Safety Assessment of Apple-Derived Ingredients As Used in Cosmetics

Safety Assessment of Apple-derived Ingredients as Used in Cosmetics

Status: Draft Final Report for Panel Review Release Date: November 20, 2015 Panel Date: December 14-15, 2015

The 2015 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 Lillian J. Gill, D.P.A. This report was prepared by Wilbur Johnson, Jr., M.S., Senior Scientific Analyst.

Commitment & Credibility since 1976

Memorandum

To: CIR Expert Panel Members and Liaisons From: Wilbur Johnson, Jr. Senior Scientific Analyst Date: November 20, 2015 Subject: Draft Final Report on Apple-derived Ingredients

At the September 21-22, 2015 Expert Panel meeting, a tentative report with two conclusions was issued. The Panel concluded that 25 apple-derived ingredients are safe in the present practices of use and concentration in cosmetics as described in this safety assessment, when formulated to be non-irritating and non-sensitizing. The Panel also concluded that the available data are insufficient for evaluating the safety of pyrus malus (apple) root extract, pyrus malus (apple) stem extract, and malus domestica (apple) stem extract, and the data that are needed to evaluate the safety of these three ingredients are:

(1) Method of manufacture and impurities (2) 28-day dermal toxicity study; if any adverse effects are noted, genotoxicity and reproductive and developmental toxicity data may be needed

The data requested have not been received. However, both updated and new use concentration data on apple-derived ingredients were received, and have been added to the safety assessment.

Included in this package for your review is the draft final safety assessment (pyrusm122015rep), the CIR report history (pyrusm122015hist), literature search strategy (pyrusm122015strat), ingredient data profile (pyrusm122015prof), minutes from the September 21-22, 2015 Panel meeting (pyrusm122015min), 2015 FDA VCRP data (pyrusm122015FDAdata), comments received from the Council (pyrusm122015pcpc1), and use concentration data received from the Council (pyrusm122015data1, pyrusm122015data 2, and pyrusm122015 data3).

The summary and discussion from the safety assessment on plant-derived fatty acid oils [pyrus malus (apple) seed oil included] were added to the current report for use in the safety evaluation of malus domestica (apple) seed oil and malus domestica (apple) oil. The Panel needs to determine which information from the summary and discussion is most useful in assessing the safety of these two ingredients, and, also, agree that pyrus malus (apple) seed oil should be removed from the report conclusion because its safety in cosmetics was previously determined. The use concentration data on pyrus malus (apple) seed oil is included in the safety assessment because there are no use concentration data on malus domestica (apple) seed oil and malus domestica (apple) oil, and these data are likely very similar to the actual use concentrations of these two ingredients in cosmetics.

Comments on the tentative report that were received from the Council during the 60-day comment period have been addressed. Two of the comments received relate to protein components in product formulations and the potential for cross- reactivity with plant constituents, and the skin bleaching function of apple root bark powder. The Panel will need to determine whether these comments warrant changes in the report discussion.

After considering the data included in this safety assessment, the Panel will need to determine whether a final report with the conclusions stated at the beginning of this memorandum should be issued.

______1620 L Street, N.W., Suite1200, Washington, DC 20036 (Main) 202-331-0651 (Fax) 202-331-0088 (Email) [email protected] (Website) www.cir-safety.org SAFETY ASSESSMENT FLOW CHART

INGREDIENT/FAMILY _____Pyrus malus (apple)-derived ingredients______

MEETING ______Dec 2015______

Public Comment CIR Expert Panel Report Status

Priority List INGREDIENT PRIORITY LIST

SLR

Mar 2015

DRAFT REPORT Draft Report June 2015 60 day public comment period

Table

Table IDA TR

IDA Notice IDA June 19, 2015

DRAFT TENTATIVE

Draft TR REPORT 60 day public comment period Sept 2015

Table

Tentative Report Issue TR Oct 4, 2015

Draft FR DRAFT FINAL REPORT

Dec 2015 60 day Public comment period

Table

Table Different Conclusion

PUBLISH Final Report Issue FR

Distributed for comment only -- do not cite or quote

CIR History of:

Pyrus malus (apple)- and Malus domestica (apple)-derived Ingredients

A scientific literature review (SLR) on the Centella Asiatica-derived ingredients was issued on March 2, 2015. Unpublished data were received during the 60-day comment period.

Draft Report, Belsito and Marks Teams/Panel: June 15-16, 2015

Comments received from the Council have been addressed and the following unpublished data were added to the draft report:

• Chemistry data on pyrus malus (apple) fruit extract • Chemistry data on pyrus malus (apple) root bark powder • Use concentration data • Method of manufacture of malus domestica (apple) fruit water • Chemistry data on pyrus malus (apple) fiber • Chemistry data on pyrus malus (apple) fruit extract • Method of manufacture and ocular/skin irritation data on pyrus malus (apple) fruit extract

Draft Report, Belsito and Marks Teams/Panel: June 15-16, 2015

Pyrus malus and Malus domestica are two genus and species names for apple. The Panel agreed that the available data are sufficient for evaluating the safety of the following 19 apple-derived ingredients in cosmetics, the ingredient source being either the fruit or seed. pyrus malus (apple) carpel powder pyrus malus (apple) pulp extract pyrus malus (apple) fiber pyrus malus (apple) seed extract pyrus malus (apple) fruit extract pyrus malus (apple) seed oil pyrus malus (apple) fruit malus domestica (apple) fiber pyrus malus (apple) fruit water malus domestica (apple) fruit extract pyrus malus (apple) juice malus domestica (apple) fruit water pyrus malus (apple) pectin extract malus domestica (apple) fruit cell culture extract pyrus malus (apple) peel extract malus domestica (apple) juice pyrus malus (apple) peel powder malus domestica (apple) seed oil pyrus malus (apple) peel wax

However, the Panel issued an insufficient data announcement on the following 9 apple-derived ingredients: pyrus malus (apple) bark extract pyrus malus (apple) stem extract pyrus malus (apple) flower extract malus domestica (apple) oil pyrus malus (apple) leaf extract malus domestica (apple) stem extract pyrus malus (apple) root bark powder malus domestica (apple) callus extract pyrus malus (apple) root extract

The data that are needed to evaluate the safety of these 9 ingredients are: • Method of manufacture and impurities • 28-day dermal toxicity study; if absorbed, genotoxicity and reproductive and developmental toxicity data may be needed • Skin irritation and sensitization data

A memorandum (citing references from the published literature) addressing some of the data needs was received from the Council, and will be included in the Wave 2 data submission.

Distributed for comment only -- do not cite or quote

Draft Tentative Report, Belsito and Marks Teams/Panel: September 21-22, 2015

Comments received from the Council have been addressed.

The Panel issued a tentative report for public comment with the conclusion that the following 25 apple-derived ingredients are safe in the present practices of use and concentration in cosmetics as described in this safety assessment, when formulated to be non-irritating and non-sensitizing.

Pyrus Malus (apple) bark extract* Pyrus Malus (apple) pulp extract* Pyrus Malus (apple) carpel powder* Pyrus Malus (apple) root bark powder* Pyrus Malus (apple) fiber* Pyrus Malus (apple) seed extract Pyrus Malus (apple) flower extract Pyrus Malus (apple) seed oil* Pyrus Malus (apple) fruit extract Malus Domestica (apple) callus extract Pyrus Malus (apple) fruit Malus Domestica (apple) fiber Pyrus Malus (apple) fruit water Malus Domestica (apple) fruit extract Pyrus Malus (apple) juice Malus Domestica (apple) fruit water Pyrus Malus (apple) leaf extract* Malus Domestica (apple) fruit cell culture extract Pyrus Malus (apple) pectin extract Malus Domestica (apple) juice Pyrus Malus (apple) peel extract* Malus Domestica (apple) oil Pyrus Malus (apple) peel powder Malus Domestica (apple) seed oil Pyrus Malus (apple) peel wax*

*Not reported to be in current use. Were ingredients in this group not in current use to be used in the future, the expectation is that they would be used in product categories and at concentrations comparable to others in this group.

The Panel concluded that the available data are insufficient for evaluating the safety of the following 3 apple-derived ingredients in cosmetic products:

Pyrus Malus (apple) root extract Pyrus Malus (apple) stem extract* Malus Domestica (apple) stem extract

The data that are needed to evaluate the safety of these 3 ingredients are:

(1) Method of manufacture and impurities (2) 28-day dermal toxicity study; if any adverse effects are noted, genotoxicity and reproductive and developmental toxicity data may be needed

Because apple-derived ingredients may be obtained from different apple cultivars, the Panel noted that the composition of ingredients derived from different cultivars should be similar to the composition of ingredients reviewed in this safety assessment. The Panel discussed the study indicating that phloridzin, a component of apple root bark powder and apple leaf extract, has a stimulatory effect on melanogenesis, but agreed that a review of ingredients for drug effects (i.e., noncosmetic effects such as skin bleaching) is not within the Panel’s purview. However, the Panel noted that an effect on cutaneous pigmentation would not be expected at the use concentrations of apple-derived ingredients in cosmetic products.

The Panel agreed that the safety test data on apple polyphenol extract and procyanidin B-2 in the report appendix should be incorporated into the body of the report. Apple polyphenol extract is actually a trade name for apple fruit extract, which is being reviewed in this safety assessment. Furthermore, apple polyphenol extract contains flavonoids, and procyanidin B-2 is a flavonoid that is present in apple fruit.

Draft Final Report, Belsito and Marks Teams/Panel: December 14-15, 2015

Comments received from the Council have been addressed. The data on of pyrus malus (apple) root extract, pyrus malus (apple) stem extract, and malus domestica (apple) stem extract requested by the Panel have not been received. Distributed for comment only -- do not cite or quote

Apple-derived Ingredients Check List for December, 2015. Analyst – Wilbur Johnson

Repeated dose

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

Pyrus Malus (Apple) X X X X Fruit Extract

Pyrus Malus (Apple) Bark Extract

Pyrus Malus (Apple) Carpel Powder

Pyrus Malus (Apple) Fiber

Pyrus Malus (Apple) Flower Extract

Pyrus Malus (Apple) Fruit

Pyrus Malus (Apple) X X Fruit Water

Pyrus Malus (Apple) Juice

Pyrus Malus (Apple) Leaf Extract

Pyrus Malus (Apple) Pectin Extract

Pyrus Malus (Apple) Peel Extract

Pyrus Malus (Apple) Peel Powder

Pyrus Malus (Apple) Peel Wax

Pyrus Malus (Apple) Pulp Extract

Pyrus Malus (Apple) Root Bark Powder

Pyrus Malus (Apple) Root Extract

Pyrus Malus (Apple) X X X X X X Seed Extract

Pyrus Malus (Apple) Seed Oil

Pyrus Malus (Apple) Stem Extract

Distributed for comment only -- do not cite or quote

Apple-derived Ingredients Check List for December, 2015. Analyst – Wilbur Johnson

Repeated dose

Malus Domestica (Apple) Fruit Extract

Malus Domestica (Apple) Fruit Water

Malus Domestica (Apple) Seed Oil

Malus Domestica (Apple) Fiber

Malus Domestica (Apple) Juice

Malus Domestica (Apple) Oil

Malus Domestica (Apple) Stem Extract

Malus Domestica (Apple) Fruit Cell Culture Extract

Malus Domestica (Apple) Callus Extract Apple Polyphenol X X X X Extract Procyanidin B-2 X X X X

Distributed for comment only -- do not cite or quote

Literature Searches on Pyrus malus-derived Ingredients (9-26-2014)

SciFinder Searches

Search Terms

Pyrus Malus (Apple) Fruit Extract Pyrus Malus (Apple) Bark Extract Pyrus Malus (Apple) Carpel Powder Pyrus Malus (Apple) Fiber Pyrus Malus (Apple) Flower Extract Pyrus Malus (Apple) Fruit Pyrus Malus (Apple) Fruit Water Pyrus Malus (Apple) Juice Pyrus Malus (Apple) Leaf Extract Pyrus Malus (Apple) Pectin Extract Pyrus Malus (Apple) Peel Extract Pyrus Malus (Apple) Peel Powder Pyrus Malus (Apple) Peel Wax Pyrus Malus (Apple) Pulp Extract Pyrus Malus (Apple) Root Bark Powder Pyrus Malus (Apple) Root Extract Pyrus Malus (Apple) Seed Extract Pyrus Malus (Apple) Seed Oil Pyrus Malus (Apple) Stem Extract

Search Updates (PubMed, 8-9-2015 and 10-26-2015)

Pyrus malus-derived ingredients above

Malus Domestica (Apple) Fruit Extract Malus Domestica (Apple) Fruit Water Malus Domestica (Apple) Seed Oil Malus Domestica (Apple) Fiber Malus Domestica (Apple) Juice Malus Domestica (Apple) Oil Malus Domestica (Apple) Stem Extract Malus Domestica (Apple) Fruit Cell Culture Extract Malus Domestica (Apple) Callus Extract Distributed for comment only -- do not cite or quote

Day 1 of the June 15-16, 2015 CIR Expert Panel Meeting – Dr. Belsito’s Team

Apple-derived Ingredients

DR. BELSITO: So we had fantastic weather. Now we move on to pyrus malus. DR. KLAASSEN: Well, we wanted to give you time to type everything. DR. BELSITO: Yes, thank you. I do appreciate that, Curt. DR. KLAASSEN: That was the only reason we did it. DR. BELSITO: Thank you. My poor typing skills. Actually you should know that I've approached the AMA to do a class action suit that individuals who don't know how to type should be considered as disabled and, therefore, doing electronic medical records is illegal, or they have to pay to give me a scribe as I have to pay for translators. Thanks for giving me that time. MS. FIUME: Good luck with that. DR. BELSITO: Yes, I know. So pyrus malus. This is apple-derived ingredients and we've been told that now there's a new name for apple. So this is the first time. There are 19 ingredients and they're skin conditioning agents, emulsions, stabilizers, viscosity increasing agents, yadda, yadda, yadda, and a bleaching agent. Hmm, interesting. And now the accepted name is malus domestica, is that right? Maybe, maybe not. DR. HELDRETH: There are a number of taxonomies that are floated around for apple. I think the way we'll just look at all of these is apple. We don't know if the Nomenclature Committee will ever actually change all of these over to one or the other. But there are only two ingredients that seem to have some overlap between the two different taxonomy schemes here, but all of these things are valid. DR. BELSITO: Bur we are looking at data on malus domestica to support this or sometimes we don't even know what we're looking at? We're just looking at -- DR. ANSELL: Well, the name change is part of INCI's review of a lot of the botanicals. So they believe that the correct name is malus domestica, but they are not deleting pyrus malus. So they can be considered equivalent. DR. BELSITO: Okay, then the root bark powder is the ingredient that's the skin bleaching agent, which we can simply deal with in the discussion by saying that's not a cosmetic agent if it bleaches the skin. But then since it's there and we don't know at what concentration it bleaches, if we were to issue safety on that we'd either have to know or say that it should be used in levels below which there would be any pigmentary effects? I don't know how to handle that, like we do for sensitization? Then the apple seed oil we've already issued. Was that part of the plant oils that we did? Yeah, so the apple seed oil is out of the picture. So after looking at this I thought the fruit extract, the fruit water, and seed extract, and seed oil, which was here in this report, if we could summarize data on the oils that we previously reviewed were safe as used. And I didn't know whether the data we had could clear the fiber, the leaf extract, the peel extract based upon manufacturing and impurity data. And I thought all the others were insufficient for manufacturing impurities composition and possible sensitization and irritation, depending upon how different they were from the fruit extract, water, and seed extract. So does that make any sense? I reviewed this several weeks ago. DR. LIEBLER: I took a little different approach. I looked at these and I figured almost everything that's in use is from the fruit. So somewhere inside of the peel, including the peel, and those are probably all going to turn out to be fine so I'm including the seeds, seed oil, and so forth. What that leaves us with is the bark, the flower, the leaf, the root bark powder, the stem, root extracts, stem extract. And I suggested that we just delete those. One possible reason for deleting those is that part of the logic for the rest of these from the fruit is that these are foods that are consumed. We don't have to worry about systemic toxicity. But people don't eat the bark, the stems, the roots and branches. DR. BELSITO: Or the seeds. DR. LIEBLER: People eat the seeds. I mean not on purpose, but they do and they're part of a fruit that's consumed all the time and plus the seed oil I think was already safe. So we're okay with the seeds. But we would arguably need tox data for all these other things because Distributed for comment only -- do not cite or quote

we can't go the food grass route with those. DR. HELDRETH: Could we alternatively have a split conclusion and have the others be insufficient. And if they're truly not in use, nobody's going to be concerned if they go to zero use nonsupportive. DR. LIEBLER: That's another strategy. I did note that the root extract had I think four uses and that was the only one of my list of possible excludables that had uses. DR. BELSITO: Well, I would rather follow Bart's lead and go insufficient and list why they're insufficient so we're done with them. And if someone wants to come back, we know exactly what we were looking for rather than just getting rid of them. DR. LIEBLER: I'm fine with that also. I just wanted to sort of separate those out and handle them differently because we can't leverage the food grass aspect of those. We may have a hard time getting any data on almost all of them. DR. ANSELL: It's also an important point of principle that it's possible to have a report with more than one family in it, but all members of a single family, any data point should be useful in supporting any of the members of that family. And so I agree, I think the fruits are all going to be -- the data's all going to be useful for looking at the various cuts of it. Bark? I don't know how bark relates to any of them. So we can generate a couple of different families within the apple of the report and see what data's available. DR. BELSITO: So then basically what we're saying is safe as used for peel, for fruit, for anything from the peel inside, and insufficient for method of manufacture, composition, or the flower, the bark, the root, and any parts that are extracted from those, stem. DR. LIEBLER: Right. DR. HELDRETH: We received composition data on the root bark powder, but it's not in the report. It's on page 42 of the PDF. We got a memo with root bark powder composition, but I didn't see root bark powder in the report. We got a memo about the composition of root bark powder. DR. LIEBLER: Actually, that's just method of manufacture. It's just -- well, it's carbon dioxide with the CO2 and then 95 to 98 percent -- DR. HELDRETH: Actually, root bark powder was not in the report anywhere. DR. BELSITO: Wasn't it? Wasn't that what caused the depigmentation? Root bark powder was used as a skin bleaching agent, didn't it say? DR. LIEBLER: Yeah, it's here. It's at the bottom. DR. HELDRETH: Oh, there it is. Sorry, I didn't see it when I looked for it. DR. LIEBLER: It's at the bottom of the left-hand column in the introduction. MR. JOHNSON: Yes, it's on page 11. DR. LIEBLER: Yes, that's it. Thank you. MS. WEINTRAUB: I wanted to mention two things that I caught during my review and that is one, there is no carcinogenicity data, and two there's no mention that I saw of pesticides. DR. BELSITO: Yeah, there was some mentioned, but that would go into our boilerplate for any plant-derived product. That should have been brought out in Centella, too. So any of the botanicals that have heavy metal and pesticide residue boilerplates. DR. LIEBLER: The fruit derived ingredients, everything from the peel inwards, would be probably fine for carcinogenicity. But the root, bark, stems, et cetera, would be insufficient for method of manufacture and impurities, but I also think there's no safety data on those things. They'd be insufficient for lots of other data we normally ask for. DR. BELSITO: Well, I think going through this we're saying they're sufficient. And then what's not sufficient is bark extract, carpal powder. I'm assuming fiber is sufficient, right, Dan? DR. LIEBLER: Yes, that's from the fruit. DR. BELSITO: Flower extract, root bark powder, root extract -- DR. LIEBLER: The leaf extract on the left-hand column, too. DR. BELSITO: -- leaf extract, thank you, and stem extract, stem extract malus domestica. And are we okay now with the fruit cell culture and the callus extract based upon what we understand from Centella asiatica? DR. LIEBLER: I expect we probably are. The callus extract is also derived Distributed for comment only -- do not cite or quote

from the fruit? It's not derived from a leaf? DR. BELSITO: Well, we don't know. DR. LIEBLER: It's from a leaf? SPEAKER: Probably from a leaf. DR. LIEBLER: Okay, so we have the fruit cell culture extract, which by the name I assume is from the fruit. And then the callus extract is probably from a leaf. DR. BELSITO: So the callus we couldn't go because we don't know. DR. LIEBLER: Yeah, okay. DR. BELSITO: So callus extract is insufficient for method of manufacture, composition, 28 day dermal, sensitization and irritation, and if absorbed, geno and repro, right, I mean the usual because we have nothing. And then this is a tentative final, is that it? So the next one would go out as a tentative final, right? MR. JOHNSON: Yes. DR. BELSITO: So it's the root, flower, leaf, and bark that are insufficient, correct? And stem. Leaf, stem, callus extract. So anything from the root, flower, leaf, callus extract, stem, and bark are insufficient. MR. JOHNSON: So Dr. Belsito, an insufficient data announcement will be issued? DR. BELSITO: Uh-huh. MR. JOHNSON: Okay. DR. BELSITO: So for method of manufacture, impurities, 28 day dermal, if absorbed, and sensitization and irritation. Anything else on this? So a split conclusion that will come back next as a tentative final, is that correct? DR. LEIBLER: And, Don, will these be formulated to be nonsensitizing? DR. BELSITO: Yes. MR. JOHNSON: Dr. Belsito, you mentioned absorption data. Due to the fact it's a botanical, would those data be expected? DR. BELSITO: No. So basically, yes, Wilbur, because we're looking at parts that aren't consumed now. That's the whole issue. So we're saying root, flower, leaf, callus extract, stem, and bark are insufficient. And we want absorption on them because we have no idea since they're not foods. So we want to know if they're absorbed and then if they're absorbed, we want to know depending upon chemical composition, we may have concerns about genotoxicity and reproductive toxicity. So it's like the full gamut. MS. FIUME: But, Dr. Belsito, normally when we have a botanical and we try and ask for absorption, we're told they really can't look at it because they don't know what to look for, what is being absorbed, because it's a botanical mixture. DR. BELSITO: Right, I understand. So if they can give us some information on what the composition is, that may not be necessary if we look and see what's in there. I mean one way or the other we need the data and if they say they can't give it to us, then it remains insufficient. Then they shouldn't be using it. Am I quoting this correctly here? DR. LIEBLER: Yep.

