Melanin is the pigment responsible for the color of the eyes, hair, and skin in humans. Tyrosinase is well known to be the key enzyme in melanin biosynthesis. JKTM-12 is composed of the flowers, roots, seeds, and receptacles of Nelumbo nucifera (lotus). In this study, JKTM-12 was investigated for its inhibitory effects on tyrosinase activity and melanin biosynthesis in B16F10 melanoma cells. Moreover, two main bioactive compounds (hyperoside and astragalin) were found from the receptacles of N. nucifera, which are used as the main material of JKTM-12. JKTM-12 was shown to inhibit tyrosinase activity and melanin biosynthesis in alpha-melanocyte-stimulating hormone-stimulated B16F10 melanoma cells. Hyperoside and astragalin, which are the main bioactive compounds of JKTM-12, not only inhibited tyrosinase activity and melanogenesis but also tyrosinase-related protein 1 and tyrosinase-related protein 2 mRNA expression without cytotoxicity at various experiment doses (0.1, 1, and 10 μg/ml). These results suggest that JKTM-12 has the potential for skin whitening with hyperoside and astragalin as the main bioactive compounds.