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Enantiomeric separation of amino acids using micellar electrokinetic chromatography after pre‐column derivatization with the chiral reagent 1‐(9‐fluorenyl)‐ethyl chloroformate
Author(s) -
Chan King C.,
Muschik Gary M.,
Issaq Haleem J.
Publication year - 1995
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150160183
Subject(s) - chemistry , chloroformate , derivatization , chromatography , micellar electrokinetic chromatography , enantiomer , diastereomer , ethyl chloroformate , sodium dodecyl sulfate , sodium , reagent , detection limit , high performance liquid chromatography , organic chemistry
Direct enantiomeric separations of some racemic amino acids derivatized with 9‐fluorenylmethyl chloroformate were obtained using cyclodextrin‐modified micellar electrokinetic chromatography (CD/MEKC) with a buffer made up of 5 m M sodium borate (pH 9.2), 150 m M sodium dodecyl sulfate (SDS) and 40 m M γ‐CD. Alternatively, enantiomeric separations were also achieved indirectly using MEKC after pre‐column derivatization with (+)‐1‐(9‐fluorenyl) ethyl chloroformate (FLEC). Using either a 10 m M sodium phosphate (pH 6.8) or a 5 m M sodium borate buffer (pH 9.2), each of which contained 25 m M SDS and 10–15% of acetonitrile, FLEC‐derivatized serine, alanine, valine, methionine, leucine, phenylalanine, 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 faster than the corresponding L ‐isomers. Also, the diastereomers of aspartic acid, glutamic acid, and proline were resolved using a 10 m M sodium citrate buffer (pH 4.4). Using KrF (248 nm) laser‐induced fluorescence, the detection limit of (+)‐FLEC derivatized DL ‐amino acids was obtained at the n M level, which was about 100 × more sensitive than UV absorption at 200 nm. Analyte concentrations as low as 3 × 10 −8 M ( DL ‐Val) could be derivatized with (+)‐FLEC.