title: Determination of coumarin-type anticoagulants in human plasma by HPLC-electrospray ionization tandem mass spectrometry with an ion trap detector.
abstract: BACKGROUND
  Coumarin-type anticoagulants are used for the long-term treatment and prevention of thromboembolic disorders. The identification of these drugs is crucial in patients with an increased prothrombin time of unknown origin. The aim of this study was to develop a sensitive and specific method for the simultaneous determination of phenprocoumon, acenocoumarol, and warfarin in human plasma by HPLC-electrospray ionization tandem mass spectrometry.
  
  
  METHODS
  After addition of the internal standard, p-chlorowarfarin, plasma samples were extracted using Oasis MCX solid-phase extraction cartridges. The compounds were separated on a Symmetry C18 column (Waters) with a mobile phase of acetonitrile-1 g/L formic acid (75:25 by volume) at a flow rate of 0.5 mL/min.
  
  
  RESULTS
  Extraction and separation of the three drugs and the internal standard were accomplished in 9 min. The overall extraction efficiency was >89% for all three compounds. The limits of detection were 1 microg/L for phenprocoumon and warfarin and 10 microg/L for acenocoumarol. Regression analysis of the calibration data revealed good correlation (r(2) >or=0.995) for all compounds. Within-run accuracies for quality-control samples were +/- 1% to 7% of the target concentration, with CVs <9%.
  
  
  CONCLUSIONS
  The proposed method enables the unambiguous identification and quantification of phenprocoumon, warfarin, and acenocoumarol in both clinical and forensic specimens. This method combines a new, rapid solid-phase extraction procedure with an extremely fast chromatographic analysis, which is especially advantageous for clinical laboratories.
authors.affiliation: Institute of Forensic Medicine, Karl-Franzens University Graz, Universitätsplatz 4, A8010 Graz, Austria. manfred.kollroser@kfunigraz.ac.at
authors.forename: Manfred
authors.initials: M
authors.lastname: Kollroser
authors.forename: Caroline
authors.initials: C
authors.lastname: Schober
mesh_terms: D015110:4-Hydroxycoumarins / Q000097:blood*; D000074:Acenocoumarol / Q000097:blood; D000925:Anticoagulants / Q000097:blood*; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D010644:Phenprocoumon / Q000097:blood; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization; D014859:Warfarin / Q000097:blood
chemical_list: D015110:4-Hydroxycoumarins; D000925:Anticoagulants; D014859:Warfarin; D000074:Acenocoumarol; D010644:Phenprocoumon