates were centrifuged at 105 000 g at 4 DC for 1 hour, the supernatant was retained and stored in aliquots at -7OLIC.The dextran-coated charcoal method was used as the competitive binding assay which was performed on the cytosol. The competition between the binding of estradiol (2,4,6,7 3H estradiol) at 16 x lOe-10 M and 1 E 3 000 000-fold molar excess of unlabelled compounds was assayed. The non-specific binding was determined by assaying in the presence and absence of 100-fold molar excess of diethylstilbesterol. COMPETITIVE BINDING ASSAY TO RECOMBINANT ERa AND ERD: The full length human recombinant ERa and ERLI were diluted 1:2500 in buffer at 4DC. Competitive binding assays were performed in 96 well polystyrene plates at 4LIC for 18 hours between the binding of 2,4,6,7-3H estradiol at 0.8 nm and 1-to 3 000 000-fold molar excess of unlabelled compound using 0.8 nM ER protein. Receptor ligand complexes were separated from free ligand using

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the hydroxylapatite method. ASSAY OF STABLY TRANSFECTED ERE-CAT REPORTER GENE IN MCF7 CELLS: LIThe ERE-CAT vector consisted of the estrogen response element (ERE) of the vitellogenin A2 gene from -331 to -295 bp cloned into the pBLCAT2 vector upstream of the thymidine kinase (tk) promoterE. This assay was carried out as published by Byford et al, 2002. CELL PROLIFERATION EXPERIMENTS: For these experiments, the cells were added to the appropriate amount of phenol red-free RPMI 1640 medium with 5% DCFCS at a concentration of 0.2 x 10e5 cells/ml and plated in monolayer in 0.5 ml aliquots to a 24 well plastic tissue culture dish. The medium was changed after 24 hours to the same medium with the appropriate concentration of test article. The culture medium was changed routinely every 3 LI4 days. Cell counts were performed on a ZBI Coulter Counter by counting released nuclei as per Daly and Darbre, 1990. REAL TIME RT-PCR ANALYSIS: The MCF7 cells were added to phenol red-free RPMI 1640 medium with 5% DCFCS at a concentration of 0.8 x 10e5 cells/mi and were then plated in a monolayer in 16 ml aliquots in plastic cell culture dishes. The cells were also plated on 0.5 ml aliquots in 24 well plastic tissue culture dishes. The cells were then left for 7 days, after which the medium was changed to phenol red-free RPMI 1640 medium with 5% DCFCS and with the required concentration of the test article. The cells were then left for 24 hours. Three wells of cells in the 24 well plates were counted for each test article and the culture dishes were washed in situ with isotonic saline, harvested into ice-cold isotonic saline using a rubber policeman and pelleted by centrifugation. The cell pellets were then stored at -70 DC for RNA preparation. The RNeasy kit with on-column DNase treatment was used to prepare the whole cell RNA. The concentration of the RNA was assessed using a GeneQuant spectrophotometer. The first strand cDNA was synthesized using the ABgene kit as per manufacturerEls instructions. A GeneAmp 5700 sequence detector was used to perform the real-time PCR and the reactions were carried out in a 96- well plate using Absolute QPCR ROX mix as per manufacturer Lis instructions. The primers and probes were designed using Primer Express Software and BLAST searches were carried out to confirm the specificity of the chosen nucleotide sequences. Sigma Genosys was used to synthesize the primers and probes and the probes were labelled with fluorescent dye at the 5 LIend (FAM) and a quencher at the 3E end (Tamra). The reactions were performed in pairs for each RNA preparation using the primers and probes for the gene of interest together with 36B4 as a non-regulated endogenous mRNA control. Quantitative values were obtained from the threshold cycle (Ct) number, the increase in fluorescent signal was associated with an exponential increase of PCR product. The Ct values for a dilution series were used to construct a standard curve and used to determine the relative amount of each mRNA. In order to determine the relative expression level, the value for pS2 mRNA was divided by the value for the endogenous non-estrogen regulated 36B4 mRNA. These pairs of reactions were carried out in triplicate in order to give an average relative expression level for each mRNA sample. The test articles were prepared as stock solutions in ethanol and then diluted 1 in 10 000 (v/v) into culture medium. Regulatory Compliance GLP No (Green & Leake, 1987; Byford, 2002; Daly&Darbre,1990) 99% pure; purchased from Sigma.Summary The results demonstrate that the test article possessed estrogenic activity in MCF7 human breast cancer cells, in vitro. The test article was able to displace 3H-estradiol from both human recombinant ERa and ER E and from ER of MCF7 cell cytosol. It also increased expression of estrogen responsive genes in the MCF7 cells and increased the growth of the estrogen dependent MCF7 cells. 3000000 Molar , Units are LIfoldmolar excess E. Inhibited the binding of 311-estradiol to both ERa (51%) and ER LI.The maximum inhibition was lower when using ER from MCF7 cytosol than the recombinant receptor proteins. 0.0004 Molar , Induced CAT gene expression with maximal effects (1.2 fold). 0.0001 Molar, Increased the expression of pS2mRNA. There was no antagonism to the stimulatory action of 10-10 M LI-estradiol on MCF7 cell growth. 0.0008 Molar , The MCF7 cells went through 2.76 LI0.06doublings within 7 days. 0.0005 Molar , There was no antagonism to the stimulatory action of 10-10 M LI-estradiol on MCF7 cell growth. (Charles,2009) Cytotoxicity

Route: in vitro. Species: hamster. The inhibition of noradrenaline induced respiration in isolated hamster brown fat cells was measured as an indication of effect on cell metabolism. The oxygen

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consumption rates of the cells were measured at 37 C using a Clark-type oxygen electrode fitted in a Perspex vessel of 1-mi volume. Brown adipocytes were isolated from adult hamsters. Cell suspensions which had been cultivated to a cell density of 10(5) cells/ml were added to a Perspex vessel (volume 1 ml) fitted with a Clark-type oxygen electrode. A buffer solution (Krebs-Ringer phosphate buffer) was added and the vessel was sealed. Test material was dissolved in ethanol or dimethyl sulfoxide and were incubated at 37 C with the cells for exactly 5 minutes during which period the oxygen consumption was registered. After this preincubation, noradrenaline was added and the oxygen consumption of the cells was registered for another 5 minutes. The noradrenaline concentration was 1 uM which is approximately twice the dose known to induce maximal respiratory rate. Materials which stimulate the basic rate of oxygen consumption, decrease the maximal noradrenaline inducible respiratory rate. 1 millimolar positive effects, 75% inhibition. (Pettersson,1980) Route: in vitro. Species: chick embryo. Ciliotoxicity was investigated using embryo chicken tracheal organ cultures. The organ cultures were prepared aseptically from 16 day to 17 day old chicken embryos. After dissection, the trachea was rinsed, then flushed with medium and subsequently cut transversely to give rings approximately 1 mm thick. The trachea rings were transferred to petri dishes containing minimum essential medium wtih Hank’s salts, Hepes and L-glutamine. One tracheal ring was then placed in a perspex testing chamber (volume 3.1 ml) that contained medium and was maintained at 37 C. A solution of test material in dimethyl sulfoxide or ethanol was then added to the chamber and time to ciliostatis was determined. Ciliary activity was studied over a 60 minute period by means of inverted microscopy using a magnification of 250. Results are time (mm) to ciliostasis at 5 mlvi concentration for all test compounds. No effects. (Pettersson,1982) Route: in vitro. Species: human 18+ yrs. The ability of the test material to increase the permeability of the membranes of human lung fibroblasts was studied by measuring the release of an intracellular nucleotide marker. Human diploid embryonic lung fibroblasts (line MRC-5) were cultivated to a cell density of 10 to the fifth cells/cm2 (approximately 7 x 10(5) cells/well). The cells were then labeled with [3H]uridine. The labelled cultures were incubated with 25 mM of the test material for 30 minutes at 37 C in Tris-buffered saline. The solution containing the leaked radioactive marker was removed and centrifuged and the radioactivity was measured. A maximal release of the radioactivity was obtained by treating control cells for 30 minutes with 0.06 M sodium borate buffer and scraping with a rubber policeman. This treatment ruptured the cell membranes leaving the nuclei intact. The results notes section indicates the percentage of nucleotide released. Positive effects, 30%. (Thelestam,1980) Route: in vitro. Species: mouse. Stem cell cultures, strain Ascites sarcoma BP8, originating from inoculated C3H mice were used to determine the toxicity of tobacco and tobacco smoke constituents on cell multiplication. The stem cell cultures were grown in test tubes in Hams FlO medium sterilized by filtration, with fetal calf serum (15w/w%), penicillin (100IU) and streptomycin (100IU) added. The test tubes were gassed with sterilized air containing 5% carbon dioxide and capped air-tight to maintain a stable pH of approximately 7.3. The cell cultures were reinoculated to a cell density of 0.1 X 10 4 cells/mi every 5th day. For the tests, the cell suspension was diluted with sterile medium to an initial cell density of 0.4 X 10 4 cells/ml. Test material was dissolved in 10 ul of ethanol, unless otherwise noted, and added to 3 ml aliquots of cell suspensions. Tests were run in duplicate. All compounds were incubated at 37 degrees for 48 hours. 10 ul of solvents were added to the controls. The growth rate of an incubated cell culture was calculated and compared to the average value of 8-10 controls performed in each series. The doubling time for control cultures was approximately 24 hours. No systematic distinction was made between viable and total cell count. The effect of the tested compound is given as the ratio between the growth rates of the incubated cell culture and the controls, expressed as a percentage. 1 millimolar positive effects, 100% inhibition. 0.1 millimolar positive effects, 22% inhibition. (Pilotti,1975)

Route: in vitro. Species: mouse. Inhibition of cell growth was investigated using Ascites sarcoma BP 8 cells. Test material was dissolved in ethanol or dimethyl sulfoxide (DMSO). After incubation for 48 hours the cell density was determined. All materials were tested at a final concentration of 1 mM and

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each was tested in duplicate. The inhibitory effect is expressed as the ratio between the growth rate of the treat cell culture and the control (solvent) and given as a percentage. Results are expressed using a 10 point scale (0-9). 0 corresponds to 0-9%, 1 corresponds to 10-19%, 2 corresponds to 20-29%, etc.) AT value is based on toxicity in all four systems e.g. ACG (cell growth of Ascites sarcoma BP8 cells); AOM (oxidative metabolism of isolated brown fat cells from adult hamsters; AMD (membrane damage of human diploid embryonic lung fibroblasts); ACA (ciliary activity of embryo chicken trachea). Some results were previously published and have been entered under the original reference. They have not been repeated here. (PILOTTI,1975) Positive effects, 9. (Curvall,1984) Route: in vitro. Species: hamster. Inhibition of the oxidative metabolism in isolated brown fat cells was determined by measuring the oxygen consumption. Brown adipocytes were isolated by collagenase treatment of brown fat from adult hamsters (Mesocricetus auratus). The addition of norepinephrine (0.6 uM) to a suspension of brown fat cells increased the oxygen consumption rate of the cells more than 10 times. While the oxygen consumption was continuously registered by polarographic methods, 1 ml of cell suspension was incubated at 37 C for 5 minutes in Krebs-Ringer phosphate buffer with the test material dissolved in ethanol or dimethyl sulfoxide. Norepinephrine was added to the oxygen consumption recorded for another 5 minutes. The toxicity was determined by comparing the norepinephrine induced oxygen consumption after addition of test material to that of the control and expressing the ratio as a percentage. All test substances were tested at a final concentration of 1 mM. Each test comprises at least five experiments performed on different cell preparations. Results are expressed using a 10 point scale (0-9). 0 corresponds to 0-9%, 1 corresponds to 10-19%, 2 corresponds to 20-29%, etc.) AT value is based on toxicity in all four systems e.g. ACG (cell growth of Ascites sarcoma BP8 cells); AOM (oxidative metabolism of isolated brown fat cells from adult hamsters; AMD (membrane damage of human diploid embryonic lung fibroblasts); ACA (ciliary activity of embryo chicken trachea). (Pettersson,et al,1980) Positive effects, 7. (Curvall,1984) Route: in vitro. Species: human 18+ yrs. Plasma membrane damage was assessed as leakage of a cytoplasmic nucleotide marker from prelabelled cells. Human diploid embryonic lung fibroblasts (line MRC-5) were cultivated in Eagle’s medium in polystyrene cells. The cells in confluent monolayers were labelled with [3H] uridine. The labelled cells were rinsed 3 times with Hank’s balanced salt solution and treated for 30 minutes at 37 C with the test material which was added as a 25 mlvi solution obtained from ethanol or dimethyl sulfoxide diluted with Tris-buffered saline. The solution was centrifuged (1000 x g, 5 minutes, 20 C) and the released radioactivity was measured. The results were expressed as a percentage of the maximal release. Each test was run in duplicate. Results are expressed using a 10 point scale (0-9). 0 corresponds to 0-9%, 1 corresponds to 10-19%, 2 corresponds to 20- 29%, etc.) AT value is based on toxicity in all four systems e.g. ACG (cell growth of Ascites sarcoma BP8 cells); AOM (oxidative metabolism of isolated brown fat cells from adult hamsters; AMD (membrane damage of human diploid embryonic lung fibroblasts); ACA (ciliary activity of embryo chicken trachea). (Thelestam et al.,1980) Positive effects, 3. (Curvall,1984)

Route: in vitro. Species: chick embryo. Ciliotoxicity was measured as time to ciliostasis in the tracheas of unborn chickens. The organ cultures were prepared aseptically from 16-day to 17-day old chicken embryos. After dissection, the trachea was rinsed, flushed with medium and cut transversely to give rings approximately 1 mm thick. The rings were transferred to plastic Petri dishes containing minimum essential medium with Hank’s salts (HMEM), Hepes (20 mM) and L-glutamine (2mM). One tracheal ring was placed in a Perspex testing chamber containing the medium (3.1 ml) and was maintained at 37oC. The test materials were added as ethanol or dimethyl sulfoxide solutions and time to cliotasis was determined. The ciliary activity was studied over a 60 minute period by means of an inverted microscope using a magnification of 250. All substances were tested at a 5 mM concentration and examined on at least 3 different occasions, each experiment involving rings from different tracheal preparations. The ciliotoxic effect was expressed as percentage using the expression 100(60-X)/60 where X is time in minutes to ciliostatis when occurring within 60 minutes. Results are expressed using a 10 point scale (0-9). 0 corresponds to 0-9%, 1 corresponds to 10-19%, 2 corresponds to 20-29%, etc.)