Distributed for comment only -- do not cite or quote

Day 1 of the June 15-16, 2015 CIR Expert Panel Meeting – Dr. Marks’ Team

Apple-derived Ingredients

DR. MARKS: Next, pyrus malus. The forbidden fruit. I don't know what Latin for "malus" is. DR. HILL: That is why I eat one every single day. (Laughter) DR. MARKS: This is the first time we have seen these forbidden apple fruit ingredients. Reading through this, there is an issue with the nomenclature, how we are going to handle nomenclature in this report, and obviously, a number of them are graphs. Tom, Ron, are the ingredients okay? If not, how do we handle the different names, like pyrus malus and then we have malus domestica. DR. EISENMANN: There are two names for the same thing. DR. MARKS: I agree, but are we going to just use one name? DR. EISENMANN: For this one, I don't know why you just don't say "apple." DR. SLAGA: That would be easier. DR. SHANK: Where are we at on the survey -- DR. SLAGA: If we go too parochial -- DR. EISENMANN: It hasn't started because I didn't know there was adding these ingredients until I read the memo. I put the two that were duplicates in the dictionary, but the two cultures, I wasn't aware they were going to be added. I haven't started the survey. I have not yet contacted the suppliers of those ingredients. DR. HILL: Beside the fact we don't have full faith in our survey, I did have a question, would that pick up the sylvestris. I don't even know if that is a different species. It seems to be another different name for certain strains of apples. DR. EISENMANN: Variety. DR. MARKS: Or cultivar. DR. HILL: The question is whether somebody was using -- we don't have a cosmetic ingredient, sylvestris, do we? DR. HELDRETH: Not yet. DR. MARKS: The nomenclature, how are we going to deal with that? DR. EISENMANN: To me, I would look at other sources and say these sources say this equals this equals this. These are all apple. How the dictionary eventually I think is going to deal with it, I think the first step is going to be to add accepted name into the definition. That is what they are working on in this first step, to define it. They are working with someone who knows all this, who is helping them to come up with what are the accepted names for all the plants. That is the first step. DR. HELDRETH: That varies, like asking two toxicologists to look at an endpoint and make a decision, you get different results. That changes from day to day. DR. MARKS: We have another one where that is going to occur later on, where we have a table with two different names. We will get to that eventually. We can do "apple," but everything has moved to the species name. Ingredients on the label are not "apple." It would be the actual species name; correct? DR. EISENMANN: It would be pyrus malus, it would be mostly pyrus malus, but there are two duplicate names, malus domestica, water and extract. DR. JOHNSON: Also, the BCRP database contains both names, malus domestica and pyrus malus. DR. MARKS: If I looked at that, what I found difficult is I looked at pyrus malus fruit extract, and that had 125 uses for the concentration of.00007, hardly toxic. Then I look at malus domestica, that has 382 uses with not a concentration of use. Is this the same stuff or not? DR. EISENMANN: I think it is. That is a question for FDA. Would somebody report they are using pyrus malus apple fruit extract, how is it put into the BCRP? Is it switched over to the current name? It is interesting that the numbers with the current name are higher. DR. DEWAN: Usually it is whoever manufactured it, that's how they enter it. Distributed for comment only -- do not cite or quote

We usually don't tend to go in and make the changes. If there is some question on the way the ingredients are listed, then we have kind of a back and forth. Usually it is the way the manufacturer wants to list it. DR. MARKS: We have irritation sensitization on the one and no tox data on the domestica. DR. HILL: Because the survey wasn't done. DR. EISENMANN: I put both in the survey, but everybody reported back under the one name. I would say those are equivalent, two names for the same thing. DR. MARKS: How are we going to reconcile the uses, one is 125. It seems like the manufacturers like domestica, since that has 382. DR. EISENMANN: Which is strange to me because I didn't get anybody reporting, but of course, there are probably a group of companies that report to the BCRP that I don't get, but it seems strange that nobody was reporting under the malus domestica name, to me. DR. DEWAN: You are correct that domestica has a higher frequency. DR. MARKS: Much higher, three times. DR. DEWAN: Yes. DR. MARKS: I'm sort of stuck on how do I reconcile that because if I look at this checklist of ingredients on the first page, the pyrus malus, not a lot there but something, and then we look at the second page and we are into malus domestica, are we going to leave this as is or are we just going to say pyrus malus and in a discussion say malus domestica, we assume, is the same as pyrus malus. DR. SLAGA: We can't use the word "assume" in the discussion. You have to sort of know. DR. EISENMANN: There are a number of references that you could cite. DR. HELDRETH: You could have a concentration of use for one and not the other, worse case scenario, the references will say the same thing. DR. MARKS: In the actual report, are we going to use pyrus malus? Is that the one we are going to use? DR. HILL: Is there an INCI dictionary entry that is malus domestica right now? DR. EISENMANN: Yes, there are four of them. DR. HILL: It would seem to me in the report title, we opt for pyrus malus and then "(malus domestica)." DR. EISENMANN: Why not just say "apple?" Later in the report, just go into it more. DR. MARKS: In the report we state pyrus malus and malus domestica is the same, and put the references that state they are the same. That takes it out of the assumed. How does that sound, Tom? DR. SLAGA: Sounds good. That would be in the introduction. DR. MARKS: Yes, I think in the introduction would be good right off the bat. It can be repeated in the discussion if you want. It is an evil fruit, isn't it? DR. SHANK: There are all kinds of apples, are those all varieties of pyrus malus? DR. MARKS: That is what she brought up with Ron Hill earlier. That is the cultivar when there are different subtypes under the species. Presumably, they are quite the same but they are not always. If you look at primula obconica, it contains an allergen which now they grow primula obconica libre, which is a cultivar and does not contain that allergen. DR. HILL: Do they grow that in Bonaire? (Laughter) DR. MARKS: No, actually in California, Goldschmidt does. They brought it specifically because when they grew obconica, the workers were getting contact dermatitis and disabled, so then they bred the libre form. It does not contain primula, which is the allergen. Ron, I hate to substantiate your concern about the different cultivars, but I don't know how to handle that to tell you the truth. We could say that with all these natural products, what is the cultivar being used, so we add not only surreal growing conditions, but then what is the cultivar within that species. I'd rather not go there, to tell the truth. (Laughter) Distributed for comment only -- do not cite or quote

DR. HILL: I think if there is something like that we know about, it is probably worth raising and putting to bed, but that is always going to be there for any botanical. That is part of our thinking. It comes back on the producers to know about these things and deal with them. DR. SLAGA: Fruit extract would be whatever apple is available. DR. EISENMANN: I think probably a lot of this is byproducts of apple juice making. DR. HILL: Sure is. DR. MARKS: How do we want to proceed with that? It's public record. I think we acknowledge there may be different cultivars, but we expect the composition to be similar. DR. HELDRETH: Why don't we say "as used." We are talking about the data we have in front of us. DR. MARKS: Correct. It is kind of interesting because that could almost become a boilerplate. DR. HILL: It's not a trivial concern because we do need to know when we are assessing safety is the material we are assessing similar to what the toxicology studies were done on. I think that is just the nature of the beast with botanicals, we have these uncertainties. DR. MARKS: We have the boilerplate that if multiple botanicals are put together, individual ingredients -- should we add a second paragraph or whatever that says we expect there are different cultivars, the composition would be similar to what we evaluated in the safety report. Is that worth bringing up as a discussion point tomorrow? The real example of primula obconica where there is a huge difference between primula obconica, the standard primula obconica and the libre cultivar, which does not contain primula or at levels that it is not sensitizing to either the workers or household. DR. SHANK: I think that is hugely important. DR. JOHNSON: I'd like to call the panel's attention to PDF page 22, Table 3, which contains composition data on different varieties of pyrus malus. DR. HILL: And then there is Table 4. What is different about 3 and 4. DR. MARKS: Minerals. Ron, you brought up this issue, did you see any significant differences reading across here of these ingredients that would be concerning? We usually don't have this, do we? DR. SHANK: There would be huge differences, but this looks like nutrition data to me. To go from almost no fat to 3.5 percent fat, fiber varies tenfold. DR. SLAGA: Most of the data is on polyphenol extracts. These are nutritional assays. DR. MARKS: I'm going to ask the question, Ron, and the rest of the team, are Table 3 and 4 helpful, should they even be included in the report? DR. SHANK: I think so because it shows there is a lot of variation in the composition between varieties, and the one you mentioned, what was it? DR. MARKS: Primula obconica. DR. SHANK: It is very important. DR. MARKS: It is certainly in this one a discussion, would you say -- how did I have that phrased? There would be similar composition with what was in the report, apples in this case or the cultivars within this. DR. SHANK: We don't have the cultivars in this report. When the food industry says here's a bottle of apple juice, is that Gravenstein apples? In California, most of those juices are Gravenstein apples. In Ohio, are they juice from Jonathans, McIntosh in Massachusetts. DR. HILL: Buy at Dollar General or this other grocery store Brazilian apples, so I have no idea -- it's a global industry. DR. SHANK: That's geographical. DR. MARKS: If I were a formulator, how would I look at this report and say okay, I'm getting an apple sourced whatever extract, leaf extract. How do I know what cultivar it came from. More importantly might be the fruit side, how do we deal with that other than saying we expect they would be similar to the ingredients in this report, the composition? Is that a good word? Sort of like "assume." Distributed for comment only -- do not cite or quote

DR. SHANK: We don't know what the composition is of the ingredients presented in the report, we have the species name, but not the variety name. Is cultivar one step below variety? DR. MARKS: No, that's the same. In my understanding botanically. Is there a botanist in the room? My understanding is "variety" and "cultivar" are synonyms. "Cultivar" is the fancier name. DR. HILL: My way of looking at this was if you look at the use concentrations, although we are missing them for a bunch of them, they are all very small, except the one that is reported for fruit water, and I assume it is actually mostly water, but they are saying the percentage. It may not matter that much from a toxicology standpoint. DR. MARKS: Grass? DR. HILL: I was going to pick on that part of it in a minute. Can we talk about we have oils in here that were reviewed in the oil report and belong in the oil report, and while I realize we might want the information for read across, why are we reviewing them again, just because they are apple seed oil, for example. Do they even belong in here with the rest of these because they are so dissimilar. DR. EISENMANN: Wax is the other one. DR. HILL: There is a wax. I didn't feel quite the same way about that one, although maybe I should have. DR. EISENMANN: You're talking about the duplicate, the pyrus malus and then the domestica. DR. HILL: The seed oil. There are three seed oils, one is just oil. I don't know if I know for sure what is the difference between oil and seed oil. The oil might need to stay in there but the seed oil, that is what was in the -- DR. JOHNSON: 244 ingredients, but there were no safety test data on the pyrus malus. DR. HILL: No, but we did all the read across based on the components of the oil and then any potential residuals. I think we might want the data in here for read across, but I wonder if we need to re-review this. We are putting the same ingredient in two different reports. I'm not sure I like that or want to see that. They are so dissimilar from the rest. DR. EISENMANN: The apple oil is something that is coming from the BCRP. I have no suppliers. DR. HILL: We don't have method of manufacture because we don't have any suppliers, I think that one ought to go out as a matter of principle. Take it out of here. DR. HELDRETH: We know it is in use. DR. EISENMANN: It's coming from the BCRP. There is one use reported to the BCRP. DR. HILL: There is no INCI name? DR. EISENMANN: Right, it's not an official INCI name. DR. HELDRETH: We have looked at ingredients before that did not have an INCI name. DR. HILL: Then it would end up being insufficient if we know nothing about what it actually is. DR. HELDRETH: If nobody is actually using it, nobody is going to care. DR. EISENMANN: Are you adding those two culture ingredients that I have not surveyed yet? DR. MARKS: I don't think we have gotten yet in terms of the final list of ingredients. DR. HILL: The sylvestris one came up in here, is that just safety data. DR. SLAGA: We had received safety test data on one of the ingredients, and I think sylvestris was listed as systematic -- DR. EISENMANN: Another name for it. DR. MARKS: Going down this list of ingredients on the checklist, are there any we should delete? We don't delete it just if it's insufficient. Is there anything that is way out of line here? This list is obviously going to be condensed because that second page where we have Distributed for comment only -- do not cite or quote

malus domestica, that's going to all disappear; is that correct? Actually, it won't disappear. We are going to be uniform in the name, the nomenclature. DR. HILL: I'm totally confused now. I thought everywhere it said "malus domestica," we would regard that as totally equivalent to pyrus malus. DR. MARKS: Yes. Are we going to have two listings like pyrus malus apple fruit extract and malus domestica apple fruit extract? DR. EISENMANN: There are only two that have two INCI names for the same thing, the water and fruit extract, if I remember. The other ones are in the BCRP but not INCI. DR. HELDRETH: If you went out and looked at labels on products -- DR. MARKS: Just leave it in and again write in the beginning of the introduction and perhaps in the discussion say this is the same. It may be worthwhile having an asterisk or double asterisk in the conclusion for these two products and saying they are the same. Again, for the reader, so that wouldn't be missed. DR. JOHNSON: One of the industry submissions on pyrus malus fruit extract, malus sylvestris was identified as the botanical name for that particular ingredient. DR. SHANK: Say that again, please. DR. JOHNSON: In one of the industry submissions, malus sylvestris was identified as a botanical name for pyrus malus apple fruit extract. DR. SHANK: Thank you. DR. MARKS: It is fruit extract and fruit water where they are identical. DR. HILL: Looking at this from the other end, for example, in the read across table, could we merge all these ones that stay the same, so the pyrus malus fruit extract, can we merge that, whatever data we may have, with the malus domestica fruit extract? DR. MARKS: I think the data can, that's easy for a read across if we say it's the same. What I was driving at was do we put both names in the conclusion. I think yes, that both are the same. DR. SLAGA: Right. DR. MARKS: One could always put a parentheses and put the other botanical name, but I think probably separate is easier or better. Of all these ingredients we have, again, small concentration of use, like the fruit extract. I had 0.00007 percent. I have a question mark. Is that correct, Carol? DR. EISENMANN: I'd have to go back and look at the original data. DR. MARKS: Did I read it correctly, that many zeros after the decimal point? DR. SHANK: Yes. DR. MARKS: Irritation and sensitization data. Where do we want to proceed? Are all the ingredients safe? Do we have needs? Carol, you mentioned two other ingredients that aren't on this list; is that right? DR. EISENMANN: They are on the list, but I haven't surveyed them. DR. MARKS: Fine. We will get that the next round. This is just the first time. DR. EISENMANN: It won't be until December. DR. MARKS: That's okay. DR. SHANK: Those that are foods, I think we do not need systemic toxicology. The question of skin sensitization, you raised an apple that caused allergic dermatitis in workers. DR. EISENMANN: That wasn't an apple. DR. MARKS: No, that wasn't an apple, that's a flower. That was just the comparison, and when you got into the cultivars, I was saying cultivars can be different. DR. HILL: I thought it was an apple. DR. MARKS: I'm sorry. I should have clarified. I did give the botanical name. It is primula obconica. That's not on this list. All the sensitization I saw in here in Wave 2 looked good. DR. SHANK: Can we cover all the varieties? DR. MARKS: I think so. Fruit water, 9 percent, seed extract, this was in Wave 2, some phototox. DR. HILL: Restricting just to food consumption, we have not gotten to like bark and leaf yet. There are eight, some of which may be duplicate, between pyrus and malus that are inhaled potentially, zero inhalation toxicology, probably doesn't matter but we don't have any. Distributed for comment only -- do not cite or quote

DR. MARKS: Can we use the boilerplate for that? DR. HILL: We have no inhalation toxicology. Where I put circles on this read across table with bark extract and leaf extract, we don't eat those. In fact, I remember growing up I was told don't eat the apple seeds, they are poisonous. I don't know if that was actually true or not. DR. SHANK: It is true. DR. HILL: There is bark extract, no toxicology, leaf extract, no toxicology. DR. EISENMANN: We did get composition of that which it surprises me why it doesn't get the name, it is 95 to 98 percent -- DR. HILL: The note I wrote here is are we really just evaluating the toxicity of thorns. DR. EISENMANN: For bark powder, from that company, yes. DR. HILL: That was my impression. That is just bark powder. I don't know about the bark extract or leaf extract or the stem extract. There is a callus extract, by the way, which I didn't know what that was exactly, but now I have a somewhat better suspicion. I wanted to see something on those. DR. MARKS: You would want it on bark? DR. HILL: What I circled was repro development and carcinogenicity or genotox, I don't know if you could do that. I didn't circle anywhere else. I guess I was thinking the read across works for everything but those. DR. MARKS: Tom? Ron? DR. SHANK: I agree, they are not foods. DR. MARKS: We need bark, anything with bark, bark extract. DR. SLAGA: Root. DR. MARKS: Root. Stems, we don't eat stems. I had tentative food ingredients and seed extracts safe, the rest insufficient. Does that summarize it? DR. SHANK: Yes. DR. SLAGA: Yes. DR. MARKS: Bark, fiber. DR. SLAGA: Fiber is within the fruit. DR. MARKS: Okay. Flower? You don't eat the flower. Fruit water, apple juice, that would be from the fruit. Leaf. Peel? We eat it. Peel is okay? DR. SHANK: Yes. DR. MARKS: Peel powder? DR. SHANK: Yes. DR. MARKS: Peel wax? DR. SHANK: Yes. DR. MARKS: Pulp extract? DR. SHANK: Yes. DR. MARKS: Root, bark, no. Root extract, no. Seed extract, okay. Seed oil, presumably read across. Would you put seed extract because of the toxicity you were talking about? DR. SHANK: We know the toxicity. You have skin sensitization. That is okay. DR. MARKS: Seed oil, okay? DR. HILL: Again, I say why is it in this report other than for the data. DR. MARKS: Have we done that before where we had two ingredients in two separate reports? DR. SHANK: Yes. DR. EISENMANN: You have but you have also taken out seed oils from other plants because you had already reviewed them in the seed oil report. I'm not sure composition of any of these other ingredients are similar. DR. MARKS: Do we want to keep the seed oil in here and duplicate it with -- DR. HILL: Yes, I think so. We have the procedure for the seed extract. The seeds not dehydrated will allow to sit in moisture for three months, and then they will extract it using 80 percent redistilled methanol with PHD added. That is it. 80 percent redistilled methanol, Distributed for comment only -- do not cite or quote

chilled, they don't say how long. Homogenizing blender, homogenates, stands overnight, no refrigerator. It doesn't say remove the oil. That is going to capture just about everything in the seeds. DR. MARKS: Seed extract, we already said okay. DR. HILL: Data. DR. MARKS: Do we want to have seed oil? DR. SHANK: We have already said that. DR. MARKS: Stem extract would be insufficient. DR. SHANK: Correct. DR. MARKS: Then we go on to the next page, malus domestica. Obviously, fruit extract. Fruit water, okay. Seed oil, okay. Fiber, okay. Juice, okay. Apple oil, okay.

