Assay method for the carboxylic acid metabolite of clopidogrel in human plasma by gas chromatography mass spectrometry

This paper describes a GC-MS method for the analysis of the carboxylic acid metabolite (SR26334, II) of methyl (+)-(S)-alpha-(o-chlorophenyl)-6,7-dihy drothieno[3,2-c]pyridine-5(4H) -acetate hydrogensulfate (clopidogrel, SR 25 990, I) in plasma and serum. The analytical procedure involves a robotic li quid-liquid extraction with diethyl ether followed by a solid-liquid extrac tion on C-18 cartridges. The derivatization process was performed using n-e thyl diisopropylethylamine and alpha-bromo-2,3,4,5,6-pentafluoro toluene. A structural analogue (III) of II, was used as internal standard. The 1/X-2 weighted calibration curve obtained in the range 5-250 ng/ml was well descr ibed by a quadratic equation. The extraction efficiency was better than 48% over the range studied; for the internal standard it averaged 51% at 50 ng /ml. Precision ranged from 3.6 to 15.8%, and accuracy was between 92 and 11 4%. Dilution has no influence on the performance of the method which could then be used to quantitate plasma samples containing up to 25000 ng/ml. The limit of quantification was 5 ng/ml. The method validation results indicat e that the performance characteristics of the method fulfilled the requirem ents for assay methods for use in pharmacokinetic studies. (C) 1998 Elsevie r Science B.V. All rights reserved.