{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,6]],"date-time":"2026-09-06T05:28:17Z","timestamp":1788672497915,"version":"build-2803163510"},"reference-count":32,"publisher":"Wiley","issue":"23","license":[{"start":{"date-parts":[[2009,11,9]],"date-time":"2009-11-09T00:00:00Z","timestamp":1257724800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Rapid Comm Mass Spectrometry"],"published-print":{"date-parts":[[2009,12,15]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>\n                    The folate antagonist methotrexate is commonly used in low dose for treatment of rheumatoid arthritis and juvenile idiopathic arthritis. Therapeutic effects are attributed to intracellular levels of various methotrexate polyglutamates. The present methodology, combining a simple preparation step with ion\u2010pairing reversed\u2010phase liquid chromatography and electrospray ionization mass spectrometry, is suitable for the measurement of methotrexate and its polyglutamates\n                    <jats:sub>2\u20137<\/jats:sub>\n                    , in human red blood cells. Sample preparation consists of perchloric acid protein precipitation followed by solid\u2010phase extraction. Baseline separation of all analytes was achieved within 10\u2009min using a Phenomenex Synergy C18 column together with a gradient solvent program obtained from blending acetonitrile with pH 7.5, 5\u2009mM aqueous dimethylhexylamine. Seven methotrexate polyglutamates were detected using multiple reaction monitoring, with the mass spectrometer operating in positive ion mode. Using 20\u2009\u00b5L injection volumes, limits of detection were 2.5\u2009nM for individual methotrexate polyglutamates, while large volume (100\u2009\u00b5L) injections led to detection limits of 0.5\u2009nM and linear calibration from 0.5 to 100\u2009nM for individual analytes. Finally, the presented methodology was applied for the analysis of methotrexate and its polyglutamates in red blood cells obtained from patients being treated for juvenile idiopathic arthritis with methotrexate. Significantly, the methodology proved suitable for determination of long\u2010chain methotrexate polyglutamates\n                    <jats:sub>5\u20137<\/jats:sub>\n                    and further, appears to be superior with respect to sensitivity, selectivity and speed as compared to all previously described approaches. Copyright \u00a9 2009 John Wiley &amp; Sons, Ltd.\n                  <\/jats:p>","DOI":"10.1002\/rcm.4300","type":"journal-article","created":{"date-parts":[[2009,11,9]],"date-time":"2009-11-09T05:25:07Z","timestamp":1257744307000},"page":"3693-3702","source":"Crossref","is-referenced-by-count":42,"title":["A novel high\u2010performance liquid chromatography\/mass spectrometry method for improved selective and sensitive measurement of methotrexate polyglutamation status in human red blood cells"],"prefix":"10.1002","volume":"23","author":[{"given":"Leon","family":"van Haandel","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mara L.","family":"Becker","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"J. 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