We use cookies to improve your experience. By continuing to browse this site, you accept our cookie policy.×
Research Article

Ultrafast selective quantification of methotrexate in human plasma by high-throughput MALDI-isotope dilution mass spectrometry

    ,
    Ethan den Boer

    Department of Clinical Chemistry, University Medical Center Rotterdam (ErasmusMC), The Netherlands

    ,
    Ron AA Mathot

    Department of Pharmacy, Clinical Pharmacology Unit, University of Amsterdam, The Netherlands

    ,
    Robert de Jonge

    Department of Clinical Chemistry, University Medical Center Rotterdam (ErasmusMC), The Netherlands

    ,
    Rob J van Klaveren

    Department of Pulmonology, University Medical Center Rotterdam, The Netherlands

    ,
    Jan Lindemans

    Department of Clinical Chemistry, University Medical Center Rotterdam (ErasmusMC), The Netherlands

    &
    Theo M Luider

    Laboratories of Neuro-Oncology & Clinical & Cancer Proteomics, Department of Neurology, University Medical Center Rotterdam (ErasmusMC), Dr. Molewaterplein 50, Room Ee-1981, 3015 GE Rotterdam, The Netherlands

    Published Online:https://doi.org/10.4155/bio.11.113

    Background: A new analytical MS method using isotope dilution combined with MALDI-triple quadrupole MS/MS has been developed and validated for the determination of methotrexate and 7-hydroxymethotrexate in plasma. Methotrexate, methotrexate-d3, 7-hydroxymethotrexate and 7-hydroxymethotrexate-d3 were monitored by selected reaction monitoring using the transitions m/z 455.2→308.2, 458.2→311.2, 471.2→324.2 and 474.2→327.2 for methotrexate, methotrexate-d3, 7-hydroxymethotrexate and 7-hydroxymethotrexate-d3, respectively. Results: The LLOQ was 1 nmol/l for methotrexate and 7-hydroxymethotrexate while the limit of detection was 0.3 nmol/l for both analytes. The new developed method was cross-validated by a fluorescence polarization immunoassay and tested for its clinical feasibility by measuring plasma samples from patients suffering from acute lymphoblastic leukemia. Plasma methotrexate concentrations ranged between 66.0 and 954 nmol/l and observed 7-hydroxymethotrexate/methotrexate ratios ranged between 0.1 and 32.4, respectively. Conclusion: The new method showed comparable analytical performances as the fluorescence polarization immunoassay, but analyte specificity and sensitivity of the newly developed method were significantly better.