DR. MARKS: Stem extract would be insufficient, fruit cell culture extract would be okay. Callus extract, presumably, is that from the fruit? SPEAKER: Yes. DR. MARKS: That's okay. Apple polyphenol extract? DR. SHANK: Okay. That one, and the procyanidin, they are not ingredients, they were added for read-across, I think. DR. HILL: Yes. DR. MARKS: Mm-hmm. DR. SHANK: Most of the data in the report are on those two, and I would suggest taking that and moving it into an appendix, as supportive material, but if you leave it in the body of the report, it's deceptive. SPEAKER: Right. DR. SHANK: And it looks like we a whole lot of data. SPEAKER: And there is. DR. SHANK: But the data isn't on -- the data are not on the ingredients. I wouldn't take it out, I'd move it into an appendix, or a separate section, or something like that. MR. JOHNSON: One question I have about this, if those data were not included in safety assessment, would the Panel have enough, you know, data to substantiate the safety of all those different ingredients that were mentioned with the exception of those that have stem bark or root, and if - - DR. SHANK: No. I think we accepted all of the others as safe, because they are food, you are right. MR. JOHNSON: Okay. DR. SHANK: And we didn't use the data from these two last entities. MR. JOHNSON: So, do they -- do they belong at all? DR. HILL: To the appendix, yes. MR. JOHNSON: They should be there? DR. MARKS: Yes. DR. HILL: They are components of the fruit. MR. JOHNSON: Yes. DR. HILL: But it makes it look like we have a whole lot more than they did, and we really do. DR. MARKS: Plus it's, I think, a little misleading to put them in a table as an ingredient when they really aren't. DR. HILL: Well, I mean, I think it's okay for the read across table, that's not going in the record report. Yes, I mean, you don't have a table in the report that has those, just a read across table just for our use. SPEAKER: Right. DR. MARKS: Oh. Okay. DR. HILL: And we do that because, for example, we have -- we list alcohols if there are esters that can be cleaved, and that sort of thing. So I think it's great to leave it there in the read across table, and you might want to separate it or something for the plant grower, but it's okay. I mean, it's for our use. DR. MARKS: I don't know that -- Is this table going in the final report? Distributed for comment only -- do not cite or quote

DR. HILL: It does not. DR. MARKS: Yes. That's what I thought, not ingredients, I'll handle in appendix, probably in the discussion too. Don't you think if we are purchasing it; or something like that? DR. HILL: Okay. That's fine. DR. MARKS: Now, so for everything else insufficient, what are our needs? Obviously we need to put that down. Since they aren't food, we need probably a 28- day dermal tox, we need -- DR. HILL: Method, impurities. DR. MARKS: Yes. Method of manufacture. DR. SHANK: Genotox. DR. SLAGA: Genotox. DR. MARKS: So you need everything, because it's nothing left there. DR. HILL: Basically. Skin sensitization. DR. MARKS: Yes. Need all -- I'm going to put need all tox. DR. HILL: Yes. DR. MARKS: Method of manufacture, reproductive, you wrote up that earlier; genotox, irritation sensitization. Okay. Good. So, let me see, tomorrow I will be making a motion; and that motion will be tentative report. DR. HILL: Can we add to that, clarification of what a pectin extract is. I guess that's the method of manufacture, even though we are saying it's safe? DR. MARKS: Hmm? DR. SLAGA: It's the extract? DR. HILL: I mean it's -- pectin, what would you extract? DR. SLAGA: To extract all pectins, isn't it? DR. HILL: I don't know. That's my point, and I can't get it -- DR. MARKS: Well can it -- I'm sure that they all are, and can you -- DR. HILL: They should be, sure. DR. MARKS: Would you go ahead and clarify that for we'll be seeing this report back, obviously. So, move that a tentative report be issued with the conclusion of fruit and seed ingredients, and there are a number of them there. I won't -- in went down the list here. Wilbur, you'll have it. MR. JOHNSON: Sure. DR. MARKS: I can repeat that tomorrow if people want it, but are safe, the rest are insufficient. We need all the toxicologic data on the non-fruit and non-seed ingredients. In the discussion we are going to handle the cultivars, and have something to the effect that they are -- we expect the cultivars of apple to be similar what was recorded in here, and that we are -- the apple was a -- the apple, polyphenol extract, and -- DR. MARKS: Hallusin. MR. JOHNSON: Cyanidin. DR. MARKS: Cyanidids are not ingredients; they'll be handled in an appendix in discussion. Does that sound good, team? DR. SHANK: It does. DR. HILL: They don't have to -- that don't have to be listed in needs, but there are also production method, general things. Could we get clarification on how the flower water is prepared because that description is not complete, and I assume I know how it's prepared, but it's not complete. And I had a question about peel extract, if they do that extraction at room temperature, or elevated temperature, and if it's at a distillation extraction, or simple soak and heat -- soak and filter, it might matter. DR. EISENMANN: It's probably not a distillation. Usually the distillations are -- when the distillation is broader, and I don't understand -- DR. HILL: Well, I mean, are they doing a steam distillation extractive of the fruit? I mean, that would be a reasonable way they might do it, and there may be multiple ways that they do it, but we've got information, and it didn't give much. I have reason for asking but I'll keep it for the moment. DR. MARKS: Okay. Any other comments? If not we'll close for today, apple. And it's -- let me save this. Distributed for comment only -- do not cite or quote

Day 2 of the June 15-16, 2015 CIR Expert Panel Meeting – Full Panel

Apple-derived Ingredients

DR. MARKS: As one of my team members says, the forbidden fruit. So, we had quite a discussion about Apple. It's the first time we've seen these 18 ingredients. There was a discussion about the nomenclature, and we were reassured that pyres malus and malus domestica are the same, particularly with relevance to the fruit extract and the fruit water, which in the table that we received are listed twice under those different species' names. We felt we could move a tentative report with the fruit and seed ingredients, and there are a number of ingredients there, if you want me to read those separately, I can, which are safe as used. The remainder ingredients from the bark leaf root, carpal powder, stem flour were insufficient, and we needed all the toxicologic data for them to see what we would find. And then, we felt -- we had some discussion about how do we handle cultivars, and it came up in Apple, but it also really is relevant to other plants. We felt there perhaps, should be a boilerplate, or at least in the discussion, that cultivars should have similar ingredients to this report, and I brought up the example of that flour called a conic -- geez, now I'm -- primula conica, which there is a cultivar which is free of the sensitizer, and they're called primum. So, you could have a cultivar with a sensitizer and another cultivar without the sensitizer. So, what we're considering in this report, we would expect in the future. Referring back, it would be the similar cultivars to what was in this report. Apple polyphenol extract and pyracyanidenen B2 is not a cosmetic ingredient, so we wanted to handle those in an appendix and discussion. So, repeat that motion that a tentative report be issued with fruit and seed ingredients, say the rest, insufficient. DR. LIEBLER: Let me follow that. So, we had root, flour, leaf, callus extracts, sound stem and bark as insufficient? DR. MARKS: Yeah. DR. LIEBLER: So peel, fruits -- eat everything else, is sufficient? DR. MARKS: Yes. DR. LIEBLER: And what we wanted was method of manufacturer impurities, 28 day dermal tox, and if absorbed, then geno and repro and sensitization and irritation on all the components that we're going insufficient for. DR. MARKS: Correct. DR. LIEBLER: Is that what you're -- okay. DR. MARKS: Yes. DR. LIEBLER: Second. DR. BERGFELD: You're a second? Any further discussion regarding the apple document and the needs? (Simultaneous discussion) DR. BERGFELD: Dan? DR. LIEBLER: Just to follow up and clarify Jim's suggestion on a boilerplate with respect to cultivars -- SPEAKER: Yeah, it's -- DR. LIEBLER: So, do you think this could be simply an edit to the botanicals boilerplate? DR. MARKS: Yes. DR. LIEBLER: Because I think that's what could be done most efficiently. DR. MARKS: Yeah. And we hadn't -- that hadn't been brought up. Actually, Ron Shank brought that up, and when I thought about it, it really makes sense, because at least my interpretation of cultivar, it's the same species, but it's a different variety within the species. DR. LIEBLER: Right. DR. MARKS: And then, when I think of primula conica, there's a significant difference between the libra cultivar and the non libra. And the libra was actually made to avoid contact dermatitis in the workers who grew these plants, because they had endemics of contact dermatitis to them. Distributed for comment only -- do not cite or quote

DR. LIEBLER: So, you could envision, and perhaps a two sentence modification to the botanicals' boilerplate; one that points out this cultivar issue. DR. MARKS: Yes. DR. LIEBLER: And the other could be a variant on -- encourage to formulate to be non (Inaudible) -- select cultivars for products in order to have a product that's non sensitizing. DR. MARKS: Right. DR. BERGFELD: Any further discussion? (No response heard) DR. BERGFELD: So, we have a tentative final. (Simultaneous discussion) DR. BERGFELD: Or an insufficient? DR. MARKS: No, no. This is the first time. DR. BERGFELD: Yeah. DR. MARKS: So, this would be a tentative. DR. BERGFELD: Tentative what? DR. MARKS: Tentative report. Yeah, tentative final. DR. BERGFELD: Final. SPEAKER: Right. DR. BERGFELD: With several insufficiencies? DR. MARKS: Correct. SPEAKER: Yeah. DR. MARKS: The fruit and seed are safe. The rest are insufficient. (Discussion off the record) DR. ANSELL: Do you think it would be an insufficient data announcement, since this is the first time? (Discussion off the record) DR. BERGFELD: So, you're changing it to insufficient data announcement? DR. MARKS: Yes. DR. BELSITO: I don't think we make those decisions (Laughter). DR. BERGFELD: Okay. Well, we're going to call the question. I think that the two teams are in agreement. Do you think that you've captured the information and the needs and those that are insufficient? DR. GILL: Yes. DR. BERGFELD: Thank you. I'm going to call the question. All those in favor of this conclusion? Thank you. Unanimous. (Motion passed unanimously) DR. BERGFELD: The conclusion has some safety parts and some insufficient parts. Okay.

Distributed for comment only -- do not cite or quote

Safety Assessment of Apple-derived Ingredients as Used in Cosmetics

Status: Draft Final Report for Panel Review Release Date: November 20, 2015 Panel Date: December 14-15, 2015

Distributed for comment only -- do not cite or quote

Abstract: The Cosmetic Ingredient Review (CIR) Expert Panel (Panel) reviewed the safety of 28 apple-derived ingredients, which function mostly as skin conditioning agents in cosmetic products. Because final product formulations may contain multiple botanicals, each containing similar constituents of concern, formulators are advised to be aware of these constituents and to avoid reaching levels that may be hazardous to consumers. Industry should use good manufacturing practices to limit impurities that could be present in botanical ingredients. The Panel reviewed relevant data relating to the safety of these ingredients, and concluded that 25 ingredients (derived from apple fruit, leaves, and flowers) are safe in the present practices of use and concentration in cosmetics as described in this safety assessment, when formulated to be non-irritating and non- sensitizing; the available data are insufficient for determining the safety of pyrus malus (apple) root extract, pyrus malus (apple) stem extract, and malus domestica (apple) stem extract.

INTRODUCTION

The safety of the following 19 Pyrus malus (apple)-derived ingredients and 9 Malus domestica (apple)-derived ingredients as used in cosmetics is reviewed in this safety assessment:

• Pyrus Malus (Apple) Fruit Extract • Pyrus Malus (Apple) Root Extract • Pyrus Malus (Apple) Bark Extract • Pyrus Malus (Apple) Seed Extract • Pyrus Malus (Apple) Carpel Powder • Pyrus Malus (Apple) Seed Oil • Pyrus Malus (Apple) Fiber • Pyrus Malus (Apple) Stem Extract • Pyrus Malus (Apple) Flower Extract • Malus Domestica (Apple) Fruit Extract • Pyrus Malus (Apple) Fruit • Malus Domestica (Apple) Fruit Water • Pyrus Malus (Apple) Fruit Water • Malus Domestica (Apple) Seed Oil • Pyrus Malus (Apple) Juice • Malus Domestica (Apple) Fiber • Pyrus Malus (Apple) Leaf Extract • Malus Domestica (Apple) Juice • Pyrus Malus (Apple) Pectin Extract • Malus Domestica (Apple) Oil • Pyrus Malus (Apple) Peel Extract • Malus Domestica (Apple) Stem Extract • Pyrus Malus (Apple) Peel Powder • Malus Domestica (Apple) Fruit Cell Culture • Pyrus Malus (Apple) Peel Wax Extract • Pyrus Malus (Apple) Pulp Extract • Malus Domestica (Apple) Callus Extract • Pyrus Malus (Apple) Root Bark Powder

The definitions of these ingredients are included in the International Cosmetic Ingredient Dictionary and Handbook (Dictionary), except for the following five:1 malus domestica (apple) seed oil, malus domestica (apple) fiber, malus domestica (apple) juice, malus domestica (apple) oil, and malus domestica (apple) stem extract. However, uses of these 5 ingredients in cosmetic products are reported in the Food and Drug Administration’s (FDA’s) Voluntary Cosmetic Registration Program (VCRP) database.2

The following functions of Pyrus malus (apple)-derived ingredients and Malus domestica-derived ingredients in cosmetic products are reported in the Dictionary:1 skin conditioning agents, binders, emulsion stabilizers, viscosity increasing agents, astringents, fragrance ingredients, antioxidants, exfoliants, and skin bleaching agents. Pyrus malus (apple) root bark powder is the only ingredient in this group that is reported to function as a skin bleaching agent (skin bleaching is not regarded as a cosmetic use in the U.S.,3 and the Panel will not evaluate safety for that use).

As stated above, Pyrus malus and Malus domestica are genus and species names for apple that appear within the 1 names of apple-derived ingredients that are listed in the Dictionary. Though most of the ingredients reviewed in this safety assessment are identified as Pyrus malus (apple)-derived ingredients, CIR has been informed by the cosmetics industry that Pyrus malus is a genus and species name for apple that is not in current use, and that the Pyrus malus-derived ingredients listed above are under consideration by the cosmetics industry for new name assignments.4 Regarding additional names for apple, Malus domestica and Malus sylvestris are listed as other names for Pyrus malus in the Germplasm Resources Information Network online database.5 The name Malus sylvestris is identified as the botanical name for pyrus malus (apple) fruit extract in data on trade name mixtures provided by industry and included in this safety assessment.6,7 Furthermore, the International Cosmetic Ingredient Nomenclature Committee (INCI), sponsored by the Personal Care Products Council (Council), has determined that the accepted scientific name for apple is Malus pumila.

Some of the ingredients (e.g., fruit/fruit-derived) reviewed in this safety assessment may be consumed as food, and daily exposure from food use would result in much larger systemic exposures than those from use in cosmetic products. The Distributed for comment only -- do not cite or quote

primary focus of the safety assessment of these ingredients as used in cosmetics is on the potential for local effects from topical exposure.

The Panel has evaluated the safety of pyrus malus (apple) seed oil and other plant-derived fatty acid oils in cosmetics, and issued a final report with the conclusion that these oils are safe in the present practices of use and concentration.8 The summary and discussion from this safety assessment are included in the current safety assessment, for consideration in the safety evaluation of malus domestica (apple) seed oil and malus domestica (apple) oil.

In the current safety assessment, if a substance tested in a study is not clearly a cosmetic ingredient, because of the absence of information on the genus and species from which the substance was derived and/or the method of extraction used, the test substance will be referred to by a common name (e.g., apple, apple juice, apple fruit extract, apple seed extract, or pyrus malus).

Data on procyanidin B-2 (epicatechin-(4β→8)-epicatechin, found in apple fruit)9 are also included for use in this safety assessment. Procyanidins are members of the procyanidin or condensed tannins class of flavonoids.10 Procyanidin B- 2 is not a cosmetic ingredient.

Data on skin depigmentation effects are included in this safety assessment; however, a review of ingredients for drug effects (i.e., noncosmetic effects) is not within the Panel’s purview.

CHEMISTRY

Definition and Structure Pyrus Malus

The definitions and functions of apple-derived cosmetic ingredients reviewed in this safety assessment are presented in Table 1.1 The ingredients in this report are related by source, as each is a derivative of apple. While the identity and concentrations of ingredient components may vary from plant part to plant part, and from extract method to extract method, those component identities and concentrations cannot be known for these industry-specific ingredients until such information is provided as outlined in the industry’s botanical framework. Differences in those components do not neccessitate the regrouping of such ingredients, but instead warrant a comparison/contrast effort as to how those differences affect safety. Those differences are likely to be informative.

Chemical and Physical Properties

Pyrus Malus (Apple) Fruit Extract

Properties of 2 trade name mixtures containing 10-25% pyrus malus (apple) fruit extract are presented in Table 2.6,7 Composition data on both mixtures are provided in the section on Composition/Impurities – Pyrus Malus (Apple) Fruit Extract.

Method of Manufacture

Pyrus Malus (Apple) Fiber

Information provided by the cosmetics industry indicates that pyrus malus (apple) fiber is made from residue formed during apple juice production. The residue is dried and mechanically ground and sieved.11

Pyrus Malus (Apple) Fruit Extract

Pyrus malus fruit extract has been prepared according to the following procedure:12 Pyrus malus fruit was peeled manually and the pericarp was separated, dried at room temperature (26-28°C), and powdered. The powder was extracted Distributed for comment only -- do not cite or quote

with 95% ethanol. The ethanol was evaporated using a rotary evaporator, and the fruit extract was stored at -20°C prior to use.

According to information provided by the cosmetics industry, pyrus malus (apple) fruit extract is made by extracting apples with a 50% vol% 1,3-butylene glycol solution (the extraction solution is 50% butylene glycol and 50% water).13 The extract is then treated as follows: filtration, sedimentation, filtration, adjustment of the concentration with additional 50% butylene glycol and 50% water solution, and packaging.

The method of manufacture of a trade name material containing 10%-25% pyrus malus (apple) fruit extract (provided by the cosmetics industry) has been described as follows:6 The plant material is extracted with 1,2-propylene glycol at “considerate” temperatures during a fixed time, and is sterile-filtered at the end of the fabrication. Lactic acid (0.1- 1%) is used for pH regulation, and the product contains 0.6% Bactiphen 250G (phenoxyethanol [75%-100%], methylparaben [10-25%], ethylparaben [1%-5%], propylparaben [1%-5%], and butylparaben [1%-5%]) as the preservative. Information provided by the cosmetics industry indicates that another trade name material containing 10%-25% pyrus malus (apple) fruit extract is manufactured using the same procedure, except that glycerin (vegetable origin) is the extraction solvent. Again, lactic acid (0.1%-1%) is used for pH regulation, but the product contains potassium sorbate (0.35%) and sodium benzoate (0.35%) as preservatives.

According to information provided by the cosmetics industry, the process of manufacturing a trade name material containing 20% pyrus malus (apple) fruit extract begins with the mechanical grinding/milling of pyrus malus fruit.14 This is followed by extraction in butylene glycol (at specific pH and temperature), the addition of phenoxyethanol, and filtration. Another tradename material containing 20% pyrus malus (apple) fruit extract is manufactured differently.15 Processing (mechanical grinding and milling), is followed by extraction in propylene glycol (at specific pH and temperature) and, then, filtration.

The method of manufacture of a product described as an aqueous solution containing 20% pyrus malus (apple) fruit extract is:16 (1) solubilization of apple powder in water; (2) separation of soluble and insoluble phases by filtration; and (3) sterilizing filtration.

A product containing pyrus malus (apple) fruit extract (18.5%), water (81.445%), and potassium sorbate (0.055%) is manufactured as follows:16 (1) solubilization of fruit of Pyrus malus in water; (2) enzymatic hydrolysis, separation of soluble and insoluble phases; (3) inactivation by thermic treatment; (4) purification of soluble phase; and (5) concentration of soluble phase and membrane sterilization.

The method of manufacture of a pyrus malus (apple) fruit extract trade name material (identified as apple polyphenol extract) has been described as follows:17 Unripe apples were crushed and pressed while 10% sodium metabisulfite solution was added. Pectolytic enzyme was used to clarify the juice obtained, and the mixture was centrifuged and/or filtered with diatomaceous earth. The clarified juice was passed through a column with aromatic synthetic adsorbents, after which the column was washed with distilled water to to remove sugars and organic acids. Apple polyphenol extract was eluted with approximately 50% ethanol and concentrated using an evaporator. The concentrated fraction was dried (using a spray drier) to obtain the apple polyphenol extract as a brown powder. Toxicity data on this material (apple polyphenol extract [brown powder]) are summarized in the Toxicology section of this safety assessment.

Malus Domestica (Apple) Fruit Water

Malus domestica (apple) fruit water is prepared by dehydrating the fruit under vacuum at low temperature.18

Apple Juice (genus and species of apple not stated)

Apple juice (from Fuji and Gala varieties; genus and species not stated) was obtained by mechanical pressure, depectinized in a water bath (2 h at 45˚C), filtered through paper, bottled, closed, and stabilized by freezing.19

Pyrus Malus (Apple) Pectin Extract

A simple procedure for the extraction of pectin has been described as follows:20 Approximately 40 g of fruit were washed with normal saline (0.90% w/v sodioum chloride in water)], after which the fruit sample was crushed and homogenized in a blender. The resulting materials were allowed to dry in a hot air (60°C) oven for approximately 2 h until the pectin extracted turned into powder. To obtain sterilized pectin, the powdered pectin was sterilized with N-saline.

