Separation of Amino Acid Enantiomers Using Precolumn Derivatization with o-Phthalaldehyde and 2,3,4,6-Tetra-O-acetyl-1-thio-β-glucopyranoside

A method is described for resolution of amino acid enantiomers. The D- and L-amino acids were reacted with o-phthalaldehyde (OPA) and the optically active 2,3,4,6-tetra-O-acetyl-1-thio-β-glucopyranoside (TATG). The reaction was complete in a few minutes at room temperature and the derivatives were quite stable. The formed diastereomers were separated by reversed phase chromatography and the selectivity was generally good, except for lysine and ornithine. A mean separation factor (α) of 1.27 was obtained with acetonitrile as an organic modifier. The fluorescence excitation and emission maxima were at 342 nm and 410 nm respectively, and the electrochemical half-wave potential E½ ≡ 0.65–0.75 V. The detection limits (for L-leucine) were 23 pmol and 1 pmol (S/N 3:1) in the respective detection modes. With laser-induced fluorescence detection (He-Cd laser, 325 nm) and microcolumns, a detection limit in the fmol range is obtainable.