title: Stable-isotope dilution liquid chromatography-electrospray injection tandem mass spectrometry method for fast, selective measurement of S-adenosylmethionine and S-adenosylhomocysteine in plasma.
abstract: BACKGROUND
  It has been postulated that changes in S-adenosylhomocysteine (AdoHcy), a potent inhibitor of transmethylation, provide a mechanism by which increased homocysteine causes its detrimental effects. We aimed to develop a rapid and sensitive method to measure AdoHcy and its precursor S-adenosylmethionine (AdoMet).
  
  
  METHODS
  We used stable-isotope dilution liquid chromatography-electrospray injection tandem mass spectrometry (LC-ESI-MS/MS) to measure AdoMet and AdoHcy in plasma. Acetic acid was added to prevent AdoMet degradation. Solid-phase extraction (SPE) columns containing phenylboronic acid were used to bind AdoMet, AdoHcy, and their internal standards and for sample cleanup. An HPLC C(18) column directly coupled to the LC-MS/MS was used for separation and detection.
  
  
  RESULTS
  In plasma samples, the interassay CVs for AdoMet and AdoHcy were 3.9% and 8.3%, and the intraassay CVs were 4.2% and 6.7%, respectively. Mean recoveries were 94.5% for AdoMet and 96.8% for AdoHcy. The quantification limits were 2.0 and 1.0 nmol/L for AdoMet and AdoHcy, respectively. Immediate acidification of the plasma samples with acetic acid prevented the observed AdoMet degradation. In a group of controls (mean plasma total Hcy, 11.2 mumol/L), plasma AdoMet and AdoHcy were 94.5 and 12.3 nmol/L, respectively.
  
  
  CONCLUSIONS
  Stable-isotope dilution LC-ESI-MS/MS allows sensitive and rapid measurement of AdoMet and AdoHcy. The SPE columns enable simple cleanup, and no metabolite derivatization is needed. The instability of AdoMet is a serious problem and can be prevented easily by immediate acidification of samples.
authors.affiliation: Laboratory of Pediatrics and Neurology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands.
authors.forename: Henkjan
authors.initials: H
authors.lastname: Gellekink
authors.forename: Dinny
authors.initials: D
authors.lastname: van Oppenraaij-Emmerzaal
authors.forename: Arno
authors.initials: A
authors.lastname: van Rooij
authors.forename: Eduard A
authors.initials: EA
authors.lastname: Struys
authors.forename: Martin
authors.initials: M
authors.lastname: den Heijer
authors.forename: Henk J
authors.initials: HJ
authors.lastname: Blom
mesh_terms: D002853:Chromatography, Liquid; D003903:Deuterium; D006801:Humans; D011865:Radioisotope Dilution Technique; D012016:Reference Values; D012435:S-Adenosylhomocysteine / Q000097:blood*; D012436:S-Adenosylmethionine / Q000097:blood*; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D012436:S-Adenosylmethionine; D012435:S-Adenosylhomocysteine; D003903:Deuterium