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AT value is based on toxicity in all four systems e.g. ACG (cell growth of Ascites sarcoma BP8 cells); AOM (oxidative metabolism of isolated brown fat cells from adult hamsters; AMD (membrane damage of human diploid embryonic lung fibroblasts); ACA (ciliary activity of embryo chicken trachea). (Pettersson,et al,1982) No effects. (Curvall, 1984) Route: in vitro. Species: human 18+ yrs. The effect of test material on lymphocyte mitogenic responses to PHA, rosette formation with sheep red blood cells and neutrophil chemotaxis was studied. DMSO with medium was used as the vehicle for whole blood cultures and ethanol with medium was used with mononuclear cell cultures. Positive effects, inhibited chemotaxis at 10 uM. (Lee,1979) Miscellaneous Route: in vitro. Species: bacteria. The effect of test chemicals on growing cultures of four bacterial strains [Bacillus subtilis, ATCC 9524, Escherichia coli ATCC 11229, Staphylococcus aureus Ox-H (penicillin sensitive) and Staphylococcus aureus ATCC 10390 (penicillin resistant)1 was studied. Dilutions of each chemical were prepared in sterile nutrient broth at 1:500. Tubes were inoculated with 0.5 ml of a 24 hour broth culture of test organism. Tubes were incubated at 37 C for 24 hours and the presence or absence of visible growth observed. Failure of growth to occur in the subculture was taken as evidence that the organisms had been killed in the original chemical-broth tubes. When this occurred, additional concentrations were prepared (1:1000, 1:2000, 1:10,000) and tested in a similar maimer. No effects, bacterial growth was noted at 1:500 dilution for all four strains tested. (Maruzzella, 1961a) Route: in vitro. Antimicrobial activity was evaluated using a standardized petri plate procedure. Organisms used were Staphylococcus aureus (SA), Escherichia coli (EC), Candida albicans and, if active in the others, a Diphtheroid (D). All materials were tested at 10% (wlv) in 95% (vlv) ethanol unless solubility problems occurred. Not all materials were tested in all 4 organisms. Controls were 3,4,4’-trichiorocarbaniiide and 2,4,4’-trichloro-2’-hydroxydiphenyl ether and hexachiorophene. No effects. (Morris, 1979) Route: in vitro. Bacteriostatic activity was tested in liquid cultures to determine minimum inhibitory concentrations (MIC). Tubes were inoculated with 50 ul of a 1:10 dilution in sterile 0.85% saline of a 24 hour shake culture (TGY broth). A minimum of 300,000 viable organisms were added to each tube. Tubes were mixed on a Vortex mixer and incubated at 37 C for 18 to 24 hours. All samples were run in duplicate and that concentration at which no growth occurred in either tube was taken as the minimum inhibitory concentration. 500 ppm positive effects, MIC in Diphtheroid.positive effects, MIC> 1000 ppm in Staph. aureus, E. coli and C. albicans. (Morris,1979) Route: in vitro. Species: fungi. Vapors of the test material were tested for antifungal properties on Candida albicans ATCC 10231, Phoma betae ATCC 6504, Geotrichum candidum Coil. No. 4762 and Oospora lactis ATCC 4798. 0.5 ml of the test organism was streaked onto the agar surface. 0.5 ml of the test material was placed in a cup in the center of a petri dish top. Dishes with culture were inverted over the top and incubated. Vapors of the test material were allowed to emanate throughout the five day incubation period at 22 degrees. After incubation, the presence of a definite clear zone of inhibition on the surface of the agar indicated that the vapor possessed antifungal activity and the larger the zone the greater the activity in this test system. All test materials were tested in triplicate with one cup per dish. No effects. (Maruzzella,1961)

Route: in vitro. Species: yeast. Candida albicans IFO 1385 was grown at 37 C, with reciprocal shaking in yeast mitogen base medium containing 250 mM glucose. Each sample of test compound was placed on the flat bottom of tissue culture plates, 50 ul of sample of yeast suspension were inoculated on each specimen and incubated at 37 C for 2 hours to settle the yeast and the specimens. After incubation, 2 ml of modified Sabouraud dextrose broth was carefully added into each well with samples, incubated at 37 C and pH changes in media were measured. After 7 days incubation, specimens were removed from each cell, washed, and processed for electron microscopic examination, powder mixed 1.2:1.0 (w/v) wI 5, 10 or 20% ETOH. Positive effects, antifungal effects varied depending upon type of plasticizer

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tested: BB > or = BS > or = DBP> = BBP = BPBG. BB = benzyl benzoate, BS = benzyl salicylate, DBP = dibutyl phthalate. (Nikawa,1995) Route: in vitro. Species: bacteria. Antibacterial activity against 9 bacteria was evaluated - Escherichia coli, Eberthella typhosa, Neisseria, Streptococcus faecalis, Streptococcus pyogenes, Staphylococcus aureus, Bacillus megatherium, Corynebacterium diphtheriae, and Oidium albicans Document in German. Data from abstract and table only. Positive effects. (Kellner,1955) Route: in vitro. Species: bacteria. Staphylococcus aureus, Escherichia coli, Proteus vulgaris, and Pseudomonas aeruginosa were used to test the bactericidal activity of test compounds No further experimental detail provided. Article in Polish. Information obtained from English synopsis and tables. Positive effects, number of colonies @ 1:500 and 1:1000-fold dilutions were “overgrown”, “overgrown”, “overgrown”, and “overgrown” for S. aureus, E. coli, P. vulgaris, and P. aeruginosa, respectively. (Munzig, 1974) Route: in vitro. Species: bacteria. The vapors of 192 aromatic chemicals were tested in vitro against growing cultures of Bacillus subtilis (var. aterrimus ATCC 6461), Senatia marcescens (ATCC 9986), Staphylococcus aureus (OX-H), Escherichia coli (ATCC 11229) and Mycobacterium avium (ATCC 4676). All of the test organisms were cultivated on nutrient agar and broth at 37 degrees C, except M. avium which was grown in nutrient agar and broth containing 5% glycerol. Using the method of Maruzzella, J.C. (see methology reference in comments section) 15 mls of nutrient agar were poured into Petri dishes and allowed to harden; the surface of the agar was streaked with 0.5 ml of a 48 hour broth culture of the test organisms (M. avium broth culture was 72 hrs old); aluminum caps with 0.5 ml of aromatic chemical were placed on the inner surface of the petri dish top; dishes were inverted and incubated 48 hours(M. avium - 72 hours). After incubation, a definate zone of inhibition on the surface of the agar indicated that the vapor possessed antibacterial activity. The larger the zone, the greater the activity. The diameters of the zones of inhibition were measured by a metric ruler with the aid of an illuminated Quebec Colony Counter. All dishes were conducted in triplicate. In some tests the vapor permitted no growth to occur on the surface of the entire dish. In this case, the zones of inhibition were recorded as 90 mm. which is the inside diameter of the Petri dish. The method used to test the effects of the vapors on bacteria was by Maruzzella, J.C., in Soap, Perfumery and Cosmetics, vol. 33, pg. 835, 1960. No effects, The vapor was completely inactive. (Maruzzella,1961b) Route: in vitro. Species: human 18+ yrs. Examined the direct effect of certain pesticides in vitro on the functions of human lymphocytes & phagocytes: lymphocyte response to phytohemagglutin (PHA) stimulation & neutrophil chemotaxis. Two methods were used in lymphocyte culture: micromethod of whole blood culture & culture of isolated mononuclear cells. Cell cultures with PHA & insecticide were maintained for 72 hr, then pulsed with [3H]TdR for 24 hr. Degree of lymphocyte response = amount [3H]TdR incorporated into acetic acid precipitable fraction. Cells were made to migrate upward against gravity for neutrophil chemotaxis. 10 micromole no effects, inhibited lymphocyte stimulation 1.5% (mononuclear cells) to 18% (whole blood); inhibited neutrophil chemotaxis 48% (not significant). (Park, 1978) Route: in vitro. Species: human 18+ yrs. Blood clotting tests were performed as described by Nour Eldin and Wilkinson and Nour-Eldin. Effect of test cpd on stypven clotting time was monitored. Solubility of fibrin was determined @ 37 C by the addition of 2 volumes of 5 M urea. Clot retraction was determined in graduated centrifuge tubes. Blood was mixed w/ 0.9% NaC1 or 0.5 ml test cpd. A roughened glass rod was inserted & clotting allowed to proceed @ 37 C. After 1 hr, clot was removed & volumes of serum in both tubes read. Positive effects, Supplementation of stypven clotting activity on platelet poor plasma noted. Actual data not given. (Nour-Eldin,1968) Route: in vitro. Species: human 18+ yrs. The effect of test cpd on the activation of plasminogen was tested on fibrinogen and on oxalated plasma by the add’n of streptokinase @ 37 C followed by thrombin. Time required for complete lysis was recorded. Positive effects, clotting time of fibrinogen was decreased relative to controls. Actual data not given. (Nour-Eldin,1968)

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Route: in vitro. Species: human 18+ yrs. Examined the activating effects of phenyl derivatives on various blood clotting factors: I (fibrinogen)-by increased sensitivity to thrombin; Vill-by increased concentration in intact blood from a case of von Willebrand’s syndrome; IX-by the amount adsorbed on aluminum hydroxide in intact blood; &, XII-by the effect on blood-clotting time of intact blood in siliconed tubes. All 4 blood-clotting factors activated. (Nour-Eldin,1967) Route: skin. Species: human 18+ yrs. Treatment of approximately 1000 cases of scabies at the Poolsbrook Treatment Center, Staveley England from 8/13/42 to 12/31/42 are described. The patients were painted from neck to soles with a 20% benzyl benzoate emulsion which was allowed to dry about 5 - 10 minutes. Over 99% of scabies cases cured if efficiently applied. Rate of failure was 0.92%. Relapses probably due to contact with untreated cases. (Graham,1943) Route: skin. Species: man. Four case histories of men who developed severe skin irritation following treatment with either benzyl benzoate or Benylate (25% benzyl benzoate emulsified in aq. soln. of triethanolamide stearate) for scabies. Irritant effects, caused pruritic dermatitis within 12 - 20 hrs after treatment that persisted for 5 - 10 days. (Daughtry,1945) Route: skin. Species: human 18+ yrs. A freshly prepared suspension of test cpd was used to treat 155 patients w/ scabies. Suspension was applied to embrocations twice for 2 days. [Article in Russian, Summary in English]. Positive effects, cure was achieved in 138 patients (89%). Good effects were obtained in patients w/ severe forms of scabies. (Bolshakova,1970) Route: in vitro. Species: guinea pig. The effect of immersion in test material on the penetration of light through stratum corneum was evaluated. The spectral transmission curve for isolated stratum corneum was measured under ambient conditions and after complete immersion of the stratum corneum in neat test material The wavelengths measured were 320 and 400 nm. 100 % skin effects, a substantial increase in transmision was noted. (Solan,1977)

Route: skin. Species: infant under 1 yr. A 2 month old infant treated (by a body wash) for scabies with an alcoholic solution of benzyl benzoate. ARTICLE IN FRENCH. Unspecified effects, infant went into convulsions 2 hours after being treated (washed) with benzyl benzoate solution. (Castot, 1980) Route: oral. Species: child 1-18 yrs. Case history of a 4 yr old girl that ingested about 50 ml of U.S. Army Insect Repellent Clothing Treatment Solution Formula M-1960. Solution Formula M-1960 was manufactured according to the following formula: benzyl benzoate, 30%; 2-butyl-2 ethyl-i, 3- propanediol, 30%; N-butylacetanilide, 30%; & sorbitan mono oleate (Tween 80), 10%. Of the 4 ingredients, it was felt that benzyl benzoate is tolerated with few ill effects & did not play a significant role in the child’s intoxication. (Boros,1972)

Route: multiple routes. Species: infant under 1 yr. The author reports experimental & clinical data on benzyl benzoate & benzyl alcohol collected over 18 months at the Pharmacology Laboratory, Johns Hopkins Univ. No toxicological cases were noted or reported, but 2 cases demonstrated the low toxicity of both materials. An infant drank 1/2 ounce of a 20% soln of benzyl benzoate & in connection with a circumcision operation, about 4 ml pure benzyl alcohol were injected instead of a 1% soln. Nonspecific effects, swelling of abdomen produced by constipation & intestinal distention by gas was relieved with a rectal tube & laxatives. (Macht,1919)

Route: unreported. Species: cat. Two cases of benzyl benzoate poisoning, 1 fatal and 1 near fatal, are reported Route, dose etc are not given. Lethal. (McCaughey,1968) A model for studying specific effects.

Kandarova,2005 . The EpiDerm protocol results are summarized in this paper and was evaluated by statistical analysis for each chemical tested.

Miller,2005 . The skin sensitization potentials for a variety of molecules available in the literature reported from LLNA experiemnts were used to develop a predictive QSAR model based on geometric, topological, electrostatic, constitutional, thermodynamic, and quantum chemical descriptors.

Corea,2006 . The calculation of the theoretical allergen deposition on fabric from laundry products

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using 24 fragrance allergens was determined. Analytical methodology. Can also refer to physical properties such as boiling temperature, flash point, etc. Schmeltz,1965. Morris,1973 Ramsey,1980. Hassan,1981 Kaiser,1988.

HaarmannReimer,1992b . The mean water solubility of benyl benzoate was 15.3 mgfL at 20 +1-0.5 C. The Flask method was used and samples were analyzed HPLC. Nath,1996. Reverchon, 1997. Rastogi,1998.

Magiatis,1999 . Quantified as a component of essential oils obtained by steam distillation of leaves, twigs and/or resin of Pistacia lentiscus var Chia.

Rastogi,2001 . Domestic and occupational products were analyzed for fragrance materials.

Ellendt,2001 . Validation of two GC-MS methods by performing recoveries from deoderants fortified with the target fragrance substances.

Congiu,2002 . Analysis of Pistacia lentiscus L. fruit and berry oil was conducted, and the products obtained by hydrodistillation and supercritical extraction were compared.

Jayaprakasha,2002 . The chemical composition of the volatile oil of Cinnamomum zeylancium buds.

Lis-Balchin,2002 . Components of jasmine absolute were reported.

Mikhaeil,2003 . The constituents of Frankincense olegum resin were determined.

Raina,2003 . A detailed GC and GC-MS examination of the oils of Ascorus calamus rhixomes and leaves was conducted.

Shin,2003 . The main components of various essential oils were determined.

Ucar,2003 . The composition of oriental spruce (Picea orientalis (L.) Link) was determined.

Cadby,2003 . A description of the analytical strategy that has been developed at Firmenich to monitor suspected allergens in fragrance concentrations.

Chaintreau,2003 . A reliable reference analytical method was developed for determining the occurrence of 24 volatile compounds in perfume concentrates that might elicit skin sensitization.

Fernandez,2003 . The chemical composition of the volatile extracts obtained from Siam and Sumatra benzoin gums by direct GC injection.

Debonneville,2004 . The quantitative capabilities of quadrupule mass spectroscopy (MS) coupled with comprehensive gas chromatography (GC) was applied to the evaluation of allergens in fragrance concentrates.

Aberchane,2004 . The chemical composition of cedarwood oils of different Moroccan provenances was determined.

Shellie,2004 . An evaluation of a method for analyzing 24 suspected allergens is presented.

Kivcak,2004 . The composition of Pistacia lentiscus and Pistacia lentiscus var. chia was determined.

Leijs,2005 . This paper describes a methodology that the authors developed to enable identification and quantification of low levels of ingrdients in presence of highly complex mixtures.

