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Estrogenic potential of Cyclohexylphenylketone as mediated through metabolic activation in rainbow trout
Citation:
Serrano, J., M. Tapper, A. Kubatova, J. Denny, T. Lahren, AND Rick Kolanczyk. Estrogenic potential of Cyclohexylphenylketone as mediated through metabolic activation in rainbow trout. SETAC, Sacramento, CA, November 02 - 08, 2018.
Impact/Purpose:
This research contributes to the development of a USEPA comprehensive program to evaluate the potential adverse effects of chemicals on vertebrate endocrine systems by further exploring the role of metabolism in estrogenic activity modulation with the use of in vitro fish assays; biotransformation data correlated to chemical exposure facilitates development of additional in vitro and short-term in vivo assays needed to better prioritize chemicals for testing and support risk assessment.
Description:
Phenones and hydroxy-benzophenones are widely used as UV radiation filters, and in the manufacturing of insecticides and pharmaceuticals. Understanding the estrogenic potential these chemicals is of interest to the US Environmental Protection Agency (USEPA). The current study sequentially combined complementary in vitro rainbow trout estrogen receptor (rtER) binding and liver slice vitellogenin (Vtg) mRNA induction assays in the context of a defined ER-mediated Adverse Outcome Pathway (AOP). Cyclohexylphenylketone (CPK) which did not bind rtER binding in cytosol was biotransformed within liver tissue to one or more metabolites that induced Vtg expression. In absence of Standards, nine CPK metabolites were identified by combining gas chromatography-mass spectrometry (GC-MS) with and without derivatization, liquid chromatography (LC) with tandem MS, and LC with high resolution time of flight (ToF) MS spectra information. LC fractionation showed relationships between LC- and GC-MS data needed to assess substitution patterns. Results supported that CPK is primarily metabolized through phase I oxidation of the cyclohexyl ring and not the phenyl group as anticipated. Two metabolites (M1 and M2; MW 186) were analyzed by GC-MS and enzyme conjugation studies and proposed to be cyclohexenyl-derivatives. M6-M9 were confirmed using derivatization techniques as hydroxylated metabolites (MW 204), with the potential for undergoing phase II conjugative metabolism to glucuronides and sulfates. Lastly, M3, M4 and M5 were identified as cyclohexanone-derivatives of CPK (MW 202), resulting from the redox interconversion of the hydroxylated metabolites. Information obtained allowed the proposal of a novel metabolic pathway for cyclic phenones in rainbow trout liver slices.