Determination of M+4 stable isotope labeled cortisone and cortisol in human plasma by mu Elution solid-phase extraction and liquid chromatography/tandem mass spectrometry

A sensitive mu Elution solid-phase extraction (SPE) liquid chromatography/tandem mass spectrometry (LC/MS/MS) method has been developed and validated for the determination of M+4 stable isotope labeled cortisone and cortisol in human plasma. In this method, M+4 cortisone and M+4 cortisol were extracted from 0.3 mL of human plasma samples using a Waters Oasis HLB 96-well mu Elution SPE plate using 70 ILL methanol as the elution solvent, and chromatographed on a Waters Symmetry C18 column (4.6 x 50 mm, 3.5 mu m). M+9 cortisone and M+9 cortisol were used as the internal standards. A PE Sciex API 4000 tandem mass spectrometer interfaced with the liquid chromatograph via a turboionspray source was used for mass analysis and detection. The selected reaction monitoring (SRM) of precursor product ion transitions were monitored at m/z 365.2 [M+H](+) -> 167.0 and at m/z 367.3 [M+H](+) -> 125.1 for M+4 cortisone and M+4 cortisol, respectively. The lower limit of quantitation was 0.1 ng mL(-1) and the linear calibration range was from 0.1 to 100 ng mL-1 for both analytes. This method demonstrated to be very reproducible and reliable. Copyright (c) 2005 John Wiley & Sons, Ltd.