Distributed for comment only -- do not cite or quote

Apple Peel Extract

Prior to analysis for anthocyanin content, apples (Malus domestica) were peeled and the peel was extracted with methanol containing 0.1% hydrochloric acid.21

Pyrus Malus (Apple) Root Bark Powder

According to information provided by the cosmetics industry, pyrus malus (apple) root bark powder is extracted with carbon dioxide, and is then purified.22 Another source indicates that the air-dried, powdered bark of Pyrus malus is extracted with petroleum, chloroform, or 90% ethanol.23 The petroleum extract yielded a faintly yellow crystalline mass, the chloroform extract yielded a dark yellow solid, and the ethanolic fraction provided a very small amount of a brown alkali- soluble residue.

Pyrus Malus (Apple) Seed Extract

Pyrus malus seed extract has been prepared according to the following procedure:24 After 3 months of stratification (steady exposure to moist environment before germination becomes possible), the seeds (not dehydrated) were extracted. The extraction procedure was performed using chilled 80% redistilled methanol, to which butylated hydroxytoluene (BHT, 10 mg) was added as an antioxidant. Extraction was followed by homogenization in a chilled blender for 10 minutes. The homogenate was allowed to stand overnight (at 4°C) in the dark, filtered, and the residue was reextracted. The filtrate was evaporated (at 35°C) in a rotary evaporator, avoiding direct sunlight, and an aqueous extract remained.

The method of manufacture of pyrus malus (apple) seed extract that is being marketed to the cosmetics industry has been described as follows:25 Ground apple seeds are extracted with alcohol (95% non-denatured alcohol). The solvent is evaporated to remove any residual alcohol. The residue is a paste that is sold in solution of various solvents.

Pyrus Malus (Apple) Seed Oil

Crude oil from the ground seeds (~ 5 g) has been extracted over a 6 h period using n-hexane.26 The solvent was removed using a rotary evaporator under reduced pressure, and the oil was flushed with a stream of nitrogen and stored at - 20°C in sealed tubes.

Composition/Impurities

The composition of various parts of the apple tree will vary based on where the tree is grown, the maturity of the tree, and storage conditions for the parts of the tree that are used to produce cosmetic ingredients. The method of extraction will also affect the composition of ingredients derived from the parts of the apple tree.

Pyrus malus

The following sugars were detected in the nectar of Pyrus malus using gas chromatography:27 fructose, glucose, and sucrose. Quantitative data were not reported.

Pyrus Malus (Apple) Bark Extract

According to information provided by the cosmetics industry, pyrus malus (apple) root bark powder extracted with carbon dioxide is purified to contain 95% to 98% phloridzin, a dihydrochalcone.22 According to another source, the following 3 compounds were isolated from the bark of Pyrus malus extracted with different solvents (petroleum, chloroform, or 90% ethanol): β-sitoserol, friedelin, and epi-friedelinol.23

Malus Domestica (Apple) Flower Extract

The extract of flowers from the redgold apple cultivar was analyzed using gas chromatography (GC) and GC-MS (mass spectrometry), and the following compounds identified were classified as making a major contribution to the odor of apple flowers:28 cis-hex-3-en-1-ol benzyl alcohol 2-phenylethyl alcohol cinnamyl alcohol Distributed for comment only -- do not cite or quote

nonanal octanol indole

Pyrus Malus (Apple) Fruit

Composition data on different varieties of Pyrus malus are included in Tables 3 and 4.29

A number of sweet and sour varieties of apples from the hilly parts of Northwest India have been found to contain malvidin monoglycoside.30 Additionally, the skin of Grimes Golden, Jonathon, and Stayman wine sap apples have been reported to contain idaein (3-β-galactosidyl cyanidin), and the related yellow varieties yielded quercetin-3-galactoside.

The following 3 predominant hydroxycinnamic derivatives in the apple (Pyrus malus L., var. Calville blanc) have been reported:31 p-coumarylquinic acid, p-coumarylglucose, and chlorogenic acid. The concentration of each derivative, some weeks after blossoming, is highest in young fruits. The amount per fruit increases during approximately 1 month and 2 months for p-coumarylquinic acid and chlorogenic acid, respectively. During this period, a relative accumulation of the o- diphenolic compound is observed, in comparison with p-coumaric derivatives.

Assay results for the presence of amino acids in Pyryus malus were as follows:32 alanine, γ-amino butyric acid, asparagine, cysteine or cystine, and glutamic acid. Quantitative data were not reported. The presence of an unidentified substance that reacted with ninhydrin was also reported.

Pyrus Malus (Apple) Fiber

Composition data on pyrus malus (apple) fiber provided by the cosmetics industry are included in Table 5.33,34

Malus Domestica (Apple) Fruit and Malus Domestica (Apple) Peel

For 20 apple cultivars, the concentrations of 5 classes of polyphenols were significantly different, across cultivar, for both the peel and flesh.35 The total polyphenol concentration ranged from 0.9 µg/g wwb in the flesh of Newtown Pippin to 453 µg/g wwb in the peel of Red Delicious. Harrison, Granny Smith, Rome, Winesap, and Black Twig cultivars contained the highest concentration of total flavan-3-ols in flesh.

Apple Fruit (genus and species not stated)

Data on the average content of phenolic compounds in fruit from apple trees (genus and species not stated; semi- dwarf and super-dwarf rootstocks) grown in Lithuania are as follows:36 Whether or not the fruit was ripe was not stated.

• Chlorogenic acid ( 729-1047 µg/g dry weight) • Phloridzin (83.7-122 µg/g dry weight) • Procyanidin B1 (33.5-81.5 µg/g dry weight) • Procyanidin B2 (504-920 µg/g dry weight) • Σ (Σ = total) Procyanidins (558-1001 µg/g dry weight) • (+)-Catechin (35.6-77.3 µg/g dry weight) • (-)-Epicatechin (217-329 µg/g dry weight) • Σ Catechins (254-406 µg/g dry weight) • Hyperoside (87-147 µg/g dry weight • Isoquercitrin (15.4-23.3 µg/g dry weight) • Rutin (15.1-21.6 µg/g dry weight) • Avicularin (59.9-94 µg/g dry weight) • Quercitrin (86.1-130 µg/g dry weight) • Σ Quercetin glycosides (264-416 µg/g dry weight) • Σ Phenolic compounds (1976-2943 µg/g dry weight)

Super-dwarf rootstocks had the highest content of all phenolic compounds tested, and semi-dwarf rootstocks had the lower content of all phenolic compounds tested. The content of (+)-catechin, procyanidin B1, and total procyanidins in apple fruits depended on the rootstock genotype. The rootstock genotype had a lesser effect on the content of quercitrin, (-)-epicatechin, Distributed for comment only -- do not cite or quote

total catechins, phloridzin, and chlorogenic acid. It was noted that conditions during the growing season, yield, and fruit weight had an impact on the content of phenolic compounds.

Pyrus Malus (Apple) Fruit Extract

Using paper chromatography, the extracts (extractant not stated) of fruits of Pyrus malus have been found to contain chlorogenic and isochlorogenic acids.23 Some glycosides of quercetin were also detected. The total polyphenols (~ 200 mg/100 g dry weight) and total flavonoids (~ 25 mg/100 g dry weight) content of pyrus malus fruit extract have also been reported.12 According one of the suppliers of pyrus malus (apple) fruit extract for use in cosmetic products, this ingredient contains sugar and organic acids.13

According to information provided by the cosmetics industry, Malus sylvestris has been identified as the botanical name for pyrus malus (apple) fruit extract in 2 pyrus malus (apple) fruit extract trade name mixtures containing 10%-25% pyrus malus (apple) fruit extract.6,7 Composition data on Malus sylvestris include (quantitative data not reported):

• Quercetin • Arabane • Mineral substances • Galactane • Essential oil • Tannins • Enzymes • Pectin • Fruit acids • Sugars • Amino acids • Wax • Vitamins

One trade name mixture contains pyrus malus (apple) fruit extract (10%-25%) and propylene glycol (75%-100%),6 and the other contains pyrus malus (apple) fruit extract (10%-25%), glycerin (75%-100%), and aqua (water) (10%-25%).7

The composition (polyphenol profiles) of a pyrus malus (apple) fruit extract trade name material (identified as apple polyphenol extract) has been described as follows:17 procyanidins (63.8%), which comprised 11.1% dimers, 12.3% trimers, 8.7% tetramers, 5.9% pentamers, 4.9% hexaers, and 20.9% other polymers. It also contained 12.4% flavan-3-ols (monomers), 6.5% other flavonoids, 10.8% non-flavonoids. In addition to the polyphenols, apple polyphenol extract also contained 1.8% moisture, 2.1% protein, and 0.4% ash.

Pyrus Malus (Apple) Juice

Juice from the Fuji and Gala apple varieties (genus and species not stated) in Brazil has the following composition:19 malic acid (0.18 to 0.389 g/100 mL), total reducing sugar (8.65 to 15.18 g/100 mL), and total phenolic compounds (100 to 400 mg/L). In comparison, apple juice from the Golden delicious variety contains: malic acid (0.312 g/100 mL), total reducing sugar (10.533 g/100 mL), and total phenolic compounds (535.082 mg/L).

Pyrus Malus (Apple) Leaf and Pyrus Malus (Apple) Leaf Extract

The partition of nitrogen in various parts of Pyrus malus throughout a one-year cycle has been studied.37 The study involved seedling apple trees that received heavy applications of sodium nitrate at regular intervals throughout the vegetative period. Positive tests for nitrates (or nitrites) were reported for one tissue only, the leaf buds, just as they were opening. Flavone glycosides have also been detected in the leaves of Pyrus malus.23 The sugar alcohol sorbitol has been detected in Pyrus malus leaves at a concentration of 0.45%.38

Although data on the composition of the cosmetic ingredient, pyrus malus leaf extract were not found, composition data on the volatile oil obtained from fresh leaves of the Malus domestica tree are available. This oil is a complex mixture of mono-, sesqui-, and di-terpenes, phenolics, and aliphatic hydrocarbons, and has been classified as cytotoxic to animal and human cancer cell lines.39 The major compounds of this oil have been characterized as: eucalyptol, (43.7%), phytol (11.5%), α-farnesene (9.6%), and pentacosane (7.6%).

Malus Domestica (Apple) Leaf Extract

A study relating to the composition and content of phenolic compounds in ethanol extracts of apple (Malus domestica) leaves was performed.40 Phloridzin (a phloretin 2'-glucoside) was a predominant component in the ethanol extracts of apple leaves from all of the cultivars that were analyzed by high-performance liquid chromatography (HPLC). Additionally, the following quercetin glycosides were identified in these extracts:

Distributed for comment only -- do not cite or quote

hyperoside isoquercitrin avicularin rutin quercitrin (major compound among the quercetin glycosides identified)

Using an ultra-performance liquid chromatography-diode array detection (UPLC-DAD) quantification method, the following 4 compounds were identified in apple leaves of two different varieties, golden and royal:41 rutin 3-hydroxyphloridzin phloridzin quercetin-3-O-arabinoside

Pyrus Malus (Apple) Pectin Extract

In the absence of data on the extract, it should be noted that pectin is a complex mixture of polysaccharides that comprises approximately one third of the cell wall dry substance of higher plants.20 The highest concentrations of pectin are found in the middle lamella of the cell wall, with concentrations gradually decreasing from the primary wall toward the plasma membrane.

Malus Domestica (Apple) Peel Extract

The total phenolic and anthocyanin content of apple peels from 6 apple (Malus domestica) cultivars grown in southern Brazil was studied.21 Total phenolic content varied from 105.4 to 269.7 mg gallic acid equivalents (GAE)/100 g of fresh mass (FM). The differences in phenolic content among the apple cultivars were statistically significant (p < 0.05). Similarly, the differences in anthocyanin composition (in apple peels) among the apple cultivars were statistically significant, and values ranged from 4.79 to 41.96 mg cyanidin-3-galactoside (cy-3-gal)/100 g of FM. Cy-3-gal is the major anthocyanin that is present in red or partially red genotype apples.

In another study, variations in the content of phenolics, antioxidant activity, and minerals in the peel and pulp of 5 apple (Malus domestica Borkh.) cultivars from Pakistan were studied.42 The mean extract yield of antioxidant components obtained with 80:20 methanol-water (v/v) was determined to be 22.1 g/100 g for the peel and 14.2 g/100 g for pulp on a dry weight (DW) basis. Ranges of total phenolics (1,907.5-2,587.9 mg GAE/100 g DW) and total flavonoids (1,214.3-1,816.4 mg catechin equivalent (CE)/100 g DW) have been reported for the peel of different cultivars of apple. Similarly, ranges of total phenolics (1,185.2-1,475.5 mg GAE/100 g DW) and total flavonoids (711.8-999.3 mg CE/100 g DW) have been reported for the pulp of different cultivars of apple. An analysis for minerals content was also performed. In both the peel and pulp, potassium (K)-containing minerals content was highest, followed by minerals containing Mg, Ca, Fe, Na, and Zn.

Pyrus Malus (Apple) Seed Extract

Endogenous levels of both cis and trans isomers of free and bound abscisic acid were studied in dormant and after- ripened Pyrus malus embryos.24 In bioassays, the level of free abscisic acid was very high in dormant embryos, mainly in cotyledons (4800 ng), but became very low after 3 months of stratification. A considerable increase in bound abscisic acid (1450 ng) was noted, and bound abscisic acid became dormant in after-ripened embryos. Bound trans-abscisic acid was detected only in the cotyledons (100 pg) of after-ripened embryos.

The extracts prepared from dry, dormant Pyrus malus L. seed (120 g) were found to contain gibberellins (GAs, 43 which are growth promoting hormones). These extracts contained GA4 (3.080 mg total; 26 µg/g dry seed) and GA7 (0.960 mg total; 8 µg/g dry seed). The ratio of GA4 to GA7 was 76:24. The ratios for seeds stratified for 35 days and 90 days were 81:19 and 64:36, respectively. Another study was performed to identify free GAs in dormant embryos of Pyrus malus L. cV Golden delicious using different extraction procedures.44 Using an ethanolic extraction procedure, minute quantities of free gibberellins (traces to 50 pg/embryo) were detected. Extraction with Tris buffer (pH 7.2) yielded slightly higher quantities of gibberellins (traces to 134 pg). Very large amounts of gibberellins, especially GA1 and GA4 (560 and 1560 pg/embryo, respectively), were detected when the embryos were crushed in Tris buffer and treated with Triton X 100.

Residual level of pesticides in pyrus malus (apple) seed extract is controlled to comply with European food limits.25

Distributed for comment only -- do not cite or quote

Apple Seed Extract

Royal Gala apple seeds (genus and species not stated) were collected from apple pomace.45 The methylated hexane extract of the seeds consisted mainly of fatty acids (80.9%) in its volatile fraction and benzaldehyde (< 0.2%) was also detected. Of the fatty acids identified, linoleic acid content was highest (51.2%), followed by palmitic acid (10.5%), linolenic acid (5.6%), stearic acid (4.3%), and oleic acid (4.1%). Fatty acid composition data are included in Table 6.

Further extraction of the seed with 70% aqueous acetone yielded the following 2 major compounds: [(6-O-β-D- glucopyranosyl)oxy]benzeneacetonitrile (amygdalin) and phloretin-2'-β-D-glucopyraide (phloridzin). The minor polyphenols identified were: chlorogenic acid, p-coumarylquinic acid, 3-hydroxyphloridzin, phloretin-2'-xyloglucoside, and quercetin glycosides.45

Pyrus Malus (Apple) Seed Oil

Data on the composition of pyrus malus seed oil are included in Table 7.26 The fatty acid profile of the seed oil was based on gas chromatography-mass spectrometry (GC-MS) analysis. GC-MS analysis of the unsaponifiable fraction of the seed oil for bioactive constituents, such as sterols, tocopherols, hydrocarbons and other components, was also performed.

USE

Cosmetic

The safety of Pyrus malus (apple)-derived ingredients is evaluated based on the expected use of these ingredients in cosmetics. The Panel uses data received from FDA and the cosmetics industry to determine cosmetic use. Use frequencies of individual ingredients in cosmetics are collected from manufacturers and reported by cosmetic product category in FDA’s VCRP database. Use concentration data are submitted by Industry in response to surveys of maximum reported use concentrations, by product category, that are conducted by the Council. Collectively, the use frequency and use concentration data indicate that 19 of the 28 apple-derived ingredients are used in cosmetic products.

According to information supplied to the FDA’s VCRP by industry, and the results from a survey of ingredient use concentrations conducted by the Council, 10 Pyrus malus (apple)-derived ingredients and 9 Malus domestica (apple)-derived ingredients are being used in cosmetic products, and malus domestica (apple) fruit extract has the highest reported use frequency (382 products)2,46 The following 9 ingredients are not reported as being used in cosmetic products:

Pyrus malus (apple) bark extract Pyrus malus (apple) peel wax Pyrus malus (apple) carpel powder Pyrus malus (apple) pulp extract Pyrus malus (apple) fiber Pyrus malus (apple) root bark powder Pyrus malus (apple) leaf extract Pyrus malus (apple) stem extract Pyrus malus (apple) peel extract

The Council survey data also indicate that apple-derived ingredients are being used in leave-on cosmetic products at maximum ingredient use concentrations up to 9% (i.e., for pyrus malus (apple) fruit water in face and neck products [not sprays]), and in rinse-off cosmetic products at maximum ingredient use concentrations up to 0.75% (i.e., for pyrus malus (apple) fruit extract in hair conditioners).46 Frequency of use/use concentration data for Pyrus malus (apple)- and Malus domestica (apple)-derived ingredients are summarized in Table 8.

Cosmetic products containing Pyrus malus (apple)-derived or Malus domestica (apple)-derived ingredients may be applied to the skin and 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 or hair for variable periods following application. Daily or occasional use may extend over many years.

The following ingredients are being used in products that may be inhaled: pyrus malus (apple) fruit extract, pyrus malus (apple) flower extract, pyrus malus (apple) fruit, pyrus malus (apple) seed oil, malus domestica (apple) fruit extract, and malus domestica (apple) fruit water. The highest maximum use concentration that is being reported for Pyrus malus (apple)- or Malus domestica (apple)-derived ingredients in these types of products is 0.1% for pyrus malus (apple) fruit extract in perfumes and in body and hand sprays. The product types and maximum use concentrations reported for all of the apple-derived ingredients that are used in products that may be inhaled are presented in Table 8. In practice, 95% to 99% of the droplets/particles released from cosmetic sprays have aerodynamic equivalent diameters >10 µm, with propellant sprays yielding a greater fraction of droplets/particles below 10 µm, compared with pump sprays.47,48,49,50 Therefore, most Distributed for comment only -- do not cite or quote

droplets/particles incidentally inhaled from cosmetic sprays would be deposited in the nasopharyngeal and bronchial regions and would not be respirable (i.e., they would not enter the lungs) to any appreciable amount.47,48

Noncosmetic

Apples are among the 20 most frequently consumed raw fruits in the United States.51

TOXICOKINETICS

Apple Juice

Unfiltered apple juice (500 ml) was consumed by human subjects, and blood and urine samples were analyzed for total phenolic content and the concentration of selected individual polyphenolic compounds.52 Large differences in apple polyphenol pharmacokinetics between subjects were observed. Pharmacokinetic data were classified into subgroups according to fast or slow rates of polyphenol metabolism. For some of the subjects, metabolism was undetectable within the time frame of the study. No differences in renal excretion were detected when male and female subjects were compared. However, relative concentrations of polyphenolic compounds were slightly higher in male subjects. Thus, apple-derived polyphenols can be readily detected in human blood and urine after apple juice consumption. The authors noted that the existence of subpopulations with different pharmacokinetics was suggestive of significant variations in individual metabolism rates of polyphenolic substances.

TOXICOLOGY

Except for data on antimicrobial activity and in vitro skin and ocular irritation data, toxicity data on the apple- derived ingredients reviewed in this safety assessment were not found in the published literature nor were unpublished data provided. However, as noted earlier, some of the ingredients (e.g., fruit/fruit-derived) reviewed in this safety assessment may be consumed as food, and daily exposure from food use would result in much larger systemic exposures than those from use in cosmetic products. Some of the ingredients (e.g., fruit/fruit-derived) reviewed in this safety assessment may be consumed as food, and daily exposure from food use would result in much larger systemic exposures than those from use in cosmetic products. The primary focus of the safety assessment of these ingredients as used in cosmetics is on the potential for local effects from topical exposure.

Toxicity data on procyanidin B-2 (epicatechin-(4β→8)-epicatechin, found in apple fruit) were identified in the published literature, and are included for use in the safety assessment of apple fruit extract. As mentioned previously, procyanidins are members of the procyanidin or condensed tannins class of flavonoids.10 Procyanidin B-2 is not a cosmetic ingredient.

