Journal
BIOANALYSIS
Volume 3, Issue 12, Pages 1369-1378Publisher
FUTURE SCI LTD
DOI: 10.4155/BIO.11.113
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Funding
- Dutch Reumafonds [09-1-402]
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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.
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