title: A simple, rapid and reliable liquid chromatography-mass spectrometry method for determination of methotrexate in human plasma and its application to therapeutic drug monitoring.
abstract: A simple, rapid and reliable liquid chromatography-electrospray ionization tandem mass spectrometry method was established and validated for the determination of methotrexate in human plasma. After a straightforward protein precipitation by acetonitrile-water (70:30, v/v), methotrexate (MTX) and p-aminoacetophenone (used as internal standard, IS) were separated on a Column C18 column (50 × 2.1 mm, 3 µm; Column Technology, Fremont, CA, USA) using a gradient elution with mobile phase of acetonitrile and 0.03% acetic acid aqueous solution at a flow rate of 0.5 mL/min. The total chromatographic runtime was 5 min for each injection. Quantification detection was performed in a triple-quadruple tandem mass spectrometer under positive mode monitoring the following mass transitions: m/z 455.3 → 308.3 for MTX and m/z 136.1 → 94.4 for IS. The calibration curve was linear over the range of 0.05-25.0 µmol/L with a lower limit of quantification of 0.05 µmol/L. The intra- and interday precisions were <5.2%, the accuracy varied from -4.1 to 4.5%. The recovery was >94%. The LC-MS/MS method showed an excellent agreement with the existing HPLC-UV method using Passing-Bablok regression and Bland-Altman difference plot analysis. The validated LC-MS/MS can be successfully applied to the routine therapeutic drug monitoring of MTX in clinical laboratories.
authors: Department of Medical Oncology, Fudan University Shanghai Cancer Center.|Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032, China.
authors: Dan
authors: D
authors: Wu
authors: Department of Medical Oncology, Fudan University Shanghai Cancer Center.
authors: Yixuan
authors: Y
authors: Wang
authors: Department of Medical Oncology, Fudan University Shanghai Cancer Center.
authors: Yan
authors: Y
authors: Sun
authors: Department of Medical Oncology, Fudan University Shanghai Cancer Center.
authors: Nian
authors: N
authors: Ouyang
authors: Department of Medical Oncology, Fudan University Shanghai Cancer Center.|Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, 200032, China.
authors: Jun
authors: J
authors: Qian
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000097:blood*; D002851:Chromatography, High Pressure Liquid / Q000191:economics / Q000379:methods*; D016903:Drug Monitoring / Q000191:economics / Q000379:methods; D006801:Humans; D057230:Limit of Detection; D008727:Methotrexate / Q000097:blood*; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000191:economics / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000191:economics / Q000379:methods
chemical_list: D000964:Antimetabolites, Antineoplastic; D008727:Methotrexate
keywords: LC-MS/MS; method comparison; methotrexate; therapeutic drug monitoring