personal care | sun care

High Levels of Free Radicals in Suncare Products Induce Aestivalis in Sensitive Subjects

K. Jung*, U. Heinrich**, H. Tronnier**, M. Schnyder***, B. Herzog***, T. Herrling* PROOF READ VERSION ONLY abstract

he market segment of suncare products for protection of sensitive skin against is growing. Specifically, pro- Ttection against Acne aestivalis (“Mallorca-Acne”) is perceived as a strong claim for this product category. There is evidence that peroxide free radicals, deriving from certain formulation components, are involved in the induction of photo-toxic skin reactions. These UV-induced free radicals can be assessed by an ESR-based method called Potential (RP), by exposing formulations to UV radiation. The aim of this study was to find a correlation between the amount of UV inducible peroxides of products and the clinical symptoms of Acne aestivalis. Two sunscreen formulas (SPF 30) containing organic UV-filters only, with an identical protection in the UV B region and a comparable protection in the UV A spectral range, have been pro- duced. For both formulations the amount of UV-inducible free radicals was assessed by the RP (Radical Potential). In addition, both formulas were tested in a clinical study with subjects with a history of Acne aestivalis. Very different RP values (levels of UV-induced free radicals) and completely different skin reactivity have been obtained, where an excellent correlation between the in vitro-parameter (RP) and the clinical symptoms could be shown. To our knowledge these results for the first time provide direct evidence for a strong correlation between UV inducible free radicals of formulations and the potential to induce A. aesti- valis in sensitive subjects. Furthermore it offers a rationale for the development of low risk sunscreen formulations by selection of suitable filter systems combined with in situ ESR analysis.

Introduction

Cosmetic products may undergo oxidation, leading to rancid- formulation quality. Furthermore, the presence of hydroxyper- ity, discoloration, and destabilization. This oxidation process oxides and other peroxide free radicals is known to induce can be minimized by the control of raw materials quality, pres- photo-toxic/chemo-toxic skin reactions [1]. ervation, and appropriate storage times and conditions. Nev- Photo allergic dermatitis is affecting 1 to 2 % of the popula- ertheless, when applied on the skin, the product is exposed tion. One of the most common form of this UV-induced skin to enviro nmental factors as sunlight and oxygen, which can reaction is given by Acne aestivalis (also known as Mallorca easily and quickly start oxidation and rancidity processes. acne), a special kind of polymorphous light eruption. It is The rancidity process can be roughly divided into an initiation, a monomorphous eruption consisting of multiple, uniform, a propagation, and a termination step, with different radical red, papular lesions, reported to occur after sun exposure. species involved in each step. Acne aestivalis is a photo-induced severe skin reaction, be- UV radiation can lead to radical chain reactions inside the formulation. Some active ingredients inside the cosmetic formulation might trigger these radical chain reactions or might act as catalyzer:

• Some organic UV-filters;

• Photocatalytically active TiO2 (uncoated)

• Perfume components containing hydroperoxides

• Unsaturated lipids of natural oils

Although these free radicals should not be harmful for the consumers (they will not penetrate into the living lay- Fig. 1 Case report of Acne aestivalis (Source: Institute Dermatronnier, [4]) ers of the skin), they can lead to a deterioration of the

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longing to the terminus “polymorpheous light dermatosis” (PLD), with immune- and autoimmune co-involvement. It is often correlated to , since the affected skin areas are exposed to frequent sun irradiation, especially UV A ra- diation. There is evidence, that peroxide reactions are involved in the pathophysiologic mechanism. have been shown to ameliorate the symptoms, but the entire molecu- lar mechanism is still not understood. [2,3]. The aim of the present investigation was to find a correla- tion between the amount of UV-inducible free radicals inside a sunscreen formulation and the clinical symptoms of Acne aestivalis.

Fig. 2 Example of free radical generation in a cosmetic formulation RP Method (Radical Potential) (placebo) spiked with natural oils or perfume oils (PO)

