Separation of amino acid and peptide stereoisomers by nonionic micelle- mediated capillary electrophoresis after chiral derivatization

Enantiomers of amino acids and peptides were derivatized with a fluorescent chiral reagent 4-(3-isothiocyanatopyrrolidin-1-yl)-7-nitro- 2,1,3-benzoxadiazole [R-(-)- or S-(+)-NBD-PyNCS] and the resulting diastereomeric derivatives separated by capillary electrophoresis (CE). The CE running buffer consisted of 25 mM acetate buffer (pH 4) and 10 mM of the nonionic surfactant Triton X-100. The excitation maximum of NBD-PyNCS at 480 nm matches the major Ar-ion emission line at 488 spiked (at 10^-4 M and 10^{- 5} M concentrations, respectively) into complex biological matrices (rabbit serum and homogenate of Aplysia californica buccal ganglion) are detected without matrix interferences. This method has also been applied to the determination of D- and L-amino acid residues in peptides after acid hydrolysis. Results from the chiral analysis of the naturally-occuring peptide, gramicidin D, are shown.