Development of an assay for methotrexate and its metabolites 7-hydroxy methotrexate and DAMPA in serum by LC-MS/MS Journal Article

  


Authors: Schofield, R. C.; Ramanathan, L. V.; Murata, K.; Fleisher, M.; Pessin, M. S.; Carlow, D. C.
Article Title: Development of an assay for methotrexate and its metabolites 7-hydroxy methotrexate and DAMPA in serum by LC-MS/MS
Abstract: Methotrexate (MTX) is a folic acid antagonist that is widely used as an immunosuppressant and chemotherapeutic agent. After high-dose administration of MTX serum levels must be monitored to determine when to administer leucovorin, a folic acid analog that bypasses the enzyme inhibition caused by MTX and reverses its toxicity. We describe a rapid and simple turbulent flow liquid chromatography (TFLC) method implementing positive heated electrospray ionization (HESI) for the accurate and precise determination of MTX, 7-hydroxymethotrexate (7-OH MTX), and 4-amino-4-deoxy-N-10-methylpteroic acid (DAMPA) concentrations in serum. MTX is isolated from serum samples (100 mu L) after protein precipitation with a methanolic solution containing internal standard (MTX-D-3) followed by centrifugation. The supernatant is injected into the turbulent flow liquid chromatography which is followed by electrospray positive ionization tandem mass spectrometry (TFLC-ESI-MS/ MS) and quantified using a six-point calibration curve. For MTX, 7-OH MTX, and DAMPA the assays were linear from 20 to 1000 nmol/L. Dilutions of 10-, 100-, and 1000-fold were validated giving a clinically reportable range of 20 to 1.0 x 10(6) nmol/L. Within-day and between-day precisions at concentrations spanning the analytical measurement ranges were less than 10 % for all three analytes.
Keywords: methotrexate; plasma; tandem mass-spectrometry; high-dose methotrexate; mass; spectrometry; therapeutic drug monitoring; carboxypeptidase-g2; turbulent flow liquid chromatography
Journal Title: Methods in Molecular Biology
Volume: 1383
ISSN: 1064-3745
Publisher: Humana Press Inc  
Publication Place: New York, NY
Date Published: 2016-01-01
Start Page: 213
End Page: 222
Language: English
ACCESSION: WOS:000368837100024
PROVIDER: wos
DOI: 10.1007/978-1-4939-3252-8_23
PUBMED: 26660190
Notes: Article; Book Chapter 23 in "Clinical Applications of Mass Spectrometry in Drug Analysis: Methods and Protocols" (ISBN: 978-1-4939-3251-1)-- 999 RIVERVIEW DR, STE 208, TOTOWA, NJ 07512-1165 USA -- Source: Wos
Altmetric
What is Altmetric?
Citation Impact
What is Dimensions Citation Badge?
BMJ Impact Analytics
MSK Authors
  1. Kazunori Murata
    33 Murata
  2. Martin Fleisher
    312 Fleisher
  3. Melissa Sue Pessin-Minsley
    50 Pessin-Minsley
  4. Lakshmi Vulimiri Ramanathan
    77 Ramanathan
  5. Ryan Schofield
    29 Schofield
  6. Dean Christian Carlow
    42 Carlow
Related MSK Work