Enantioselective HPLC determination of E‐6087, a new COX‐2 inhibitor, in human plasma: Validation and pharmacokinetic application

E‐6087 is a nonsteroidal anti‐inflammatory compound that selectively inhibits cyclooxygenase‐2. Because E‐6087 has a chiral center, this compound is a racemic mixture of two stereoisomers, (+)‐( R )‐E‐6087 (E‐6231) and (−)‐( S )‐E‐6087 (E‐6232). A normal‐phase liquid‐chromatographic method for the enantioselective determination of E‐6087 in human plasma was developed and validated. The samples were extracted using solid‐phase extraction cartridges containing C 18 sorbent, and the extracts were redissolved in absolute ethanol and injected into the chromatographic system. The enantiomeric separation was achieved on a chiral stationary‐phase column of derivatized amylose, and the enantiomers were quantified by fluorescence detection. The method was validated for drug concentrations ranging from 5 to 400 ng/ml for both enantiomers. No peaks interfering with the quantification of enantiomers were observed. The limit of quantification was 5 ng/ml, with precision expressed as a coefficient of variation lower than 10.6% and accuracy expressed as relative error lower than 12.2%. The utility of this method was demonstrated by analysis of plasma samples from healthy volunteers given an oral dose of rac ‐E‐6087. Peak plasma levels of E‐6231 were higher than levels obtained for E‐6232. Results were consistent with those obtained with a conventional reversed‐phase method used for determination of the racemic compound. Chirality 16:302–308, 2004. © 2004 Wiley‐Liss, Inc.