Acute Toxicity

Oral

Apple Fruit Extract

Apple polyphenol extract (dissolved in 0.5% sodium carboxymethyl cellulose solution) was administered intragastrically to Sprague-Dawley (Crj:CD) rats (5 males, 5 females) at an oral dose of 2,000 mg/kg body weight (dose volume = 10 ml/kg).17 The production of apple polyphenol extract is described in the Method of Manufacture section of this safety assessment. The animals were necropsied after a 14-day observation period. None of the animals died, and the authors noted that there were no significant changes in organs examined. It was concluded that the acute oral minimum fatal dose of apple polyphenol extract was > 2,000 mg/kg body weight.

Pyrus Malus (Apple) Seed Extract

The acute oral toxicity of pyrus malus (apple) seed extract (1% w/v in hexyldecanol) was studied using 10 male mice (IFFA CREDO OF 1).25 The test substance was administered at a maximal oral dose of 20 ml/kg, and dosing was followed by an 8-day observation period. Gross necropsy was performed on day 8. There was no evidence of test substance-related clinical signs in the study, and the behavior of animals was considered normal. Gross necropsy did not Distributed for comment only -- do not cite or quote

reveal any evidence of modification of the main organs. Particularly, there were no signs of necrosis or ulceration of the digestive tract.

Subcutaneous

Procyanidin B-2

Procyanidin B-2 (> 94% pure) was administered to groups of ten 6-week-old Sprague-Dawley rats of the Crj: CD, SPF/VAF strain (5 males, 5 females/group).10,9 The test material (in purified water) was administered subcutaneously to 3 groups at doses of 500 mg/kg, 1,000 mg/kg, and 2,000 mg/kg, respectively (dose volume = 10 ml/kg). The single dose was injected into the neck. A fourth group (control) was dosed with purified water. None of the animals died. Hair loss and crust formation (in all 3 dose groups), and swelling (2,000 mg/kg group) were reported. The following observations were made at necropsy on day 14: thickening of the subcutis with granulomatous inflammation in the 1,000 mg/kg group (4 of 5 males; 2 of 5 females) and 2,000 mg/kg group (all animals); accumulation of pigment-laden macrophages in the duodenal mucosa in the 500 mg/kg group (1 of 5 females), 1,000 mg/kg group (4 of 5 males; all females), and 2,000 mg/kg group (all animals).

Repeated Dose Toxicity

Oral

Apple Fruit Extract

Apple polyphenol extract was administered intragastrically to 3 groups of Sprague-Dawley (Crj:CD) rats (10 males, 10 females) at doses of 500; 1,000; and 2,000 mg/kg/day (dose volume = 10 ml/kg).17 The doses were administered daily for 90 days. A fourth group served as the untreated control. Necropsies were performed on all animals at the end of the study. Body weight gain was unaffected by treatment, and changes in food consumption were attributed to normal biological variation. Statistically significant differences (P < 0.05 or P < 0.01) in hematological, clinical chemistry, and urinary tests were reported. However, these changes were within the normal range of physiological background data and were not correlated with the apple polyphenol extract dosage. Gross necropsy findings were not indicative of adverse changes in any of the organs examined. A statistically significant increase in lung weight was found in the 1,000 mg/kg/day dose group, but this change was within the normal physiological range. Histopathological examination of organs did not reveal any changes that were related to dosing with the test material.

Antimicrobial Activity

Pyrus Malus (Apple) Pectin Extract

Pectin was extracted from several varieties of apple, i.e., American, Delicious, and Maharaj-ji (all obtained from Kashmir).20 Pectin content was found to be maximum in Maharaj-ji (20.60%), followed by Delicious (14.4%) and American (11.60%). The pectin extracted was evaluated for in vitro antibacterial activity against different pathogenic bacterial cultures and antifungal activity. Pectin extracted from the Delicious variety had potent antibacterial activity against Klebsiella pneumonia (mean inhibitory concentration [MIC] = 0.8 mg/ml), followed by Streptococcus pyogenes (MIC = 0.3 mg/ml), E. coli (MIC = 0.7 mg/ml), and Lactococcus sp. (MIC = 0.7 mg/ml). There was no evidence of antibacterial activity in any of the bacterial strains tested with pectin extracted from the other varieties. Regardless of the variety from which pectin was extracted, there was no evidence of antifungal activity against the following fungal strains: Aspergillus niger, Candida albicans, or Saccharomyces cerevisiae.

REPRODUCTIVE AND DEVELOPMENTAL TOXICITY

Data on the reproductive and developmental toxicity of apple-derived ingredients were not found in the published literature nor were unpublished data provided.

GENOTOXICITY

In Vitro

Apple Fruit Extract Distributed for comment only -- do not cite or quote

The genotoxicity of apple fruit extract was evaluated in the Ames test using Escherichia coli strain WP2uvrA and Salmonella typhimurium strains TA100, TA98, TA1535, and TA1537.10 The test material was evaluated at doses up to 5,000 µg/plate with and without metabolic activation. Slight genotoxicity was observed at a dose of 2,500 µg/plate without metabolic activation, but not at the other doses, with or without metabolic activation.

Apple fruit extract (concentrations up to 0.313 mg/ml) was evaluated for genotoxic activity in Chinese hamster CHL/IU mammalian cells using a chromosomal aberrations assay with and without metabolic activation.10 Genotoxicity was not observed, with or without metabolic activation.

Pyrus Malus (Apple) Seed Extract

The genotoxicity of pyrus malus (apple) seed extract (1% w/v in hexyldecanol) was studied using the following S. typhimurium strains: TA98, TA100, TA1535, TA1537, and TA1538.25 The test substance was evaluated at doses up to 5,000 µg/plate with and without metabolic activation. Revertant frequencies in treated strains tested were similar to vehicle control values. All positive control data were within acceptable ranges. The test substance was classified as non-genotoxic.

Procyanidin B-2

The genotoxicity of procyanidin B-2 (> 94% pure) was evaluated using S. typhimurium strains TA98, TA100, TA1535, and TA1537, and E. coli strain WP2uvrA.9 Procyanidin B-2 was tested at doses up to 5,000, µg/plate with or without metabolic activation. Test results were negative.

In the chromosome aberrations assay using Chinese hamster lung cells, procyanidin B-2 (> 94% pure) was evaluated with (concentrations up to 4.8 mM) and without (concentrations up to 1.8 mM) metabolic activation.9 Neither structural aberrations such as chromosome-type aberrations or chromatid-type aberrations were observed with or without metabolic activation. However, polyploidy was observed with metabolic activation.

In Vivo

Apple Fruit Extract

The genotoxicity of apple fruit extract was evaluated in the micronucelus test.17 Three groups of 5 male Sprague- Dawley rats (7 weeks old) were dosed orally with 500 mg/kg body weight; 1,000 mg/kg body weight; and 2,000 mg/kg body weight (dose volume = 10 ml/kg), respectively. Genotoxicity was evaluated by measuring the frequency of polychromatic erythrocytes cells in bone marrow. Test results were negative.

Procyanidin B-2

In a micronucleus test, groups of 8-week-old mice [Crj: CD-1 (ICR), specific pathogen free (SPF)/virus antibody free (VAF)] received single subcutaneous injections of 500 mg/kg, 1,000 mg/kg, and 2,000 mg/kg procyanidin B-2, respectively.9 The frequency of micronucleated polychromatic erythrocytes in the bone marrow of mice dosed with procyanidin B-2 was not significantly different from that of the negative control. Whether or not a statistical analysis of the data was performed was not stated.

CARCINOGENICITY

Data on the carcinogenicity of apple-derived ingredients were not found in the published literature nor were unpublished data provided.

ANTICARCINOGENICITY

In in vitro studies, apple extracts and components, especially oligomeric procyanidins, have been shown to influence multiple mechanisms that are relevant for cancer prevention.53 Additionally, apple products have been shown to prevent Distributed for comment only -- do not cite or quote

skin, mammary, and colon carcinogenesis in animal models, and epidemiological observations indicate that regular consumption of one or more apples a day may reduce the risk for lung and colon cancer.

IRRITATION AND SENSITZATION

Skin Irritation and Sensitization

In Vitro

Pyrus Malus (Apple) Fruit Extract

The skin irritation potential of a tradename material containing 20% pyrus malus (apple) fruit extract was evaluated using the EpidermTM model, a reconstructed human epidermal model consisting of human-derived epidermal keratinocytes.54 This is a European Center for the Validation of Alternative Methods (ECVAM)-validated test method.55 The keratinocytes have been cultured to form a multilayer, highly differentiated model of the human epidermis. In the EpidermTM model assay, a chemical is classified as an irritant if the mean relative tissue viability of exposed tissues is reduced by 50% of the mean viability of the negative controls. A non-irritant’s viability is > 50%. Sterile Dulbecco’s phosphate-buffered saline (DPBS) and sterile deionized water served as negative controls. Sodium dodecyl sulfate solution (5%) served as the positive control. The test material was classified as a non-irritant in this assay. The positive control was classified as an irritant.54 In the same assay, negative results were also reported for another tradename material containing 20% pyrus malus (apple) fruit extract.56

Non-Human

Pyrus Malus (Apple) Fruit Extract

A product described as an aqueous solution containing 20% pyrus malus (apple) fruit extract was applied for 4 h (under a dressing [type not stated]) to the skin of 3 albino rabbits according to the Organization for Economic Co-operation and Development (OECD) 404 protocol.16 Test sites were examined for up to 72 h after removal of the dressing. The test substance was classified as having slight skin irritation potential.

The skin sensitization potential of a product described as an aqueous solution containing 20% pyrus malus (apple) fruit extract was studied in the maximization test (OECD 406 protocol) using albino guinea pigs (number not stated).16 Positive reactions were not observed. However, it was concluded that the test substance had slight skin sensitization potential, but did not induce skin irritation.

Pyrus Malus (Apple) Seed Extract

The skin irritation potential of pyrus malus (apple) seed extract (1% w/v in hexyldecanol) was evaluated using 6 New Zealand white rabbits.25 Prior to application, the test substance was diluted to a concentration of 23% in sterile distilled water (effective test substance concentration = 1% x 23% = 0.23%). An occlusive patch containing the diluted test substance (0.5 ml) was applied for 24 h to abraded and intact skin on the right and left side, respectively. The dose per cm2 was not stated. Reactions were scored according to the Draize scale at 24 h, 48 h, and 72 h post-application. Mild erythema was observed at intact and abraded test sites of all animals, but was completely reversible between 48 h and 72 h post-application. Edema was not observed. The test substance was classified as a slight skin irritant (index of cutaneous irritation = 0.58).

The guinea pig maximization test was used to evaluate the skin sensitization potential of pyrus malus (apple) seed extract (1% w/v in hexyldecanol) in 20 male guinea pigs (strain not stated).25 During induction, the animals were injected intradermally with the test substance, followed by topical dermal application of the test substance (25% in Vaseline) under an occlusive patch for 48 h. The effective concentration of the test substance applied dermally during induction was 0.25% (1% x 25% = 0.25%). The dose per cm2 was not stated. During the challenge phase, the test substance (0.25%) was applied to the flank, under an occlusive, patch for 24 h. An additional 20 guinea pigs served as controls (treated similarly, but without test substance application). Reactions were scored at 24 h and 48 h post-application. Mild erythema was observed in treated and control animals, and reactions had cleared by 48 h. The erythema observed was interpreted as a consequence of an irritation phenomenon, rather than as a sensitization phenomenon. This was so because the erythema observed at 24 h was reported for an equal number of treated and control animals, and because the percentage of the reaction observed was extremely low. The test substance was classified as a non-sensitizer.

Procyanidin B-2

Distributed for comment only -- do not cite or quote

Procyanidin B-2 (vehicle not stated) was applied topically to dorsal abraded and intact skin of 12 New Zealand white rabbits (Kbs: NZW, SPF) using an occlusive patch (0.5 ml on 2.5 x 2.5 cm patch).9 Sodium lauryl sulfate (8% aqueous) and physiological saline served as positive and negative controls, respectively. The patches remained in place for 24 h, and reactions were scored at 24.5 h and 72 h post-application. Skin irritation was not observed at abraded or intact sites treated with procyanidin B-2, the vehicle, or saline. Significant inflammation (severe irritation) was observed at sites treated with 8% sodium lauryl sulfate.

The skin sensitization potential of procyanidin B-2 was evaluated in a maximization test using 24 male guinea pigs (Crj: Hartley,SPF/VAF).9 A 2 x 4 cm area of dorsal skin was used in the induction phase of the study, and the test procedure involved a combination of subcutaneous injections and topical applications. During the induction phase, procyanidin B-2 (1%) was injected subcutaneously, and, after 24 h, an occlusive patch containing 0.2 ml of 10% procyanidin B-2 was applied for 24 h. The challenge phase was initiated after a 2-week non-treatment period. A 2 x 2 cm occlusive patch containing 10% procyanidin B-2 (0.1 ml) was applied for 24 h. Reactions were scored at 24.5 h, 48 h, and 72 h post-application. 2,4- Dinitrochlorobenzene (1% DNCB) served as the positive control. Procyanidin B-2 did not induce sensitization. DNCB was a strong sensitizer.

Human

Pyrus Malus (Apple) Fruit Water

The skin irritation and sensitization potential of a face and neck product containing 8.7835% pyrus malus (apple) fruit water was evaluated in a human repeated insult patch test involving 49 male and female subjects.57 During induction, an occlusive patch (2 cm2) containing the test substance (0.2 ml) was applied for 24 h to an area between the scapula and waist that was adjacent to the midline. A total of 9 induction applications were made. A 2-week non-treatment period was followed by the challenge phase. A challenge patch was applied for 24 h to a new test site. Reactions were scored at 24 h and 72 h post-application. Transient, barely perceptible (+) to mild (1-level) reactions (all non-specific patch test responses) were observed in 5 or 49 subjects during the induction and/or challenge phase of the study. It was concluded that the test substance did not induce skin irritation or cause allergic contact dermatitis.

Phototoxicity

Pyrus Malus (Apple) Seed Extract

The phototoxicity of pyrus malus (apple) seed extract (1% w/v in hexyldecanol) was studied using albino guinea pigs.25 The animals (10 per group) received an application of the test substance (1 ml) with or without UVA irradiation. The dose per cm2 was not stated. Test sites were examined at 1 h, 6 h, and 24 h post-irradiation. There was no evidence of dermal reactions in irradiated or non-irradiated animals, and the test substance was classified as non-phototoxic.

Ocular Irritation

In Vitro

Pyrus Malus (Apple) Fruit Extract

The ocular irritation potential of a tradename material containing 20% pyrus malus (apple) fruit extract was evaluated using the EpiocularTM model, a multilayer, highly differentiated model of the human corneal epithelium.54 In the EpiocularTM model assay, a chemical is classified as an irritant if the mean relative tissue viability of the exposed tissues is reduced by 60% of the mean viability of the negative controls. A non-irritant’s viability is > 40%. Sterile DPBS and sterile deionized water served as negative controls. Methyl acetate served as the positive control. The test material was classified as a non-irritant in this assay. The positive control was classified as an irritant. In the same assay, negative results were also reported for another tradename material containing 20% pyrus malus (apple) fruit extract.56

Using the method of neutral red release, the ocular irritation potential of a product described as an aqueous solution containing a maximum percentage of 20% pyrus malus (apple) fruit extract was determined using a monolayer of rabbit corneal fibroblasts.16 The product was tested at concentrations up to 50% (effective test substance concentration = 20% x 50% = 10%) In this assay, cytotoxicity was evaluated by determining the test concentration that caused 50% cell death (IC50). Test results indicated that the IC50 was above a concentration of 50%. The % cell death that was observed at a dilution of 50% was 15%, and, thus, cytotoxicity was said to have been negligible.

Distributed for comment only -- do not cite or quote

In Vivo

Pyrus Malus (Apple) Fruit Extract

A product described as an aqueous solution containing a maximum percentage of 20% pyrus malus (apple) fruit extract (0.1 ml) was instilled into one eye of each of 3 rabbits according to the OECD 405 protocol.16 Instillation was not followed by ocular rinsing. Untreated eyes served as controls. The test substance was slightly irritating to the eye and did not cause lesions of the ocular mucosa that were considered significant.

Pyrus Malus (Apple) Seed Extract

Pyrus malus (apple) seed extract (0.1 ml; 1% in hexyldecanol) was instilled into the right eye of each of 6 New Zealand white rabbits, and remained for 24 h.25 Ocular irritation was scored according to the method of Kay and Calandra at 24 h, 48 h, 72 h, 96 h, and 12 h post-instillation. At 24 h post-instillation, conjunctival redness, swelling, and discharge were observed in 6 rabbits. These reactions were completely reversible at 48 h and 96 h post-instillation. Lesions of the cornea or iris were not observed. The test substance was classified as slightly irritating to the eyes (accurate ocular irritation index at 24 h = 0.58).

Procyanidin B-2

The ocular irritation potential of procyanidin B-2 (> 94% pure) was evaluated using twelve male New Zealand white rabbits (Kbs: NZW, SPF).9 The test material (100 µl) was instilled in the right eye of each animal. The left eye served as the untreated control. Half of the animals were subjected to ocular rinsing after instillation. Ocular reactions were scored for up to 120 h post-instillation. No changes were observed in the cornea or iris, but slight irritation of the conjunctiva was observed. Procyanidin B-2 was classified as minimally irritating to the eyes of rabbits.

MISCELLANEOUS STUDIES

Phloridzin

A study on phloridzin-induced melanogenesis was performed using B16 melanoma cells.58 Phloridzin (found in many parts of the apple tree) induced a dose-dependent increase in tyrosinase activity and melanin content, and these changes were accompanied by an increase in the levels of tyrosinase and the tyrosine-related proteins, TRP-1 and TRP-2. Also, the cAMP-dependent protein kinase A (PKA) inhibitor H89 impaired the response of the tyrosinase activity and melanin synthesis to phloridzin. Collectively, the results of this study indicated that phloridzin increased tyrosinase gene expression through the cAMP signaling pathway, leading to stimulation of melanogenesis.

SUMMARY AND DISCUSSION FROM FINAL REPORT ON PLANT-DERIVED FATTY ACID OILS AS USED IN COSMETICS

Summary

The report addresses the safety of Plant-Derived Fatty Acid Oils. These oils, which are derived from vegetable and fruit plants, are composed of mono-, di-, and, primarily, triglycerides, free fatty acids and other minor components, including natural antioxidants and fat-soluble vitamins. The percentage of chemical constituents and nutritional content of individual oil types is dependent on region where the oil plant is grown, individual cultivars, and plant genetics. Oils used in cosmetics are likely produced in the same manner as those used in the food industry. Oils may be expressed through mechanical or solvent extraction. The oils may undergo further refining, such as neutralizing, bleaching, and deodorizing, to remove pigments, odors, unsaponifiable materials, and other undesirables.

Individuals who have food allergies to a plant protein rarely exhibit allergic reactions when exposed to refined oils of the same plant. Data evaluation by the CIR Expert Panel regarding method of manufacture indicates that protein constituents do not partition into the refined oils. The CIR Expert Panel also has found a general lack of clinical effects for Distributed for comment only -- do not cite or quote

fatty acid oils that they have already reviewed; however, other researchers have raised concerns about the presence of residual proteins in oils, such as peanut and soy.

Glycidol fatty acid esters are possible impurities in refined vegetable oils. While the amount of glycidol that may be present with glycidol fatty acid esters is not known, the IARC has noted that glycidol is probably carcinogenic to humans and that glycidol fatty acid esters are not classifiable as to carcinogenicity in humans. Peanuts and soy may contain aflatoxins, metabolic products of certain molds that are carcinogenic to humans.

Of the oils described in this report, Butyrospermum Parkii (Shea) Butter has the most reported uses in cosmetic and personal care products with a total of 1950 and is used at a maximum concentration of 60%. Oils are used in a wide variety of cosmetic products, including use in hair spray and other aerosolized products. None of the oils, or the related counterparts, described in this report are restricted from use in the European Union.

Anacardium Occidentale (Cashew) Seed Oil was not a tumor promoter in a DMBA skin test system.

The safety focus of use of these oils as cosmetic ingredients is on the potential for irritation and sensitization. Undiluted, technical grade, Arachis Hypogaea (Peanut) Oil was moderately irritating to rabbits and guinea pig skin, and 5% aq. solutions of a bar soap containing 13% sodium cocoate had irritation scores of 1.6-4.0/8 in animal studies. However, the remaining animal and clinical irritation and/or sensitization studies conducted on a large number of the oils included in this report, primarily in formulation, did not report any significant irritation or sensitization reactions, indicating that refined oils derived from plants are not dermal irritants or sensitizers.

The phototoxic potential of formulations containing Butyrospermum Parkii (Shea) Butter and Elaeis Guineensis (Palm) Oil and of Oryza Sativa (Rice) Bran and (Rice) Germ Oil, neat, was evaluated in animal studies, and the phototoxic potential of Cocos Nucifera (Coconut) Oil, Sodium Cocoate, Prunus Amygdalus Dulcis (Sweet) Almond Oil, and Oryza Sativa (Rice) Bran Oil was examined clinically. None of these ingredients were phototoxic.

