Determination of the carboxylic acid metabolite of clopidogrel in human plasma by liquid chromatography–electrospray ionization mass spectrometry

A rapid and specific liquid chromatographic–mass spectrometric method has been developed and validated for the determination of the carboxylic acid metabolite of clopidogrel in human plasma. Sulphafurazole was used as internal standard. The samples were subjected to a solid phase extraction procedure using Hypercarb cartridges. The chromatographic separation was performed on a reversed phase porous graphitized carbon column using a mobile phase consisting of 70% methanol in water containing 0.1% (v/v) trifluoroacetic acid, pumped at a flow rate of 0.25 ml min^-1. The analytes were detected after positive electrospray ionization using the selected ion monitoring mode of the species at m/z 308 for the carboxylic acid metabolite of clopidogrel, m/z 322 for clopidogrel and m/z 268 for sulphafurazole. Calibration graphs were linear (r>0.9994, n=6), in the range 100–1000 ng ml^-1 for the carboxylic acid metabolite of clopidogrel. The intra- and inter-day R.S.D. values were <3.1%, while the relative error Er was less than -9.6% (n=6). The limits of detection (3.3(sigma)) and quantitation (10(sigma)) for the carboxylic acid metabolite of clopidogrel were found to be 28 and 93 ng ml^-1, respectively. The efficiency of the solid phase extraction procedure for the carboxylic acid metabolite of clopidogrel averaged 74.6%.