A semistable spin probe PCA (2,2,5,5-tetramethyl pyrrolidine N-oxyl, 0.01 mM final concentration) was added to the di- Clinical Study luted cosmetic product. The samples are inserted in capillary quartz tubes, the concentration of the spin marker is moni- The test subjects, all being sensitive to Acne aestivalis, are tored by ESR spectroscopy before and after defined UV radia- irradiated using only UV A radiation, on 5 successive days. A tion doses. The PCA spin probe is photostable and resistant Solar Simulator (SOL 5, Hönle AG, Germany) was used. The to antioxidants, but it promptly reacts with the UV generated irradiation dosis was 20 J/cm2, far below the erythema thresh- free radicals inside the samples (mainly hydroxyl radicals, lipid old. The test formulations, as well as the positive and negative peroxides and lipidic radicals). The amount of UV generated control, are applied 10 minutes prior to the irradiation on the free radicals can be quantitatively detected from a calibra- décolleté area. The skin reactions are evaluated directly after tion curve and the results are expressed in percentage of UV- each irradiation and again after 24 hours, prior to the next inducible free radicals. irradiation dosis. The UV irradiation of the samples was performed with a UV A market sunscreen gel (SPF 30) served as a negative control, solar simulator 300 W Oriel (Newport). The irradiances as in- which has never provoked a skin reaction in sensitive subjects. tegrated value over the spectral ranges were E (UVB=280- A sunscreen formulation served as a positive control, which 320) = 23.5 W/m2 and E (UVA=320-400 nm) = 180 W/m2. provokes Acne aestivalis in disposed subjects. Increase of UV doses was achieved by increasing irradiation times. The emitting Intensity is controlled before each measurement. Results A cut-off filter 320 nm (ITOS W-NG-320, Schott) was used to obtain only UV A radiation. An example of RP data is given in Fig. 2. A cosmetic formula- ESR measurements were performed with a high sensitive X- tion was spiked with natural oils or perfume oils (PO). The band bench top Electron Spin Resonance Spectrometer Mi- Almond oil showed higher radical generation compared to niScope MS300, supplied from Magnettech GmbH Berlin, Olive oil, probably due to differences in the fatty acid compo- Germany [5]. sition and the oxidation grade. The two perfume oils showed

AD

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very different reactions: PO “A” showed very low radical gen- The two sunscreens 12-1 and 12-2 were tested regarding eration (RP = 6.6 %), whereas PO “B” showed a strong radi- their capacity to induce Acne aestivalis in sensitive subjects: cal generation under UV (RP = 63.9 %). Sensorial properties had not changed. The added perfume oils differ with respect Positive Control to terpene composition, oxidation state, and especially in the content of hydroperoxide-terpenes, what is most likely the The most intensive reactions were observed after application reason for their different behaviours. of the positive control. This internal standard serves as a con- Two sunscreen formulations have been produced, both hav- ing an SPF of 30 and the same UVA/UVB ratio. The UV-filter composition differed (see Tab. 1). % of induced free % of induced free Formulation 12-1 contained only photostable UV-filters, where- Product radicals UVAB radicals UVA as formulation 12-2 contained also photo-unstable filters. The Radical Potential (RP) of both formulations was detected, using 12-1 0 ± 0,7 0 ± 0,1 the full spectrum UV light (280-400 nm), and the UV A region of the spectrum (320-400 nm) only, since Acne aestivalis is report- ed to be induced mainly by UV A radiation. Formulation 12-1 12-2 45,3 ± 1.0 21,9 ± 0.2 did not induce free radicals, whereas formulation 12-2 showed a high radical generation, both under full-spectrum UV and UV A Tab. 2 Relative amount of free radicals. N = 2 radiation (45.3 % and 21.9 %, respectively). (see Tab. 2).

Phase INCI 12-1 12-2 Sucrose Polystearate (and) Cetyl Palmitate 3.00 3.00 Disodium Cetearyl Sulfosucchinate 1.00 1.00 Cetearyl Alcohol 1.00 1.00 Lauryl Lactate 5.00 5.00 C12-C15 Alkyl Benzoate 10.00 10.00 Dibutyl Adipate 10.00 10.00 A Phenoxyethanol (and) Ethylhexylglycerin 1.00 1.00 Ethylhexyl salicylate (EHS) 5.00 5.00 Butyl methoxydibenzoylmethane (BMDBM) 4.00 Diethylhexyl butamido triazone (DBT) 3.00 Bis-ethylhexyloxyphenol methoxyphenyl triazine (BEMT) 2.00 2.00 Ethylhexyl triazone (EHT) 3.00 2.50 Diethylamino Hydroxybenzoyl Hexyl Benzoate (DHHB) 3.00 Aqua 44.60 39.10 Glycerine 2.00 2.00 B Xanthan Gum 0.20 0.20 Disodium EDTA 0.20 0.20 Phenylbenzimidazol sulfonic acid (PBSA) 2.00 C Aqua 5.00 Tromethamine 2.00 D Methylene bis-benzotriazolyl tetramethylbutylphenol (MBBT) 3.50 E Undecan, Tridecan 2.00 2.00

Tab. 1 INCI of the two SPF 30 sunscreen formulas

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trol of the inducibility of Acne aestivalis in the selected test persons.

