Separation of Clenpenterol enantiomers in Capillary Electrophoresis with cyclodextrin-type chiral selectors and study of structure of selector-selectand complexes by using NMR spectroscopy

Author: Elene Tatunashvili
Keywords: cyclodextrin, capillary electrophoresis, NMR spectroscopy, enantiomers, migration order, nuclear overhouser enhancement
Annotation:

The existences of chiral organic molecules have long been known. In parmaceuptical industries approximately 90% of chiral drags are marked as racemates, consisting of an equimolar mixture of two enantiomers. Although the enantiomers have the same chemical connectivity of atoms, they differ in their interactions with enzymes, proteins and receptors. These differences in interactions lead to differences in the biological activities. With this tendency the development of new improved methods for the separation and determination of enantiomers became a permanent necessity and also a challenge. Capillary electrophoresis (CE) is beginning to play a major role in the separation of chiral compounds. In capillary electrophoresis enantioseparation can be achieved only using chiral selectors. Cyclodextrins (CDs) are the important and frequently used class of chiral selectors in CE. In process of enantioseparation, cyclodextrins discriminate between the enantiomers by an enantioselective complexation of the analyte and the chiral selector, giving rise to differences in the electrophoretic mobilities of the enantiomers. In the present work enantioseparation and enantiomer migration order of the chiral β-adrenergic agonist drug clenpenterol in the presence of native cyclodextrins and charged and uncharged derivatives of β-CD as chiral selectors was studied in CE. The reversal of enantiomer migration order(EMO) was observed in case of heptakis(2,3-diacetyl)- β-cyclodextrin and heptakis (2,6-disulfo-3-methyl)-β-cyclodextrin compared with native β-cyclodextrin. 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 complexes enantiomer-chiral selector was rationalized by using NMR spectroscopy experiments, in particular rotating frame nuclear Overhauser experiment (ROESY) and molecular modeling methods. The possible mechanisms of the EMO reversal of the clenpenterol enantiomers depending on the structure of the CD were studied by using H1, HSQC, COSY and 1 D and 2D ROESY experiments. Selective ROESY experiments were carried out to all systems in which reversal of enantiomer migration order was seen in CE runs. Selective irradiations were done with the aim to detect intermolecular NOE interactions, and to detail the structure of the complexes. Thereby, significant differences were observed between the structure of β-cyclodextrin and HDA-β-CD complexes with Clenpenterol and also in case of total energies of binding forces of each enantiomer with CDs.

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