Capillary electrophoretic determination of amino acids improvement by cyclodextrin additives

Abstract

Previously, we have developed methods for the capillary electrophoretic determination (without derivatization) of twenty common amino acids with indirect absorbance detection [Y.-H. Lee and T.I. Lin, J. Chromatogr. A, 680 (1994) 287]. Here, we report a further improvement in the resolution, resulting from the use of various cyclodextrin (CD) additives. Capillary electrophoresis (CE) was performed at pH 11.0 with either p-aminosalicylic acid (PAS) or 4-(N,N′-dimethylamino)-benzoic acid (DMAB) as the background electrolyte (BGE). The effects on the CE separation brought about by various CD additives, e.g. α- and β-cyclodextrins, methyl-, hydroxypropyl-2 2,6-dimethyl-, and 2,3,6-trimethyl-β-cyclodextrins were investigated. These CD additives form inclusion complexes with the BGE and also with amino acids, altering the migration behavior of the analytes. For some amino acids which have previously proved difficult or impossible to separate, an improvement in the separation selectivity has been attained. Association constants of BGEs for cyclodextrins were determined on the basis of the CE mobility changes caused by the CD additive. The concentration of CD additive needed to achieve a suitable separation was calculated and optimized experimentally. Using 10 m M PAS or DMAB, in the presence of 20 m M α-cyclodextrin at pH 11.0, eighteen amino acid peaks were baseline-resolved in under 35 min. Leu and Ile could also be separated in the presence of 15–20 m M β-CD under similar CE conditions. The performance of CE in separating amino acids in the presence of various CD additives is discussed.