The comedogenicity of Corylus Avellana (Hazel) Seed Oil was evaluated using rabbits, and a slight difference in the number and size of the pilosebaceous follicles and a slight excess of sebum and a dilation of the follicles was observed. In clinical testing with an eye mask containing 0.2% Ribes Nigrum (Black Currant) Seed Oil (undiluted), the formulation was non-comedogenic.

The ocular irritation potential of a number of the oils, mostly in formulation, was evaluated in testing using animals or alternative assays. The majority of the test results did not report significant ocular irritation. A lotion containing 1.5% Elaeis Guineensis (Palm) Oil was moderately irritating to rabbit eyes, and a mascara containing 9.4% Linum Usitatissimum (Linseed) Seed Oil was moderately irritating in an alternative assay.

In human testing, a mascara containing 9.4% Linum Usitatissimum (Linseed) Seed Oil did not produce ocular irritation or adverse effects in contact lenses wearers or subjects with sensitive eyes. An eye mask containing 0.2% Ribes Nigrum (Black Currant) Seed Oil (undiluted) was tested and considered safe for contact lens wearers.

DISCUSSION

Plant-derived fatty acid oils, oils which have been hydrogenated to reduce or eliminate unsaturation, fatty acid salts, and oil unsaponifiables were reviewed by the CIR Expert Panel. Most of theses ingredients in this report are mixtures of triglycerides containing fatty acids and fatty acid derivatives, the safety of which in cosmetics has been established. Upon review of these ingredients, the Panel expressed concern regarding pesticide residues and heavy metals that may be present in botanical ingredients. They stressed that the cosmetics industry should continue to use the necessary procedures to limit these impurities in the ingredient before blending into cosmetic formulations.

Additionally, the Panel considered the safety of glycidol and glycidol fatty acid esters in refined vegetable oils. While the Panel recognizes that these impurities may be carcinogenic, absorption through the skin would be very low and likely does not pose a significant hazard. Nonetheless, suppliers should take steps to eliminate or reduce the presence of glycidol and glycidol fatty acid esters in plant-based fatty acid oils that are used in cosmetic products. Aflatoxins, which are potent carcinogens, may be present in moldy nuts and coconut copra, but are not found in oils expressed from these nuts and copra. The Panel adopted the U.S. Department of Agriculture designation of <15 ppb as corresponding to “negative” aflatoxin content.

Certain of the plant-derived oils are used in cosmetic products that may be inhaled during their use. In practice, however, the particle sizes produced by the cosmetic aerosols are not respirable. Distributed for comment only -- do not cite or quote

The Panel discussed the relationship between food allergies and exposure to refined oils. Individuals who have food allergies to a plant protein rarely exhibit allergic reactions when exposed to refined oils of the same plant. The Panel has found a general lack of clinical effects for plant-derived fatty acid oils already reviewed.

Fatty acid composition data were available for the majority of the oils included in this review and the Panel agreed that the composition data, in combination with the available data on method of manufacture, impurities, safety test data, a long history of safe use in foods, and an absence of adverse reactions in clinical experience, was a sufficient basis for determining safety. The Expert Panel did note that vegetable oil is a blend of a number of different oils, and that a specific composition of vegetable oil was not available. The Expert Panel determined that the safety of vegetable oil as used in cosmetic formulations has been established, providing that the blend contains oils for which the fatty acid composition is known.

Additionally, while data on the fatty acid composition of Fragaria Vesca (Strawberry) Seed Oil and Fragaria Virginiana (Strawberry) Seed Oil were not available, data were available for Fragaria Ananassa (Strawberry) Seed Oil and Fragaria Chiloensis (Strawberry) Seed Oil. In that the fatty acid compositions of Fragaria Ananassa and Fragaria Chiloensis (Strawberry) Seed Oil were similar to each other, it was assumed that Fragaria Vesca and Fragaria Virginiana (Strawberry) Seed Oils would also have similar fatty acid compositions.

The Expert Panel also noted that arachidonic acid is a fatty acid constituent of Lycium Barbarum Seed Oil, Oryza Sativa (Rice) Germ Oil, and Sclerocarya Birrea Seed Oil. Although a previously published CIR evaluation concluded that insufficient data exist to support the safety of arachidonic acid in cosmetic products, the Panel was of the opinion that the concentration of use of these ingredients was sufficiently low that the amount of free arachidonic acid from these oils would not warrant concern.

Finally, the conclusion reached by the Panel on the safety of the plant-derived fatty acid oils supersedes the 2001 conclusion of insufficient data for Corylus Americana and Corylus Avellana (Hazel) Seed Oil.

SUMMARY

Most of the ingredients reviewed in this safety assessment are Pyrus malus (apple)-derived ingredients, and pyrus malus is part of each ingredient name. However, CIR has been informed by the cosmetics industry that Pyrus malus is not the current accepted name for apple that is used by botanists, and that the ingredients reviewed in this safety assessment are under consideration by the cosmetics industry for new name assignments.

The following functions of Pyrus malus (apple)-derived ingredients and Malus domestica-derived ingredients in cosmetic products are being reported: skin conditioning agents, binders, emulsion stabilizers, viscosity increasing agents, astringents, fragrance ingredients, antioxidants, exfoliants, and skin bleaching agents (skin bleaching is not regarded as a cosmetic use in the U.S.) Pyrus malus (apple) root bark powder is the only ingredient in this group that is reported to function as a skin bleaching agent in cosmetic products.

According to information supplied to FDA by industry as part of the VCRP, and the results from a survey of ingredient use concentrations conducted by the Council, the following 10 Pyrus malus (apple)-derived ingredients and 9 Malus domestica (apple)-derived ingredients are being used in cosmetic products, and malus domestica (apple) fruit extract has the highest reported use frequency (382) products):

Pyrus malus (apple) fruit extract Malus domestica (apple) fruit extract Pyrus malus (apple) flower extract Malus domestica (apple) fruit water Pyrus malus (apple) fruit Malus domestica (apple) seed oil Pyrus malus (apple) fruit water Malus domestica (apple) fiber Pyrus malus (apple) juice Malus domestica (apple) juice Pyrus malus (apple) pectin extract Malus domestica (apple) oil Pyrus malus (apple) peel powder Malus domestica (apple) stem extract Pyrus malus (apple) root extract Malus domestica (apple) fruit cell culture extract Pyrus malus (apple) seed extract Malus domestica (apple) callus extract Pyrus malus (apple) seed oil

Distributed for comment only -- do not cite or quote

Pyrus malus (apple)-derived ingredients are being used in leave-on cosmetic products at maximum ingredient use concentrations up to 9% (i.e., for pyrus malus (apple) fruit water in face and neck products [not sprays]), and in rinse-off cosmetic products at maximum ingredient use concentrations up to 0.75% (i.e., for pyrus malus (apple) fruit extract in hair conditioners).

The available information on the composition of apples indicates that phenolic compounds are present in the fruit, juice, leaves, peel, and seeds. Phloridzin (found in many parts of the apple tree) induced a dose-dependent increase in tyrosinase activity and melanin content, and these changes were accompanied by an increase in the levels of tyrosinase and the tyrosine-related proteins, TRP-1 and TRP-2.

Apple-derived polyphenols were readily detected in human blood and urine after apple juice consumption.

In an acute oral toxicity study of pyrus malus (apple) seed extract (1% w/v in hexyldecanol), there was no evidence of test substance-related clinical signs. At gross necropsy, there were no signs of necrosis or ulceration of the digestive tract, or modification of the main organs. In acute oral and repeated dose toxicity tests (rats) on apple fruit extract, there were no significant hematological, clinical, chemical, histopathological, or urinary effects at a dose of 2000 mg/kg.

A single subcutaneous injection of procyanidin B-2 into rats did not produce signs of significant injury, and a lethal dose of > 2,000 mg/kg was reported.

In the Ames test without metabolic activation, apple fruit extract was slightly genotoxic when tested at a high concentration of 2500 µg/plate, but significant genotoxic activity was not found in the chromosomal aberration test or the micronucleus test. In another Ames test, pyrus malus (apple) seed extract (1% w/v in hexyldecanol) was non-genotoxic in the following Salmonella typhimurium strains, with and without metabolic activation: TA98, TA100, TA1535, TA1537, and TA1538. Results were negative for procyanidin B-2 in the Ames test and in a chromosome aberrations assay involving Chinese hamster lung cells.

An aqueous solution containing 20% pyrus malus (apple) fruit extract was slightly irritating to the skin of rabbits. In the guinea pig maximization test, this solution did not induce skin irritation, but had slight skin sensitization potential. The skin irritation potential of a tradename material containing 20% pyrus malus (apple) fruit extract was evaluated using the EpidermTM model, and results were negative.

Pyrus malus (apple) seed extract (1% w/v in hexyldecanol) caused slight skin irritation when diluted with water to an ingredient concentration of 0.23% and applied to the skin of rabbits. In the guinea pig maximization test, pyrus malus (apple) seed extract (1% w/v in hexyldecanol) caused skin irritation, but not sensitization, when tested at an ingredient concentration of 0.25% during induction and challenge. A preparation containing procyanidin B-2 did not cause skin irritation in rabbits. Additionally, procyanidin B-2 did not induce skin sensitization in guinea pigs in the maximization test.

A face and neck product containing 8.7835% pyrus malus (apple) fruit water did not cause skin irritation or allergic contact dermatitis in an RIPT involving 49 subjects.

Pyrus malus (apple) seed extract (1% w/v in hexyldecanol) was classified as non-phototoxic in a study involving albino guinea pigs.

An aqueous solution containing a maximum percentage of 20% pyrus malus (apple) fruit extract (0.1 ml) was slightly irritating to the eyes of rabbits. The same was true after instillation of pyrus malus (apple) seed extract was (1% in hexyldecanol) into the eyes of rabbits. In primary ocular irritation tests involving rabbits, both a preparation containing procyanidin B-2 and the vehicle induced slight conjunctival irritation; however, ethanol was presumed to have been the causative agent.

The ocular irritation potential of an aqueous solution containing a maximum percentage of 20% pyrus malus (apple) fruit extract, tested at ingredient concentrations up to 10%, was evaluated using the method of neutral red release. Cytotoxicity was evaluated in this assay and found to be negligible; thus, ocular irritation was negligible. A tradename material containing 20% pyrus malus (apple) fruit extract was evaluated for ocular irritation potential using the EpiocularTM model, and results were negative.

DISCUSSION

Distributed for comment only -- do not cite or quote

Because final product formulations may contain multiple botanicals, each possibly containing similar constituents of concern, formulators are advised to be aware of these constituents and to avoid reaching levels that may be hazardous to consumers. Therefore, when formulating products, manufacturers should avoid reaching levels of plant constituents that may cause sensitization or other adverse health effects. Protein components should also be minimized, given the potential for cross-reactivity with plant constituents. Considering that apple-derived ingredients may be obtained from different apple cultivars, the Panel noted that the composition of ingredients derived from different cultivars should be similar to the composition of ingredients reviewed in this safety assessment.

The Panel discussed the study included in this report that indicated that phloridzin, a component of apple fruit, apple bark extract, apple root bark powder, apple leaf extract, and apple seed extract, has a stimulatory effect on melanogenesis, but agreed that a review of ingredients for drug effects (i.e., noncosmetic effects such as skin bleaching) is not within the Panel’s purview. However, the Panel noted that an effect on cutaneous pigmentation would not be expected at the use concentrations of apple-derived ingredients in cosmetic products.

The Panel expressed concern about pesticide residues and heavy metals that may be present in botanical ingredients. They stressed that the cosmetics industry should continue to use current good manufacturing practices (cGMPs) to limit impurities. The Panel noted the USDA designation of ≤ 15 ppb as corresponding to “negative” aflatoxin content. Additionally, the Panel stated that aflatoxins would not be present at levels of toxicological concern in apple-derived ingredients.

Additionally, the Panel discussed the potential for incidental inhalation exposures to apple-derived ingredients in products that are sprayed or in powder form and agreed that, based on likely airborne particle size distributions and concentrations in the breathing zone and ingredient use, incidental inhalation would not lead to local respiratory effects or systemic effects.

The Panel determined that the following data are needed to evaluate the safety of pyrus malus (apple) root extract, pyrus malus (apple) stem extract, and malus domestica (apple) stem extract in cosmetic products:

• Method of manufacture and impurities • 28-day dermal toxicity study; if any adverse effects noted, genotoxicity and reproductive and developmental toxicity data may be needed

CONCLUSION

The CIR Expert Panel concluded that the following 25 apple-derived ingredients are safe in the present practices of use and concentration in cosmetics as described in this safety assessment, when formulated to be non-irritating and non- sensitizing.

Pyrus Malus (apple) bark extract* Pyrus Malus (apple) pulp extract* Pyrus Malus (apple) carpel powder* Pyrus Malus (apple) root bark powder* Pyrus Malus (apple) fiber* Pyrus Malus (apple) seed extract Pyrus Malus (apple) flower extract Pyrus Malus (apple) seed oil* Pyrus Malus (apple) fruit extract Malus Domestica (apple) callus extract* Pyrus Malus (apple) fruit Malus Domestica (apple) fiber Pyrus Malus (apple) fruit water Malus Domestica (apple) fruit extract Pyrus Malus (apple) juice Malus Domestica (apple) fruit water Pyrus Malus (apple) leaf extract* Malus Domestica (apple) fruit cell culture extract Pyrus Malus (apple) pectin extract Malus Domestica (apple) juice Pyrus Malus (apple) peel extract* Malus Domestica (apple) oil Pyrus Malus (apple) peel powder Malus Domestica (apple) seed oil Pyrus Malus (apple) peel wax*

The Panel concluded that the available data are insufficient for evaluating the safety of the following 3 apple-derived ingredients in cosmetic products:

Pyrus Malus (apple) root extract Pyrus Malus (apple) stem extract* Malus Domestica (apple) stem extract Distributed for comment only -- do not cite or quote

1 Table 1. Definitions and functions of the ingredients in this safety assessment. Ingredient/CAS No. Definition Function Pyrus malus (apple) fruit Pyrus malus (apple) fruit extract is the extract of the fruit of Pyrus malus. Skin- extract Conditioning Agents - Miscellaneous Pyrus malus (apple) bark Pyrus malus (apple) bark extract is the extract of the bark of Pyrus malus. Cosmetic extract Astringents Pyrus malus (apple) carpel Pyrus malus (apple) carpel powder is the powder obtained from the dried, ground Skin- powder carpels of Pyrus malus. Conditioning Agents - Miscellaneous Pyrus malus (apple) fiber Pyrus malus (apple) fiber is the finely ground fiber obtained from the dried fruit of Binders; Emuls Pyrus malus. ion Stabilizers; Vis cosity Increasing Agents - Aqueous Pyrus malus (apple) flower Pyrus malus (apple) flower extract is the extract of the flowers of Pyrus malus. Skin- extract Conditioning Agents - Miscellaneous Pyrus malus (apple) fruit Pyrus malus (apple) fruit is the fruit of the apple, Pyrus malus. Cosmetic Astringents Pyrus malus (apple) fruit Pyrus malus (apple) fruit water is an aqueous solution of the steam distillate obtained Fragrance water from the fruit of Pyrus malus. Ingredients Pyrus malus (apple) juice Pyrus malus (apple) juice is the liquid expressed from the fresh pulp of the apple, Pyrus Skin- malus. Conditioning Agents - Miscellaneous Pyrus malus (apple) leaf Pyrus malus (apple) leaf extract is the extract of the leaves of Pyrus malus. Skin- extract Conditioning Agents - Miscellaneous Pyrus malus (apple) pectin Pyrus malus (apple) pectin extract is the extract of the pectin of Pyrus malus. Skin- extract Conditioning Agents - Miscellaneous Pyrus malus (apple) peel Pyrus malus (apple) peel extract is the extract of the peel of Pyrus malus. Antioxidants extract Pyrus malus (apple) peel Pyrus malus (apple) peel powder is the powder obtained from the dried, ground peel of Exfoliants powder Pyrus malus. Pyrus malus (apple) peel wax Pyrus malus (apple) peel wax is a wax obtained from the peel of the apple, Pyrus Not Reported malus. Pyrus malus (apple) pulp Pyrus malus (apple) pulp extract is the extract of the pulp of Pyrus malus. Skin- extract Conditioning Agents - Miscellaneous Pyrus malus (apple) root bark Pyrus malus (apple) root bark powder is the powder obtained from the dried, ground Skin Bleaching powder root bark of Pyrus malus. Agents Pyrus malus (apple) root Pyrus malus (apple) root extract is the extract of the roots of Pyrus malus. Skin- extract Conditioning Agents - Miscellaneous Pyrus malus (apple) seed Pyrus malus (apple) seed extract is the extract of the seeds of Pyrus malus. Skin- extract Conditioning Agents - Miscellaneous Pyrus malus (apple) seed oil Pyrus malus (apple) seed oil is the oil expressed from the seeds of Pyrus malus. Skin- Conditioning Agents - Emollient Pyrus malus (apple) stem Pyrus Malus (Apple) Stem Extract is the extract of the stems of Pyrus malus. Skin- extract Conditioning Agents - Miscellaneous Distributed for comment only -- do not cite or quote

1 Table 1. Definitions and functions of the ingredients in this safety assessment. Ingredient/CAS No. Definition Function Malus domesatica (apple) Malus Domestica Fruit Extract is the extract of the fruit of Malus domestica. Antioxidants fruit extract Malus domestica (apple) fruit Malus Domestica Fruit Water is an aqueous solution of the steam distillates derived Fragrance water from the fruit of Malus domestica. Ingredients; Sk in- Conditioning Agents - Emollient Malus domestica (apple) fruit Malus Domestica Fruit Cell Culture Extract is the extract of a culture of the fruit cells Skin- cell culture extract of Malus domestica. Conditioning Agents - Miscellaneous Malus domestica (apple) Malus Domestica Callus Extract is the extract of the callus of Malus domestica grown Antioxidants; S callus extract in culture. kin Protectants

Table 2. Properties of Pyrus Malus (Apple) Fruit Extract Trade Name Mixtures.6,7 A mixture containing 10-25% pyrus A mixture containing 10-25% pyrus malus (apple) fruit extract and 75- malus (apple) fruit extract, 10-25% 100% propylene glycol water, and 75-100% glycerin Clear, brownish yellow colored Clear, brownish yellow colored liquid liquid Faint fruity odor Faint fruity odor

Table 3. Composition Data on Different Varieties of Pyrus malus.29

Non- Fat Fiber Protein Pectin Reducing reducing Total Variety (%) (%) (%) (%) Sugar Sugar Sugar Amri 0.22 5.89 0.47 0.58 10.34 3.87 14.21 Bluish Golden 5.15 3.61 0.29 1.29 9.71 2.58 11.83 Bonza 0.21 4.41 0.74 1.63 11.18 4.28 15.46 Discovery 0.66 8.6 0.44 1.6 10.59 6.16 16.75 Elster 0.25 7.46 0.36 0.88 9.51 7.88 16.59 Florina 0.24 4.17 0.48 0.62 11.84 5.93 17.77 Gala 0.26 4.35 0.27 0.53 11.95 5.78 17.73 Gloster 69 3.38 4.03 0.17 1.07 10.76 3.75 14.51 Golden Delicious 0.23 5.06 0.44 0.51 11.29 4.41 15.7 Granny Smith 0.28 3.8 0.48 0.99 11.14 4.32 15.46 Green Sleeves 0.24 6.14 0.37 1.05 11.37 7.92 19.29 Ida Red 0.21 1.91 0.36 0.69 9.95 3.57 13.52 Kaja 0.08 1.13 0.38 0.43 7.1 0.57 7.67 Kids Orange 0.21 1.16 0.35 1.07 11.37 7.51 18.88 Mushadi 0.23 5.55 0.45 0.52 8.69 3.75 12.44 Red Delicious 0.07 2.56 0.47 0.58 10.74 4.09 14.83 Samootree 0.02 3.92 0.34 1.29 13.14 6.99 20.13 Summer Red 0.13 5.26 0.46 0.69 12.88 6.28 19.16 Tyees Men Early 0.23 2.94 0.38 0.5 11.66 6.47 18.13 Spartan 0.28 3.8 0.48 0.99 10.28 1.97 12.25

Distributed for comment only -- do not cite or quote

Table 4. Mineral Composition/Chemistry Data on Different Varieties of Pyrus malus.29 Vitamin Ascorbic Sodium Potassium Calcium Iron Phosphorus Magnesium A Acid Moisture Ash pH Variety (mg/100 (mg/100 (mg/100 (mg/100 (mg/100 g) (mg/100 g) (mg/100 (mg/100 (%) (%) Value g) g) g) g) g) g)

Amri 8.69 82.52 15.09 15.31 8.11 5.63 0.94 3.44 80.5 1.29 4.07 Bluish Golden 8.31 75.19 8.34 16.29 14.88 7.47 0.89 3.78 83.39 3.07 3.78 Bonza 14.27 127.34 4.2 13.99 10.97 4.2 0.91 3.91 71.46 1.84 3.76 Discovery 11.01 109.36 16.34 20.79 14.62 7.23 0.95 4.2 75.91 1.14 3.87 Elster 10.14 96.94 11.25 19.89 11.7 7.09 0.99 5.93 79.72 1.51 3.75 Florina 12.8 114.63 16.98 17.85 12.19 2.46 0.8 4.28 75.4 1.71 4.19 Gala 11.4 82.27 13.31 21.99 15.58 7.6 0.79 3.22 81 1.29 4.22 Gloster 69 11.84 93.76 16.75 17.04 14.82 10.86 0.92 3.29 80.26 1.53 3.79 Golden Delicious 7.75 63.43 8.2 15.51 13.69 3.88 0.84 3.51 84.49 1.06 4.03 Granny Smith 10.37 73.82 13.1 14.55 11.73 10.73 0.87 3.91 82.71 1.73 3.94 Green Sleeves 9.12 75.19 13.59 7.87 4.44 10.92 0.92 2.74 81.75 1.61 3.79 Ida Red 7.7 65.76 9.44 9.06 6.26 3.08 0.94 3.32 84.6 1.63 3.78 Kaja 9.79 102.87 15.25 14.74 10.26 1.57 0.96 3.33 83.67 1.82 3.74 Kids Orange 8.2 72.98 11.97 16.41 14.4 1.25 0.99 7.38 83 1.26 4.44 Mushadi 11.2 64.89 9.58 13.28 7.12 7.08 0.9 5.33 84.25 2.01 4.08 Red Delicious 12.49 73.01 4.56 16.11 7.72 4.46 0.97 4.32 82.15 1.69 4.59 Samootree 10.49 84.69 16.66 13.74 11.63 1.96 0.79 3.95 78.12 1.45 3.93 Summer Red 6.4 79.72 12.24 17.21 10.74 5.16 0.88 1.38 87.1 2.6 3.89 Tyees Men Early 9.01 76.44 13.95 17.33 9.07 1.8 0.76 3.66 81.5 1.63 3.57 Spartan 10.37 73.82 13.14 14.55 11.73 10.73 0.81 2.99 82.71 1.73 4.07 Mean Values 10.06 84.43 12.19 15.67 11.08 5.78 0.89 3.8 Median Values 81.95 1.63 3.91

Distributed for comment only -- do not cite or quote

33,34 Table 5. Composition Data on Pyrus Malus (Apple) Fiber.

