Quantitative determination of rivastigmine and its major metabolite in human plasma by liquid chromatography with atmospheric pressure chemical ionization tandem mass spectrometry

A sensitive, specific, accurate and reproducible LC–MS–MS method was developed and validated for the simultaneous determination of rivastigmine and its major metabolite NAP 226-90 in human plasma according to International Regulatory Requirements. After addition of their respective labelled internal standards, the compounds were extracted from plasma using methyl-tert.-butyl ether at basic pH with a simultaneous derivatization of NAP 226-90 with propionic anhydride, and backextracted into an acidic solution. After re-extraction the compounds were analyzed on a 3-μm Purospher Star RP-18 column interfaced with a MDS Sciex API 3000 triple quadrupole mass spectrometer. Positive atmospheric chemical ionization was employed as the ionization source. The analytes and their internal standards were detected by use of multiple reaction monitoring mode. Intra- and inter-day accuracy and precision were found suitable over the range of concentrations between 0.200 and 30.0 ng/ml. The LC–MS–MS method was crossvalidated with a previously developed in-house GC–MS method by the analysis of plasma samples obtained from patients after administration of Exelon® capsules and showed excellent correlation between the methods.