A Validated Stable Isotope Dilution Liquid Chromatography Tandem Mass Spectrometry Assay for the Trace Analysis of Cocaine and Its Major Metabolites in Plasma

Abstract

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 isotope dilution liquid chromatography/atmospheric pressure chemical ionization/tandem mass spectrometry. Previously reported methods do not have the sensitivity and specificity that can be attained with this method. Plasma samples required no cleanup apart from protein precipitation, and no derivatization was required. Selected reaction monitoring was performed on the transitions 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 (norcocaine). The standard curves were linear over the range from 2 ng/mL (benzoylecgonine, cocaine, and norcocaine) or 5 ng/mL (ecgonine methyl ester) to 1000 ng/mL in rat plasma. The lower limit of quantitation (LLQ) for benzoylecgonine, cocaine, and norcocaine was 2 ng/mL, and for ecgonine methyl ester, the LLQ was 5 ng/mL for plasma. This simple, rapid, reliable, and sensitive method of quantitation had excellent accuracy and precision for the four analytes. The method was sensitive enough to permit a detailed study of the pharmacokinetics of cocaine and its metabolites after administration of a bolus intravenous dose to rats.

Quantitative analysis of cocaine and its metabolites in whole blood and urine by high-performance liquid chromatography coupled with tandem mass spectrometry

“…Several LC/MS/MS applications for cocaine have been published, but they have either been qualitative analysis of only COC and BZE in plasma [9,10] or quantitative analysis of the cocaine and metabolites in other matrices such as urine, oral fluid, hair and meconium [11][12][13][14][15]. Published methods in blood are mainly performed in plasma and typically limited to cocaine and one or two metabolites [3,8,[16][17][18]. Cailleux et al [19] published a LC/MS/MS method for whole blood where BZE and EME coeluted and the sample preparation involved the hazard trichloromethane.…”

Analysis of protein ions in the range 3000–12000 Th under partial (no discharge) atmospheric pressure chemical ionization conditions using ion trap mass spectrometry

“…Another advantage is that APCI is not as much affected by limitations on signal/noise ratio due to matrix effects 16,. 17…”

Cocaine Metabolism Accelerated by a Re-Engineered Human Butyrylcholinesterase

“…Plasma samples were prepared for LC-MS as described by Singh et al (1999). Frozen plasma was quickly thawed, mixed, and microcentrifuged for 2 min at 14,000 rpm; 300-l supernatant aliquots were then transferred to clean tubes with 1 ml of acetonitrile, mixed for 10 s, and centrifuged for 10 min at 2,500g, 4°C.…”