Abstract
CYP2E1 is expressed in adult (1, 2) and fetal (3) human liver in addition to extrahepatic tissues such as lung and placenta (4). Treatment of primary cultures of human hepatocytes with ethanol induces CYP2E1 protein (5) and this is consistent with the finding that hepatic CYP2El protein (6) and mRNA (7) amounts are increased in alcoholics. Although only a few drugs (e.g., acetaminophen [8]) have been identified as substrates for CYP2E1, many low molecular weight procarcinogens are activated by this P450 (9). Chlorzoxazone 6-hydroxylation (19–12), N-nitrosodimethylamine N-demethylation (11, 13, 14), and p-nitrophenol hydroxylation (12, 14) reactions can be used to measure the catalytic activity of cDNA-expressed CYP2El. Each of these activities is also frequently used as an enzyme-selective catalytic monitor for human hepatic CYP2El (see Notes 1 and 2). Experiments with inhibitory antiCYP2E1 antibodies and CYP2E1-selective chemical inhibitors suggest that CYP2E1 contributes extensively to these activities in human liver microsomes (9, 15–18). Recently, lauric acid 11 -hydroxylation was identified as an alternative probe for human hepatic CY2El (19). However, an advantage of using p-nitrophenol hydroxylation to measure CYP2E1 activity is that the formation of p-nitrocatechol can be measured by a simple spectrophotometric assay, although high-performance liquid chromatographic (HPLC) assays have also been developed to quantify thep-nitrocatechol metabolite (20, 21). This chapter describes a modification of a spectrophotometric method (22) for the determination of p-nitrophenol hydroxylase activity.
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K. H. Chang, T., L. Crespi, C., J. Waxman, D. (1998). Spectrophotometric Analysis of Human CYP2E1-Catalyzed p-Nitrophenol Hydroxylation. In: Phillips, I.R., Shephard, E.A. (eds) Cytochrome P450 Protocols. Methods in Molecular Biology™, vol 107. Humana Press. https://doi.org/10.1385/0-89603-519-0:147
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DOI: https://doi.org/10.1385/0-89603-519-0:147
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