Effects of alkylphenols on the biotransformation of diuron and enzymes involved in the synthesis and clearance of sex steroids in juvenile male tilapia ( Oreochromus mossambica )

Abstract

Previous studies using in vivo bioassay guided fractionation indicated that the herbicide diuron (3-(3,4-dichlorophenyl)-1,1-dimethylurea) and alkylphenol (AP)-containing surfactants were detected in fractions of extracts that induced the estrogenic biomarker, vitellogenin in fish exposed to surface water extracts from the United States. However, when the compounds were evaluated individually using in vivo estrogenic assays or in vitro estrogen receptor assays, estrogenic activity was not observed. Since APs have been shown to alter activity and content of cytochrome P450s (CYP) which convert diuron to potential estrogenic metabolites, the hepatic biotransformation of diuron was measured with and without a 7day pretreatment of p-Octylphenol (OP) and p-Nonylphenol (NP) at low (OP 13ng/L+NP 91ng/L), and high concentrations (OP 65ng/L+NP 455ng/L) in juvenile male Nile tilapia (Oreochromus niloticus). Pre-treatment with the OP/NP (AP) mixture caused elevated levels of NADPH-catalyzed formation of 3,4-dichlorophenyl-N-methylurea (DCPMU) but not 3,4-dichlorophenylurea (DCPU). Fish were also treated with nominal concentrations of low (40ng/L) and high (200ng/L) diuron and each of its three degradates/metabolites: DCPMU, DCPU and 3,4-dichloroaniline (DCA). Additional treatments were conducted with APs and Diuron as a mixture at the low concentrations which mimicked concentrations observed in surface waters. Hepatic vitellogenin (Vtg) mRNA was induced by exposure to the high concentrations of Diuron, as well as DCPMU and DCPU in both concentrations. Brain cytochrome P450 aromatase activity was generally diminished by diuron, its metabolites, and the AP/diuron mixtures. 17β-Hydroxysteroid dehydrogenase (17βHSD) levels were also reduced by DCPMU and DCA in the lower concentrations, but not by higher concentrations. While the AP mixture reduced 17βHSD, the AP/diuron mixture induced testosterone (T) biosynthesis at the single concentration tested. Although CYP3A expression was induced by all diuron metabolites, it was unchanged by the AP mixture. These data indicate that mixtures of AP and diuron enhanced the formation of the metabolite (DCPMU) which induced vitellogenin, and reduced T biosynthetic enzymes (17βHSD inhibition). Overall, these data showed that APs may have induced the biotransformation of diuron to at least one metabolite, that may disrupt androgen biosynthesis and potentially alter steroid feedback pathways in the central nervous system.

Estrogenic and anti-androgenic effects of the herbicide tebuthiuron in male Nile tilapia (Oreochromis niloticus)

“…Then, Pereira et al (2016) demonstrated that diuron metabolites, but not diuron, have estrogenic effects in female Nile tilapias, evidenced by increased estradiol (E 2 ) plasma levels, gonadosomatic indices and elevated percentage of final vitellogenic oocytes, despite decreasing the amount of germinative cells. Felício et al (2015) further showed that exposure of Nile tilapia to diuron and its metabolites at similar concentrations in combination with alkylphenols resulted in increases in vitellogenin levels and decreased testosterone levels, confirming the estrogenic and anti-androgenic effects of diuron and its metabolites in fish.…”

“…Diuron, another phenylurea herbicide extensively used worldwide for weed control was also found in surface waters at concentrations up to 200 ng/L in the San Francisco Bay-USA (Schlenk et al, 2012). Based on this information, studies regarding alterations in sexual hormones and gametogenesis in tilapias were previously done by our group, using 40, 100 and 200 ng/L od diuron (Pereira et al, 2015(Pereira et al, , 2016Felício et al, 2015;Boscolo et al, 2017). Therefore, tebuthiuron concentrations used in this study was chosen considering realistic tebuthiuron concentration found in natural environments but also considering other studies previously published by our laboratory with diuron, for comparative purposes.…”

“…Diuron, another urea-derived herbicide, was reported to cause morphological (Scheil et al, 2009;Mhadhbi and Beiras, 2012), biochemical (Sanchez-Muros et al, 2013) and physiological changes in fish (Miranda et al, 2008), besides causing also deficiency in fish growth (Nebeker and Schuytema, 1998). Diuron at concentrations similar to those commonly found in most contaminated aquatic environments (up to 200 ng/L; Schlenk et al, 2012;Köck-Schulmeyer et al, 2013;Masiá et al, 2015) have also been recently shown to alter the metabolism of sexual hormones in Nile tilapia, leading to alterations in gametogenesis and gonadosomatic index (Pereira et al, 2015(Pereira et al, , 2016Felício et al, 2015) and aggressive behavior (Boscolo et al, 2017). Schlenk et al (2012) first proposed that diuron in combination with alkylphenol ethoxylates at environmental concentrations would exert estrogenic effects by increasing vitellogenin levels in Japanese medaka (Oryzias latipes) using in vivo bioassays.…”

Herbicide Diuron as Endocrine Disrupting Chemicals (EDCs) through Histopathalogical Analysis in Gonads of Javanese Medaka (Oryzias javanicus, Bleeker 1854)

“…In a study by Felício et al [34], it was observed that Diuron has shown to antagonize estrogenic and androgenic receptors of steroid hormones which can potentially impair the gonadal steroidogenesis and the reproductive system of teleosts. Diuron has also shown to be anti-androgenic, inducing vitellogenin in juvenile male tilapia (Oreochromis mossambica) [35] and anti-androgenic compounds (AAC) with key enzymatic activities [36]. Another effective herbicide called Atrazine also demonstrated the reduction in the reproduction of Japanese medaka (O. latipes) [29] and fathead minnow (Pimephales promales) [30].…”

Review of and Recommendations for Monitoring Contaminants and their Effects in the San Francisco Bay−Delta

“…Alkylphenol ethoxylates and alkylphenols enhanced the estrogenic activity of diuron in male Fathead Minnows . Subsequent studies in male tilapia also indicated the induction of cytochrome P450 by nonylphenol ethoxylates (NPE) and nonylphenols NPs, presumably enhancing the formation of more active metabolites of diuron such as 3,4-dichlorophenyl-N-methylurea (DCPMU), which was more potent than the parent compound, diuron, in males and females (Felicio et al 2016;Pereira et al 2016). Anti-androgenic activity was also noted in males, in addition to the estrogenic responses that indicate multiple targets that result in overall feminization (Pereira et al 2015).…”