Negative Control

In the test areas treated with the negative control (market sunscreen gel- formulation) no skin reactions were observed in any of the subjects. (Reference for Positive and Negative control: confidential in- formation DermaTronnier)

12-1

In the test areas treated with this sunscreen formula, no Fig. 3 Score evaluation of the clinical study skin reactions could be observed dur- ing the 5 days treatment in none of the 6 subjects. This sunscreen formula Score value: offered complete protection against 0 = no reaction 0 ± = light reaction 1 Acne aestivalis. + = distinct reaction 2 ++ = severe reaction 3 +++ = very severe reaction 4 12-2 n.t. = no treatment Test product Observation time P1 P2 P3 P4 P5 P6 UV14-132-12-1 Beobachtungszeit 0 0 0 0 0 0 This test product provoked skin re- 1. Day before irradiation 0 0 0 0 0 0 actions classified to Acne aestivalis 1. Day after irradiation 0 0 0 0 0 0 in 5 out of 6 subjects. Subject n°4 2. Day before irradiation 0 0 0 0 0 0 showed only slight symptoms, which 2. Day after irradiation 0 0 0 0 0 0 3. Day before irradiation 0 0 0 0 0 0 occurred already after the third ex- 3. Day after irradiation 0 0 0 0 0 0 position day. Subjects n° 1, 5, and 6 4. Day before irradiation 0 0 0 0 0 0 showed light to distinct reaction dur- 4. Day after irradiation 0 0 0 0 0 0 5. Day before irradiation 0 0 0 0 0 0 ing the exposition time, so that after UV14-132-12-2 5. Day after irradiation 0 0 0 0 0 0 the third and fourth day no further ir- 1. Day before irradiation 0 0 0 0 0 0 radiation and treatment could be ap- 1. Day after irradiation 0 0 0 0 0 0 2. Day before irradiation 0 0 ± 0 0 0 plied for ethical reasons. Subject n° 3 2. Day after irradiation 0 0 ± ± 0 0 showed already severe skin reactions 3. Day before irradiation ± 0 ± ± 0 ± after the second treatment day, so 3. Day after irradiation ± 0 ++ n.t. ± 0 + n.t. that no further irradiation could be 4. Day before irradiation + 0 ++ ± ± + 4. Day after irradiation + n.t. 0 ++ n.t. ± + n.t. + n.t. applied after day 4. 5. Day before irradiation + 0 ++ ± + + Negative control 5. Day after irradiation 0 0 0 0 0 0 The results in Tab. 3 were quantified 1. Day before irradiation 0 0 0 0 0 0 1. Day after irradiation 0 0 0 0 0 0 using a score system, which is illustrat- 2. Day before irradiation 0 0 0 0 0 0 ed in Fig. 3 2. Day after irradiation 0 0 0 0 0 0 3. Day before irradiation 0 0 0 0 0 0 3. Day after irradiation 0 0 0 0 0 0 4. Day before irradiation 0 0 0 0 0 0 Discussion 4. Day after irradiation 0 0 0 0 0 0 5. Day before irradiation 0 0 0 0 0 0 Two sunscreen formulas (SPF 30) have Positive control 5. Day after irradiation 0 0 0 0 0 0 1. Day before irradiation ± 0 0 0 0 0 been under investigation. Although 1. Day after irradiation 0 0 ± 0 0 0 both contained organic UV-filters only 2. Day before irradiation ± 0 ± 0 0 0 and provide a comparable protection 2. Day after irradiation + n.t. 0 + n.t. ± ± ± 3. Day before irradiation + ± + ± ± + in the UV A spectral range, very dif- 3. Day after irradiation + n.t. ± + n.t. ± ± + n.t ferent RP values (levels of UV-induced 4. Day before irradiation + ± + + + + free radicals) and completely different 4. Day after irradiation + n.t. + n.t. + n.t. + n.t. + n.t. + n.t. skin reactivity have been obtained. An 5. Day before irradiation + + + + + + excellent correlation between the in Tab. 3 Evaluation of the adverse skin reaction of treated subjects where 24 is the sum of the vitro-parameter (RP) and the clinical highest possible skin reaction (score value 4 x 6 subjects) symptoms could be shown.

