Development and validation of a sensitive LC-MS/MS method for simultaneous quantification of sinotecan and its active metabolite in human blood

摘要

Sinotecan is a camptothecin analog, currently under clinical testing as an antitumor medication. We developed and validated a rapid, specific and reliable liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous quantification of sinotecan and its active metabolite, 7-hydroxyethyl-camptothecin (7-HEC), in human blood. Aliquots (200 μL) of heparinized blood samples were processed by deproteinization with 400 μL acetonitrile each. Chromatographic analyte separation used an Agilent Zorbax SB C_8 column (4.6 mm × 150 mm, 5 μm) and methanol/10 mM ammonium acetate/formic acid (70/30/0.14, v/v/v) as mobile phase, at a flow rate of 0.60 mL/min. A Thermo Finnigan TSQ Quantum Ultra tandem mass spectrometer was operated in multiple-reaction monitoring mode. The precursor-to-product ion transitions m/z 493 → m/z (331 + 375) for sinotecan, m/z 393 → m/z (233 + 261) for 7-HEC, and m/z 396 → m/z 352 for d3-SN38 (IS) were used for quantification. The method was validated for 1.0-500 ng/mL for sinotecan and 0.5-250 ng/mL for 7-HEC using 200 μL of blood sample. Total time for each chromatograph was ～6.0 min. The intra- and inter-day precision and accuracy of the quality control samples at low, medium, and high concentration levels exhibited relative standard deviations (RSD) < 13.8% and the accuracy values ranged from -5.3% to 2.4%. The method was successfully applied to a pharmacokinetic study of sinotecan in cancer patients