RESUMO
OBJECTIVE To estab lish the pre paration method of clopidogrel active thiol metabolite （CATM），and to provide reference for the synthesis of cis-CATM. METHODS CATM was prepared ，separated and purified with isolated rat liver perfusion and ChromCore 120 C18 preparative column ，using（S）-2-oxo-clopidogrel as substrate. The target compounds were identified by mass spectrometry and nuclear magnetic resonance spectroscopy. The retention time of the active configuration of CATM in the human body （cis-CATM）were compared to confirm the proportion of active configuration in the target product. RESULTS The conversion rate of the target product was 11.71％. The target products were identified as CATM by MS and 1H-NMR. Peak 2-peak 5 of CATM were four stereoisomers. The retention time of them were 21.3，22.3，26.5，27.3 min. The peak area ratios of them were 7.13％，7.23％，63.52％，14.97％，respectively. Based on that retention time of the active configuration of CATM in human body was 26.3 min，the active cis-stereoisomer in the target product CATM accounted for 63.52％. CONCLUSIONS This method is low-cost ，simple，and can prepare CATM with higher active configuration.
RESUMO
A sensitive and selective liquid chromatography–tandem mass spectrometric (LC ? MS/MS) method was established to determine 2-oxo-clopidogrel, a crucial intermediate metabolite in human plasma. A chromatographic separation was performed on a Sapphire C18 column following a liquid–liquid extraction sample preparation with methyl t-butyl ether. Detection was carried out on a triple quadrupole mass spectrometer operated in multiple reaction monitoring (MRM) with an electrospray ionization (ESI) mode. The method was validated in terms of specificity, accuracy, precision and limit of quantification. The calibration curves ranged from 0.50 to 50.0 ng/mL with good linearity. The stability was fully validated with addition of 1,4-dithio-DL-threitol (DTT) into the plasma sample prior to and in the preparation procedure. The validated method was proved to be suitable for use in pharmacokinetic study after single oral administration of 75 mg clopidogrel tablets in human subjects, which could make contribution to intensive study of the clinical drug–drug interactions of clopidogrel and individual treatment.