The conjugation between bovine serum albumin (BSA) and three antioxygenic micromolecules of ferulic acid, calycosin and formononetin was investigated here, using fluorescence spectroscopy (FS), ultraviolet spectroscopy (UV) and ultrafiltration (UF). The binding constants K_LB and distances r of these compounds and BSA were calculated respectively, according to Lineweaver-Burk equation and energy transfer theory. And every type of conjugation effort was estimated by the calculation of thermodynamic parameters. Besides, the binding constants K_a and binding site numbers n in terms of Scatchard equation were also obtained. The results were showed as follows: the ferulic acid, calycosin and formononetin could form non-covalent compounds with BSA, which led to the static quenching of endogenous fluorescence of BSA; but the enlargement of polarity and volume of the molecules caused the decrement of the affinity, which was mainly on the electrostatic interaction of these compounds with BSA. In the concentration of our experiment, the results indicated the competitive binding reaction of ferulic acid, formononetin and calycosin with BSA.

Key words: antioxygenic micromolecule, bovine serum albumin, competition binding, fluorescence spectroscopy