Ka,2005 . Volatile compounds were identified in extracts of Angelic, Pepermint, Pine Needles and Sweet Flag.

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Sacchetti,2005 . Compound found in c. odorata.

Samarasekera,2005 . The mosquitocidal compounds from cinnamomum zeylanicum bark oil were identified.

Baier,2005 . Fast and high resolution GCMS analysis is a suitable method for routine analysis of allergens in fragrance products. Chericoni,2005. 1.4% composition in Cinnamon leaf oil.

Jantan,2005 . The composition of the essential oils of Ciimamomum sintoc was determined. Jantan,2005a. The percentage composition of the leaf oil of Cinnamomum pubescens was determined.

Begum,2007 . The chemical composition of the essential oil of Cinnamomum verum leaves was provided.

Villa,2007 . The aim of this work was to develop a simple, selective and reliable HPLC analytical procedure suitable for the simultaneous determination of fragrance allergens potentially present in essential oils and finished scented products. 24 fragrance allergens were selected.

Kilic,2008 . The percent composition of the leaf oil of Ribes nigrum was determined.

Lamas,2010 . A fast, simple and inexpensive method was developed for the determination of 24 fragrance allergens in indoor air based on the use of a very low amount of adsorbent to retain the compounds. Article in foreign language; no English translation available.

Meyer,1960 . No English abstract or translation available. Schaaf,1961

Bochdalek,1964 . Article in Polish. No English translation available.

Etzweiler,1984 . Article in German. A method for the measurement of headspace analysis was presented.

Aoyama,1988 . No English translation available.

Pitche,2002 . Article in French; only brief English abstract available. The acute elimination of Scabies by benzyl benzoate-sulfiram was investigated. Biological tests other than classical toxicology testing and pharmacological tests. Hagley,1965.

Ishikawa,1965 . No English translation available.

Shamukhamedova,1972 . Benzyl benzoate was reported to be effective in treating latent atypical scabies.

Cook,1975 . Regulation of the growth of Acinetobacter calcoaceticus was studied McGlinn,1976.

Hadani,1977 . Repellent and acaricidal properties against ticks were evaluated

Beggs, 1977 . Regulation of the synthesis of benzyl alcohol dehydrogenase in Acinetobacter calcoaceticus NCIB 8250 was studied Yamada,1988.

Woodfolk,1995 . The test material was evaluated as a treatment for the reduction of allergens in carpet Citations. No original data; the authors cite others work.

Ishihara,1978 . The results of a closed patch test was reported. See Location #7 188 for the original study. Comparison of test methodologies, models.

RIFM,2008 [Api,2008] . Comparison of murine local node assay EC3 values with the NOELs from from confirmatory human sensitization tests are shown using data for 31 fragrance ingredients.

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Computational methodology including use of computers in toxicology or safety evaluation. Also includes articles on databases. ECETOC,1995 A database has been developed for 176 chemicals for which rabbit skin irritation corrosion data is available.

Smith,2000 . Database of esters causing skin irritation in humans. Editorial comments, including letters to editors, authors responses, etc.

Roberts,2002 . A new procedure by IFRA for handling inquiries by dermatologists. Enzymatic Processes

Natsch,2008 . In this study two model systems were used. The first one was the ARE-regulated quinone reductase (QR) activity in Hepa 1C1C7 cells and the second was the ARE-regulated luciferase activity in the cell line AREc32, which contains an eightfold repeat of the ARE sequence upstream of a luciferase reporter gene. These models were used to assess activation of the Keap/Nrf2/ARE regulatory pathway by a collection of 102 different chemicals of known skin sensitization potential. Estrogen or estrogenic effects. Includes endocrine assays,endocrine disruptors, androgen and androgenic effects and thyroid assays (THYROID).

Klopman,2003 . HPV (high production volume) chemicals were screened for the presence of substructures (biophores) responsible for estrogen receptor binding. How materials are used. A use test/study conducted in humans as a means of measuring the elicitation potential of a material in an actual consumer product. Somogyi,1996.

Buckley,2007 . This survey assessed the use of fragrance chemicals by EU manufacturers of cosmetics and detergents intended for the UK market. It also examined the frequency with which fragrances from fragrance mix I and fragrance mix II were found in products in current use. Laws, legislation, regulation, etc.

Steinberg,2003 . A review of the 7th Amendment to the Cosmetics Directive regarding the labeling requirements of the 26 allergies. Local Lymph Node Assay

Gerberick,2005 . A compilation of an extensive chemical data set that covers a wide range of chemicals and sensitization activity.

RIFM,2005 [Lalko,20051 . A comparison of LLNA results for 29 fragrance materials and how they correlate with human data. Methods for determining priorities. Johnson,1984. Mixture.

Landegren,1979 . The efficacy of 2 types of Tenutex, a proprietary preparation, which is an aqueous emulsion was compared. Tenutex containing DDT, disulfiram and benzyl benzoate, was compared to Tenutex which did not contain DDT.

Wetzel,1999 . A mixture of 100 different odorants (Henkel 100) elicited a transient increase in intracellular [Ca(2+)j in human embryonic kidney 293 cells (HEK293) stably or transiently transfected with the plasmid pORl7-40. It was later found that only helional was effective; the remaining 99 odorants were ineffective.

Roemer,2000 . The flavor components of cigarettes are listed.

Baker,2004 . The potential chemical changes and biological activity of smoke from cigarettes with added ingredients was investigated. The ingredients of the experimental cigarettes were listed.

Baker,2004a . The effects of flavoring and additives on smoke chemistry was investigated. The

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individual ingredients used in the experimental ingredient mixtures were reported. News articles or pseudo-reviews. An article that reviews or discusses a topic and mentions a number of materials that are of interest. The article does not cite any new or original data. News clippings fall in this category.

Roberts,2002a . A procedure for dermatologists to seek and trace information with regard to possible fragrance allergy.

Vey,2003 . A procedure for dermatologists to seek and trace information with regard to possible fragrance allergy. Papers that are a review on a general subject, not a specific material. TREV papers may contain one or more RIFM materials. TREV papers do not contain original data.

Hjorth,1962 . This paper is a capsule preview of two detailed papers to be published in Arch Pharm Chemi Spiegel, 1963 Jordan,1973. Mitchell,1975a. Collins,1975. Atkins,1975. Fisher,1976. Marzulli,1976. Schorr,1978. Fisher,1980. Fisher,1981 Bronaugh,1984. Stofberg,1987.

Reifenrath,1995 . Skin penetration and evaporation studies Scheinman,1996.

DeGroot,1997 . A clinical review of adverse reactions to fragrances deGroot,1998. Hostynek,1998. Guba,2000.

Zhai,2001 . A topic review on the effect of occlusion on percutaneous absorption and summarizes related details.

Steinberg,2001 . The draft 7th Amendment to the Cosmetic Directive 76/768 EEC is discussed. In addition, 26 fragrance allergens that the European Parliament is requesting be labeled in the ingredient list of a fragrance they are present in (at a certain level) are also mentioned.

Steinberg,2002 . A review of the 7th amendment of the European Cosmetics Directive and the materials that are affected.

Loden,2002 . The authors discuss using animal testing and maximum permitted concentrations to reduce contact allergy to fragrances in cosmetic products.

Schroeder,2009 . The Scientific Committee on Cosmetic Products and Non-Food Products Intended for Consumers (SCCNFP) in the EU issued an opinion concerning fragrance allergy in consumers, identifying the most frequently reported and consumer-recognized fragrance allergens. Perfume. Butterfield, 1963

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Pharmacological effects not clearly toxic.

Macht,1921 . Three benzyl preparations were pharmacologically examined. Photochemistry. See also phototoxicity (PTOX) and photosensitization (PSEN). Shibamoto,1985. Shibamoto,1985a. Schunk,1986.

Schunk,1988 . All materials were tested after they were irradiated with UV & sunlight. Toda,1988. Reports on patients such as patch testing.

Fisher,1975 . Patch test concentrations and vehicles are discussed

Sugawara,1984 . Results not specified by material in English abstract Reproductive effects. See also teratogenicity (TERAT) and fetal (FETAL) effects. Material is given to mother and father before or during gestation to see how it affects the ability to reproduce viable offspring. Toxic effects may be noted Eibs,1982 Used as part of the vehicle

Puri,1987 . A mixture of benzyl benzoate and cator oil was tested as vehicle control. Dose not given. Review articles. Papers that are reviews on a material or a class of materials. A review paper may be on one or more RIFM materials. Review papers do not contain original data.

Macht, 1918 . Clinical applications of the smooth muscle relaxation effect of the test material were discussed. Data on pharmacology & toxicity in many specifies is presented but not given in detail Kutob,1956.

Freese,1979 . Review of Teratogenicity and LD5O information. Cadby,1983. WHO,2002. Summary of results of safety evaluation of benzyl derivatives used as flavouring agents, annual volumes, acute toxicity, short term studies of toxicity and genotoxicity was shown.

Heydorn,2003 . A review of the literature to investigate an association between fragrance allergy and hand eczema.

Adams,2005 . The key data relevant to the safety evaluation of benzyl alcohol, benzaldehyde, or benzoic acid and 34 structurally related substances are presented.

Natsch,2009 . A comparison of LLNA data with modelling predictive data. Sensitization studies and effects. Distinguish those with patients (PAT).

Griem,2003 . A comparison of LLNA EC3 values with HRIPT and HMT LOEL, and NOEL values was carried out and revealed that the EC3 can be used as a surrogate for the human NOEL in risk assessment.

Gerberick,2007 . An examination into whether the reactivity of several materials with lysine, cysteine and GSH peptides correlates with sensitization potency in the LLNA.

RIFM,2009 [Api,2009] . A detailed analysis of the dermal sensitization data for approximately 50 fragrance ingredients that have exhibited dermal sensitization has been conducted.

Keller,2009 . NESIL and AEL values for fragrance allergens from the IFRA!RIFM dataset are shown.

Nardelli,2009 . These fragrance allergens were identified in 127 patients reacting to the 48 specific topical pharmaceutical products. Skin absorption. For other types of absorption use ABS as with mucous membranes (MUC).

Bucks,1989 . A review of literature was conducted to illustrate that occlusion does not necessarily increase the percutaneous absorption of all chemicals.

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Hostynek,1995. Skin or respiratory irritation.

Bagley,1996 . Skin irritation data bank was developed by ECETOC using available in vivo irritation data. Structure activity relationships such as studies of a group of aldehydes or other similar structures. Cronin,1994.

Nishimura,1994 . Mouse LD5O values were taken from RTECS. Paper in Japanese Barratt,1996. Hostynek, 1997. Devillers,2000.

Serra,2001 . Three quantitative structure toxicity relationships were developed, to predict 50% population growth impairment concentrations for industrially important aromatic solvents.

Schultz,2003 . The data for hydrophobicity and electrophilicity along with toxicity values were reported using these materials.

Debska,2003 . Structure-smell relationships for chemical compounds were determined. Studies between acute (ACU) and Chronic (CHRONIC). Not applicable to environmental studies. (28-90 days)

Fukayama,1999 . Subchronic inhalation studies of fragrance mixtures in rats and hamsters; individual ingredients were not tested separately. Studies of odor.

Bergstrom,2000 . The most common volatile compounds of floral odor belonging to various chemical classes are presented. Studies on cosmetic products or directly related thereto. Maibach,1980. Test methodology (new or altered classical), testing procedures, guidelines, etc. If applicable, see also biological tests (BlOTS). Harkiss,1973

Nishimura,1994b . The molecular weight, solubility parameter, molar volume, log P and LD5O value of 56 chemicals were analyzed in this study.

Patlewicz,2008 . This article compares and contrasts two schemes of reactivity rules. In particular they use the mechanistic domain approach. The approach taken for the comparison focused on a set of LLNA data for 210 chemicals.

Achilleos,2009 . A method to screen contact allergens in a non-animal test is described.

Environmental Data

Acute toxicity Route: unclassified. Species/Media: insect. Cultures of adult copra mites (Tyrophagus putrescentiae) were maintained 25 +1- bC and 75% relative humidity. The test material in methanol was applied to black cotton fabric pieces, and after drying for 1 minute in a fume hood, each fabric piece was placed at the bottom of a Petri dish. Groups of 25 adult mites were placed in each Petri dish that was then covered. The controls were placed in plates with fabric pieces containing 20 ul methanol. Mortality was determined 24 hours after the treatment under a binocular microscope and the mites were considered to be dead if their appendages did not move when they were prodded with a pin. The treatments were

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replicated 4 times, and the LD5O values were calculated by probit analysis. LD50 10.03 UGIC2 10.03 Dg/cm2 calculated LD5O, The 95% confidence limit was 3.42 - 16.13 ug/cm2. (Kim,2004) Route: unclassified. Species/Media: insect. Cultures of adult copra mites (Tyrophagus putrescentiae) were maintained 25 +I bC and 75% relative humidity. The test material in methanol was applied to black cotton fabric pieces, and after drying for 1 minute in a fume hood, each fabric piece was placed at the bottom of a Petri dish. Groups of 25 adult mites were placed in each Petri dish that was then covered. The controls were placed in plates with fabric pieces containing 20 ul methanol. 50.9 Dg/cm2 lethal, Within 1 hour after the treatment, > 90% mortality was produced. Uncoordinated behavior without knockdown was produced before death, with a common feature in death being the forelegs extended side to side. (Kim,2004) Route: unclassified. Species/Media: insect. To the bottom of each well of 24-well tissue cultures plates, 30 ul dilutions of the test material in ethanol were applied at concentrations ranging from 0.01 D 0.640 g/m2. Only ethanol was applied to the control wells, and all were allowed to dry for 24 hours before 100 mites were added to each well. To prevent the mites form escaping, the upper rims of the wells were covered with Glue Pelton. Incubation was conducted for 1, 5 and 24-hour periods in darkness at 25oC, with 75% relative humidity. Mortality was observed with a stereo binocular microscope after each incubation period, and the lowest concentration that killed 50% of the mites was recorded as the EC5O. Two experiments were conducted, each in triplicate. Summary At 1, 5 and 24 hours, the EC5O was calculated to be 0.33 +1- 0.03, 0.23 +1- 0.01, and 0.06 +1- 0.005 g/m2, respectively. Positive effects, At 1, 5 and 24 hours, the EC5O was calculated to be 0.33 +1- 0.03, 0.23 +1- 0.01, and 0.06 +1- 0.005 g/m2, respectively. (Raynaud,2000) Route: Freshwater. Species/Media: sludge. The respiration rate of an activated sludge (a mixed population of aquatic microorganisms) was measured after a contact time of 3 hours. The respiration of the same activated sludge in the presence of various concentrations of the test substances (1000, 1800, 3200, 5600, and 10,000 mg/L) under otherwise identical conditions was also measured. The inhibitory effect of the test substance at a particular concentration was expressed as a percentage of the mean respiration rates of two controls. An EC5O value was calculated from determinations at different concentrations. Reference substance was 3,5-dichlorophenol. Paper in German. Data obtained from tables and ‘similar” translated study. Study Length: 3 hours Regulatory Compliance ISO 8192; OECD Guideline 209 GLP Yes 1800 mg/i positive effects, benzyl benzoate showed 29.4% respiration inhibition of activated sludge at a test substance concentration of 1800 mg/L. 1000 mg/i positive effects, benzyl benzoate showed 18.3% respiration inhibition of activated sludge at a test substance concentration of 1000 mg/L. 10000 mg/i positive effects, benzyi benzoate showed 47.4% respiration inhibition of activated sludge at a test substance concentration of 10000 mg/L. EC5O exceeds 10000 mg/L. 3200 mg/i positive effects, benzyl benzoate showed 35.0% respiration inhibition of activated siudge at a test substance concentration of 3200 mg/L. 5600 mg/i positive effects, benzyl benzoate showed 38.1% respiration inhibition of activated sludge at a test substance concentration of 5600 mg/L. (HaarmannReimer, 1992)

