Determination of plasma and serum homocysteine by high-performance liquid chromatography with fluorescence detection

G. Minniti, A. Piana, U. Armani, R. Cerone

Research output: Contribution to journalArticle

Abstract

Determination of homocysteine in plasma or serum is becoming an important diagnostic procedure. Accurate, rapid and low cost methods for measuring homocysteine are therefore required. We have improved an HPLC method and made it suitable for clinical application. The total homocysteine in plasma consists of free homocysteine (i.e., reduced plus oxidized homocysteine in the non-protein fraction of plasma) and protein-bound homocysteine. The method consists of the following steps: reduction of the sample with tri-n-butylphosphine, precipitation of proteins with trichloroacetic acid (10%) and derivatization with ammonium 7-fluorobenzo-2-oxa-1,3-diazole-4-sulfonate. The derivatives are separated by reversed-phase high-performance liquid chromatography followed by fluorescence detection. The concentrations (mean±S.D.) of total homocysteine in plasma from 77 normal subjects, 44 male and 33 female adults, were 8.4±2.15 and 7.1±1.18 μmol/l, respectively. Serum concentrations were 8.8±2.6 μmol/l in males and 7.6±1.5 μmol/l in females. Copyright (C) 1998 Elsevier Science B.V.

Original languageEnglish
Pages (from-to)401-405
Number of pages5
JournalJournal of Chromatography A
Volume828
Issue number1-2
DOIs
Publication statusPublished - Dec 18 1998

Keywords

  • Amino acids
  • Derivatization, LC
  • Homocysteine

ASJC Scopus subject areas

  • Analytical Chemistry

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