The interaction of genistein and human serum albumin (HSA) was investigated by fluorescence quenching spectra, synchronous fluorescence spectra and ultra-violet absorption spectra. The results showed that the quenching mechanism of the intrinsic fluorescence of HSA by genistein is due to the formation of genistein-HSA complex, resulting in a static quenching procedure. The binding constants (KA) were 1.00 106 (27 ℃), 1.66 106 (37 ℃) and 5.25 106 (47 ℃), respectively. According to the Frster theory of non-radiation energy transfer, the binding distances (r) were 2.59 nm (27 ℃), 2.65 nm (37 ℃) and 2.90 nm (47 ℃), respectively. The thermodynamic parameters showed that the binding power between genistein and HSA is mainly the electrostatic interaction. Synchronous spectrum was used to investigate the conformational change of HSA.