title: Measurement of methotrexate polyglutamates in human erythrocytes by ion-pair UPLC-MS/MS.
abstract: BACKGROUND
  Low-dose methotrexate is used for the treatment of rheumatoid arthritis and juvenile idiopathic arthritis, but its effectiveness greatly varies between individuals. Therapeutic drug monitoring of intracellular methotrexate metabolites, the γ-polyglutamates (MTXGlu(n)), in human erythrocytes has shown promise in providing a basis for individualization of therapy.
  
  
  RESULTS
  This work presents expedient methodology for the analysis of MTXGlu(1-7) in human erythrocytes by ion-pair UPLC with detection by tandem MS (UPLC-ESI-MS/MS). The use of N,N-dimethylheptylamine as an ion-pair agent was found to be favorable over others. Thermal extraction of erythrocyte lysates provides a simple one-step extraction procedure. The entire chromatographic run time is 6 min and the assay was validated within the therapeutic range of these metabolites
  
  
  CONCLUSIONS
  The developed sample preparation procedure in combination with ion-pair UPLC-ESI-MS/MS analysis allowed for expedient quantitation of MTXGlu(1-7) in human erythrocytes. The rapid analysis time would enable therapeutic drug monitoring of MTXGlu(1-7) in the clinic.
authors.affiliation: Division of Clinical Pharmacology & Medical Toxicology, Children's Mercy Hospitals & Clinics, 2401 Gillham Road, Kansas City, MO 64108, USA.
authors.forename: Leon
authors.initials: L
authors.lastname: van Haandel
authors.forename: Mara L
authors.initials: ML
authors.lastname: Becker
authors.forename: T D
authors.initials: TD
authors.lastname: Williams
authors.forename: J Steven
authors.initials: JS
authors.lastname: Leeder
authors.forename: John F
authors.initials: JF
authors.lastname: Stobaugh
mesh_terms: D002851:Chromatography, High Pressure Liquid; D003903:Deuterium / Q000737:chemistry; D004912:Erythrocytes / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007477:Ions / Q000737:chemistry; D007553:Isotope Labeling; D008727:Methotrexate / Q000097:blood*; D010627:Phenethylamines / Q000737:chemistry; D011099:Polyglutamic Acid / Q000097:blood*; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry
chemical_list: D007477:Ions; D010627:Phenethylamines; C043554:N,N-dimethylphenylethylamine; D011099:Polyglutamic Acid; D003903:Deuterium; D008727:Methotrexate