High-Performance Frontal Analysis for the Determination of the Free Drug Concentration in Protein Binding Equilibrium Using Porous Polymer Stationary Phase

Abstract

A high-performance frontal analysis method using a polyvinyl alcohol gel as a stationary phase has been developed for the determination of the free concentration of relatively hydrophilic drugs (salicylate and acetazolamide) in the state of a drug-protein binding equilibrium. When an excess volume of a drug-albumin mixed solution was injected directly to this column, the drug was eluted as a broad peak with a plateau region following the protein peak. When the free drug fraction was higher, a larger injection volume was necessary to obtain the plateau region. The free drug concentration was determined from the plateau height; the results agreed well with those obtained by the ultrafiltration method. The total salicylate concentration could be simultaneously determined from the peak area. The present method was applied to an estimation of the salicylate-albumin binding parameters. The obtained values agreed well with the reported values. The present method enables a simple and rapid determination of free drugs in protein binding equilibrium.