Route: Freshwater. Species/Media : Daphnia magna. The acute toxicity of test msterial to the daphnid, Daphnia magna, was performed under static-renewal conditions in sealed vials without headspace to minimize loss of test substance to the atmosphere. The water used for the acclimation of the test organism and for all toxicity testing was carbon filtered, deionized water adjusted to a hardness of 160 to 180 mg/L as CaCO3 and adjusted to a pH of <8.0 with 5% H3P04. The test material was mixed in dilution water, and duplicate samples from each concentration were used for testing. The test vessels were filled to capacity with the test solution to eliminate headspace, and sealed with Teflon caps. Juvenile daphnids, Daphnia magna, less than 24 hours old were employed as the test organisms and were produced from a culture apparently free of disease, injuries and abnormalities. This culture had been maintained in 100% dilution water under static renewal conditions for more than 7 days. Water quality was within normal limits for the duration of the study. The minimum dissolved oxygen concentration was 8.1 mg/L, which is 89% at 20C. Ten daphnids were added to each vial, which were

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filled to capacity to eliminate headspace. Nominal concentrations of test material, 1.3, 2.2, 3.7, 6.1, 10, and 17 mg/L, plus control, OmgIL,were used for testing. Fresh test solutions were prepared after 24 hours and used to renew the media in the test vessels. The range of test concentrations was established prior to the definitive test using range-finding experiments of concentrations 0.10 to lOOmg/L.Test vessels were randomly arranged during the 48 hour test, incubated at 20C +I 1C, with a 16 hour light and 8 hour dark photoperiod. The number of surviving organisms, the occurrence of immobility and sublethal effects, and the presence of insoluble material were determined visually and recorded after 0, 24, and 48 hours. Vials remained sealed from 0 to 24 hours (when media renewal occurred) and again from 24 to 48 hours to prevent loss of test material to the atmosphere. Dead test organisms were removed at 24 hours. Results of the toxicity test were interpreted by standard statistical techniques. The 24 and 48 hour LC5O and EC5O values were calculated using the binomial or probit methods. The NOEC is the highest tested concentration of test substance that allowed 90% or greater survival without any sublethal effects. Subjects: 10 Unspecified Sex Study Length: 48 hours Regulatory Compliance This study was performed in compliance with OECD (1997) Good Laboratory Practice Standards. Characterization of the test substance was not conducted in compliance with GLP rules. GLP Yes Methodology: OECD 202 1997. Subjects: 10 Unspecified Sex Study Length: 48 hours Regulatory Compliance This study was performed in compliance with OECD (1997) Good Laboratory Practice Standards. Characterization of the test substance was not conducted in compliance with GLP rules. GLP Yes (OECD 202) T.R. Wilbury sample number 1755 The NOEC was determined by examination of the data rather than a “t” test because only two replicates were used for each concentration. The continuously recorded temperature was not always 20+/-iC (the individually recorded temperatures were always within the specified range). The deviations did not effect the outcome of the study and no other protocol deviations occurred during the study.LC5O 7.77mg/i 11.0 mg/i calculated LC5O, The LC5O at 24 hours. 1.73 mg/i reported No Effect Concentration, The NOEC calculated at 48 hours. 7.77 mg/l calculated LC5O, The LC5O at 48 hours. 3.09 mg/i calculated EC5O value, The EC5O at 48 hours. 4.26 mg/i calculated EC5O vaiue, The EC5O at 24 hours. (Boeri,2003) Inhalation Route: Freshwater. Species/Media: fish. A 96-hour acute toxicity study was conducted with 10 Zebra fish per dose levei. Fish were approximately 2.5-3.5 cm in length. The study was conducted under semi-static conditions with a photoperiod of 16 hours lightJ8 hours dark in test vesseis containing 5 L of test medium. Test compound concentrations were 0, 3.9, 5.5, 7.8, and 11.0 mg/L. Temperature, oxygen content and pH value were measured at 0 and 24 hours. Fish mortality was observed at 2, 24, 48, 72 and 96 hours Paper in German. Data obtained from tables and a “similar” translated study. Finai LCO/LC 100 was per an amendment that was provided to RIFM. Subjects: 40 Unspecified Sex Study Length: 96 hours LC5O Geometric mean: LCO/LC100: 2.32 mg/iSummary LCO: 1.90 mg/i (arithmetic mean of analytical values after 24 hours of exposure); LC100: 2.84 mg/i (arithmetic mean of anaiyticai values after 24 hours of exposure); Geometric mean: LCO/LC100: 2.32 mg/i 3.9 mg/i no effects. 11 mg/i iethal, 10/10 fish dead after 24 hours. 7.8 mg/I lethal, a total of 10/10 fish died. 9/10 fish dead after 24 hours; 10/10 fish dead after 48 hours.positive effects, LCO: 1.90 mg/i (arithmetic mean of analytical vaiues after 24 hours of exposure); LC100: 2.84 mg/i (arithmetic mean of analytical values after 24 hours of exposure); Geometric mean: LCO/LC100: 2.32 mg/i. 5.5 mg/i lethal, a total of 10/10 fish died. 3/10 fish dead after 24 hours; 6/10 fish dead after 48 hours; 10/10 fish dead after 72 hours. (HaarmamiReimer, 1993) Miscellaneous Route: skin. Species/Media: insect. Brains of diapausing fly pupae (Sarcophage crassipaipis) were extirpated by making a small incision in the dorsal region of the head, flushing out the fat body and removing the exposed brain. The ring gland was then pushed towards the thorax and incision sealed with a thread ligature. Operated pupae were held in a Petri dish containing moist filter paper. Test compounds were applied to debrained fly pupae on the day after surgery. The anterior cap of the puparium was removed and 5 ul of test material was appiied directly to the pupai cuticle. A controi

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group received distilled water instead of test material. The effect on stimulation of development was based on percent mortality during diapause, percent diapause termination and percent adult emergence Flys were 20-25 days in diapause at 20 C prior to experimentation. 5 Dl no effects, % mortality during diapause was 0, % diapause termination was 0. (Denlinger,1980) Route: in vitro. Species/Media: insect. A repellancy chamber was constructed from plexiglas above which fluorescent tubes were housed. Tick larvae were used, unfed, when they were 14-21 d old. Test substances were weighed & diluted w/ acetone to give testing concentrations. Test paper (11 cm Whatman No. 1) was impregnanted w/ test or control substance, allowed to air dry, and placed in repellancy chamber. A stainless steel disc was placed on the paper & approx. 100 larval ticks were placed on the disc and observed for 10 mm. Any ticks moving from the disc, past the circle of impregnated paper were crushed & counted. Positive effects, data were not suitable for probit analysis, hence no repellancy values were calculated. Approx. 30 mg/ml resulted in 95% repellancy. (Matthewson, 1981) Route: unclassified. Species/Media: insect. Test substance(s) dissolved in acetone was applied to the skin & hair of the perianal region of a calf in a stationary stall. Fifteen minutes after treatment 50 male & 50 female adult ticks were released on sawdust bedding beneath the calf. The progress & pattern of the infestation were recorded @ 24, 48, & 72 h after releasing the ticks. 200 mg/mi positive effects, insects in treated area vs. those in controls @ 24 h. 3M & 3F vs. 76M & 76F. Effect decreased w/ time. (Matthewson,1981) Route: in vitro. Species/Media: insect. An olfactometer was used to measure the attractiveness of various odors to silkworm iarvae. A series of 3 to 5 repetitions of each experiment were made in which larvae were exposed to test sustance and their movement monitored. 0.1 ml of each test substance was dissolved in 100 ml of distilled water prior to testing Article in Japanese. Information obtained from English summary and tables. Positive effects, the number of moved larvae at the odor site and odorless site was 152/250 and 54/250 , respectively. The ratio of odor/odorless was 2.8. (Hirao,1964) Route: in vitro. Species/Media: insect. An olfactometer was used to measure the attractiveness of various odors in combination with beta-gamma hexanol to silkworm larvae. A series of 3-5 repetitions of each experiment were made in which larvae were exposed to test substance and their movement monitored. 0.1 ml of each of test substance was dissolved in 100 ml distilled water for use in treatment Article in Japanese. Information obtained from English summary and tables. Positive effects, the number of moved larvae at the odor site and the site containing odor plus beta-gamma hexenol was 32 and 91, respectively. Total number of larvae used was 150. (Flirao,1964) Route: in vitro. Species/Media: insect. An olfactometer was used to measure the attractiveness of various odors in combination with steam distillate of mulberry leaves. A series of 3 to 5 repetitions of each experiment were made in which larvae were exposed to test material and their movement monitored. 0.1 ml of each test substance was dissolved in 100 ml distilled water for use in treatment. Positive effects, the number of moved larvae and the odor site and the site containing odor plus distillate was 40 and 92, respectively. Total number of larvae used was 150. (Hirao,1964) Environmental

Route: Freshwater. Species/Media: algae. The acute toxicity of test material to the algae, Selenastrum capricornutum, was performed under static conditions in sealed containers from which all the headspace was removed to minimize loss of test substance from test vessel. Six concentrations of test material, 0.033, 0.065, 0.13, 0.25, 0.50, and 1.Omg/L,plus control, Omg/L,were used for testing. The range of test concentrations was established prior to the definitive test using range-finding experiments of concentrations 0.01 to lOOmg/L.The concentration of test substance in the test vessels was measured at time zero using HPLC and found to be 0.0332, 0.0647, 0.127, 0.247, 0.472, and 0.972mg/L respectively. These initial measured concentrations were used for calculations. The test material was diluted in freshwater AAP medium (USEPA, 1978; T.R.Wilbury Standard Operating Procedure No. 6) adjusted to pH 7.5, and the test vessels were filled to capacity with the test solution to eliminate

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headspace, and sealed with Teflon caps. The test algae, Selenastrum capricornutum, (UTEX 1648) was obtained from the Culture Collection of Algae from the University of Texas at Austin and acclimated to test conditions for more than 14 days. The subsample of algae used for experimentation was a seven day old, actively growing culture and was distributed among twenty replicates for the control and 11 replicates for the test solutions at approximately 10,000cells/mL. Controls also included a stability test with the test substance at 3.Omg/L in experimental vials without inoculum. Test vessels were incubated at 24C +1-2C on a rotary shaker (repositioned daily) with a 24 hour light and 0 hour dark photoperiod. At 24, 48, and 72 hours, three treatment vessels were sacrificed for analysis of algal cell number and observations by direct microscopic examination with a hemocytometer. At the end of the study, a 0.5mL aliquot from each test vessel at the maximum growth inhibition level (0.972mg/L) was combined with lOOmLof fresh media to determine algistatic or algicidal effects of the test substance. The effective concentrations (EC5O) and the NOECs were determined using the mean measured concentration of benzyl benzoate and the number of cells/mL, average specific growth rate, and the area under the growth curve. Subjects: 10000 Unspecified Sex Study Length: 72 hours Regulatory Compliance This study was performed in compliance with OECD (1997) Good Laboratory Practice Standards. Characterization of the test substance was not conducted in compliance with GLP rules. GLP Yes (OECD 201) T.R. Wilbury sample number 1755 No protocol deviations occurred.EC5O 0.31 lmg/LSummary The toxic effects of benzyi benzoate were aigistatic rather than algicidal to Selenastrum. 0.311 mg/i calculated EC5O value, This is the 72 hour EC5O calculated using the area under the growth curve. 0.280 mg/i calculated EC5O value, This is the 48 hour EC5O calculated using the number of celis/mL. 0.787 mg/l calculated EC5O value, This is the 48 hour EC5O caicualted using the average specific growth rate. 0.363 mg/i calculated EC5Ovalue, This is the 72 hour EC5O calculated using the number of cells/mL. 0.475 mg/i calculated EC5O value, This is the 72 hour EC5O calculated using the average specific growth rate. 0.337 mg/i calculated EC5O value, This is the 48 hour EC5O calculated using the area under the growth curve. > 0.972 mg/i calculated EC5O value, This is the 24 hour EC5O calculated using the average specific growth rate. 0.247 mg/I reported No Effect Concentration, This is the 72 hour NOEC calculated using the number of ceiis/niL and the average specific growth rate. 0.6 17 mg/i calculated EC5O value, This is the 24 hour EC5O calculated using the area under the growth curve. 0.928 mg/i calcuiated EC5O value, This is the 24 hour EC5O caiculated using the number of cells/mL. 0.0647 mg/I reported No Effect Concentration, This is the 72 hour NOEC calculated using the area under the growth curve. (Ward,2003) Species/1’Iedia: sludge. A mixed population of aquatic microorganisms (activated sludge) originating from an aeration tank of a waste water plant treating predominantly domestic sewage was used. A suspension of 100 mg/L test material in a mineral medium was inoculated and incubated under aerobic conditions in the dark. Allowance was made for the endogenous activity of the inoculum by running parallel bianks with inoculum but without test substance. A reference compound, aniline, was run in parallel to check the operation of the procedures. Degradation was followed by the determination of oxygen uptake and measurements were taken at frequent intervals to allow the identification of the beginning and end of biodegradation. The test lasted for 28 days and was performed at a temperature of 20 C. Because of the nature of the biodegradation and of the mixed bacterial populations used as inocuia, determinations were performed at least in duplicate. The oxygen uptake was calculated from readings taken at regular and frequent intervals, using the method given by the manufacturer of the equipment. At the end of incubation the pH of the contents of the flasks was measured Paper in German. Data obtained from tables and a ‘similar’ translated study. Study Length: 28 days Regulatory Compliance Council Directive 79/83 1 EEC Annex V (1990) Method C.4-D “Manometric Respirometry Test”; OECD Guideline 301 F. GLP Yes 100 mg/i readily biodegradable, within the test period of 28 days, a degradation of 94% was determined for benzyi benzoate. Benzyl benzoate was classified as LlReadily Biodegradable LI.In parallel preparations with the reference substance Aniline, a degradation of 75% was achieved within 28 days. (HaarmannReimer,1992a)

Species/Media: sludge. A study was conducted to determine the ultimate biodegradability of the test

