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The microsomal metabolism of phenol (11 °C) over an annual reproductive cycle from June to December was studied using fall spawning adult brook trout (Salvelinus fontinalis). Hepatic microsomes were isolated from three male and three female fish each month. Incubations were optimized for time, cofactor concentration, pH, and microsomal protein concentration. The formation of phase I ring-hydroxylation metabolites, i.e., hydroquinone (HQ) and catechol (CAT), was quantified by HPLC with dual-channel electrochemical detection. Sample preparation and chromatographic conditions were optimized to achieve the separation and sensitivity required for the analysis of these labile products. Biotransformation of phenol over a range of substrate concentrations (1 to 150 mM) was quantified for the calculation of Michaelis–Menten constants (Km and Vmax) for each month. Results indicate a nearly equal production of HQ and CAT among males and females in late June. At the peak of maturity in October, there was an approximate ten-fold greater production of ring-hydroxylation metabolites noted in females in comparison with males on a total liver basis. In vitro phase II biotransformation of phenol glucuronidation was assessed by determining the Michaelis–Menten constants (Km, Vmax) using brook trout hepatic microsomes over a range of substrate concentrations (1 to 60 mM). Initially, there were no significant differences in the glucuronide rate of formation (pmol/min/mg protein) or total capacity (nmol/min/liver) between females and males. At the peak of maturation, the maximum rate of glucuronide formation was 4-fold less in females; however, the total capacity was 2-fold less in females due to the increased liver size in the females. The alterations in biotransformation coincided with increases in the hepatic and gonadal somatic indices and with changes in plasma hormone concentrations. These experiments provide insight into the metabolic deactivation of xenobiotics and to provide data for the prediction of altered hepatic biotransformation rates and pathways during the reproductive cycle. [ABSTRACT FROM AUTHOR]

The in vitro biotransformation of phenol at 11 °C was studied using pre-spawn adult rainbow (Oncorhynchus mykiss) (RBT), brook (Salvelinus fontinalis) (BKT), and lake trout (Salvelinus namaycush) (LKT) hepatic microsomal preparations. The incubations were optimized for time, cofactor concentration, pH, and microsomal protein concentration. Formation of Phase I ring-hydroxylation and Phase II glucuronidation metabolites was quantified using HPLC with dual-channel electrochemical and UV detection. The biotransformation of phenol over a range of substrate concentrations (1 to 180 mM) was quantified, and the Michaelis–Menten kinetics constants, Km and Vmax, for the formation of hydroquinone (HQ), catechol (CAT), and phenylglucuronide (PG) were calculated. Species differences were noted in the Km values for Phase I enzyme production of HQ and CAT, with the following rank order of apparent enzyme affinity for substrate: RBT > BKT = LKT. However, no apparent differences in the Km for Phase II metabolism of phenol to PG were detected. Conversely, while there were no apparent differences in Vmax between species for HQ or CAT formation, the apparent maximum capacity for PG formation was significantly less in LKT than that observed for RBT and BKT. These experiments provide a means to quantify metabolic activation and deactivation of xenobiotics in fish, to compare activation and deactivation reactions across species, and to act as a guide for future predictions of new chemical biotransformation pathways and rates in fish. These experiments provided the necessary rate and capacity (Km and Vmax) inputs that are required to parameterize a fish physiologically based toxicokinetic (PB-TK) model for a reactive chemical that is readily biotransformed, such as phenol. In the future, an extensive database of these rate and capacity parameters on important fish species for selected chemical structures will be needed to allow the effective use of predictive models for reactive, biotransformation chemicals in aquatic toxicology and environmental risk assessment. [ABSTRACT FROM AUTHOR]

Providing an alternative to pyrethroids, organophosphates, and carbamates, the neonicotinoids are now the most widely used insecticides in the world. They are water soluble and relatively stable in soil and water which allows for run-off through surface waters and thus potentially impacting aquatic species and environments. While the mammalian metabolism of neonicotinoids has been studied extensively, there is a lack of understanding of their metabolism in fish species. The current study constitutes the first report of the metabolism of imidacloprid (IMI) and acetamiprid (AC) in rainbow trout. Formation of respective metabolites 5-hydroxy-imidacloprid and N-desmethyl-acetamiprid was conserved across orders of biological organization in both microsomal and liver slice assays. Michaelis–Menten kinetics were determined for the microsomal conversion of IMI to 5-hydroxy-imidacloprid in rainbow trout (Km = 79.2 µM; Vmax = 0.75 pmole/min/mg) and rat (Km = 158.7 µM; Vmax = 38.4 pmole/min/mg). Kinetics for the microsomal demethylation of AC to N-desmethyl-acetamiprid were determined in the rat (Km = 70.9 µM; Vmax = 10 pmoles/min/mg). N-desmethyl-acetamiprid was found in detectable but below quantifiable levels across the range of test concentrations which precluded a calculation of kinetic rate constants in rainbow trout (RBT). Ultimately, the formation of the metabolites 5-hydroxy-imidacloprid and N-desmethyl-acetamiprid was conserved across RBT and rat species. [ABSTRACT FROM AUTHOR]

1. Cyclic phenones are chemicals of interest to the USEPA due to their potential for endocrine disruption to aquatic and terrestrial species. 2. Prior to this report, there was very limited information addressing metabolism of cyclic phenones by fish species and the potential for estrogen receptor (ER) binding and vitellogenin (Vtg) gene activation by their metabolites. 3. The main objectives of the current research were to characterize rainbow trout (rt) liver slice-mediated in vitro metabolism of model parent cyclic phenones exhibiting disparity between ER binding and ER-mediated Vtg gene induction, and to assess the metabolic competency of fish liver in vitro tests to help determine the chemical form (parent and/or metabolite) associated with the observed biological response. 4. GC-MS, HPLC and LC-MS/MS technologies were applied to investigate the in vitro biotransformation of cyclobutyl phenyl ketone (CBP), benzophenone (DPK), cyclohexyl phenyl ketone (CPK) mostly in the absence of standards for metabolite characterization. 5. It was concluded that estrogenic effects of the studied cyclic phenones are mediated by the parent chemical structure for DPK, but by active metabolites for CPK. A definitive interpretation was not possible for CBP and CBPOH (alcohol), although a contribution of both structures to gene induction is suspected. [ABSTRACT FROM AUTHOR]

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 and other international environmental organizations. 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). Cyclic phenones, branched phenones, and hydroxybenzophenones bound to rtER with relative potency ranging from no affinity to high binding affinity of 0.11%, and many induced Vtg gene expression in rt liver slices. In addition, cyclohexylphenylketone which did not bind rtER binding in cytosol was biotransformed within liver tissue to a chemical that induced Vtg expression.