    Papers of special note have been highlighted as: ▪ of interest

    Bibliography

    • Jolivet J, Cowan KH, Curt GA, Clendeninn NJ, Chabner BA. The pharmacology and clinical use of methotrexate. N. Engl. J. Med.309(18),1094–1104 (1983).
    • Warren RB, Chalmers RJ, Griffiths CE, Menter A. Methotrexate for psoriasis in the era of biological therapy. Clin. Expl Dermatol.33(5),551–554 (2008).
    • Luqmani R, Hennell S, Estrach C et al. British Society for Rheumatology and British Health Professionals in rheumatology guideline for the management of rheumatoid arthritis (the first two years). Rheumatology45(9),1167–1169 (2006).
    • Pincus T, Sokka T. Should aggressive therapy for rheumatoid arthritis require early use of weekly low-dose methotrexate, as the first disease-modifying anti-rheumatic drug in most patients? Rheumatology45(5),497–499 (2006).
    • Lennard L. Therapeutic drug monitoring of cytotoxic drugs. Br. J. Clin. Pharmacol.52(Suppl. 1),75S–87S (2001).
    • Li H, Luo W, Zeng Q, Lin Z, Luo H, Zhang Y. Method for the determination of blood methotrexate by high performance liquid chromatography with online post-column electrochemical oxidation and fluorescence detection. J. Chromatogr. B Analy. Technol. Biomed. Life Sci.845(1),164–168 (2007).▪ [6–22] describe common analytical techniques for methotrexate determination in biological matrices.
    • Albertioni F, Pettersson B, Beck O, Rask C, Seideman P, Peterson C. Simultaneous quantitation of methotrexate and its two main metabolites in biological fluids by a novel solid-phase extraction procedure using high-performance liquid chromatography. J. Chromatogr. B Biomed. Appl.665(1),163–170 (1995).
    • Nelson JA, Harris Ba, Decker WJ, Farquhar D. Analysis of methotrexate in human plasma by high-pressure liquid chromatography with fluorescence detection. Cancer Research37(11),3970–3973 (1977).
    • Belz S, Frickel C, Wolfrom C, Nau H, Henze G. High-performance liquid chromatographic determination of methotrexate, 7-hydroxymethotrexate, 5-methyltetrahydrofolic acid and folinic acid in serum and cerebrospinal fluid. J. Chromatogr. B Biomed. Appl.661(1),109–118 (1994).
    • 10  Barbieri A, Sabatini L, Indiveri P, Bonfiglioli R, Lodi V, Violante FS. Simultaneous determination of low levels of methotrexate and cyclophosphamide in human urine by micro liquid chromatography/electrospray ionization tandem mass spectrometry. Rapid Commun. Mass Spectrom.20(12),1889–1893 (2006).
    • 11  Przybylski M, Preiss J, Dennebaum R, Fischer J. Identification and quantitation of methotrexate and methotrexate metabolites in clinical high-dose therapy by high pressure liquid chromatography and field desorption mass spectrometry. Biomed. Mass Spectrometry9(1),22–32 (1982).
    • 12  Turci R, Fiorentino ML, Sottani C, Minoia C. Determination of methotrexate in human urine at trace levels by solid phase extraction and high-performance liquid chromatography/tandem mass spectrometry. Rapid Commun. Mass Spectrom.14(3),173–179 (2000).
    • 13  Kuo Cy, Wu HL, Kou Hs, Chiou SS, Wu DC, Wu SM. Simultaneous determination of methotrexate and its eight metabolites in human whole blood by capillary zone electrophoresis. J. Chromatogr. A1014(1–2),93–101 (2003).
    • 14  Hayashi H, Fujimaki C, Tsuboi S, Matsuyama T, Daimon T, Itoh K. Application of fluorescence polarization immunoassay for determination of methotrexate-polyglutamates in rheumatoid arthritis patients. Tohoku J. Exp. Med.215(1),95–101 (2008).
    • 15  Paxton JW, Rowell FJ. A rapid, sensitive and specific radioimmunoassay for methotrexate. Clinica Chimica Acta80(3),563–572 (1977).
    • 16  Hendel J, Sarek LJ, Hvidberg EF. Rapid radioimmunoassay for methotrexate in biological fluids. Clin. Chem.22(6),813–816 (1976).
    • 17  Stout M, Ravindranath Y, Kauffman R. High-performance liquid chromatographic assay for methotrexate utilizing a cold acetonitrile purification and separation of plasma or cerebrospinal fluid. J. Chromatogr.342(2),424–430 (1985).
    • 18  Steinborner S, Henion J. Liquid–liquid extraction in the 96-well plate format with SRM LC/MS quantitative determination of methotrexate and its major metabolite in human plasma. Anal. Chem.71(13),2340–2345 (1999).
    • 19  Cociglio M, Hillaire-Buys D, Alric C. Determination of methotrexate and 7-hydroxymethotrexate by liquid chromatography for routine monitoring of plasma levels. J. Chromatogr. B Biomed. Appl.674(1),101–110 (1995).
    • 20  Mccrudden EA, Tett SE. Improved high-performance liquid chromatography determination of methotrexate and its major metabolite in plasma using a poly(styrene-divinylbenzene) column. J. Chromatogr.721(1),87–92 (1999).
