SKIN-WHITENING EFFECTS OF MEDITERRANEAN HERBAL EXTRACTS 317 Figure 3. Percentage of Inhibition of skin pigmentation (PI) of formulation A (hydroquinone), formulation B (kojic acid), and formulation C (mixture of herbal extracts) obtained by in vivo study over the monitoring period of 4 weeks. Figure 4. Increase in skin sensitivity to UVB irradiation expressed by photosensitivity percentage after 6 weeks of treatment with formulation A (hydroquinone), formulation B (kojic acid), and formulation C (mix- ture of herbal extracts) vs. control (no topical treatment). In our study, caper buds, blood orange, rice grains, and olive leaf can be considered im- portant natural sources of anti-melanogenic substances for the high level of polyphenolic content. In fact, it is possible to suppose that skin-whitening effect of herbal combination may be attributed to polyphenols and their association (8, 22–25). It is known that oxidative species are involved in skin UV-induced pigmentation and antioxidant substances such as polyphenols can affect melanogenesis processes by improv- ing skin oxidative stress defense (8). Moreover, from the data obtained in the in vitro study, we observed a direct inhibition of tyrosinase enzyme activity by herbal extracts. Our results are in agreement with data reported in several studies that attributed interest- ing anti-melanogenic activity to some group of polyphenolic compounds commonly con- tained in these plants. Flavonoids such as hesperidin, naringenin, and eriodictyol are considered the most abundant fl avanones contained in citrus fruits (8). The effects of

JOURNAL OF COSMETIC SCIENCE 318 these fl avonoids on melanin synthesis have been already evaluated (8,9). Hesperidin and naringenin have a chemical structure similar to hydroquinone and it was supposed that they can act as a substrate competitor for tyrosinase (26). Hesperidin showed to induce inhibition of tyrosinase activity in human primary melanocytes in a dose-dependent man- ner and to protect fi broblasts and collagen against oxidative and UVA-induced damages (8,27). Moreover, fl avonoids such as quercetin, kaempferol, and their derivates, typically contained in Capparis spinosa extract (25,28), showed to possess a strong tyrosinase inhibitory activity (23). Further investigation provided that the anti-melanogenic effect of quercetin and kaempferol was due to different mechanism of actions. Quercetin was able to decrease the intracellular tyrosinase activity not only by inhibition of enzyme ac- tivity but also by reduction of its protein expression (29), and kaempferol could act by copper chelation of tyrosinase enzyme as long as its 3-hydroxyl group was free (23). Similar activity was associated to hydroxycinnamic acid group, such as ferulic acid and caffeic acid, mainly contained in Oryza sativa extracts, by inactivation of tyrosinase effect (30,31). Finally, the skin protective action against UV light of olive leaf extract and its polyphenols, such as oleuropein and luteolin, was already known (22,24,32). In addition, one of the main drawbacks of skin-whitening agents, such as hydroquinone and kojic acid, is the skin-sensitizing effect to the sunlight exposure due to the repeated application of these substances. To simplify treatment regimen and reduce the risk of side effects, application of skin-whitening ingredients usually needs to avoid sun exposure and to use protective ingredients, such as sunscreens (32–36). The in vivo model used in this work can predict the skin photosensitizing effect induced by cosmetic ingredients after repeated topical applications (21). Results showed a safer activity of the natural mixture than hydroquinone and kojic acid. As previously reported in literature (11), we can sup- pose that polyphenols could act as cutaneous sunscreen and they can protect the skin against the radiation-induced infl ammation, oxidative stress, DNA damage, and other diseases induced by UV-light exposure. In conclusion, in this study we evaluated the anti-melanogenic effects and the safety of herbal extracts obtained from typical Mediterranean species such as Capparis spinosa, Cit- rus sinensis, Oryza sativa, and Olea europaea by in vitro and in vivo models. Further studies could be useful to better understand the mechanisms of action and the future application of these polyphenols as skin-whitening agents. REFERENCES (1) T. Hanamura, E. Uchida, and H. Aoki, Skin-lightening effect of a polyphenol extract from Acerola (Malpighia emarginata DC.) Fruit on UV-induced pigmentation, Biol. Pharm. Bull., 72, 3211–3218 (2008). (2) Y. J. Kim and T. Yokozama, Modulation of oxidative stress and melanogenesis by proanthocyanidins, Biol. Pharm. Bull., 32, 1155–1159 (2009). (3) U. Panich, K. Kongtaphan, T. Onkoksoong, K. Jaemsak, R. Phadungrakwittaya, A. Thaworn, P. Akarasereenont, and A. Wongkajornsilp, Modulation of antioxidant defense by Alpinia galangal and Curcuma aromatica extracts correlates with their inhibition of UVA-induced melanogenesis, Cell Biol. Toxicol., 26, 103–116 (2010). (4) K. H. Wang, R. D. Lin, F. L. Hsu, Y. H. Huang, H. C. Chang, C. Y. Huang, and M. H. Lee, Cosmetic applications of selected traditional Chinese herbal medicines, J. Ethnopharmacol., 106, 353–359 (2006). (5) G.R. Kanthraj, Skin-lightening agents: New chemical and plant extracts-ongoing search for the holy grail!, Indian J. Dermatol. Venereol. Leprol., 76, 3–6 (2010).