Composition Impurities Water (1.5%) Ash (1.3%) Not dioxin-like polychlorinated biphenyls (PCBs, 3 µg/kg; not detected) Fat (1.9%) Lead (0.36 mg/kg) Saturated fatty acids (0.3%, calculated as triglyceride) Cadmium (< 0.01 mg/kg) Monounsaturated fatty acids (0.6%, calculated as triglyceride) Mercury (< 0.30 mg/kg) Polyunsaturated fatty acids (1%, calculated as triglyceride) Total protein content (4.5%) Arsenic (0.02 mg/kg) Carbohydrate (25.5%, calculated) Ochratoxin A (< 0.5 µg/kg) Sucrose (enzymatical, 1609 mg/100 g) Patulin (< 20 µg/kg) Glucose (enzymatical, 2989 mg/100 g) Aflatoxin B1 (< 0.7 µg/kg) Fructose (enzymatical, 7406 mg/100 g) Aflatoxin B2 (< 0.2 µg/kg) Sodium (< 50 mg/kg) Aflatoxin G1 (< 0.7 µg/kg) Dietary fibers (65.3 g/100 g) Aflatoxin G2 (< 0.2 µg/kg) Total aflatoxin (B1, B2, G1, and G2, not detected)

Nitrite (as Na-nitrite, <3 mg/kg)

Nitrate (as Na-nitrate, 29 mg/kg)

Total nitrite/nitrate (as NaNO2, 23.5 mg/kg)

Total nitrite/nitrate (as NaNO3, 29 mg/kg)

Pesticide Residues

Boscalid (0.164 mg/kg)

Chlorantraniliprole (0.024 mg/kg)

Chlorpyrifos (0.072 mg/kg)

Cyprodinil (0.032 mg/kg)

Etofenprox (0.020 mg/kg)

Fenoxycarb (0.032 mg/kg)

Fludioxonil (0.028 mg/kg)

Hexythiazox (0.020 mg/kg)

Indoxacarb (0.020 mg/kg)

Methoxyfenozide (0.052 mg/kg)

Myclobutanil (0.028 mg/kg)

Pendimethalin (0.020 mg/kg)

Phosalon (0.020 mg/kg)

Primicarb (0.032 mg/kg)

Propargite (0.080 mg/kg)

Pyraclostrobin (0.116 mg/kg)

Tebufenpyrad (0.024 mg/kg)

Trifloxystrobin (0.160 mg/kg)

Captan (0.628 mg/kg)

Iprodion (0.036 mg/kg)

Distributed for comment only -- do not cite or quote

Table 6. Fatty Acid Composition Data on Apple Seed Extract.45 Fatty Acids % Composition Butyl linoleate 1.50% 2-Dodecenal < 0.2% Deca-2,4-dienal 0.49% Ethyl linoleate 4.31% Ethyl oleate < 0.2% Ethyl palmitate 0.56% Ethyl stearate < 0.2% Hexyl hexanoate 0.54% Hexyl linoleate 3.30% Hexyl octanoate 0.49% Hexyl palmitate 0.61% Linoleic acid 51.15% Linolenic acid 5.60% Methyl docosanoate 0.72% Methyl eicosanoate 2.18% Methyl eicosenoate 1.05% Methyl heneeicosenoate < 0.2% Methyl heptadecanoate 0.28% Methyl linoleate 37.71% Methyl linolenate 5.60% Methyl nonodecanoate < 0.2% Methyl nonanoate < 0.2% Methyl octanoate < 0.2% Methyl oleate 4.12% Methyl palmitate 9.93% Methyl palmitoleate < 0.2% Methyl pentadecanoate < 0.2% Methyl pentanoate < 0.2% Methyl stearate 4.33% Methyl tetracosanoate < 0.2% Methyl tricosanoate < 0.2% Myristyl myristate < 0.2% Nonacosane 3.59% Oleic acid 4.12% Palmitic acid 10.49% Squalene 3.40% Stearic acid 4.33% Tricosene 4.29% Total fatty acids 80.91%

Distributed for comment only -- do not cite or quote

Table 7. Composition Data on Pyrus Malus (Apple) Seed Oil.26 Seed Oil and Fatty Acids Unsaponifiables Hexadecanoic Acid, Ethyl Ester: 6.6 ± Seed Oil: 28.9 ± 0.9% 0.3% Palmitic Acid (C16:0): 6.1 ± 0.4% Phytol: 0.6 ± 1.1% Palmitoleic Acid (C16:1): 0.2% Ethyl Oleate: 34.6 ± 3.1% Heptadecanoic Acid (C17:0): Not detected 9-Hexadecenal: 0.7 ± 0.4% Stearic Acid (C18:0): 2.0 ± 0.4% 3-Eicosene: 0.8 ± 0.2% Octadecanoic Acid, Ethyl Ester: 2.9 ± Oleic Acid (C18:1): 38.7 ± 1.7% 1.1% Linoleic Acid (C18:2): 49.6 ± 2.2% 1-Docasene: 2.4 ± 0.3% Linolenic Acid (C18:3): 0.4% Docosane: 0.9 ± 0.5% Eicosanoic Acid (C20:0): 0.9 ± 0.1% 1-Hexacosene: 1.2 ± 1.2% 11-Eicosanoic Acid (C20:1): 0.6% Octacosane: 0.8 ± 0.4% Docosanoic Acid (C22:0): 0.7% Squalene: 5.8 ± 0.8% Nonacosane: 0.9 ± 0.3%

β-Tocopherol: 1.7 ± 0.5%

α-Tocopherol: 6.1 ± 0.6%

Campesterol: 0.7 ± 0.2%

Avenasterol: 0.6 ± 0.1%

β-Sitosterol: 13.6 ± 1.4%

9,19-Cyclolanost-24-en-3-ol: 3.6 ± 1.1%

Stigmast-4-en-3-one: 3.8 ± 0.5%

Distributed for comment only -- do not cite or quote

Table 8. Current Frequency and Concentration of Use According to Duration and Type of Exposure.2,46 Pyrus Malus (Apple) Fruit Malus Domestica (Apple) Fruit Extract Extract Pyrus Malus (Apple) Fruit # of # of Uses Conc. (%) # of Uses Conc. (%) Uses Conc. (%) Totals/Conc. Range 125 0.000002-1 382 0.8 NR 0.00005-0.03 Duration of Use Leave-On 116 0.000002-1 283 NR NR 0.00005 Rinse off 9 0.000075-0.75 98 0.8 NR 0.0018-0.03 Diluted for (bath) Use NR 0.1 1 NR NR NR Exposure Type Eye Area 4 0.018-0.29 28 NR NR NR Incidental Ingestion 3 0.001-0.0036 6 NR NR NR Incidental Inhalation- Sprays 106 0.00083-0.1 1 NR NR NR Incidental Inhalation- Powders 106 0.00003-0.12** 1 NR NR 0.00005 Dermal Contact 121 0.00003-1 90 0.8 NR 0.00005 Deodorant (underarm) 1* NR NR NR NR NR Hair - Non-Coloring NR 0.0002-0.75 73 NR NR 0.0018-0.03 Hair-Coloring NR 0.1 NR NR NR NR Nail NR 0.000002-0.00007 1 NR NR NR Mucous Membrane 7 0.001-0.1 19 NR NR NR Baby Products NR NR NR NR NR NR Pyrus Malus (Apple) Fruit Water Pyrus Malus (Apple) Juice Malus Domestica (Apple) Juice # of # of Uses Conc. (%) # of Uses Conc. (%) Uses Conc. (%) Totals/Conc. Range NR 0.008-9 NR 0.0003-0.7 11 NR Duration of Use Leave-On NR 0.008-9 NR 0.07 2 NR Rinse off NR 0.008-0.5 NR 0.0003-0.7 9 NR Diluted for (bath) Use NR NR NR 0.007 NR NR Exposure Type Eye Area NR 0.2 NR NR 2 NR Incidental Ingestion NR 0.008 NR NR NR NR Incidental Inhalation- Sprays NR NR NR NR NR NR Incidental Inhalation- Powders NR 0.76-9** NR 0.07** NR NR Dermal Contact NR 0.008-9 NR 0.0025-0.07 1 NR Deodorant (underarm) NR NR NR NR NR NR Hair - Non-Coloring NR NR NR 0.0003-0.07 7 NR Hair-Coloring NR NR NR NR 1 NR Nail NR NR NR NR NR NR Mucous Membrane NR 0.008 NR 0.0025-0.007 1 NR Baby Products NR NR NR NR NR NR Pyrus Malus (Apple) Peel Pyrus Malus (Apple) Root Pyrus Malus (Apple) Seed Powder Extract Extract # of # of Uses Conc. (%) # of Uses Conc. (%) Uses Conc. (%) Totals/Conc. Range 2 NR 4 NR 16 0.001-0.6 Duration of Use Leave-On 1 NR 1 NR 13 0.001-0.6 Rinse off 1 NR NR NR 3 NR Diluted for (bath) Use NR NR 3 NR NR NR Exposure Type Eye Area NR NR NR NR 2 0.6 Incidental Ingestion NR NR NR NR 2 0.001 Incidental Inhalation- Sprays 1*** NR 2*** NR 2*** NR Incidental Inhalation- Powders 1*** NR 2*** NR 2*** NR Dermal Contact 1 NR 4 NR 9 0.0015-0.6 Deodorant (underarm) NR NR NR NR NR NR Hair - Non-Coloring NR NR NR NR NR NR Hair-Coloring 1 NR NR NR NR NR Nail NR NR NR NR NR NR Mucous Membrane NR NR 2 NR 2 0.001 Baby Products NR NR NR NR NR NR

Distributed for comment only -- do not cite or quote

Table 8. Current Frequency and Concentration of Use According to Duration and Type of Exposure.2,46 Pyrus Malus (Apple) Flower Pyrus Malus (Apple) Pectin Extract Extract Pyrus Malus (Apple) Seed Oil # of # of Uses Conc. (%) # of Uses Conc. (%) Uses Conc. (%) Totals/Conc. Range NR 0.0005 9 NR NR NR Duration of Use Leave-On NR 0.0005 5 NR NR 0.001 Rinse off NR NR 4 NR NR NR Diluted for (bath) Use NR NR NR NR NR NR Exposure Type Eye Area NR NR NR NR NR NR Incidental Ingestion NR NR NR NR NR NR Incidental Inhalation- Sprays NR 0.0005 3* NR NR NR Incidental Inhalation- Powders NR NR 1 NR NR 0.001 Dermal Contact NR 0.0005 5 NR NR 0.001 Deodorant (underarm) NR NR NR NR NR NR Hair - Non-Coloring NR NR 1 NR NR NR Hair-Coloring NR NR NR NR NR NR Nail NR NR NR NR NR NR Mucous Membrane NR NR NR NR 10 NR Baby Products NR NR 1 NR NR NR Malus Domestica (Apple) Seed Malus Domestica (Apple) Fruit Oil Water Malus Domestica (Apple) Oil # of # of Uses Conc. (%) # of Uses Conc. (%) Uses Conc. (%) Totals/Conc. Range 12 NR 113 2 1 NR Duration of Use Leave-On 12 NR 106 2 1 NR Rinse off NR NR 7 NR NR NR Diluted for (bath) Use NR NR NR NR NR NR Exposure Type Eye Area NR NR 1 NR 1 NR Incidental Ingestion 10 NR NR NR NR NR Incidental Inhalation- Sprays 1*** NR 2 2 NR NR Incidental Inhalation- Powders 1*** NR 2 NR NR NR Dermal Contact 2 NR 105 2 1 NR Deodorant (underarm) NR NR NR NR NR NR Hair - Non-Coloring NR NR NR NR NR NR Hair-Coloring NR NR NR NR NR NR Nail NR NR NR NR NR NR Mucous Membrane 10 NR 2 NR NR NR Baby Products NR NR NR NR NR NR Malus Domestica (Apple) Stem Malus Domestica (Apple) Fruit Malus Domestica (Apple) Callus Extract Cell Culture Extract Extract # of # of Uses Conc. (%) # of Uses Conc. (%) Uses Conc. (%) Totals/Conc. Range 1 NR 29 0.00001-1 NR NR Duration of Use Leave-On 1 NR 27 0.00001-1 NR NR Rinse off NR NR 2 0.00001-0.2 NR NR Diluted for (bath) Use NR NR NR NR NR NR Exposure Type Eye Area NR NR 9 0.009-0.089 NR NR Incidental Ingestion NR NR NR NR NR NR Incidental Inhalation- Sprays 1*** NR 9*** NR NR NR Incidental Inhalation- Powders 1*** NR 9*** 0.0001-1 NR NR Dermal Contact 1 NR 28 0.00001-1 NR NR Deodorant (underarm) NR NR NR NS NR NR Hair - Non-Coloring NR NR NR 0.18-0.2 NR NR Hair-Coloring NR NR NR NR NR NR Nail NR NR NR NR NR NR Mucous Membrane NR NR NR NR NR NR Baby Products NR NR NR NR NR NR

Distributed for comment only -- do not cite or quote

Table 8. Current Frequency and Concentration of Use According to Duration and Type of Exposure.2,46 Malus Domestica (Apple) Fiber # of Uses Conc. (%) Totals/Conc. Range 3 NR Duration of Use Leave-On 3 NR Rinse off NR NR Diluted for (bath) Use NR NR Exposure Type Eye Area NR NR Incidental Ingestion NR NR Incidental Inhalation- Sprays 3*** NR Incidental Inhalation- Powders 3*** NR Dermal Contact 3 NR Deodorant (underarm) NR NR Hair - Non-Coloring NR NR Hair-Coloring NR NR Nail NR NR Mucous Membrane NR NR Baby Products NR NR NR = Not Reported; Totals = Rinse-off + Leave-on Product Uses. *It is possible that these products may be sprays, but it is not specified whether the reported uses are sprays. **It is possible that these products may be powders, but it is not specified whether the reported uses are powders. ***Not specified whether a powder or spray, so this information is captured for both categories of incidental inhalation. 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.

Distributed for comment only -- do not cite or quote

References

1. Nikitakis, J. and Breslawec H. P. International Cosmetic Ingredient Dictionary and Handbook. 14 ed. Washington, DC: Personal Care Products Council, 2014.

2. Food and Drug Administration (FDA). Information supplied to FDA by industry as part of the VCRP FDA database. 2015. Washington, D.C.: FDA.

3. Food and Drug Administration (FDA). Rulemaking history for OTC skin bleaching products. http://www.fda.gov/Drugs/DevelopmentApprovalProcess/DevelopmentResources/Over-the- CounterOTCDrugs/StatusofOTCRulemakings/ucm072117.htm. Last Updated 2015.

4. Personal Care Products Council. Pyrus malus-derived Ingredients. Communication with the Personal Care Products Council on 10-23-2014. 2014.

5. United States Department of Agriculture. Germplasm Resources Information Network. Malus domestica. http://www.ars-grin.gov4/cgi-bin/npgs/html/tax_search.pl. Last Updated 2015.

6. Grau Aromatics GmbH & Co.KG. Apple Extract HS 1806 AT (propylene glycol extract). Unpublished data submitted by the Personal Care Products Council on 10-17-2014. 2013. pp.1-2.

7. Grau Aromatics GmbH & Co.KG. Apple Extract HS (glycerin/water extract). Unpublished data submitted by the Personal Care Products Council on 10-17-2014. 2013. pp.1-2.

8. Burnett, C. Heldreth B. Bergfeld W. F. Belsito D. V. Hill R. A. Klaassen C. D. Liebler D. C. Marks J. G. Jr. Shank R. C. Slaga T. J. and Snyder P. W. Final report. Plant-derived fatty acid oils as used in cosmetics. 2011. pp.1-100. Washington, D.C.: Cosmetic Ingredient Review.

9. Takahashi, T. Yokoo Y. Inoue T. and Ishii A. Toxicological studies on procyanidin B-2 for external application as a hair growing agent. Food and Chemical Toxicology. 1999;37:545-552.

10. MDidea Extracts Professional. Apple polyphenol: toxicology and safety. www.MDidea.com. Last Updated 2013. Date Accessed 11-14-2014.

11. Personal Care Products Council. Method of manufacture and composition: Pyrus malus (apple) fiber. Unpublished data submitted by the personal Care Products Council on 11-10-2014. 2014. pp.1

12. Jayakumar, R. and Kanthimathi M. S. Inhibitory effects of fruit extracts on nitric oxide-induced proliferation in MCF-7 cells. Food Chemistry. 2011;126:956-960.

13. Personal Care Products Council. Method of manufacture and composition: Pyrus Malus (Apple) Fruit Extract. Unpublished data submitted by the Personal Care Products Council on 12-3-2014. 2014. pp.1

14. Active Concepts. Manufacturing flow chart 10392-ABS apple extract BG (pyrus malus (apple) fruit extract). Unpublished data submitted by the Personal Care Products Council on 3-30-2015. 2015. pp.1

15. Active Concepts. Manufacturing flow chart. 10307-ABS apple extract PG (pyrus malus (apple) fruit extract). Unpublished data submitted by the Personal Care Products Council on 3-30-2015. 2015. pp.1

16. Anonymous. Summary of information on pyrus malus (apple) fruit extract prepared for the CIR safety assessment of pyrus malus (apple)-derived ingredients as used in cosmetics. Unpublished data submitted by the Personal Care Products Council on 5-27-2015. 2015. pp.1-2.

17. Shoji, T. Akazome Y. Kanda T. and ikeda M. The toxicology and safety of apple polyphenol extract. Food Chem.Toxicol. 2004;42(6):959-967.

18. FruitOFood. Dehydration process for the preparation of apple fruit water. Unpublished data submitted by the Personal Care Products Council on 11-3-2014. 2014. pp.1-2. Distributed for comment only -- do not cite or quote

19. Vieira, R. G. Prestes R. A. Denardi F. Nogueira A. and Wosiacki G. Chemical pattern of Brazilian apples. A chemometric approach based on the Fuji and Gala varieties. Cienc.Tecnol.Aliment., Campinas. 2011;31(2):418-426.

20. Mathur, A. Sharma V. Bhardwaj A. Yousuf S. Verma S. K. Singh S. K. and Dua V. K. Pectin content as an index for screening different varieties of apple (Pyrus Malus L.) of Kashmir (J & K) on the basis of antimicrobial activity. J.Chem.Pharm.Res. 2011;3(2):886-891.