    • 21  Emara S, Askal H, Masujima T. Rapid determination of methotrexate in plasma by high-performance liquid chromatography with online solid-phase extraction and automated precolumn derivatization. Biomed. Chromatogr.12(6),338–342 (1998).
    • 22  Aboleneen H, Simpson J, Backes D. Determination of methotrexate in serum by high-performance liquid chromatography. J. Chromatogr. B Biomed. Appl.681(2),317–322 (1996).
    • 23  Meesters RJ, Cornelissen R, Van Klaveren RJ et al. A new ultrafast and high-throughput mass spectrometric approach for the therapeutic drug monitoring of the multi-targeted anti-folate pemetrexed in plasma from lung cancer patients. Anal. Bioanal. Chem.398,2943–2948 (2010).▪▪ [23–34] describe the use of MALDI-triple quadrupole mass spectrometry in biological application.
    • 24  Wagner M, Varesio E, Hopfgartner G. Ultra-fast quantitation of saquinavir in human plasma by matrix-assisted laser desorption/ionization and selected reaction monitoring mode detection. J. Chromatog.872(1–2),68–76 (2008).
    • 25  Volmer Da, Sleno L, Bateman K et al. Comparison of MALDI to ESI on a triple quadrupole platform for pharmacokinetic analyses. Anal. Chem.79(23),9000–9006 (2007).
    • 26  Meesters RJ, Van Kampen JJ, Reedijk ML et al. Ultrafast and high-throughput mass spectrometric assay for therapeutic drug monitoring of antiretroviral drugs in pediatric HIV-1 infection applying dried blood spots. Anal. Bioanal. Chem.398(1),319–328 (2010).
    • 27  Van Kampen JJ, Reedijk ML, Burgers PC et al. Ultrafast analysis of plasma and intracellular levels of HIV protease inhibitors in children. a clinical application of MALDI mass spectrometry. PloS One5(7),e11409 (2010).
    • 28  Meesters RJ, Van Kampen JJ, Scheuer RD, van der Ende ME, Gruters RA, Luider TM. Determination of the antiretroviral drug tenofovir in plasma from HIV-infected adults by ultrafast isotope dilutionMALDI-triple quadrupole tandem mass spectrometry. J. Mass Spectrom.46,282–289 (2011).
    • 29  Hatsis P, Brombacher S, Corr J, Kovarik P, Volmer DA. Quantitative analysis of small pharmaceutical drugs using a high repetition rate laser matrix-assisted laser/desorption ionization source. Rapid Commun. Mass Spectrom.17(20),2303–2309 (2003).
    • 30  Sleno L, Volmer DA. Some fundamental and technical aspects of the quantitative analysis of pharmaceutical drugs by matrix-assisted laser desorption/ionization mass spectrometry. Rapid Commun. Mass Spectrom.19(14),1928–1936 (2005).
    • 31  Porta T, Grivet C, Knochenmuss R, Varesio E, Hopfgartner G. Alternative CHCA-based matrices for the analysis of low molecular weight compounds by UV-MALDI-tandem mass spectrometry. J. Mass Spectrom.46(2),144–152 (2011).
    • 32  Rathore R, Corr J, Scott G, Vollmerhaus P, Greis KD. Development of an inhibitor screening platform via mass spectrometry. J. Biomol. Screen.13(10),1007–1013 (2008).
    • 33  Wolfe RR, Chinkes DL. Determination of isotope enrichment. In: Isotope Tracers in Metabolic Research – Principles and Practice of Kinetic Analysis (Second Edition). John Wiley & Sons, Hoboken, NJ, USA, 93–113 (2005).
    • 34  Van Kampen JJ, Burgers Pc, Gruters RA et al. Quantitative analysis of antiretroviral drugs in lysates of peripheral blood mononuclear cells using MALDI-triple quadrupole mass spectrometry. Anal. Chem.80(13),4969–4975 (2008).
    • 35  Farquhar D, Loo TL. Synthesis and biologic evaluation of 7-hydroxymethotrexate, 7-methylaminopterin, and 7-methylmethotrexate. J. Med. Chem.15(5),567–569 (1972).
    • 36  Altman DG, Bland Jm. Measurements in medicine: the analysis of method comparison studies. The Statistician32(3),307–317 (1983).
    • 37  Bland JM, Altman DG. Comparing methods of measurement. Why plotting difference against standard method is misleading. Lancet346(8982),1085–1087 (1995).
    • 38  Rubino FM. Separation methods for methotrexate, its structural analogues and metabolites. J. Chromatogr. B Biomed. Sci. Appl.764(1–2),217–254 (2001).
    • 39  Khoo SH, Gibbons SE, Back DJ. Therapeutic drug monitoring as a tool in treating HIV infection. AIDS15(Suppl. 5),S171–S181 (2001).
    • 40  Erttmann R, Bielack S, Landbeck G. 7-Hydroxy-methotrexate and clinical toxicity following high-dose methotrexate therapy. J. Cancer Res. Clin. Oncol.109(1),86–88 (1985).
    • 101  CDER. Center for Drug Evaluation and Research. Guidance for Industry. Bioanalytical Method Validation (2001). www.fda.gov/downloads/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/UCM070107.pdf