OBJECTIVE:To study the interaction mechanism between flavonoids and human serum albumin (HSA),and to compare the effects of different B-ring substitutions(hydroxyl,methoxyl group)of flavonoids on macromolecular receptor. METH-ODS:The interaction regularity between three flavonoids with different B-ring substitutions(quercetin,hesperetin,methyl hespere-tin) and HSA was studied with fluorescence spectroscopy,the fluorescence quenching types between 3 flavonoids and HSA were determined and analyzed,and the velated binding constant,binding site and thermodynamic parameters were calculated. RE-SULTS:The quenching constants (Ksv) and binding constants (KA) were decreased with the increase of temperatures. The number of binding site(n)was approximately equal to one,and the thermodynamic parameters ΔH<0,ΔS>0,the binding interaction of these compounds with macromolecules was influenced because of the difference of the B-ring substituents. CONCLUSIONS:The quenching mechanism between three flavonoids and HSA was static quenching;the number of binding site was one;the interaction force of the three compounds with HSA was mainly static electricity,and hydroxyl group in the B-ring was likely the major active group and exerted stronger binding force than methoxyl group to connect with macromolecules.