Novel metabolic pathways of p-n-nonylphenol catalyzed by cytochrome P450 and estrogen receptor binding activity of new metabolites

Abstract

Nonylphenol, which is used industrially as a surfactant, is an endocrine-disrupting chemical (EDC) which has estrogenic activity. The novel biotransformation of nonylphenol was investigated, based on our previously reported ipso-metabolism of para-substituted phenols by cytochrome P450 (P450). Three novel metabolites of nonylphenol, i.e., nonylquinol, 4′- hydroxynonanophenone (CO-NP) as benzyl-oxidized nonylphenol, and hydroquinone, were detected in a rat liver microsome reaction mixture. On the other hand, production of 1-(4′-hydroxyphenyl)nonan-1-ol (OH-NP), namely benzyl-hydroxylated nonylphenol, was detected in a human liver microsome reaction mixture. The formation of all these metabolites was suppressed by the addition of P450 inhibitor. This showed that all nonylphenol metabolism was catalyzed by P450. To identify which P450 isoenzyme is involved in each reaction, fourteen human P450 (CYP) isozymes, CYP1A1, 1A2, 2A6, 2B6, 2C8, 2C9, 2C18, 2C19, 2D6, 2E1, 3A4, 3A5, 3A7, and CYP4A11, were examined. CYP1A1, 1A2, and CYP2B6 effectively catalyzed the production of nonylquinol. CYP2B6 also catalyzed the benzyl-hydroxylation to give OH-NP. Hydroquinone was formed mainly from OH-NP, not via CO-NP. We examined the estrogenic activity of these new metabolites by estrogen receptor (ER)-binding reporter gene assay. Nonylquinol, OH-NP and hydroquinone have no ER-binding activity. However, CO-NP showed the same level of estrogen receptor binding activity as nonylphenol. Moreover, the amount of CONP formed was small. Therefore, the novel metabolic pathways led overall to metabolic inactivation, as concerns the estrogenic activity of nonylphenol through the ER.