AIM: To develop an analytical method for simultaneous quantification of 5-hydroxyomeprazole (5-OH-OP) and omeprazole sulfone (OPS), and explore whether omeprazole (OP) is an appropriate phenotypic probe for CYP2C19 and CYP3A4 in Chinese liver microsomes. METHODS: OP metabolism in vitro was conducted in Chinese liver microsomes, and the major metabolites 5-OH-OP and OPS were determined using high pressure liquid chromatography (HPLC). Monoclonal antibodies anti-CYP2C8/9/19 and anti-CYP3A4 were employed to conduct inhibition experiments. The protein contents of CYP2C19 and CYP3A4 were quantified using Western blot analysis and densitometric scanning. RESULTS: 5-OH-OP and OPS gave a baseline resolution in the HPLC analysis. The detection limits for both compounds were 0.01 nmol and the recovery (98%-102%) had good precision with relative standard deviation of < 9.5%. Both anti-CYP2C8/9/19 and anti-CYP3A4 had a significant inhibitory effect (P < 0.05) on the 5-OH-OP formation in a substrate concentration-dependent manner, and anti-CYP3A4 alone could almost abolish the formation of OPS (> 87%). At a substrate concentration of 2 mumol/L OP, good correlations were found between OP 5-hydroxylation and S-mephenytoin 4'-hydroxylation activities (r = 0.72, P < 0.01), OP 5-hydroxylation activities and CYP2C19 contents (r = 0.82, P < 0.01), and OP sulfoxidation activities and CYP3A4 contents (r = 0.78, P < 0.01) in Chinese liver microsomes. CONCLUSION: OP metabolism is mediated mainly by CYP2C19 and CYP3A4, and OP can be used to probe CYP2C19 and CYP3A4 activities in Chinese liver microsomes at appropriate substrate concentrations with the HPLC method presently developed.
AIM: To develop an analytical method for simultaneous quantification of 5-hydroxyomeprazole (5-OH-OP) and omeprazole sulfone (OPS), and explore whether omeprazole (OP) is an appropriate phenotypic probe for CYP2C19 and CYP3A4 in Chinese liver microsomes. METHODS: OP metabolism in vitro was conducted in Chinese liver microsomes, and the major metabolites 5-OH-OP and OPS were determined using high pressure liquid chromatography (HPLC). Monoclonal antibodies anti-CYP2C8/9/19 and anti-CYP3A4 were employed to conduct inhibition experiments. The protein contents of CYP2C19 and CYP3A4 were quantified using Western blot analysis and densitometric scanning. RESULTS:5-OH-OP and OPS gave a baseline resolution in the HPLC analysis. The detection limits for both compounds were 0.01 nmol and the recovery (98%-102%) had good precision with relative standard deviation of < 9.5%. Both anti-CYP2C8/9/19 and anti-CYP3A4 had a significant inhibitory effect (P < 0.05) on the 5-OH-OP formation in a substrate concentration-dependent manner, and anti-CYP3A4 alone could almost abolish the formation of OPS (> 87%). At a substrate concentration of 2 mumol/L OP, good correlations were found between OP 5-hydroxylation and S-mephenytoin 4'-hydroxylation activities (r = 0.72, P < 0.01), OP 5-hydroxylation activities and CYP2C19 contents (r = 0.82, P < 0.01), and OP sulfoxidation activities and CYP3A4 contents (r = 0.78, P < 0.01) in Chinese liver microsomes. CONCLUSION: OP metabolism is mediated mainly by CYP2C19 and CYP3A4, and OP can be used to probe CYP2C19 and CYP3A4 activities in Chinese liver microsomes at appropriate substrate concentrations with the HPLC method presently developed.
Authors: Miaoying Shi; Yumeng Dong; Hans Bouwmeester; Ivonne M C M Rietjens; Marije Strikwold Journal: Arch Toxicol Date: 2022-05-23 Impact factor: 6.168