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material using the sealed vessel test. The test was conducted in 160 ml vessels (hypovials) containing 100 ml mineral salts medium inoculated with secondary effluent and the respective test or reference substance. The inoculum used was 10% by volume of activated sludge plant secondary effluent, filtered through a Whatman filter paper (541) to remove coarse particulate matter. The level of dissolved inorganic carbon (DIC) was reduced by sparging the filtered effluent with nitrogen after prior adjustment of the pH to 6.5. Test concentration was nominal 10 mg/l organic carbon. Test temperature range was 17-22 C. Multiple vessels were prepared per test material sealed with a butyl rubber septum and an aluminium crimp seal. The headspace in each vessel had a volume of 60 ml and when filled with air, contained approximately 6 times the mass of oxygen required for the complete oxidation of the test material. The sealed vessels were incubated at 20 C on a rotary shaker. At intervals during the 28 day test period a vessel was removed and concentration of carbon dioxide in the headspace gas determined. The seal is then broken and the concentration of inorganic carbon in the test medium was determined. Analysis of both the headspace gas and the liquid medium for C02/DIC was performed on day numbers: 3, 7, 10, 14, 18, 21, 24 and 28 using the Tonics 555 Inorganic Carbon Analyser. The total inorganic carbon in the vessel was calculated and corrected by subtracting the inorganic carbon produced in the control. The control vessels were identical to the test vessels except for the omission of the test material. From a knowlege of the initial organic carbon concentration added as test substance, the extent of mineralisation was determined. A test substance was considered readily and ultimately biodegradable if the material exceeded 60% biodegradation within a 10 day window over 28 days Reference material not provided. Nominal carbon concentrations of the test materials were used based on the calculated percentage carbon (from molecular formula) assuming 100% purity of the test material. Study Length: 28 days Regulatory Compliance OECD, Paris, 1981. Test guideline 301B, Decision of the Council C(81) 30, final. Draft Ecotoxicology Section Standard Operating Procedure Number 158.01. Operation of the Sealed Vessel Test. GLP Yes (Birch,1991) R441/5 11498 On day 28 the mean percentage biodegradation of the test material was calculated from four vessels not five because only a partial analysis was performed on the fifth vessel (liquid analysed only). There is no explanation in the study file to explain how this omission occurred.Summary Biodegradation on day 28 was 93.0% (76.6%-109.5%) 95% Confidence Interval. The requirements for ready and ultimate biodegradability were met. 10 mg/l readily biodegradable, Biodegradation on day 28 was 93.0% (76.6%-109.5%) 95% Confidence Interval. The requirements for ready and ultimate biodegradability were met. (Quest,1994)

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Heydorn S. , Menne T. and Johansen J.D. (2003) Fragrance allergy and hand eczema - a review. Contact Dermatitis, 48(2), 59-66.

Heydom S. , Johansen J.D. , Andersen K.E. , Bruze M. , Svedman C. , White I.R. , Basketter D.A. and Menne T. (2003a) Fragrance allergy in patients with hand eczema - a clinical study. Contact Dermatitis, 48(6),3 17-323. Hirao T. and Ishikawa S. (1964) Studies on olfactory sensation in the larvae of the silkworm, Bombyx mon. I. Chemotaxis of hatched larvae by olfactometer method. Nippon Sanshigako Zasshi, 33(4),

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277-285. Hjorth N. (1961) Reactions to single components in balsam of Peru. In Eczematous Allergy to Balsams, Ch.6, 43-48. Hjorth N. (1961a) Other Aromatic Substances Related to Balsams. In Eczematous Allergy to Balsams, Chapter 10, 94-111. Hjorth N. (1961b) The Development and Persistence of Cross-Sensitization. In Eczematous Allergy to Balsams, Chapter 17, 144-165. Hjorth N. (1962) Skin reactions to preservatives in creams. Archives of Pharmacology, 77, 43-46. Hostynek J.J. (1995) Predicting absorption of fragrance chemicals through human skin. Journal of the Society of Cosmetic Chemists Japan, 46(4), 221-229. Hostynek J.J. and Magee P.S. (1997) Fragrance allergens: Classification and ranking by QSAR. Toxicology in Vitro, 11(4), 377-384. Hostynek J.J. (1998) Exposure to fragrances: Their absorption and potential toxicity. Drugs and the Pharmaceutical Sciences, 91, 601-623. International Flavors and Fragrances (1982) Acute eye irritation test in rabbits with benzyl benzoate. Unpublished. International Flavors and Fragrances (1982a) Acute oral toxicity study in rats with benzyl benzoate (14 days). Unpublished. International Flavors and Fragrances (1982b) Acute oral toxicity study in rats with benzyl benzoate (14 days). Unpublished. International Flavors and Fragrances (1982c) Acute oral toxicity study in rats with benzyl benzoate (14 days). Unpublished. International Flavors and Fragrances (1982d) Acute oral toxicity study in rats with benzyl benzoate (14 days). Unpublished. June 09a June 09b Ishihara M. (1977) Problems of closed patch tests with ingredients of cosmetic products. Journal of Japanese Cosmetic Science Society, 1, 87-102. Ishihara M. (1978) The environment and the skin. Journal of the Medical Society of Toho University, 25(5-6), 750-766.

Ishihara M. , Itoh S. , Hayashi S. and Satake T. (1979) Methods of diagnosis in cases of cosmetic dermatitis and facial melanosis in females. Nishinthon J. Dermatology, 41(3), 426-439.

Ishihara M. , Itoh M. , Hosono K. and Nishimura M. (1981) Some problems with patch tests using fragrance materials. Skin Research, 23(6), 808-8 17.

Ishihara M. , Itoh M. , Nishimura M. , Kinoshita M. , Kantoh H. , Nogami T. and Yamada K. (1986) Closed epicutaneous test. Skin Research, 28(suppl 2), 230-240. Ishilcawa S. and Hirao T. (1965) Studies on olfactory sensation in the larvae of the silkworm, Bombyx mon. III. Attractants and repellents of hatched larvae. Bull. Sericul. Exp. Sta., 20(1), 21-36. [Sanshi Shikenjo] Itoh M. (1982) Sensitization potency of some phenolic compounds - with special emphasis on the relationship between chemical structure and allergenicity. Journal of Dermatology, 9(3), 223-233.

Itoh M. , Ishthara M. , Hosono K. , Kantoh H. , Kinoshita M. , Yamada K. and Nishimura M. (1986) Results of patch tests conducted between 1978 and 1985 using cosmetic ingredients. Skin Research, 28(suppl.2), 110-119.

Itoh M. , Hosono K. , Kantoh H. , Kinoshita M. , Yamada K. , Kurosaka R. and Nishimura M. (1988) Patch test results with cosmetic ingredients conducted between 1978 and 1986. Journal Society Cosmetic Science, 12(1), 27-41. binJantan I. , Yalvema M.F. , Ayop N. and Ahmad A.S. (2005) Constituents of the essential oils of

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Cinnamomum sintoc Blume from a mountain forest of Peninsular Malaysia. Flavour and Fragrance Journal, 20(6), 60 1-604.

Jantan I.b. , Yalvema M.F. , Abu Bakar B. , Muhammad K. , Ayop N. and Ahmad A.S. (2005a) Constituents of the leaf oil of cinnamomum pubescens kochummen. Journal of Essential Oil Research, 17(5), 513-515. Jaulmes P. (1946) A case of poisoning by impure benzyl benzoate [containing benzyl alcohol]. Tray. Soc. Pharm. Montpellier, 6, 47-50.

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Johnson O.H. , Casey S. , Doeltz M.K. , McCaleb K.E. , Miller A.M. , Papa P.A. , Swett L.B. Valentini M.A. and Helmes C.T. (1984) A study of biocides for the selection of candidates for carcinogen bioassay. Journal of Environmental Science and Health, A19(1), 1-25.

Jolanki R. , Suhonen R. , Henriks-Eckerman M.-L. , Estlander T. and Kanerva L. (1997) Contact allergy to salicyl alcohol in aspen bark. Contact Dermatitis, 37(6), 304-205. Jordan Jr. W.P. (1973) Resorcinol monobenzoate, steering wheels, peruvian balsam [Letter to the Editor]. Archives of Dermatology, 108, 278.

Ka M.-H. , Choi E.H. , Chun H.-S. and Lee K.-G. (2005) Antioxidative activity of volatile extracts isolated from Angelica tenuissimae roots, peppermint leaves, pine needles, and sweet flag leaves. Journal of Agricultural and Food Chemistry, 53(10), 4124-4129. Kaiser R. (1988) New volatile constituents of Jasminum sambac (L.) Aiton. Devl. Food Sci., 18 (Flavors/Fragrances), 669-684.

Kandorova H. , Liebsch M. , Gerner I. , Schmidt E. , Genschow E. , Traue D. and Spielman H. (2005) The epiderm test protocol for the upcoming ECVAM validation study on in vitro skin irritation tests-an assessment of the performance of the optimised test. Alternatives to Laboratory Animals (ATLA), 33(4), 35 1-367.

Kato Y. , Sugiura M. , Hayakawa R. , Sakaida T. and Hirose 0. (1999) Two cases of contact dermatitis due to propolis - Patch testing with fragrances detected in propolis by GC-MS-. Environmental Dermatology, 6(4), 231-236. Katz A.E. (1946) Dermal irritating properties of essential oils and aromatic chemicals. The Spice Mill, 69, 46-47, 50-5 1..

Keller D. , Krauledat M. and Scheel J. (2009) Feasibility study to support a threshold of sensitization concern concept in risk assessment based on human data. Archives of Toxicology, 83(12), 1049- 1060. Kellner W. and Kober W. (1955) The possibility of using ethereal oils for the disinfection of rooms. Arzneimittel-Forschung (Drug Research), 5, 224-229.

Kilic C.S. , Koyuncu M. , Ozek T. and Baser K.H.C. (2008) Essential oil of the leaves of Ribes nigrum L. from Turkey. Journal of Essential Oil Research, 20(6), 512-5 14. Kim H.-K. , Kim J.-R. and Ahn Y.-J. (2004) Acaricidal activity of cinnamaldehyde and its congeners against tyrophagus putrescentiae(Acari: Acaridae). Journal of Stored Products Research, 40(1), 55-63.

Kivcak B. , Akay S. , Demirci B. and Baser H.C. (2004) Chemical composition of essential oils from leaves and twigs of pistacia lentiscus, pistacia lenticus var: chia, and Pistacia terebinthus from Turkey. Pharmaceutical Biology, 42(4-5), 360-366.

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Kiecak G. (1979) The open epicutaneous test (OET), a predictive test procedure in the guinea pig for estimation of allergenic properties of simple chemical compounds, their mixtures and of finished cosmetic preparations. International Federation Societies Cosmetic Chemists, 9/18/79. Kiecak G. (1985) The Freund’s Complete Adjuvant Test and the Open Epicutaneous Test. In: Current Problems in Dermatology, Vol. 14, 152-17 1. Klopman G. and Chakravarti S.K. (2003) Screening of high production volume chemicals for estrogen receptor binding activity (II) by the Mu1tiCASE expert system. Chemosphere, 51(6), 461-468. Koch H.P. , Hofeneder M. and Bohne B. (1993) The yeast test: An alternative method for the testing of acute toxicity of drug substances and environmental chemicals. Methods and Findings in Experimental and Clinical Pharmacology, 15(3), 141-152. Kutob S. and Burlage H.M. (1956) Benzyl benzoate. A review. Amer. Prof. Pharmacist, 22(11), 1011- 1016. Research Institute for Fragrance Materials, mc, (2005). Potency estimations for several fragrance materials in the local lymph node assay and their correlation to human studies. Unpublished report 47900 from Lalko J. and Api A.M.

Lamas J.P. , Sanchez-Prado L. , Garcia-Jares C. and Llompart M. (2010) Determination of fragrance allergens in indoor air by active sampling followed by ultrasound-assisted solvent extraction and gas chromatography-mass spectrometry. Journal of Chromatography A, 1217(12), 1882-1890. Landegren I. , Borglund E. and Storgards K. (1979) Treatment of scabies with disulfiram and benzyl benzoate emulsion: A controlled study. Acta Dermato-Venereologica, 59(3), 274-276. Larsen W.G. (1977) Perfume dermatitis: A study of 20 patients. Archives of Dermatology, 113, 623- 626. Larsen W.G. (1983) Allergic contact dermatitis to the fragrance material lilial. Contact Dermatitis, 9(2), 158-159. T-P.Lee, R.Moscati and B.H.Park (1979) Effects of pesticides on human leukocyte functions. Research Communications in Chemical Pathology and Pharmacology, 23(3), 597-609. Lehman A.J. (1955) Insect repellents. Association Food Drug Officials United States, 19(2), 87-99. Leijs H. , Broekhans J. , Van Pelt L. and Mussinan C. (2005) Quantitative analysis of the 26 allergens for cosmetic labeling in fragrance raw materials and perfume oils. Journal of Agricultural and Food Chemistry, 53(14), 5487-5491.

Lis-Balchin M. , Hart S. and Lo B.W.H. (2002) Jasmine absolute (Jasminum grandiflora L.) and its mode of action on guinea-pig ileum in vitro. Phytotherapy Research, 16(5), 437-439. Ljunggren B. (1981) Contact dermatitis to estradiol benzoate. Contact Dermatitis, 7(3), 141-144. Loden M. and Edlund F. (2002) Strategies to reduce contact alleregy to fragrances. Cosmetics and Toiletries, 117(2), 39-46. Lynn E.V. and Gaston D. (1926) The relaxing action of some aromatic esters. Journal of the American Pharmaceutical Association, 15(7), Macht D.I. (1918) On the relation between the chemical structure of the opium alkaloids and their physiological action on the smooth muscle with a pharmacological and therapeutic study of some benzyl esters. II. A pharmacological and therapeutic study of some benzyl esters. Journal of Pharmacology, 11(6), 419-446. Macht D.I. (1919) Further experiences, experimental and clinical, with benzyl benzoate and benzyl alcohol. The Journal of Pharmacology and Experimental Therapeutics, 13, 509-511. Macht D.I. (1920) On the use of benzyl benzoate in some circulatory conditions. New York Medical Journal, 112(9), 269-27 1.

Macht D.I. (1921) Pharmacological examination of cinnamein, benzyl succinate and benzyl nitrite.

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Proceedings of the Society for Experimental Biology and Medicine, 18, 177-179. Macht D.I. and Leach H.P. (1929) Concerning the antipyretic properties of benzyl benzoate. The Journal of Pharmacology and Experimental Therapeutics, 35, 281-296.

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Maibach H.I. , Akerson J.M. , Marzulli F.N. , Wenninger J. , Greif M. , Hjorth N. , Andersen K.E. and Wilkinson D.S. (1980) Test concentrations and vehicles for dermatological testing of cosmetic ingredients. Contact Dermatitis, 6, 369-404.

