高速液体クロマトグラフィーによるタンパク結合の定量的研究 : 多孔性ガラスビーズに固定化したアルブミンを用いる方法

抄録

Application of high performance liquid chromatographic (HPLC) technique was attempted to the study of drug-protein interactions by immobilizing human serum albumin monomer (HSAm) to glyceryl porous blass beads (Glyceryl-CPG). Experiments were all performed at 4°C in 1/15 M phosphate buffer, pH 7.40, containing 0.01% ethylene diamine tetraacetic acid (EDTA) at a flow rate of 2 ml/min per column by both single and double column method. This flow rate was approximately ten times greater than that of the previous affinity chromatographic technique employing Sepharose 4B as a solid support. Thereby, effective time reduction in the determination of binding parameters was possible without apparent adverse effects of immobilization. Binding parameters were determined for the interactions with salicylic acid, warfarin, and diazepam. Diazepam, in particular, could not be previously studied with Sepharose 4B support due to its extensive binding to the support itself.