A validated stable isotope dilution liquid chromatography tandem mass spectrometry assay for the trace analysis of cocaine and its major metabolites in plasma

A validated method has been developed for the simultaneous quantitation of cocaine and its major metabolites (ecgonine methyl ester, benzoylecgonine, and norcocaine) in rat plasma. The method is based upon the use of stable i sotope dilution liquid chromatography/atmospheric pressure chemical ionizat ion/tandem mass spectrometry, Previously reported methods do not have the s ensitivity and specificity that can be attained with this method. Plasma sa mples required no cleanup apart from protein precipitation, and no derivati zation was required. Selected reaction monitoring was performed on the tran sitions of m/z 200 to m/z 182 (ecgonine methyl ester), m/z 290 to m/z 168 ( benzoylecgonine), m/z 304 to m/z 182 (cocaine), and m/z 290 to m/z 168 (nor cocaine), The standard curves were linear over the range from 2 ng/ml(benzo ylecgonine, cocaine, and norcocaine) or 5 ng/mt (ecgonine methyl ester) to 1000 ng/mL in rat plasma. The lower limit of quantitation (LLQ) for benzoyl ecgonine, cocaine, and norcocaine was 2 ng/mL, and for ecgonine methyl este r, the LLQ was 5 ng/mL for plasma. This simple, rapid, reliable, and sensit ive method of quantitation had excellent accuracy and precision for the fou r analytes, The method was sensitive enough to permit a detailed study of t he pharmacokinetics of cocaine and its metabolites after administration of a bolus intravenous dose to rats.