title: Rapid measurement of total plasma homocysteine by HPLC.
abstract: BACKGROUND
  Determination of plasma homocysteine has gained increasing interest during the past few years. Several HPLC methods for determination of homocysteine are available. Based on these methods, we developed a new HPLC assay for rapid and sensitive measurement of total plasma homocysteine.
  
  
  METHODS
  As a reducing reagent tris-(2-carboxylethyl)-phosphine is used, ammonium 7-fluorobenzo-2-oxa-1,3-diazole-4-sulfonate serves as the derivatization agent. Separation is performed by reversed-phase HPLC using a precolumn and a 55-mm RP(18) cartridge; mobile phase: 0.1 mol/l KH(2)PO(4) with 5% methanol, adjusted to pH 2.7 with ortho-phosphoric acid, flow-rate 1.1 ml/min.
  
  
  RESULTS
  Homocysteine is clearly separated from other thiols, the retention time being 2.2 min, total analysis time is 6 min. Tests for linearity, recovery and precision are satisfactory, as well as the comparison with a commercial available assay method. Detection limit of the method is 0.5 micro mol/l, it could be further enhanced for measurements of even lower homocysteine concentrations in, e.g., cell culture supernatants.
  
  
  CONCLUSIONS
  The described method is well suited for analysis of thiols in blood specimens. It is more convenient and more rapid than methods described earlier.
authors.affiliation: Institute for Medical Chemistry and Biochemistry, University of Innsbruck, A-6020, Innsbruck, Austria.
authors.forename: Barbara
authors.initials: B
authors.lastname: Frick
authors.forename: Katharina
authors.initials: K
authors.lastname: Schröcksnadel
authors.forename: Gabriele
authors.initials: G
authors.lastname: Neurauter
authors.forename: Barbara
authors.initials: B
authors.lastname: Wirleitner
authors.forename: Erika
authors.initials: E
authors.lastname: Artner-Dworzak
authors.forename: Dietmar
authors.initials: D
authors.lastname: Fuchs
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006710:Homocysteine / Q000097:blood*; D006801:Humans; D007202:Indicators and Reagents; D010084:Oxidation-Reduction; D010720:Phosphines; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013438:Sulfhydryl Compounds / Q000097:blood
chemical_list: D007202:Indicators and Reagents; D010720:Phosphines; D013438:Sulfhydryl Compounds; D006710:Homocysteine