Quantitation of amino acids in plasma by high performance liquid chromatography: Simultaneous deproteinization and derivatization with 9-fluorenylmethyloxycarbonyl chloride

A. Jámbor, I. Molnár-Perl

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

This paper, as a novelty to this field, presents the deproteinization and derivatization of plasma's free amino acids (PFAAs), simultaneously, in a single step, with the acetonitrile (ACN) containing 9-fluorenylmethyloxycarbonyl chloride (FMOC) reagent. Deproteinization and derivatization, were studied with 22 amino acids, applying photodiode array (DAD) and fluorescence (FL) detection, simultaneously. Model investigations have been carried out as a function of the FMOC concentration, reaction time and reaction conditions: with standard solutions, with human plasma samples in its initial condition and fortified with standard amino acids (excluding tryptophan because it co-elutes with the hydrolyzed FMOC). Reproducibilities of 22 amino acids, including both histidine and tyrosine derivatives, obtained under optimum derivatization conditions are presented (at 3.0 mM FMOC concentration, at pH 9; derivatization time = 20 min), and characterized with the relative standard deviation percentages of their responses (≤4.4%, RSD). Quantitation limit (LOQ) of amino acid FMOC derivatives proved to be 2.5 pmol, except for cystine, ornithine (5 pmol) and for the total of tyrosines (N-FMOC-tyrosine and N,O-FMOC-tyrosine 10 pmol).

Original languageEnglish
Pages (from-to)6218-6223
Number of pages6
JournalJournal of Chromatography A
Volume1216
Issue number34
DOIs
Publication statusPublished - Aug 21 2009

Keywords

  • 9-Fluorenylmethyloxycarbonyl chloride
  • Amino acids
  • Deproteinization
  • Derivatization
  • HPLC-UV(DAD)/fluorescence detection
  • High performance liquid chromatography
  • Plasma

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Organic Chemistry

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