The validation of a bioanalytical method for the determination of clopidogrel in human plasma

Abstract

A fast, sensitive and specific LC–MS/MS bioanalytical method for the determination of unchanged clopidogrel in human plasma has been developed and validated over the range of 10–12,000 pg mL −1 ( r 2 0.9993) by the Contract Research group at HFL. Samples (0.3 mL) were buffered (pH 6.8), extracted using diethyl ether and 10 μL of the sample extract was injected onto the LC–MS/MS system. Analysis was performed using a C8 column (temperature controlled to 50 °C) by gradient elution at a flow rate of 0.9 mL min −1 over a 3 min run time. Retention times of 1.61 and 1.59 min were observed for clopidogrel and 2H 3-clopidogrel (I.S.), respectively. Detection was achieved using a Sciex API 4000, triple quadrupole mass spectrometer, in positive TurboIonspray™ (electrospray) ionisation mode. Ion transitions were monitored using MRM (multiple reaction monitoring) for clopidogrel ( m/ z 322–212) and for 2H 3-clopidogrel ( m/ z 327–217). This validated method was used to support a pharmacokinetic study in healthy volunteers.