J. Cosmet. Sci., 64, 89–98 (March/April 2013) 89 Address all correspondence to Youn-Jung Kim at yj129@khu.ac.kr. and Mu-Hyoung Lee at mhlee@khmc. or.kr. Inhibitory effect of Gastrodia elata extract on melanogenesis in HM3KO melanoma cells HAK-JU KIM, JONG-HO LEE, MIN-KYUNG SHIN, KANG HYUN LEEM, YOUN-JUNG KIM, and MU-HYOUNG LEE, Department of Dermatology, College of Medicine, Kyung Hee University, Seoul 130-701 (H.-J.K., J.-H.L., M.-K.S., M.-H.L.), College of Oriental Medicine, Semyung University, Jecheon 390-711 (K.H.L.), and College of Nursing Science, Kyung Hee University, Seoul 130-701 (Y.-J.K.), Republic of Korea. Accepted for publication June 19, 2012. Synopsis Effective and safe antimelanogenic agents derived from natural products get interest continuously for both medical and cosmetic purposes. Melanin synthesis is regulated by melanogenic enzymes such as tyrosinase, tyrosinase-related protein 1 (TRP-1), and tyrosinase-related protein 2 (TRP-2) in mammals. Gastrodia elata (GE) has been shown to have multiple therapeutic actions related to antioxidation in many diseases. In this study, we investigate whether water extraction of GE has inhibitory effects on melanogenesis in vitro and on the expression of mRNA and protein of tyrosinase, TRP-1, and TRP-2 in HM3KO melanoma cells. To examine the inhibitory effect of GE on melanogenesis, mushroom tyrosinase inhibition assay reverse tran- scription polymerase chain reaction and western blotting of tyrosinase, TRP-1, and TRP-2 were performed using HM3KO melanoma cells. In this study, the GE extract was found to signifi cantly inhibit mushroom tyrosinase activity (69.3 ± 7.2% of the control, p 0.05), and the expression of mRNA and protein of tyrosinase, TRP-1, and TRP-2 was reduced signifi cantly. These results suggest that the antimelanogenic effect of GE extract is mainly due to the decreased expression of mRNA and protein of tyrosinase, TRP-1, and TRP-2 in the process of melanin synthesis. INTRODUCTION Melanin, synthesized in the melanosome of melanocytes in mammals, is transferred to neighboring keratinocytes. The role of melanin is to protect the skin from ultraviolet (UV) damage by absorbing UV sunlight and removing reactive oxygen species (1). The color of skin and hair is determined by a number of factors, but most important is thede- gree and distribution of melanin pigmentation. However, its excessive accumulation

JOURNAL OF COSMETIC SCIENCE 90 causes hyperpigmentation induced by UV irradiation or medical conditions such as melasma, postinfl ammatory hyperpigmentation, and solar lentigo. Therefore, the control of melanogenesis is an important strategy in the treatment of abnormal skin pigmenta- tion for cosmetic and medical purposes (2). Most controllers of melanogenesis act not only by altering tyrosinase gene expression but also by altering the mechanisms responsible for the transfer of melanosomes to keratino- cytes (3). The melanocyte-specifi c enzymes, tyrosinase and tyrosinase-related protein 1 and 2 (TRP-1 and TRP-2), are involved in converting tyrosine into melanin pigments (4,5). In particular, tyrosinase catalyzes two rate-limiting reactions involved in melano- genesis: the hydroxylation of tyrosine to 3,4-dihydroxyphenylalanine (DOPA), and the oxidation of DOPA, which results in the formation to dopaquinone (6). Since tyrosinase is a key enzyme in melanogenesis, it has been the target during the screening of new pigmentation inhibitors from natural products. Some examples of these products include kojic acid, arbutin, ascorbic acid derivatives, retinoic acid, azelaic acid, hydroquinone, catechins, aloesin, resveratrol, oxyresveratrol, and 4,4′-dihydroxybiphenyl (7–16). Various plant extracts have recently been studied to fi nd new natural antimelanogenic products. Gastrodia elata (GE), of the Orchidaceae family, is a traditional herbal medicine in East Asian countries. Zhao et al. (17) identifi ed the components of GE Blume using the capillary zone electrophoresis, the fi ve constituents—gastrodin (GA), 4-hydroxybenzyl alcohol (4-HBA), vanillyl alcohol, 4-hydroxybenzaldehyde (HD), and vanillin—in the extracts of GE Blume roots. It has been used for centuries as an anticonvulsant, analgesic, and sedative for the medical treatment of headaches, epilepsy, dizziness, rheumatism, neuralgia, paralysis, hyperten- sion, and other neuralgic disorders (18,19). A previous study showed that the anticonvul- sive effect of GE was attributed to its antioxidant activity, which was due to the antioxidant actions of 4-HBA, one of several phenolic components of GE extract (20). Since GE extract has several phenolic components that have antioxidant activity, this plant has been suggested to also have a depigmenting effect. Our group is currently trying to fi nd new antimelanogenic substances from natural sources. In this regard, GE has long been used in the treatment of many diseases, espe- cially skin diseases, and is well known for its antioxidant effects. In this study, we evaluate the antimelanogenic effects of GE in HM3KO melanoma cells by suppressing the expres- sion of tyrosinase and TRP-1 mRNA and protein levels. MATERIALS AND METHODS GE EXTRACT Plant extracts were purchased from the PURIMED Co., Ltd (Seoul, Korea). The pharma- ceutical name was Gastrodiae Rhizoma, which is the tuberous root extract of the GE Blume. The dried herb recommended in the protocol provided by PURIMED Co. Ltd was purchased from a Chinese market (Hong Kong, China) and used in this study. For the extraction of water-soluble fraction, 200 mg of the plants were cut into small pieces and boiled in 200 ml of distilled water under refl ux conditions. After 1 h, the mixture was fi ltered and the fi ltrate was collected. An additional 250 ml of water was added to the