Maruzzella J.C. , Chiaramonte J.S. and Garofalo M.M. (1961) Effects of vapours of aromatic chemicals on fungi. Journal of Pharmaceutical Sciences, 50(8), 665-668. Maruzzella J.C. and Bramnick E. (1961a) The antibacterial properties of perfumery chemicals. Soap, Perfumery and Cosmetics, 743-745.

Maruzzella J.C. , Garofalo M.M. and Chiaramonte J.S. (1961b) How vapors of aromatic chemicals affect bacteria. American Perfumer, 76(2), 35-39. Marzulli F.N. and Maibach H.I. (1976) Contact allergy: Predictive testing in man. Contact Dermatitis, 2, 1-17. Mason E.C. and Pieck C.E. (1920) A pharmacological study of benzyl benzoate. Journal of Laboratory and Clinical Medicine, 6, 62-77. Masuck I. , Hutzler C. and Luch A. (2010) Investigations on the emission of fragrance allergens from scented toys by means of headspace solid-phase microextraction gas chromatography Dmass spectrometry. Journal of Chromatography A, 1217(18), 3136-3 143.

Matthewson M.D. , Hughes G. , Macpherson I.S. and Bernard C.P. (1981) Screening techniques for the evaluation of chemicals with activity as tick repellents. Pestic. Sci., 12(4), 455-462. McCaughey H. (1968) Poisoning of cats with benzyl benzoate. Australian vet. Journal, 44(2), 82. McGlinn S.M. , Shepherd B.A. and Martan J. (1976) Effects of cyproterone acetate on associations of spermatozoa and seminal vesicles in the guinea pig. Contraception, 14(2), 183-19 1. Mclii M.C. , Giorgini S. and Sertoli A. (1983) Occupational dermatitis in a bee-keeper. Contact Dermatitis, 9(5), 427-428.

Meneghini C.L. , Vena G.A. and Angelini G. (1982) Contact dermatitis to scabicides. Contact Dermatitis, 8(4), 285-286. Meyer F. and Kerk L. (1960) Percutaneous absorption of physostigmine from benzene and some related solvents. Uber die percutane resorption von eserin aus benzol und einigen verwandten iosungsmitteln. Archives of Toxicology, 18, 131-139. Meyer F. (1965) Penetrating agents. Patent, British, 1,001,949, M49750IVa/30h, 7/20/61.

Meynadier J.-M. , Meynadier J. , Colmas A. , Castelain P.-Y. , Ducombs G. , Chabeau G. , Lacroix M., Martin P. and Ngangu Z. (1982) Preservative sensitivity. Annales Dermatologie Venereologie, 109(12), 1017-1023.

Meynadier J.M. , Meynadier J. , Peyron J.L. and Peyron L. (1986) Clinical forms of skin manifestations in allergy to perfume. Annales Dermatologie Venereologie, 113(1), 31-39. Migaily N. (1979) Effect of castor oil and benzyl benzoate used as a vehicle for antiandrogens on the adrenal cortex. Archives Andrology, 2(4), 365-369.

Mikhaeil B.R. , Maatooq G.T. , Badria F.A. and Amer M.M.A. (2003) Chemistry and iminunomodulatory activity of frankincense oil. Journal of Biosciences, 58(3-4), 230-238.

Miller M.D. , Yourtee D.M. , Glaros A.G. , Chappelow C.C. , Eick J.D. and Holder A.J. (2005) Quantum mechanical structure-activity relationship analyses for skin sensitization. Journal of

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Chemical Information and Modeling, 45(4), 924-929. Mitchell J.C. (1975) Patch testing with some components of balsam of Peru. Contact Dermatitis, 1, 39 1-392. Mitchell J.C. (1975a) Contact hypersensitivty to some perfume materials. Contact Dermatitis, 1, 196- 199.

Mitchell J.C. , Calnan C.D. , Clendenning W.E. , Cronin E. , Hjorth N. , Magnusson B. , Maibach H.I., Meneghini C.L. and Wilkinson D.S. (1976) Patch testing with some components of Balsam of Peru. Contact Dermatitis, 2(1), 57-58.

Mitchell J.C. , Adams R.M. , Glendenning W.E. , Fisher A. , Kanof N. , Larsen W. , Maibach H.I., Rudner E.J. , Schnorr W. , Storrs F. and Taylor J.S. (1982) Results of standard patch tests with substances abandoned. Contact Dermatitis, 8(5), 336-337.

Morita S. , Yamada A. , Ohgaki S. , Noda T. and Taniguchi S. (1980) Safety evaluation of chemicals for use in household products. I. Teratological studies on benzyl benzoate and 2-(morpholinothio) benzothiazole in rats. Annual Report Osaka City Inst.Public Health Environment Sci., 43, 90-97. Morris W.M. (1973) High resolution Infrared spectra of fragrance and flavor compounds. Journal Ass. off. analyt. Chem., 56(5), 1037-1064.

Morris J.A. , Khettry A. and Seitz E.W. (1979) Antimicrobial activity of aroma chemicals and essential oils. Journal of the American Oil Chemists Society, 56(5), 595-603. Munzig H.P. and Schels H. (1974) Potential replacement of preservatives in cosmetics by essential oils. Pollena-Tluszcze Srodki Piorace Kosmetyki, 18(8/9), 355-361.

Nardelli A. , DEiHooghe E. , Drieghe J. , Dooms M. and Goossens A. (2009) Allergic contact dermatitis from fragrance components in specific topical pharmaceutical products in Belgium. Contact Dermatitis, 60(6), 303 D313. Nath S.C. , Pathak M.G. and Baruah A. (1996) Benzyl benzoate, the major component of the leaf and stem bark oil of Cinnamomum zeylanicum Blume. Journal of Essential Oil Research, 8(3), 327- 328.

Natsch A. , Gfeller H. , Rothaupt M. and Ellis G. (2007) Utility and limitations of a peptide reactivity assay to predict fragrance allergens in vitro. Toxicology In Vitro, 21(7), 1220-1226. Natsch A. and Emter R. (2008) Skin sensitizers induce antioxidant response element dependent genes: Application to the in vitro testing of the sensitization potential of chemicals. Toxicological Sciences, 102(1-2), 110-119.

Natsch A. , Emter R. and Ellis G. (2009) Filling the concept with data: Integrating data from different in vitro and in silico assays on skin sensitizers to explore the battery approach for animal-free skin sensitization testing. Toxicological Sciences, 107(1), 106-121. Nethercott J.R. , Nield G. and Holness D.L. (1989) A review of 79 cases of eyelid dermatitis. Journal of the American Academy of Dermatology, 21, 223-230. Nielsen C. and Higgins J.A. (1921) Further observations on the pharmacology of benzyl compounds. Journal of Laboratory and Clinical Medicine, 7(2), 69-83. Nielsen N.M. and Bundgaard H. (1987) Prodrugs as drug delivery systems. 68. Chemical & plasma- catalyzed hydrolysis of various esters of benzoic acid: A reference system for designing prodrug esters of carboxylic acid agents. International Journal of Pharmacology, 39, 75-85. Nikawa H. , Yamamoto T. and Hamada T. (1995) Effect of components of resilient denture-lining materials on the growth, acid production and colonization of Candida albicans. Journal of Oral Rehab., 22(11), 817-824.

Nishimura M. , Ishihara M. , Itoh M. , Hosono K. and Kantoh H. (1984) Results of patch tests on cosmetic ingredients conducted between 1979 and 1982. Skin Research, 26(4), 945-954.

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Nishimura H. , Saito S. , Kishida F. and Matsuo M. (1994) Analysis of acute toxicity (LD5O-value) of organic chemicals to mammals by solubility parameter (delta). 2. Acute oral toxicity to mice. Sangyo Igaku, 36(6), 421-427.

Nishimura FT., Saito S. , Kishida F. and Matsuo M. (1994a) Analysis of acute toxicity (LD5O-value) of organic chemicals to mammals by solubility parameter (delta). 1. Acute oral toxicity to rats. Sangyo Igaku, 36(5), 314-323. [Japanese Journal Industrial Health] Nishimura H. , Saito S. , Kishida F. and Matsuo M. (1994b) Analysis of acute toxicity (LD5O-value) of organic chemicals to mammals by solubility parameter (delta). 3. Acute dermal toxicity to rabbits Sangyo Igaku, 36(6), 428-434. Nnoruka E.N. and Agu C.E. (2001) Successful treatment of scabies with oral ivermectin in Nigeria. Tropical Doctor, 31(1), 15-18. Nour-Eldin F. (1967) Chemical activation of blood-clotting factors and thrombosis. Nature (London), 214, 1362-1363. Nour-Eldin F. (1968) Phenols and blood coagulation. Journal of Biomedical Materials Research, 2, 23- 42.

Nunns D. , Ferguson J. , Beck M. and Mandal D. (1997) Is patch testing necessary in vulval vestibulitis? Contact Dermatitis, 37(2), 87-89. Ornellas M.R. (1965) Chemotherapeutic activitty of benzyl benzoate. Bulletin Cancer, 52(4), 371-378. Pack G.T. and Underhill F.P. (1922) The influence of benzyl benzoate upon nitrogenous metabolism. Journal Metabolic Research, 2, 73-105. Park B.H. and Lee T.P. (1978) Effects of pesticides on human leukocyte function. Proc. 4th FDA Symp. U.S.N.A. 8-28/30-78.

Patlewicz G. , Roberts D.W. and Uriarte E. (2008) A comparison of reactivity schemes for the prediction skin sensitization potential. Chemical Research in Toxicology, 21(2), 521-541.

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Pitche P. , Woilcenstein P. , Cremer G. , Foulet F. , Lascaux A-S. and Revuz J. (2002) Profuse scabies: Kinetic curve of parasitologic cure with an association of benzyl benzoate and sulfiram. Annales Dermatologie Venereologie, 129(3), 287-289.

Placzek M. , Fromel W. , Eberlein B. , Gilbertz K.-P. and Przybilla B. (2007) Evaluation of phototoxic properties of fragrances. Acta Dermato-Venereologica, 87(4), 312-316 Postel E. (1943) Benzyl benzoate, a component of Peru balsam. Klinische Wochenschrift, 22, 362-364. Pun C.P. , Elger W.G. and Pongubala J.M.R. (1987) Induction of menstruation by antiprogesterone ZK 98.299 in cycling bonnet monkeys. Contraception, 35(4), 409-421. Quest International (1977) Guinea pig skin irritation test with benzyl benzoate. Unpublished. August 08 Quest International (1981) Guinea pig skin irritation test with benzyl benzoate. Unpublished. April 21 Quest International Ltd. (1994) Biodegradation study of benzyl benzoate in a sealed vessel test. Unpublished. September 27

Raina V.K. , Srivastava S.K. and Syamasunder K.V. (2003) Essential oil composition of Acorus calamus L. from the lower region of the Himalayas. Flavour and Fragrance Journal, 18(1), 18-20.

Ramsey J.D. , Lee T.D. , Osselton M.D. and Moffat A.C. (1980) Gas-liquid chromatographic retention indices of 296 non-drug substances on SE-30 or OV-1 likely to be encountered in toxicological

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analyses. Journal of Chromatography A, 184, 185-206.

Rastogi S.C. , Johansen J.D. , Frosch P. , Menne T. , Bruze M. , Lepoittevin J.P. , Dreier B. , Andersen K.E. and White I.R. (1998) Deodorants on the European market: Quantitative chemical analysis of 21 fragrances. Contact Dermatitis, 38(1), 29-35.

Rastogi S.C. , Heydorn S. , Johansen J.D. and Basketter D.A. (2001) Fragrance chemicals in domestic and occupational products. Contact Dermatitis, 45(4), 221-225.

Raynaud S. , Fourneau C. , Laurens A. , Hocquemiller R. , Loiseau P. and Bones C. (2000) Squamocin and benzyl benzoate, acaricidal components of Uvaria pauci-ovulata bark extracts. Planta Medica, 66(2), 173-175. Reifenrath W.G. (1995) Volatile substances. Many factors affect evaporation and skin penetration. But which are the most predictive? Cosmetics and Toiletries, 110(7), 85-93. Research Institute for Fragrance Materials, mc, (1970). The contact sensitizing potential of fragrance materials in humans. Report to RIFM. Unpublished report 1760 from Kligman A.M. June 01 Research Institute for Fragrance Materials, mc, (1978). Report on human maximization studies. Report to RIFM. Unpublished report 1787 from Kligman A.M. February 28 Research Institute for Fragrance Materials, mc, (1979). Report on human maximization studies. Report to RIFM. Unpublished report 1697 from Epstein W.L. July 06 Research Institute for Fragrance Materials, mc, (1980). 30-Day dermal toxicity study of benzyl benzoate in rats. Report to RIFM. Unpublished report 4000 from Moreno O.M. , Ratcliffe H.L. and Singh H. Research Institute for Fragrance Materials, mc, (1984). Primary irritation test in rabbits. Report to RIFM. Unpublished report 1795 from Haynes G. June 01 Research Institute for Fragrance Materials, mc, (1985). Primary irritation test in rabbits. Report to RIFM. Unpublished report 3099 from Haynes G. June 01 Research Institute for Fragrance Materials, mc, (2003). Modified primary dermal irritation test (MPDI) with benzyl benzoate. Unpublished report 44230 from Harrison L.B. and Stolman L.P. November 24 Research Institute for Fragrance Materials, mc, (2004). Repeated insult patch test with benzyl benzoate. Unpublished report 47159 from Harrison L.B. and Spey D.R. July 07a July 07b Research Institute for Fragrance Materials, mc, (2004). Rechallenge of repeated insult patch test with benzyl benzoate. Unpublished report 47160 from Harrison L.B. and Spey DR. Research Institute for Fragrance Materials, mc, (2005). Benzyl benzoate diluted with vehicle 1:3 ETOH:DEP: Local Lymph Node Assay Unpublished report 47377 from Betts C.J. January 20 Reverchon E. and DellaPorta G. (1997) Tuberose concrete fractionation by supercritical carbon dioxide. Journal of Agricultural and Food Chemistry, 45(4), 1356-1360. Roberts G. (2002) Procedures for supplying fragrance information to dermatologists. American Journal of Contact Dermatitis, 13(4), 206-2 10.

Roberts G. , Vey M. and Troy W. (2002a) Skin reaction to fragrances: Procedures for promptly supplying fragrance information to dermatologists. Exogenous Dermatology, 1(5), 266-268.

Roberts D.W. , Patlewicz G. , Kern P.S. , Gerberick F. , Kimber I. , Dearman R.J. , Ryan C.A. Basketter D.A. and Aptula A.O. (2007) Mechanistic applicability domain classification of a local lymph node assay dataset for skin sensitization. Chemical Research in Toxicology, 20(7), 1019- 1030.

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Romaguera C. , Camarasa J.M.G. , Alomar A. and Grimalt F. (1983) Patch tests with allergens related to cosmetics. Contact Dermatitis, 9(2), 167-168. Rudner E.J. (1977) North American group results. Contact Dermatitis, 3(3), 208-209. Rudner E.J. (1978) Patch test results of the North American Contact Dermatitis Group. Cosmetics and Toiletries, 93, 53-54. Rudzki E. and Grzywa Z. (1983) Dermatitis from propolis. Contact Dermatitis, 9, 40.

