Analysis of microsomal metabolic stability using high-flow-rate extraction coupled to capillary liquid chromatography-mass spectrometry

A method is described for on-line high-speed extraction of microsomal samples and analysis by capillary liquid chromatography-mass spectrometry (LC-MS) for the determination of metabolic stability in connection with the development of positron emission tomography (PET) tracers. The method allowed direct injections of large sample volumes at a fast extraction rate, providing a gain in both sensitivity and sample preparation time. The calibration curve of the test compound flumazenil (Ro 15-1788) was linear in the concentration range of 1-150 nM, with a correlation coefficient exceeding 0.999. The accuracy of the method ranged from 98 to 101%. A high precision was obtained, with mean intra-assay and inter-assay relative standard deviations of at most 1.4 and 1.5%, respectively, for quality control (QC) samples. The extraction efficiency was determined to be 99.4%, the total recovery 96% and the carryover to