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Despite the relatively low number of subjects, this result pro- • The RP value seems to be a suitable predictive tool for creat- vides an excellent correlation between the in vitro-parameter ing sunscreen formulations with low potential of UV induc- (RP) and the clinical symptoms, corroborating the hypothesis ible peroxide radicals and elicitation of Acne aestivalis. of involvement of free radical species in Acne aestivalis. With an RP value of 0, test formula 12-1 had no UV inducible • As a sound formulation strategy it can be recommended to free radical potential, whereas 12-2 had a high one, with RP avoid any raw materials with UV inducible peroxide radical values of 45.3 % in the UVB/A range and 21.9 % under pure potential and/or by adding effective antioxidants to reduce UVA irradiation. In parallel, the low free radical potential 12-1 the amount of free radicals. formula did not elicit Acne aestivalis symptoms in the clinical study, in contrast to 12-2, with a very high incidence rate of References 5 out of 6 subjects. The differences in the two formulas only lay in different UV- [1] Andersch Björkman Y (1), Hagvall L, Siwmark C, Niklasson B, Karlberg AT, Brå- filter combination. Formula 12-1 contained highly stable or- red Christensson J. Air-oxidized linalool elicits eczema in allergic patients - a ganic, oil soluble or particulate UV-filters, whereas formula repeated open application test study. Contact Dermatitis. (2014) 70(3):129-38. 12-2 contained in addition non-stabilized BMDBM and water- soluble PBSA. The first one is known to undergo photoreac- [2] Tronnier H, Heinrich U. Die polymorphen Lichtdermatosen. Studie zur Pathoge- tions under UV radiation leading to intermediate radical spe- nese, Prophylaxe und Therapie. Akt. Dermatol. (1994) 20, 220-226. cies [6]; the second one converts into an organic semistable free radicals under UV radiation [7]. [3] Rippke F, Wendt G, Bohnsack K, Dörschner A, Stäb F, Hölzle E, Moll I. Results of In the present study Acne aestivalis has been provoked by a photoprovocation and field studies on the efficacy of a novel topically applied formulation containing high levels of UV-inducible peroxides in polymorphous light eruption. J Dermatolog Treat. (2001) 12, 3-8. and organic free radicals. Exemplarily, the free radicals are [4] Tronnier H. Zur Pathogenese der Mallorca-Akne. Parfümerie und Kosmetik generated via photounstable organic UV filters. Similar high (1991) 72, 286-292. levels of peroxides and lipid radicals can be also generated due to other components of a skin care formulation, such as [5] Herrling T, Seifert M, Sandig G, Jung K. The Radical Power RP of Products. perfumes, or oils. These components are thought to induce SOFW-Journal (2015) 141,8-12. Acne aestivalis and photo-and chemo-toxic reactions when exposed to higher levels of UV radiation in sensitive persons [6] Schwack W, Rudolph T. Pho- [8]. However, in no case the presence of the pure components contact tochemistry of dibenzoyl- is responsible for a high Radical Power: Nor the terpene com- methane UVA filters. Part I. ponents of a perfume (e.g. linalool, geraniol, citronellol, limo- J. Photochem. Photobiol. B nene, among others), nor the pure natural oils (almond oil, *Katinka Jung (1995) 28, 229-234. sunflower oil, avocado oil, among others) will induce radicals Thomas Herrling under UV irradiation or adverse skin reactions. In all cases, as [7] Bastien N, Millau JF, Rouabhia Gematria Test Lab GmbH to our best knowledge, the hydroperoxide-adducts present in M, Davies JRH, Drouin R. The Parkstr. 23 | 13187 Berlin | Germany the terpenes or the beginning of rancidity in oils are respon- Sunscreen Agent 2-Phenyl- sible for high RP values. **Ulrike Heinrich benzimidazole-5-Sulfonic Acid Hagen Tronnier Photosensitizes the Formation of Oxidized Guanines In Cellulo DermaTronnier GmbH & Co. KG Conclusion after UV-A or UV-B Exposure. Institut für Experimentelle Dermatologie Journal of Investigative Derma- an der Universität Witten/Herdecke • Sunscreen formulations using non photo stable filter sys- tology (2010) 130, 2463–2471. Alfred-Herrhausen-Str. 44 | 58455 Witten |Germany tems may have a high potential of UV inducible peroxide ***Marcel Schnyder radicals, expressed by a high RP value. [8] Fotiades J1, Soter NA, Lim HW. Bernd Herzog Results of evaluation of 203 • Clinical data presented here indicate a clear correlation be- patients for photosensitivity in BASF Grenzach GmbH tween the level of UV inducible free radicals and the elicita- a 7.3-year period. J Am Acad 79639 Grenzach-Wyhlen | Germany tion of Acne aestivalis in sensitive individuals. Dermatol. (1995) 33, 597-602.

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