21. Vieira, F. G. K. Borges G. D. C. Copetti C. Amboni R. D. D. C. Denardi F. and Fett R. Physico-chemical and antioxidant propereties of six apple cultivars (Malus domestica Borkh) grown in southern Brazil. Scientia Horticulture. 2009;122:421-425.

22. Personal Care Products Council. Information on pyrus malus (apple) root bark powder. Unpublished data submitted by the Personal Care Products Council on 10-23-2014. 2014. pp.1-2.

23. Chakaravarti, D. Chakaravarti N. N. and Chakaravarti A. Chemical investigation of Pyrus malus. Journal of the Indian Chemical Society. 1965;42(8):573-574.

24. Barthe, P. and Bulard C. Bound and free abscisic acid levels in dormant and after-ripened embryos of Pyrus malus L. cv Golden delicious. Zeitschrift fuer Pflanzenphysiologie. 1978;90(3):201-208.

25. Anonymous. Summary of information on pyrus malus (apple) seed extract. Unpublished data submitted by the Personal Care Products Council on 5-28-2015. 2015. pp.1-3.

26. Arain, S. Sherazi S. t. H. Bhanger M. I. Memon N. Mahesar S. A. and Rajput M. T. Prospects of fatty acid profile and bioactive composition from lipid seeds for the discrimination of apple varieties with the application of chemometrics. Grasas Y Aceites. 2012;63(2):175-183.

27. Battaglini, M. B. Characterization of the sugar composition of nectars of some species and cultivars of fruit trees. Apicultural Abstracts. 1974;25(2):71-76.

28. Omata, A. Yomogida K. and Nakamura S. Volatile components of apple flowers. Flavour and Fragrance Journal. 1990;5(1):19-22.

29. Aziz, M. Jadoon S. and Sh Z. Apples pulp (Pyrus malus) nutritional profiling evaluation of various varieties of balochistan. Pakistan Journal of Nutrition. 2014;12(3):239-243.

30. Sharma, J. N. and Seshadri T. R. Survey of anthocyanins from Indian sources: Part II. Journal of Scientific and Industrial Research. 1955;14B:211-214.

31. Machiex, J. J. Concentration changes in esters of p-coumaric and caffeic acids in the fruit of Pyrus malus during growth. Physiologie Vegetale. 1974;12(1):25-33.

32. Datta, S. C. Free amino acids of indian fruits. Bulletin of the Botanical Society of Bengal. 1963;17(1-2):8-9.

33. Institut Dr.Erdmann. Examination report: Apple fiber. Unpublished data submitted by the Personal Care Products Council on 11-10-2014. 2009. pp.1-2.

34. Erdmann Analytics. Examination report: Apple fiber. Unpublished data submitted by the Personal Care Products Council on 11-10-2014. 2014. pp.1-4.

35. Thompson-Witrick, K. A. Goodrich K. M. Neilson A. P. et al. Characterization of the polyphenol composition of 20 cultivars of cider, processing, and dessert apples (Malus x domestica Borkh.) grown in Virginia. J.Agric.Food Chem. 2014;62(41):10181-10191.

36. Kviklys, D. Liaudanskas M. Janulis V. Viskelis P. Rubinskiene M. Lanauskas J. and Uselis N. Rootstock genotype determines phenol content in apple fruits. Plant Soil Environ. 2014;60(5):234-240.

37. Thomas, W. The seat of formation of amino acides in Pyrus Malus L. Science. 1927;66(1):115-116. Distributed for comment only -- do not cite or quote

38. Strain, H. H. Sources of d-Sorbitol. Journal of the American Chemical Society. 1937;59:2264-2266.

39. Walia, M. Mann T. S. Kumar D. Agnihotri and Singh B. Chemical composition and in vitro cytotoxic activity of essential oil of leaves of Malus domestica growing in Western Himalaya (India). Evidence-based Complimentary and Alternative Medicine. 2012;2012:649-727.

40. Liaaudanskas, M. Viškelis P. Rudonis R. Kviklys D. Uselis N. and Janulis V. Phenolic composition and antioxidant activity of Malus domestica leaves. The Scientific World Journal. 2014; http://www.hindawi.com/journals/tswj/2014/306217.

41. Walia, M. Kumar S. and Agnihotri V. K. UPLC-PDA quantificatio of chemical constituents of two different varieties (golden and royal) of apple leaves and their antioxidant activity. April 27. doi: 10.100/jsfa.7239. [Epub ahead of print]. J.Sci.Food Agric. 2015;

42. Manzoor, M. Anwar F. Saari N. and Ashraf M. Variations of antioxidant characteristics and mineral contents in pulp and peel of different apple (Malus domestica Borkh.) cultivars from Pakistan. Molecules. 2012;17:390- 407.

43. Dennis, F. G. Jr. Martin G. C. Gaskin P. and MacMillan J. Gibberellins in mature apple seeds - Contaminants? Planta. 1980;147:376-377.

44. Bianco, J. Lassechere S. and Bulard C. Gibberellins in dormant embryos of Pyrus malus L. cv Golden delicious. J.Plant Physiol. 1984;116:185-188.

45. Lu, Y. and Yeap Foo L. Constitution of some chemical components of apple seed. Food Chem. 1998;61:29-33.

46. Personal Care Products Council. Concentration of use by FDA product category: Pyrus malus (apple)- and Malus domestica (apple)-derived ingredients. Unpublished data submitted by the Personal Care Products Council on 10-9-2015. 2015. pp.1

47. Rothe H, Fautz R, Gerber E, Neumann L, Rettinger K, Schuh W, and Gronewold C. Special aspects of cosmetic spray safety evaluations: Principles on inhalation risk assessment. Toxicol Lett. 2011;205(2):97-104. PM:21669261.

48. Bremmer HJ, Prud'homme de Lodder LCH, and van Engelen JGM. Cosmetics Fact Sheet: To assess the risks for the consumer; Updated version for ConsExpo 4. 20200. http://www.rivm.nl/bibliotheek/rapporten/320104001.pdf. Date Accessed 8-24-2011. Report No. RIVM 320104001/2006. pp. 1-77.

49. Rothe H. Special aspects of cosmetic spray evaluation. Unpublished information presented to the 26 September CIR Expert Panel. Washington D.C. 2011.

50. Johnsen MA. The Influence of Particle Size. Spray Technology and Marketing. 2004;14(11):24- 27. http://www.spraytechnology.com/index.mv?screen=backissues.

51. Food and Drug Administration (FDA). Food for human consumption. Specific nutrition labeling requirements and guidelines. 21 CFR 101.44. 2014.

52. Wruss, J. Lanzerstorfer P. Heumer S. et al. Differences in pharmacokinetics of apple polyphenols after standardized oral consumption of unprocessed apple juice. Nutr.J. 2015; http://www.nutritionj.com/content/14/1/32.

53. Gerhauser, C. Cancer chemopreventive potential of apples, apple juice, and apple components. Planta Med. 2008;74(13):1608-1624.

54. Active Concepts. Dermal and ocular irritation tests. ABS apple extract BG (contains 20% pyrus malus (apple) fruit extract. Unpublished data submitted by the Personal Care Products Council on 3-30-2015. 2015. pp.1-4.

55. MatTek Corporation. Skin irritation (ECVAM validated). The model and method. http://www.mattek.com/epiderm/applications/ecvam-skin-irritation-test. Last Updated 2015. Distributed for comment only -- do not cite or quote

56. Active Concepts. Dermal and ocular irritation tests. ABS Apple Extract PG (contains 20% pyrus malus (apple) fruit extract). Unpublished data submitted by the Personal Care Products Council on 3-30-2015. 2015. pp.1-4.

57. RCTS, Inc. Clinical safety evaluation repeated insult patch test of a face and neck product containing 8.7835% pyrus malus (apple) fruit water. Unpublished data submitted by the Personal Care Products Council on 5-22- 2015. 2015. pp.1-7.

58. Jung, E. Lee J. Huh S. et al. Phloridzin-induced melanogenesis is mediated by the cAMP signaling pathway. Food Chem Toxicol. 2009;47(10):2436-2440.

Distributed for comment only -- do not cite or quote

2015 FDA VCRP Data Pyrus Malus (Apple) Fruit Extract 03D - Eye Lotion 2 03G - Other Eye Makeup Preparations 2 04A - Cologne and Toilet waters 1 07C - Foundations 1 07E - Lipstick 3 10A - Bath Soaps and Detergents 3 10B - Deodorants (underarm) 1 10E - Other Personal Cleanliness Products 1 12A - Cleansing 3 12C - Face and Neck (exc shave) 6 12D - Body and Hand (exc shave) 9 12F - Moisturizing 87 12G - Night 2 12H - Paste Masks (mud packs) 2 12I - Skin Fresheners 2 Total 125

Pyrus Malus (Apple) Pectin Extract 01A - Baby Shampoos 1 07B - Face Powders 1 12A - Cleansing 2 12C - Face and Neck (exc shave) 2 12F - Moisturizing 1 12H - Paste Masks (mud packs) 1 12I - Skin Fresheners 1 Total 9

Pyrus Malus (Apple) Peel Powder 05F - Shampoos (non-coloring) 1 12D - Body and Hand (exc shave) 1 Total 2

Pyrus Malus (Apple) Root Extract 02D - Other Bath Preparations 2 12C - Face and Neck (exc shave) 1 12F - Moisturizing 1 Total 4

Pyrus Malus (Apple) Seed Extract 03G - Other Eye Makeup Preparations 2 05G - Tonics, Dressings, and Other Hair Grooming Aids 1 07E - Lipstick 2 07I - Other Makeup Preparations 1 12A - Cleansing 3 12C - Face and Neck (exc shave) 2 Distributed for comment only -- do not cite or quote

12F - Moisturizing 1 12G - Night 1 13B - Indoor Tanning Preparations 3 Total 16

Malus Domestica (Apple) Fruit Extract 02D - Other Bath Preparations 1 03C - Eye Shadow 2 03D - Eye Lotion 15 03F - Mascara 2 03G - Other Eye Makeup Preparations 9 04E - Other Fragrance Preparation 1 05A - Hair Conditioner 31 05F - Shampoos (non-coloring) 27 05G - Tonics, Dressings, and Other Hair Grooming Aids 1 05I - Other Hair Preparations 15 07A - Blushers (all types) 1 07B - Face Powders 1 07C - Foundations 2 07E - Lipstick 6 08B - Cuticle Softeners 1 10A - Bath Soaps and Detergents 4 10E - Other Personal Cleanliness Products 8 11A - Aftershave Lotion 1 12A - Cleansing 26 12C - Face and Neck (exc shave) 26 12D - Body and Hand (exc shave) 26 12E - Foot Powders and Sprays 1 12F - Moisturizing 93 12G - Night 5 12H - Paste Masks (mud packs) 2 12I - Skin Fresheners 3 12J - Other Skin Care Preps 17 13B - Indoor Tanning Preparations 2 Total 329

Malus Domestica (Apple) Fruit Water 03D - Eye Lotion 1 04A - Cologne and Toilet waters 1 07B - Face Powders 2 07I - Other Makeup Preparations 1 10A - Bath Soaps and Detergents 2 12A - Cleansing 3 12C - Face and Neck (exc shave) 4 12F - Moisturizing 95 12H - Paste Masks (mud packs) 2 12J - Other Skin Care Preps 1 Distributed for comment only -- do not cite or quote

13B - Indoor Tanning Preparations 1 Total 113

Malus Domestica (Apple) Seed Oil 07E - Lipstick 10 12D - Body and Hand (exc shave) 1 12F - Moisturizing 1 Total 12

Malus Domestica (Apple) Fiber 12C - Face and Neck (exc shave) 3 Total 3

Malus Domestica (Apple) Juice 03F - Mascara 2 05A - Hair Conditioner 1 05F - Shampoos (non-coloring) 6 06D - Hair Shampoos (coloring) 1 10A - Bath Soaps and Detergents 1 Total 11

Malus Domestica (Apple) Oil 03D - Eye Lotion 1 Total 1

Malus Domestica (Apple) Stem Extract 12D - Body and Hand (exc shave) 1 Total 1

Malus Domestica (Apple) Fruit Cell Culture Extract 03D - Eye Lotion 5 03F - Mascara 1 03G - Other Eye Makeup Preparations 3 07C - Foundations 1 12A - Cleansing 1 12C - Face and Neck (exc shave) 8 12D - Body and Hand (exc shave) 1 12F - Moisturizing 4 12G - Night 3 12H - Paste Masks (mud packs) 1 12J - Other Skin Care Preps 1 Total 29 Distributed for comment only -- do not cite or quote

Memorandum

TO: Lillian Gill, D.P.A. Director - COSMETIC INGREDIENT REVIEW (CIR)

FROM: Beth A. Lange, Ph.D. Industry Liaison to the CIR Expert Panel

DATE: October 9, 2015

SUBJECT: Concentration of Use Information Apple-Derived Ingredients

Updated use information from 2014 concentration of use survey

New use information from 2015 survey Distributed for comment only -- do not cite or quote

Concentration of Use by FDA Product Category – Pyrus malus-Derived Ingredients*

(Note: Pyrus malus and Malus domestica are genus species names that are both used for apple)

Malus Domestica Fruit Extract Pyrus Malus (Apple) Pectin Extract Pyrus Malus (Apple) Fruit Extract Pyrus Malus (Apple) Peel Extract Pyrus Malus (Apple) Bark Extract Pyrus Malus (Apple) Peel Powder Pyrus Malus (Apple) Carpel Powder Pyrus Malus (Apple) Peel Wax Pyrus Malus (Apple) Fiber Pyrus Malus (Apple) Pulp Extract Pyrus Malus (Apple) Flower Extract Pyrus Malus (Apple) Root Bark Powder Pyrus Malus (Apple) Fruit Pyrus Malus (Apple) Root Extract Pyrus Malus (Apple) Fruit Water Pyrus Malus (Apple) Seed Extract Malus Domestica Fruit Water Pyrus Malus (Apple ) Seed Oil Pyrus Malus (Apple) Juice Pyrus Malus (Apple) Stem Extract Pyrus Malus (Apple) Leaf Extract Ingredient Product Category Maximum Concentration of Use Pyrus Malus (Apple) Fruit Extract Bubble baths 0.1% Pyrus Malus (Apple) Fruit Extract Other bath preparations 0.1% Pyrus Malus (Apple) Fruit Extract Eye shadow 0.29% Pyrus Malus (Apple) Fruit Extract Eye lotion 0.048-0.18% Pyrus Malus (Apple) Fruit Extract Other eye makeup preparations 0.018% Pyrus Malus (Apple) Fruit Extract Colognes and toilet water 0.00083% Pyrus Malus (Apple) Fruit Extract Perfume 0.1% Pyrus Malus (Apple) Fruit Extract Hair conditioner 0.0002-0.75% Pyrus Malus (Apple) Fruit Extract Shampoos (noncoloring) 0.0002-0.5% Pyrus Malus (Apple) Fruit Extract Tonics, dressings and other hair grooming 0.25% aids Pyrus Malus (Apple) Fruit Extract Other hair preparations (noncoloring) 0.75% Pyrus Malus (Apple) Fruit Extract Hair dyes and colors 0.1% Pyrus Malus (Apple) Fruit Extract Foundation 0.00009-0.0015% Pyrus Malus (Apple) Fruit Extract Lipstick 0.001-0.0036% Pyrus Malus (Apple) Fruit Extract Nail creams and lotions 0.000002% Pyrus Malus (Apple) Fruit Extract Nail polish and enamel 0.00007% Pyrus Malus (Apple) Fruit Extract Bath soaps and detergents 0.002-0.011% Pyrus Malus (Apple) Fruit Extract Aftershave lotions 0.0005-0.01% Pyrus Malus (Apple) Fruit Extract Preshave lotions 0.002% Pyrus Malus (Apple) Fruit Extract Skin cleansing (cold creams, cleansing 0.000075-0.25% lotions, liquids and pads) Pyrus Malus (Apple) Fruit Extract Face and neck products not spray 0.00003-1% Pyrus Malus (Apple) Fruit Extract Body and hand products not spray 0.053-0.12% spray 0.06-0.1% Pyrus Malus (Apple) Fruit Extract Moisturizing products Distributed for comment only -- do not cite or quote

not spray 0.00016% Pyrus Malus (Apple) Fruit Extract Night products Not spray 0.0006-0.015% Pyrus Malus (Apple) Fruit Extract Paste masks and mud packs 0.034-0.35% Pyrus Malus (Apple) Fruit Extract Skin fresheners 0.15% Pyrus Malus (Apple) Fruit Extract Other skin care preparations 0.00006% Pyrus Malus (Apple) Fruit Extract Suntan products not spray 0.00009% Pyrus Malus (Apple) Fruit Extract Other suntan preparations 0.0044% Pyrus Malus (Apple) Flower Body and hand products Extract spray 0.0005% Pyrus Malus (Apple) Fruit Powders (dusting and talcum) 0.00005% Pyrus Malus (Apple) Fruit Shampoos (noncoloring) 0.0018-0.03% Pyrus Malus (Apple) Fruit Water Eye lotion 0.2% Pyrus Malus (Apple) Fruit Water Lipstick 0.008% Pyrus Malus (Apple) Fruit Water Other makeup preparations 0.5% Pyrus Malus (Apple) Fruit Water Skin cleansing (cold creams, cleansing 0.008-0.1% lotions, liquids and pads) Pyrus Malus (Apple) Fruit Water Face and neck products not spray 0.75-9% Pyrus Malus (Apple) Fruit Water Body and hand products not spray 1% Pyrus Malus (Apple) Fruit Water Moisturizing products not spray 0.5-6% Pyrus Malus (Apple) Fruit Water Night products not spray 0.1% Pyrus Malus (Apple) Fruit Water Paste masks and mud packs 0.5% Pyrus Malus (Apple) Juice Bath oils, tablets and salts 0.007% Pyrus Malus (Apple) Juice Hair conditioners 0.0003% Pyrus Malus (Apple) Juice Rinses (noncoloring) 0.07% Pyrus Malus (Apple) Juice Shampoos (noncoloring) 0.005-0.07% Pyrus Malus (Apple) Juice Bath soaps and detergents 0.0025-0.003% Pyrus Malus (Apple) Juice Skin cleansing products (cold creams, 0.005% cleansing lotions, liquids and pads) Pyrus Malus (Apple) Juice Face and neck products not spray 0.07% Pyrus Malus (Apple) Seed Extract Eye lotion 0.6% Pyrus Malus (Apple) Seed Extract Foundation 0.0015% Pyrus Malus (Apple) Seed Extract Lipstick 0.001% Pyrus Malus (Apple) Seed Oil Face powder 0.001% Pyrus Malus (Apple) Seed Oil Makeup bases 0.001% *Ingredients in the title of the table but not in the table were included in the survey, but no uses were reported. Information collected in 2014 Table prepared July 30, 2014 Distributed for comment only -- do not cite or quote

Updated October 27, 2014: Pyrus Malus (Apple) Juice: added skin cleansing products; Pyrus Malus (Apple) Seed Extract: added eye lotion Updated September 23, 2015: Pyrus Malus Apple Fruit Extract: eye lotion high concentration increased from 0.06 to 0.18%; bath soaps and detergent lower concentration decreased form 0.01% to 0.002%; face and neck products high concentration increased from 0.12% to 1% Distributed for comment only -- do not cite or quote

Concentration of Use by FDA Product Category – Apple-Derived Ingredients*

Malus Domestica Callus Extract Malus Domestica Oil Malus Domestica Fruit Cell Culture Extract Malus Domestica Stem Extract Malus Domestica Fiber Malus Domestica Fruit Water Malus Domestica Juice Malus Domestica Fruit Extract Malus Domestica Seed Oil

Ingredient Product Category Maximum Concentration of Use Malus Domestica Fruit Cell Culture Eye lotion 0.009-0.089% Malus Domestica Fruit Cell Culture Mascara 0.089% Malus Domestica Fruit Cell Culture Hair conditioners 0.18-0.2% Malus Domestica Fruit Cell Culture Shampoos (noncoloring) ≤0.2% Malus Domestica Fruit Cell Culture Foundations 0.0001% Malus Domestica Fruit Cell Culture Skin cleansing (cold creams, cleansing 0.00001% lotions, liquids and pads) Malus Domestica Fruit Cell Culture Face and neck products Not spray 0.018-1% Malus Domestica Fruit Cell Culture Body and hand products Not spray 0.0001% Malus Domestica Fruit Cell Culture Moisturizing products Not spray 0.18-0.2% Malus Domestica Fruit Cell Culture Night products Not spray 0.089% Malus Domestica Fruit Cell Culture Skin fresheners 0.00001% Malus Domestica Fruit Cell Culture Other skin care preparations 0.18% Malus Domestica Fruit Extract Paste masks and mud packs 0.8% Malus Domestica Fruit Water Face and neck products Spray 2% *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 2015 Table prepared October 8, 2015 Distributed for comment only -- do not cite or quote

Distributed for comment only -- do not cite or quote