Stereospecific analysis of the major metabolites of ibuprofen in urine by sequential achiral-chiral high-performance liquid chromatography

Abstract

A sequential achiral-chiral HPLC method has been developed for the stereospecific analysis of the two major urinary metabolites of ibuprofen, namely hydroxyibuprofen and carboxyibuprofen. Achiral analysis was carried out using a Partisil column (250×4.6 mm, 5 μm) and a mobile phase of hexane:ethanol (98.2:1.8, v/v) containing trifluoroacetic acid (TFA; 0.05%, v/v) at a flow-rate of 2.0 ml/min. The HPLC eluate containing the two metabolites was separately collected, evaporated under nitrogen and the residue dissolved in the mobile phase used for chiral chromatography. Chiral-phase analysis was carried out using a Chiralpak AD CSP (250×4.6 mm, 10 μm) with a mobile phase of hexane:ethanol (92:8, v/v) containing TFA (0.05%, v/v) at a flow-rate of 1.0 ml/min. In both assays the analytes were quantified by ultraviolet detection at a wavelength of 220 nm. Modification of the mobile-phase composition allowed the resolution of all six analytes in a single chromatographic run but with an increase in run time and consequent band broadening. The analytical method described allows the direct quantitation of the stereoisomers of both metabolites of ibuprofen in urine following the administration of therapeutic doses of the racemic drug to man.