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A product resulting from the biotransformation of genistein by a recombinant Pichia pastoris was isolated and identified as 3′-hydroxygenistein, on the basis of mass, 1 H NMR, and 13 C NMR spectrophotometric analysis. The maximal product concentration and the conversion yield of the biotransformation in a 5 l fermenter were 3.5 mg/l and 14%, respectively. The inhibitory effects of 3′-hydroxygenistein on tyrosinase activity were investigated in vitro using mushroom tyrosinase. The results showed that 3′-hydroxygenistein potently inhibited tyrosinase activity with an IC 50 value of 15.9 μM. Furthermore, the inhibitory effects of 3′-hydroxygenistein on melanogenesis were also investigated in vitro in cultured B16 melanoma cells, and it was shown that 3′-hydroxygenistein dose-dependently inhibited melanogenesis in non-toxic concentrations. In summary, the 3′-hydroxygenistein that was produced from genistein by the recombinant yeast was confirmed as a potent tyrosinase inhibitor and inhibited melanogenesis in B16 cells. [ABSTRACT FROM AUTHOR]