Objective: To study the interaction of sulfamethazine (SMZ) with human serum albumin (HSA). Method: The quenching mechanism

binding constants and binding sites (n) were studied using molecular modeling

fluorescence quenching spectroscopy

three-dimensional fluorescence spectroscopy and UV-vis absorption spectroscopy under simulative physiological conditions. Result: The experiment results obtained from molecular modeling indicated that there were hydrogen bonds and hydrophobic interactions between SMX and HSA;the fluorescence quenching spectra data showed that the fluorescence intensity of HSA was quenched by the gradual addition of SMX. The binding constants of SMX with HSA at 293

298

303

308 K were calculated to be 3.42×104

3.13×104

2.63×104

2.13×104 mol·L-1

respectively

which means that the quenching mechanism was suggested as a static quenching. Meanwhile

corresponding numbers of binding sites (n) were calculated to be 0.99

0.96

0.98

1.09

respectively. The thermodynamic parameters of the reaction

namely standard enthalpy ΔH0 and entropy ΔS0

had been calculated to be -18.76 kJ·mol-1 and 22.80 kJ·mol-1

respectively

which suggested that the electrostatic and hydrophobic interactions were the predominant intermolecular forces in stabilizing the SMZ-HSA complex. Experimental results obtained from the three-dimensional fluorescence spectroscopy and UV-vis absorption spectroscopy confirmed that the secondary structure of HSA was altered in the presence of SMZ in aqueous solution. Conclusion: Molecular modeling and multi-spectroscopic approaches are good methods for investigating the interaction between SMZ and HSA. They have the advantages of high speed and high sensitivity.