ENANTIOMERIC SEPARATION OF AMINO-ACIDS USING MICELLAR ELECTROKINETIC CHROMATOGRAPHY AFTER PRECOLUMN DERIVATIZATION WITH THE CHIRAL REAGENT 1-(9-FLUORENYL)ETHYL CHLOROFORMATE

Direct enantiomeric separations of some racemic amino acids derivatize d with 9-fluorenylmethyl chloroformate were obtained using cyclodextri n-modified micellar electrokinetic chromatography (CD/MEKC) with a buf fer made up of 5 mM sodium berate (pH 9.2), 150 mM sodium dodecyl sulf ate (SDS) and 40 mM gamma-CD. Alternatively, enantiomeric separations were also achieved indirectly using MEKC after pre-column derivatizati on with (+)-1-(9-fluorenyl) ethyl chloroformate (FLEC). Using either a 10 mM sodium phosphate (pH 6.8) or a 5 mM sodium berate buffer (pH 9. 2), each of which contained 25 mM SDS and 10-15% of acetonitrile, FLEC -derivatized serine, alanine, valine. methionine, leucine, phenylalani ne, tryptophan, and their diastereomeric pairs were all separated: the L-isomers migrated faster than the corresponding D-isomers. However, when (-)-FLEC was used for derivatization, the D-isomers migrated fast er than the corresponding L-isomers. Also, the diastereomers of aspart ic acid, glutamic acid, and proline were resolved using a 10 mM sodium citrate buffer (pH 4.4). Using KrF (248 nm) laser-induced fluorescenc e, the detection limit of (+)-FLEC derivatized DL-amino acids was obta ined at the nM level, which was about 100 X more sensitive than UV abs orption at 200 nm. Analyte concentrations as low as 3 X 10(-8) M (DL-V al) could be derivatized with (+)-FLEC.