Reversed-phase liquid chromatographic behavior of the macrolide antibiotics clarithromycin and roxithromycin

Abstract

The effect of mobile-phase composition and column temperature on the reversed-phase (RP) chromatographic behavior of the macrolide antibiotics clarithromycin and roxithromycin has been extensively studied. Sets of experimental retention data for these drugs, obtained on two octadecyl silica-based columns, with buffered mobile phases of pH 7.0 modified with one, two, or three organic solvents (methanol, acetonitrile, or isopropanol), were fitted by use of appropriate equations relating the retention of a solute to the organic modifier content of the mobile phase. The physical meaning of the equation coefficients is discussed, as is the dependence of these coefficients on the type of organic modifier(s) and the composition of the mobile phase. The van't Hoff plots for the macrolides in different binary, ternary, and quaternary mobile phases were highly linear at temperatures between 25 and 70°C. The enthalpies of adsorption of these solutes calculated from the slopes of the corresponding regression lines were exothermic for all mobile phases except for use of acetonitrile or 6∶1 (v/v) acetonitrile-isopropanol as organic modifiers. The results of our study provide a satisfactory thermodynamic explanation of the mechanism of retention of these macrolides in RP HPLC and give further information enabling selection of the optimum separation conditions for the determination of clarithromycin in biomatrixes with roxithromycin as an internal standard, and vice versa.