Abstract: The interaction and mechanism between bovine serum albumin (BSA) and 24(R)-pseudo sapogenin DQ24(R)-PDQ were studied by fluorescence spectrometry, and the binding site was confirmed by displacement experiment. The results showed the binding constants were calculated to be 2.076×10³L·mol^-1 at 298 K, 1.587×10³L·mol^-1 at 303 K and 1.303×10³L·mol^-1 at 310 K respectively, and 24(R)-PDQ induced the intrinsic fluorescence quenching of BSA through a static quenching procedure. The predominant forces in the 24(R)-PDQ-BSA complex were hydrogen binding and Van der waals force. The number of binding sites in this binary system was approximately 1. The specific binding of 24(R)-PDQ in the vicinity of Site Ι of BSA was clarified.