Stereoselective Binding of Warfarin and Ketoprofen to Human Serum Albumin Determined by Microdialysis Combined with HPLC

Abstract

Microdialysis sampling technique combined with high performance liquid chromatography (HPLC) was utilized to determine the stereoselective binding of warfarin and ketoprofen to human serum albumin (HSA) in solution. The HPLC conditions for separation of warfarin and ketoprofen enantiomers on HSA column were optimized. The unbound enantiomers in the mixed buffer (0.067 M phosphate and pH 7.4) solution of the drug racemate with HAS were sampled with microdialysis probe, and the amount of them was determined by HPLC on HSA column. It was observed that the unbound concentrations of R-warfarin and R-ketoprofen are about 1.08∼1.34-and 1.08∼1.15-fold of S-warfarin and S-ketoprofen, respectively, which indicated that the S-enantiomers bind to HSA more strongly than R-enantiomers to HSA. The interaction parameters for binding of warfarin and ketoprofen enantiomers to HSA including binding constants and number of binding site were obtained by the Scatchard analysis of experimental data, and the stereoselectivity (α) calculated by the binding constants of warfarin enantiomers to HSA is 1.92, but that of ketoprofen enantiomers is very close. HSA exhibits stronger stereoselectivity to warfarin racemate than to ketoprofen racemate.