Abstract: The interaction between warfarin and bovine serum albumin (BSA) under pseudo-physiological conditions was investigated by UV-Vis absorption spectrometry and spectrofluorimetry. The quenching mechanism of BSA by warfarin was discussed. It showed that the quenching process was a dynamic quenching, and the quenching constants were 6.05×10⁴ L·mol^-1 at 16 ℃ and 6.14×10⁴ L·mol^-1 at 37 ℃, separately. Based on the Frster theory of non-radiative energy transfer, the energy transfer efficiency and the distance of BSA to warfarin were calculated, which was 0.37 and 3.15 nm, respectively.