The purpose of this study is to investigate the effect of H2O2 on the aging of melanogenesis in human melanocytes. The staining of SA-β-galactosidase, an aging marker, was remarkably increased in the cells aged with H2O2 at 62.5 µM or more compared with young cells. The intracellular H2O2 level and melanin synthesis were also reduced in both H2O2-treated cells and senescent cells compared with young cells in DCFH-DA assay. Both the senescent cells and the H2O2-treated cells showed higher expression level of Catalase than young cells in western blot and immunofluorescence staining. Furthermore, the expression levels of TRP-1, TRP-2 and p300 were reduced in both senescent cells and the H2O2-treated cells, but that of SIRT-1 was inverted compared with young cells. In addition, H2O2 reduced the expression level of MITF but increased that of Nrf2 in nucleus. Those results indicate that the expression levels of antioxidant enzymes in senescent cells and H2O2-treated cell are upregulated, but the expression levels of proteins involved in melanin synthesis are downregulated. Above findings suggest that H2O2 could play a key role in the aging process of melanogenesis through modulation of MITF and Nrf2.