Evaluation of the effect of TM208 on the activity of five cytochrome P450 enzymes using on-line solid-phase extraction HPLC-DAD: a cocktail approach.

Wensi Lin, Jianmei Zhang, Xiaomei Ling, Ning Yu, Jing Li, Haisong Yang, Runtao Li, Jingrong Cui

Index: J. Chromatogr. B. Analyt. Technol. Biomed. Life Sci. 923-924 , 29-36, (2013)

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Abstract

A rapid, simple, and sensitive on-line solid-phase extraction HPLC-DAD method for simultaneous evaluation of the activity of five CYP450 isoforms (CYP1A2, CYP2C19, CYP2D6, CYP2E1 and CYP3A4) in vivo has been developed and validated. The five specific probe substrates include caffeine (1A2), metoprolol (2D6), dapsone (3A4), omeprazole (2C19) and chlorzoxazone (2E1). Automated pre-purification of plasma and enrichment of analytes were performed using a C18 on-line solid-phase extraction cartridge. After being eluted from the cartridge, the analytes and the internal standard antipyrine were separated on a C18 RP analytical column and analyzed by DAD. The method was validated to quantify the concentration ranges of 0.05-50.0 μg/ml for dapsone and omeprazole, 0.1-50.0 μg/ml for caffeine and 0.2-50.0 μg/ml for metoprolol and chlorzoxazone. The linearity (R(2)) for all analytes tested was exceeded 0.99. The intra-day precision ranged from 0.29 to 13% and the inter-day precision ranged from 5.0 to 15%, respectively. The intra-day and inter-day accuracy were between 86.7% and 113.6%. The extraction recoveries were in the range 82.8-109.9% for all the analytes and internal standard antipyrine. This method was successfully applied to evaluate the effects of TM208 on rat five CYP450 isoforms.Copyright © 2013 Elsevier B.V. All rights reserved.