Sacchetti G. , Maietti S. , Muzzoli M. , Scaglianti M. , Manfredini S. , Radice M. and Bruni R. (2005) Comparative evaluation of 11 essential oils of different origin as functional antioxidants, antiradicals and antimicrobials in foods. Food Chemistry, 91(4),621-632.

Samarasekera R. , Kaihari K.S. and Weerasinghe 1.5. (2005) Mosquitocidal activity of leaf and bark essential oils of ceylon Cinnamomum zeylanicum. Journal of Essential Oil Research, 17(3), 301- 303. Schaaf V.F. (1961) Chemische konstitution und wirkung im akanthosetest. Dermatologica, 123, 362- 374. Scheinman P.L. (1996) Allergic contact dermatitis to fragrance: A Review. American Journal of Contact Dermatitis, 7(2), 65-76.

Schiede E. , Mischke U. , Roll R. and Kayser D. (1992) A national validation study of the acute-toxic- class method - An alternative to the LD5Otest. Archives of Toxicology, 66(7), 455-470.

Schmeltz I. , Stedman R.L. , Chamberlain W.J. and Stills C.D. (1965) Benzyl esters, indoles and carbazoles in cigarette smoke. Chemisty md., December, 2009-20 10.

Schnuch A. , Uter W. , Geier J. , Lessmarm H. and Frosch P.J. (2007) Sensitization to 26 fragrances to be labelled according to current European regulation. Contact Dermatitis, 57(1), 1-10. Schorr W.F. (1978) Testing for perfume allergy: A complicated challenge. Cosmetics and Toiletries, 93, 49-53. Schroeder W. (2009) Understanding fragrance in personal care. Cosmetics and Toiletries, 124(11), 36, 37-42, 44.

Schultz T.W. , Netzeva T.I. and Cronin M.T.D. (2003) Selection of data sets for QSARs: Analyses of Tetrahymena toxicity from aromatic compounds. SAR & QSAR in Environmental Research, 14 (1), 59-81. Schunk H. and Shibamoto T. (1986) A photochemical study on essential oil constituents: Benzyl benzoate and eugenol. In Sci. Prog. & Abs. Book, Nov. 16-20, 86.

Schunk H.H. , Shibamoto T. , Tan H.K. and C-I.Wei (1986a) Biological and chemical studies on photochemical products obtained from eugenol, benzyl acetate and benzyl benzoate. Flavors Frag., A World Persp. Proc. 10th Internat. Cong. Es. Oils., 1045-1068.

Schunk H.H. , Shibamoto T. , Tan H.K. and Wei C.-I. (1988) Biological and chemical studies on photochemical products obtained from eugenol, benzyl acetate and benzyl benzoate. Developments in Food Science, 18, 1045-1068.

Serra J.R. , Jurs P.C. and Kaiser K.L.E. (2001) Linear regression and computational neural network prediction of Tetrahymena acute toxicity for aromatic compounds from molecular structure. Chemical Research in Toxicology, 14(11), 1535-1545.

Shamukhamedova S.I. , Vainshtein A.M. and Anikyavichus V.B. (1972) Laboratory diagnosis and clinical picture of scabies in Latvian SSR. Med. Paraznol. Parazit. Bolezni., 41(5), 581-583. Shellie R. , Marriott P. and Chaintreau A. (2004) Quantitation of suspected allergens in fragrances (Part I): Evaluation of comprehensive two-dimensional gas chromatography for quality control. Flavour and Fragrance Journal, 19(2), 91-98.

Shibamoto T. and Umano K. (1985) Photochemical products of benzyl benzoate: Possible formation of

CIR Panel Book Page 273 _.1 IIt— fl Benzyl benzoate Distributed for Comment Only - Do Not Quote or Cite Page 66 of 67

skin allergens. Journal of Toxicology: Cutaneous and Ocular Toxicology, 4(2), 97-103. Shibamoto T. (1985a) Photochemistry Of A Major Essential Oil Constituent, Benzyl Benzoate. In The Shelf Life Of Foods And Beverages, 12, 745-754. Shin S. (2003) Anti-aspergillus activities of plant essential oils and their combination effects with ketoconazole or amphotericin B. Archives of Pharmacal Research, 26(5), 389-393. Simici D. and Marcu I. (1926) The vascular action of benzyl benzoate and acetate in man. Archives Mal. Coeur, 19(10), 654-662.

Smith J.S. , Macma O.T. , Sussman N.B. , Karol M.H. and Maibach 11.1.(2000) Experimental validation of a structure-activity relationship model of skin irritation by esters. Quantitative Structure -Activity Relationships, 19(5), 467-474. Snapper J. , Grunbaum A. and Sturkop S. (1925) About the fission and oxidation of benzyl alcohol and benzyl esters in the human organism. Biochemische Zeitschrift, 155, 163-173. Solan J.L. and Laden K. (1977) Factors affecting the penetration of light through stratum corneum. Journal of the Society of Cosmetic Chemists Japan, 28, 125-137. Somogyi L.P. (1996) The flavour and fragrance industry: Serving a global market. Chemisty md., March 4, 170-173. Spiegel A.J. and Noseworthy M.M. (1963) Use of nonaqueous solvents in parenteral products. Journal of Pharmaceutical Sciences, 52(10), 917-927. Stater J.W. (1922) The action of benzyl benzoate and morphine on the vesical sphincter. Journal of Urology, 8, 239-245. Steinberg D.C. (2001) Regulatory review. Cosmetics and Toiletries, 116(11), 26-34. Steinberg D.C. (2002) The European Cosmetic Directive: The impact on fragrances. Global Cosmetic Industry (Gd), 170(2), 29-34. Steinberg D.C. (2003) The labeling of fragrance allergens in the European Union. Cosmetics and Toiletries, 118(11), 69-72. Stofberg J. and Grundschober F. (1987) Consumption ratio and food predominance of flavoring materials. Perfumer and Flavorist, 12(4), 27-68.

Stransky L. , Vasileva S. and Mateev G. (1996) Contact bullous pemphigoid? Contact Dermatitis, 35 (3), 182.

Sugawara M. , Nakayama H. , Araki Y. , Watanabe S. , Someya T. and Muraki S. (1984) Contact hypersensitivity to ylang-ylang oil components. Skin Research, 26(4), 912-919. [Hiful Takasago International Corporation (1981) Toxicology studies of benzyl benzoate in the guinea pig. Unpublished. November 18 November 18A November 18B

Takenaka T. , Hasegawa E. , Takenaka U. , Saito F. and Odaka T. (1986) Fundamental studies of safe compound perfumes for cosmetics. Part 1. The primary irritation of compound materials to the skin. Unknown Source, pp. 313-329. Thelestam M. , Curvall M. and Enzell C.R. (1980) Effect of tobacco smoke compounds on the plasma membrane of cultured human lung fibroblasts. Toxicology, 15(3), 203-217.

Toda H. , Mihara S. and Shibamoto T. (1988) Studies on photochemistry of flavor chemicals. Devi. Food Sci., 17, 585-602.

Ucar G. , Balaban M. and Usta M. (2003) Volatile needle and wood extrracts of oriental spruce Picea orientalis (L.) Link. Flavour and Fragrance Journal, 18(5), 368-375. Vey M. (2003) Procedures for supplying information promptly to dermatologists. [Letter to the Editor] Contact Dermatitis, 48(1), 56-58.

Villa C. , Gambaro R. , Mariani E. and Dorato S. (2007) High-performance liquid chromatographic method for the simultaneous determination of 24 fragrance allergens to study scented products.

CIR Panel Book Page 274 Benzyl benzoate Distributed for Comment Only - Do Not Quote or Cite Page 67 of 67

Journal of Pharmaceutical and Biomedical Analysis, 44(3), 755 E762.

Ward T.J. , Wyskiel D.C. and Boeri R.L. (2003) Growth and reproduction toxicity test with benzyl benzoate and the freshwater alga, Selenastrum capricornutum. Unpublished.

Wetzel C.H. , Oles M. , Wellerdieck C. , Kuczkowiak M. , Gisselmann G. and Hatt H. (1999) Specificity and sensitivity of a human olfactory receptor functionally expressed in human embryonic kidney 293 cells and Xenopus laevis oocytes. The Journal of Neurosciences, 19(17), 7426-7433. World Health Organization (2002) Benzyl derivatives. WHO Food Additives Series, 48, 227-271.

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Yamada T. , Ohsawa K. and Ohno H. (1988) The usefulness of alkaline solutions for clearing the uterus and staining implantation sites in rats. Exp. Anim.(Tokyo), 37(3), 325-331. Zhai H. and Maibach H.I. (2001) Effects of skin occulusion on percutaneous absorption: An overview. Skin Pharmacology and Applied Skin Physiology, 14(1), 1-10.

No data added since 27-Oct-10.

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PersonalCare ProductsCouncil Committedto Safety, Quality& Innovation Memorandum

TO: F. Alan Andersen, Ph.D. Director - COSMETIC INGREDIENT REVTEW (CIR)

FROM: John Bailey, Ph.D. Industry Liaison to the CIR Expert Panel

DATE: May 19, 2010

SUBJECT: Concentration of Use Benzyl Alcohol, Benzoic Acid, Sodium Benzoate and Potential Additions

11011 7th Street, N.W, Suite 300 Washington, D.C. 20036-4702 202.331.1770 202.331.1969 (fax) www.personalcarecouncil.org

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Concentration of Use Benzyl Alcohol, Benzoic Acid, Sodium Benzoate, Calcium Benzoate*, Magnesium Benzoate*, Potassium Benzoate*, Benzyl Benzoate9[

Ingredient Product Category Concentration of Use

Benzyl Alcohol Baby shampoos 0.3%

Benzyl Alcohol Baby lotions, oils, powders and creams 0.3%

Benzyl Alcohol Other baby products 0.004%

Benzyl Alcohol Bath oils, tablets and salts 0.009-0.9%

Benzyl Alcohol Bubble baths 0.00 1-0.3%

Benzyl Alcohol Eyebrow pencil 0.00002-0.2%

Benzyl Alcohol Eyeliner 0.000008-0.8%

Benzyl Alcohol Eye shadow 0.00003-0.4%

Benzyl Alcohol Eye lotion 0.3-0.6%

Benzyl Alcohol Eye makeup remover 0.000009-0.5%

Benzyl Alcohol Mascara 0.00003-0.7%

Benzyl Alcohol Colognes and toilet waters 0.4-2%

Benzyl Alcohol Perfumes 0.04-3%

Benzyl Alcohol Powders (dusting and talcum) 0.04-0.05%

Benzyl Alcohol Sachets 2%

Benzyl Alcohol Other fragrance preparations 0.02-0.9%

Benzyl Alcohol Hair conditioners 0.002-1%

Benzyl Alcohol Hair sprays (aerosol fixatives) 0.0003%

Benzyl Alcohol Permanent waves 0.0002%

Benzyl Alcohol Rinses (noncoloring) 3%

Benzyl Alcohol Shampoos (noncoloring) 0.006-1%

Benzyl Alcohol Tonics, dressings and other hair grooming aids 0.0008-1%

Benzyl Alcohol Other hair preparations (noncoloring) 0.005-2%

Benzyl Alcohol Hair dyes and colors (all types requiring caution statement and patch 2-10% testing)

Benzyl Alcohol Hair tints 0.0004%

Benzyl Alcohol Hair bleaches 0.00007-4%

Benzyl Alcohol Other hair coloring preparations 10%

Benzyl Alcohol Blushers (all types) 0.00005-0.0005%

Page 1 of 7

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Benzyl Alcohol Face powders 0.0001-002%

Benzyl Alcohol Foundations 0.0006-1%

Benzyl Alcohol Leg and body paints 0,05-0.4%

Benzyl Alcohol Lipstick 0.02-0.9%

Benzyl Alcohol Makeup bases 0.0002%

Benzyl Alcohol Other makeup preparations 0.00002%

Benzyl Alcohol Basecoats and undercoats (manicuring preparations) 0.00002%

Benzyl Alcohol Cuticle softeners 0.2%

Benzyl Alcohol Nail creams and lotions 0.00006-0.09%

Benzyl Alcohol Nail polish and enamel 0.002-0.5%

Benzyl Alcohol Nail polish and enamel removers 0.0009-0.3%

Benzyl Alcohol Dentifrices (aerosol, liquid, pastes and powders) 0.00002-0.2%

Benzyl Alcohol Mouthwashes and breath fresheners (liquids and sprays) 0.000006%

Benzyl Alcohol Bath soaps and detergents 0.005-0.9%

Benzyl Alcohol Deodorants (underarm) 0.03-0.5%

Benzyl Alcohol Feminine hygiene deodorants 1%

Benzyl Alcohol Other personal cleanliness products 0.009-0.09%

Benzyl Alcohol Aftershave lotions 0.0002-0.5%

Benzyl Alcohol Preshave lotions (all types) 0.02%

Benzyl Alcohol Shaving cream (aerosol, brushless and lather) 0.0001-0.02%

Benzyl Alcohol Shaving soaps (cakes, sticks, etc.) 0.04%

Benzyl Alcohol Other shaving preparations 0.00005-0.4%

Benzyl Alcohol Skin cleansing (cold creams, cleansing lotions, liquids and pads) 0.00003-1%

Benzyl Alcohol Depilatories 0.003-0.3%

Benzyl Alcohol Face and neck creams, lotions and powders 0.04-1%

Benzyl Alcohol Body and hand creams, lotions and powders 0.02-0.1%

Benzyl Alcohol Moisturizing creams, lotions and powders 0.0008-1%

Benzyl Alcohol Night creams, lotions and powders 0.06-0.6%

Benzyl Alcohol Paste masks (mud packs) 0.0002-0.7%

Benzyl Alcohol Skin fresheners 0.5%

Benzyl Alcohol Other skin care preparations 0.003-0.009%

Benzyl Alcohol Suntan gels, creams and liquids 0.0005%

Page 2 of 7

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Benzyl Alcohol Indoor tanning preparations 0.0002%

Benzyl Alcohol Other suntan preparations 0.0009%

Benzoic Acid Baby lotions, oils powders and creams 0.000002%

Benzoic Acid Bath oils, tablets and salts 0.000003-0.08%

Benzoic Acid Eyebrow pencil 0.1%

Benzoic Acid Eyeliner 0.00005-0.03%

Benzoic Acid Eye shadow 0.2%

Benzoic Acid Eye lotion 0.005-0.2%

Benzoic Acid Eye makeup remover 0.2%

Benzoic Acid Mascara 0.002%

Benzoic Acid Colognes and toilet waters 0.000003%

Benzoic Acid Perfumes 0.000003-0.05%

Benzoic Acid Powders (dusting and talcum) 0.001%

Benzoic Acid Hair conditioners 0.00002-1%

Benzoic Acid Hair sprays (aerosol fixatives) 0.02-0.08%

Benzoic Acid Rinses (noncoloring) 0.00005%

Benzoic Acid Shampoos (noncoloring) 0.00002-0.5%

Benzoic Acid Tonics, dressings and other hair grooming aids 0.0004-0.3%

Benzoic Acid Hair dyes and colors (all types requiring caution statement and patch 0.004% testing)

