Quantitation of amino acids in plasma by high performance liquid chromatography: simultaneous deproteinization and derivatization with 9-fluorenylmethyloxycarbonyl chloride.
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 (<or=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).
Jámbor
A
A
Institute of Chemistry, Department of Analytical Chemistry, L. Eötvös University, H-1518, Budapest 112, Hungary.
Molnár-Perl
I
I
D000368:Aged; D000596:Amino Acids / Q000097:blood*; D053000:Analytic Sample Preparation Methods / Q000379:methods*; D001943:Breast Neoplasms / Q000097:blood / Q000601:surgery; D002851:Chromatography, High Pressure Liquid; D005260:Female; D005449:Fluorenes; D006801:Humans; D007202:Indicators and Reagents; D008875:Middle Aged; D015203:Reproducibility of Results
C496789:9-fluorenylmethoxycarbonyl; D000596:Amino Acids; D005449:Fluorenes; D007202:Indicators and Reagents