The Inhibitory Effect of Polyphenon 60 from Green Tea on Melanin and Tyrosinase in Zebrafish and A375 Human Melanoma Cells

Abstract

Polyphenon 60 (PP60) from green tea has long been used as an antioxidant, anticancer, antimicrobial, and antimutagenic. Aim of the Study. To investigate tyrosinase inhibition-related kinetic mechanism and antimelanogenesis potential of PP60. Materials and Methods. The effect of PP60 on melanin and tyrosinase was evaluated in A375 melanoma cells and zebrafish embryos. The melanoma cells were treated with 20, 40, and 60 µg/mL of PP60, and tyrosinase expression was induced by using L-DOPA. The western blot method was used for the evaluation of tyrosinase expression. Cell lysates were prepared from treated and untreated cells for cellular tyrosinase and melanin quantification. Furthermore, zebrafish embryos were treated with 20, 40, and 60 µg/mL of PP60 and reference drug kojic acid for determination of depigmentation and melanin quantification. In vitro assays were also performed to examine the impact of PP60 on mushroom tyrosinase activity. To determine cytotoxicity, MTT was used against melanoma cell line A375. Results. PP60 showed good tyrosinase inhibitory activity with an IC50 value of 0.697 ± 0.021 µg/mL as compared to kojic acid a reference drug with an IC50 value of 2.486 ± 0.085 µg/mL. Kinetic analysis revealed its mixed type of inhibition against mushroom tyrosinase. In addition, western blot analysis showed that at 60 µg/mL dose of PP60 significantly reduced L-DOPA-induced tyrosinase expression in melanoma cells. PP60 significantly inhibits the cellular tyrosinase ( $p<0.05$ ) and reduces the melanin ( $p<0.05$ ) contents of melanoma cells. Furthermore, PP60 was found to be very potent in significantly reducing the zebrafish embryos’ pigmentation ( $p<0.05$ ) and melanin ( $p<0.05$ ) content at the dose of 60 µg/mL. Conclusions. Our results demonstrate that PP60 has a strong potency to reduce pigmentation. It may be useful for the cosmetic industries to develop skin whitening agents with minimal toxic effects.