Micellar Electrokinetic Capillary Chromatography as a Tool for Cytochrome P450 2C9 Activity Study
| Authors | |
|---|---|
| Year of publication | 2007 |
| Type | Article in Proceedings |
| Conference | HPLC 2007. Abstract Book. |
| MU Faculty or unit | |
| Citation | |
| Web | 31st International Symposium on High Performance Liquid Phase Separations and Related Techniques - HPLC 2007 |
| Field | Analytic chemistry |
| Keywords | MEKC cytochrome P450 |
| Description | Cytochrome P450 2C9 (CYP2C9) is one of the most important isoform in human liver involved in the metabolism of a large number of therapeutic agents. Predominantly high performance liquid chromatography (HPLC) with UV and/or fluorescence detection is chosen for these studies. By reason of sample saving and minor time demands, the possibility of automation, CE analysis is suitable for studies comprising work with enzymes, substrates as well as inhibitors. The applicability of capillary electrophoresis (CE) for the determination of enzymatic activity of CYP2C9 with diclofenac as a probe substrate is demonstrated. Micellar electrokinetic capillary chromatography (MEKC) with SDS as a pseudostationary phase was used for this purpose. The kinetic study on the given enzymatic reaction was also performed since basic characterization of drug biotransformation generally begins with the enzyme kinetic analysis of metabolite formation. As a result Michaelis constant and maximum reaction velocity were evaluated, the values 3.44 uM respectively 19.78 nmol.min-1.nmol-1 were in agreement with the literature data. Moreover, the proposed method is suitable for assessing of substrate inhibition study using sulfaphenazole as an inhibitor of CYP2C9 activity. As a result the IC50 value and inhibition constant for sulfaphenazole were determinated 0.29 uM and 0.09695 uM respectively which are in consistence with those reported in literature. |
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