Benzoic Acid Hair lighteners with color 0.03%

Benzoic Acid Other hair coloring preparations 0.0 1%

Benzoic Acid Face powders 0.000005%

Benzoic Acid Foundations 0.00005-0.2%

Benzoic Acid Lipstick 0.003-0.3%

Benzoic Acid Makeup bases 0.6%

Benzoic Acid Cuticle softeners 0.2%

Benzoic Acid Nail creams and lotions 0.0005%

Benzoic Acid Mouthwashes and breath fresheners (liquids and sprays) 0.005-2%

Benzoic Acid Bath soaps and detergents 0.000005-0.5%

Benzoic Acid Deodorants (underarm) 0.03-0.5%

Benzoic Acid Other personal cleanliness products 0.000003-0.2%

Page 3 of 7

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Benzoic Acid Aftershave lotions 0.00005-0.1%

Benzoic Acid Shaving cream (aerosol, brushless and lather) 0.06-0.2%

Benzoic Acid Skin cleansing (cold creams, cleansing lotions, liquids and pads) 0.000005-5%

Benzoic Acid Depilatories 0.008-0.09%

Benzoic Acid Face and neck creams, lotions and powders 0.0003-0.2%

Benzoic Acid Body and hand creams, lotions and powders 0.0006-1%

Benzoic Acid Moisturizing creams, lotions and powders 0.00005-0.2%

Benzoic Acid Night creams, lotions and powders 0.07-0.3%

Benzoic Acid Paste masks (mud packs) 0.0002-0.2%

Benzoic Acid Skin fresheners 0.2%

Benzoic Acid Other skin care preparations 0.0006-0.2%

Benzoic Acid Suntan gels, creams and liquids 0.00005%

Benzoic Acid Other suntan preparations 0.2%

Sodium Benzoate Baby shampoos 0.3%

Sodium Benzoate Baby lotions, oils, powders and creams 0.3%

Sodium Benzoate Other baby products 0.5-0.9%

Sodium Benzoate Bath oils, tablets and salts 0.3-1%

Sodium Benzoate Bubble baths 0.004-0.3%

Sodium Benzoate Eyeliner 0.0001-0.3%

Sodium Benzoate Eye shadow 0.1-0.2%

Sodium Benzoate Eye lotion 0.02-0.3%

Sodium Benzoate Eye makeup remover 0.2%

Sodium Benzoate Mascara 0.00001%

Sodium Benzoate Other eye makeup preparations 0.4%

Sodium Benzoate Perfumes 0.0001-0.05%

Sodium Benzoate Hair conditioners 0.000005-1%

Sodium Benzoate Hair sprays (aerosol fixatives) 0.3%

Sodium Benzoate Permanent waves 0.1%

Sodium Benzoate Rinses (noncoloring) 0.3-1%

Sodium Benzoate Shampoos (noncoloring) 0.00001-0.6%

Sodium Benzoate Shampoos (noncoloring) 0.1-0.5%

Sodium Benzoate Other hair preparations (noncoloring) 0.003%

Page 4 of 7

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Sodium Benzoate Hair dyes and colors (all types requiring caution statement and patch 0.008-0.5% testing)

Sodium Benzoate Hair bleaches 0.4%

Sodium Benzoate Other hair coloring preparations 0.2%

Sodium Benzoate Blushers (all types) 0.02%

Sodium Benzoate Face powders 0.3%

Sodium Benzoate Foundations 0.2%

Sodium Benzoate Leg and body paints 0.2-0.5%

Sodium Benzoate Lipstick 0.002%

Sodium Benzoate Makeup bases 0.0002%

Sodium Benzoate Cuticle softeners 0.00 1%

Sodium Benzoate Nail polish and enamel 0.00001%

Sodium Benzoate Dentifrices (aerosol, liquid, pastes and powders) 0.003-0.6%

Sodium Benzoate Mouthwashes and breath fresheners (liquids and sprays) 0.3-0.6%

Sodium Benzoate Other oral hygiene products 0.001%

Sodium Benzoate Bath soaps and detergents 0.0004-0.5%

Sodium Benzoate Deodorants (underarm) 0.5%

Sodium Benzoate Other personal cleanliness products 0.005-0.6%

Sodium Benzoate Aftershave lotions 0.00 1-0.002%

Sodium Benzoate Shaving creams (aerosol, brushless and lather) 0.1-0.3%

Sodium Benzoate Other shaving preparations 0.3%

Sodium Benzoate Skin cleansing (cold creams, cleansing lotions, liquids and pads) 0.000001-0.6%

Sodium Benzoate Depilatories 0.00005-0.2%

Sodium Benzoate Face and neck creams, lotions and powders 0.002-0.4%

Sodium Benzoate Face and neck sprays 0.5%

Sodium Benzoate Body and hand creams, lotions and powders 0.0001-1%

Sodium Benzoate Moisturizing creams, lotions and powders 0.0004-0.3%

Sodium Benzoate Night creams, lotions and powders 0.009-0.3%

Sodium Benzoate Paste masks (mud packs) 0.0004-0.5%

Sodium Benzoate Other skin care preparations 0.003-0.2%

Sodium Benzoate Suntan gels, creams and liquids 0.0002%

Sodium Benzoate Other suntan preparations 0.003%

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Calcium Benzoate Face powders 0002%

Calcium Benzoate Foundations 0.003%

Calcium Benzoate Suntan gels, creams and liquids 0004%

Potassium Benzoate Cuticle softeners 0.0003%

Potassium Benzoate Bath soaps and detergents 0.0002%

Benzyl Benzoate Bath oils, tablets and salts 0.007-2%

Benzyl Benzoate Bubble baths 0.0005%

Benzyl Benzoate Eye lotion 0.0006-0.002%

Benzyl Benzoate Perfumes 05-4%

Benzyl Benzoate Powders (dusting and talcum) 03%

Benzyl Benzoate Other fragrance preparations 0.002%

Benzyl Benzoate Hair conditioners 0.009-2%

Benzyl Benzoate Hair sprays (aerosol fixatives) 0.006%

Benzyl Benzoate Shampoos (noncoloring) 0.3-2%

Benzyl Benzoate Tonics, dressings and other hair grooming aids 0.05-4%

Benzyl Benzoate Other hair preparations (noncoloring) 0.01%

Benzyl Benzoate Hair dyes and colors (all types requiring caution statement and patch 0.5-2% testing)

Benzyl Benzoate Other hair coloring preparations 0.07%

Benzyl Benzoate Face powders 0.001-0.02%

Benzyl Benzoate Foundations 0.0001-0.005%

Benzyl Benzoate Leg and body paints 0.002%

Benzyl Benzoate Lipstick 0.000005-0.2%

Benzyl Benzoate Nail creams and lotions 0.2%

Benzyl Benzoate Nail polish and enamel 0.8%

Benzyl Benzoate Dentifrices (aerosol, liquid, pastes and powders) 0,0002%

Benzyl Benzoate Bath soaps and detergents 0.006-0.4%

Benzyl Benzoate Deodorants (underarm) 0.006-0.6%

Benzyl Benzoate Other personal cleanliness products 0.00 14-0.2%

Benzyl Benzoate Aftershave lotions 0.00 1-0.4%

Benzyl Benzoate Preshave lotions (all types) 0.0008%

Benzyl Benzoate Shaving cream (aerosol. brushless and lather) 0.0001-0.4%

Page 6 of 7

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Benzyl Benzoate Shaving soaps (cakes, sticks, etc.) 0.02%

Benzyl Benzoate Other shaving preparations 0.05%

Benzyl Benzoate Skin cleansing (cold creams, cleansing lotions, liquids and pads) 0.009-0.02%

Benzyl Benzoate Depilatories 0.007%

Benzyl Benzoate Face and neck creams, lotions and powders 0.009-0.6%

Benzyl Benzoate Body and hand creams, lotions and powders 0.006-0.6%

Benzyl Benzoate Body and hand sprays 0.007%

Benzyl Benzoate Night creams, lotions and powders 0.004-0.007%

Benzyl Benzoate Other skin care preparations 0.02%

Benzyl Benzoate Indoor tanning preparations 0.004%

Benzyl Benzoate Other suntan preparations 0.008% *Under consideration for addition to the report on Benzyl Alcohol, Benzoic Acid and Sodium Benzoate. ¶Ingredients included in the title of the table but not found in the table were included in the concentration of use survey, but no uses were reported. Information collected in 2010 Table prepared May 19. 2010

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PersonalCare iProductsCouncil Committedto Safety, Quality& Innovation Memorandum

TO: F. Alan Andersen, Ph.D. Director - COSMETIC 1NGREDIENT REVIEW (Cifi)

FROM: John Bailey, Ph.D. Industry Liaison to the CIR Expert Panel

DATE: March 24, 2011

SUBJECT: Comments on the Scientific Literature Review on Benzyl Alcohol, Benzoic Acid and its Salts and Ester

p.3, 18 - Please update the USP (more recent volumes in John Krowka’s office) and Food Chemical Codex (more recent volume in Carol Eisenmann’s office) specifications. Sodium Benozate is also in the Food Chemical Codex. The Food Chemical Codex limits for lead are now not more than 2 mg/kg lead (rather than 10 mg/kg as stated in the Scientific Literature Review). p.4 - In the cosmetic use section, please also cite the 2005 SCCP opinion which can be found at http://ec.europa.eu/health/ph risklcommittees/04 sccp/docs/sccp o 015.pdf (a link to this opinion is also available from the On-Line). p.5 - If Benzyl Benzoate is in category ifi which means “insufficient data available to permit final classification”, it is not clear why the report says “benzyl benzoate has been approved for use in OTC pedulicides”. p.5 - In the second paragraph under Benzyl Benzoate, please delete “species not stated”. Dogs and cats were used. If you need to include something, “breed not stated” would be more accurate. p.6 - Where is the in vivo guinea pig study described in the summary of the Dermal Studies section? Is the summary actually referring to the in vitro study of Benzoic Acid penetration through guinea pig skin? p.6 - The paragraph on the rhesus monkey dermal penetration study cited to the RJCLID dataset (reference 3) needs to be deleted. This is the FDA study (reference 56 in the Cifi report) described in the previous paragraph. The TUCLID dataset cites the rhesus monkey dermal penetration study to reference 83. Reference 83 in the reference section of the RJCLID dataset is given as “Bronaugh, R.L. et al., Fd. Chem. Toxicol. 28, 369-373 (1990).” which is the same as reference 56 in the CIR report. p.7 - The sentence “The killing of remaining animals was scheduled for the day immediately after the last exposure” suggests that some animals died. The next sentence starts with: “None of the animals died...” Which sentence is correct? p.7 - In the summary of the Acute Oral Exposure section it states that oral 50LD values in mice were 2,800 mg/kg. Where is that value for mice coming from as it is not in the text or Table 4? p.7 - For which species was the oral 50LD for Benzyl Alcohol 1610 mg/kg?

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p.9 - The Benzyl Benzoate study in rats, cats and rabbits should be moved into the Benzyl Benzoate section. It should be made clear that this is an abstract cited to a secondary reference. p.9 - In the summary of the Repeated Dose Exposure section, please include the dermal doses of Benzyl Benzoate that were associated with effects. p.9 - The doses used in the study described in the second paragraph under Benzyl Benzoate are not clear. p.10 - Please provide the doses in the summary of the Repeated Dose Exposure section. p.10 - In the summary of the Ocular Irritation/Toxicity section, please provide the concentrations that were irritating. p.10 - What was the vehicle used in the ocular toxicity study of Benzyl Alcohol in rabbits? p.11 - Please include the concentrations of Benzyl Alcohol, Benzoic Acid and Sodium Benzoate that were irritating to the eyes of rabbits. p.12 - The second paragraph under Benzyl Alcohol, Sodium Benzoate and Benzyl Benzoate in the Photohemolysis section says: “The radiation sources described in the preceding study were used.” But the sources used are not described in the previous paragraph. It just says that the substances were exposed to UVA or UVB radiation. p.13 - In the summary of the Reproductive and Developmental Toxicity section, please provide the doses. What were the NOAEL or NOEL values for developmental toxicity? Please revise the following sentence. “There was no evidence of adverse effects on pregnancy outcome was observed in a study of the safety of topical application of benzyl benzoate (25%) lotion in pregnant women.” p.13 - In the last paragraph of this page, it is not clear what the 0.01 g and 0.0125 g doses represent, a dietary level, a dose per rat, a total dose, or a daily dose? p.14- - The second and third paragraphs of this page appear to be describing the same study. Both sources (RIFM synopsis and IIJCLID dataset) indicate that the study was completed in Wistar rats. RIFM cites this study to Morita et al. (1980) and the UJCLID cites to Morita et al. (1981). Despite the differences in the date, the source of the reference (Annual Report Osaka City Inst.Public Health Environment Sci., 43, 90-97) is the same in both sources. p.15 - Please include the concentration of Benzoic Acid that was negative in the recombinant yeast human estrogen receptor assay (first paragraph). p.15 - Please include the concentration of Benzyl Benzoate that produced estrogenic responses in cultured human breast cancer cells (fourth paragraph). p.17 - In the summary of the Carcinogenicity section, please include the doses used in the NTP study. Based on what effect did EPA consider the NTP study results equivocal? p.17 - Please include the route of exposure used in the NTP study. The NTP study should be cited to NTP (reference 77) rather than EPA (reference 76). p.18 - In the Summary, Please include the doses and species used in the uterotrophic assay. At what concentration relative to estrogen did Benzyl Benzoate induce an estrogenic response in human breast cancer cells? p.18 - Please include the inhalation exposure concentrations of Benzyl Alcohol and Benzoic Acid that caused death in rats, and the concentration in the repeated exposure study that was not associated with any effects. p.19 - What dermal doses of Benzyl Benzoate were associated with effects in rats and rabbits?

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p.19 - Please provide the concentrations of Benzyl Alcohol, Benzyl Benzoate, Benzoic Acid and Sodium Benzoate that were irritating to the eyes of rabbits. p.19 - At what concentration was Benzyl Benzoate “slightly phototoxic’? p.19 - Please provide the dietary dose of Sodium Benzoate that resulted in marked maternal toxicity. Please provide the dermal dose of Benzyl Benzoate that was associated with testicular atrophy in rabbits. p.19 - At what concentrations were Benzoic Acid and Sodium Benzoate photogenotoxic? What doses were used in the NTP carcinogenicity study? Based on what tumor type did EPA consider the results of the NTP study equivocal? p.22, Table 4 - Please define GLP at the end of the Table. p.32 - Please update references 20 (USP) and 21 (Food Chemical Codex). p.34 - References 54 and 60 are the same reference, the RIFM Benzyl benzoate synopsis.

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