Binding of sulfamethoxazole to human serum albumin studied by a combined technique of microdialysis with liquid chromatography

Abstract

A new, simple and fast method for the determination of the interaction parameters of sulfamethoxazole (SMZ) to human serum albumin (HSA) has been developed by utilizing a microdialysis sampling technique combined with liquid chromatography (LC). The drug and protein were mixed in different molar ratios in 0.067 M potassium phosphate buffer, pH 7.4, and incubated at 37 °C in a water-bath. Then the microdialysis probe was put in the SMZ-HSA solution and sampled at a perfusion rate of 1 μl/min. The concentration of SMZ in the microdialysates was determined by reversed-phase liquid chromatography. The recovery ( R) was also determined in vitro under similar conditions, R is about 41.8% with an RSD of about 2.3%. The association constant ( K) and the number of the binding sites on one HSA molecule ( n) are calculated by three equations, the values of nK estimated by three methods are quite similar. The values found for K and n are 3.24 × 10 3 M −1 and 3.04, respectively.