SEPARATION OF THE STEREOISOMERS OF THE MAIN METABOLITE OF A NONSTEROIDAL ANTIINFLAMMATORY DRUG, FLOBUFEN, BY CHIRAL HIGHPERFORMANCE LIQUID-CHROMATOGRAPHY

The major metabolite of a novel non-steroidal anti-inflammatory drug, 2',4-difluorobiphenyl-4-yl)-4-oxo-2-methylbutanoic acid (flobufen, I), namely ifluorobiphenyl-4-yl)-2-methyl-gamma-butyrolactone (4-dihydrof lobufen lactone, III), has four stereoisomers consisting of two racemi c pairs of enantiomers. Of three chiral stationary phases tested, Cycl obond I beta-RSP (Astec) (beta-cylodextrin derivatized with R,S-hydrox ypropyl) was best able to separate the (++)(--) racemate, with a liqui d phase containing acetonitrile as modifier and triethylamine acetate as buffer. Using the Box-Wilson Central Composite Design for three fac tors, an optimum combination of pH and concentrations of the modifier and buffer was eventually obtained. A chromatographic response functio n based on a combination of the Kaiser peak separation function, P-i, and retention time of the second eluting enantiomer, t(RL), served as a response criterion for the process of optimization. The optimum cond itions developed for the (++)(--) racemate were also found to be suita ble for separating the (+-)(-+) racemate, for which earlier studies ha d shown the separation to be more facile. Separation of the four stere oisomers of III, for which the chiral chromatographic system optimized in this study is proposed as the second stage, is targeted at a bioch emical study of the stereoisomeric metabolism of I.