Separation of enantiomers of tetramisole by capillary electrophoresis method and study of separation mechanisms

A signficant part of biological processes is based on chiral recognition. Therefore,these recognition mechanisms are widely studied. One of the valuable instrumental methods for such studies is capillary electrophoresis (CE). The addition of chiral selector is necessary to separate the enantiomers. For this purpose, native and substituted cyclodextrins (CD) can be used. CDs can bind enantiomers selectively and thus lead to a difference in their migration speed. Thus, the thermodynamic selectivity of recognition 1.01 is sufficient for observing baseline resolved peaks in CE while this is not the case in chromatographic techniques even with the most advanced packing materials, column technologies and instrumentation. In the present study CE was used for separation of enantiomers of tetramisole with β-CD and its derivative, heptakis-(2,3-di-O-methyl)-β-CD, heptakis-(2,3-di-O-acethyl)-β-CD and heptakis-(2-O-methyl-3-O-acethyl)-β-CD as chiral selectors. Separation of enantiomers was performed in fused-silica capillary of 50 µm ID and 24 and 32.5 cm, effective and total lengths, respectively. The background electrolyte was 100 mM triethanolamine phosphate with pH=3.0. The reversal of enantiomer migration order(EMO) was observed in case of heptakis(2,3-di-O-acetyl)- β-cyclodextrin and heptakis (2,3-di-O-methyl)-β-cyclodextrin compared with native β-cyclodextrin and heptakis-(2-O-methyl-3-O-acethyl)-β-CD. Reversal of EMO can be achieved by changes in the electrophoretic mobilities of the complexes formed between the chiral selector and each enantiomer which can be caused by modifications in the molecular recognition mechanisms between enantiomer and chiral selector. On the next stage of our investigation, the attempt was made to explain complexation mechanism and evaluate the intermolecular forces which take part in chiral recognition. Therefore, in order to understand the fine intermolecular interactions between analyte and selector and evaluate nature of intermolecular forces, the structure of the enantiomer-chiral selector complexes was rationalized by using NMR spectroscopy experiments, in particular rotating frame nuclear Overhauser experiment (ROESY) and molecular modeling methods.