A Low Picomole Fluorometric Detection System for Amino Acid Analysis

An amino acid detection system was developed for low picomole analysis of amino acids including proline, based on OPA-post-labeling with a non-switching NaCIO flow system. A computer-controlled HPLC equipped with a three-solvent gradient mixer was assembled with the detection system to analyze 17 kinds of amino acids in 85 min with a stable base line. The optimum conditions of the NaCIO and OPA solutions were determined as 0.002%–0.22ml/min and 0.16%–0.26ml/min, respectively. The use of NaCIO caused only approximately 30% decrease in fluorescence response of the usual amino acids, except for Pro and Cys, the latter even being enhanced by about 10 times. The detection limit for Pro and Cys was 500 and 1000 fmol, respectively, and that of the usual amino acids was 100 fmol. The calibration curves of all amino acids showed good linearity in a range between 5 and 500 pmol. This system was used as a detector for enantiomeric analysis of glutamic acid and cysteic acid with a reversed phase HPLC during stereochemical studies on the natural peptide derivative, oxycadystin.