Human risk assessment through development and application of a physiologically based toxicokinetic model for 4-tert-octylphenol.

4-tert-octylphenol (4-tert-OP) is an ecologically hazardous substance, and exposure to it in the environment has been consistently reported in the past. Despite the hazards and widespread exposure to 4-tert-OP, tools for scientific assessment of 4-tert-OP exposure risk level in humans are lacking. The main purpose of this study was to develop a physiologically-based-toxicokinetic (PBTK) model for 4-tert-OP and to perform quantitative risk assessment of 4-tert-OP in various population groups using the established model. Based on the results of toxicokinetic experiments on male rats, the PBTK model for 4-tert-OP was established and verified, and this was converted to a model for humans through interspecies extrapolation. Based on the previously reported no-observed-adverse-effect-levels for rats, it was possible to estimate the 4-tert-OP reference dose in humans through reverse dosimetry using the model. Biomonitoring data derived from various population groups were applied to the human PBTK model to calculate external exposures and margin of safety for 4-tert-OP for each population group. The PBTK model established in this study adequately explained the toxicokinetic experimental values at acceptable levels and was able to quantitatively predict the 4-tert-OP exposure level in the testes related to male reproductive toxicity. In addition, the degree of external exposure to 4-tert-OP could be scientifically estimated based on biomonitoring values derived from various biological matrices. The reference doses for systemic and reproductive toxicity caused by 4-tert-OP in male humans were calculated to be 0.16 and 1.12 mg/kg/day, respectively. The mean external exposure to 4-tert-OP in each population group estimated based on plasma and urine biomonitoring data was 0.04-66.24 mg/kg/day, showing very large exposure diversity between groups. Exposure risks to 4-tert-OP in populations ranged from safe to risky, suggesting the need for continued monitoring and risk management of 4-tert-OP worldwide. This study provides valuable scientific insight regarding the 4-tert-OP human risk assessment.

Effects of crosstalk between steroid hormones mediated thyroid hormone in zebrafish exposed to 4-tert-octylphenol: Estrogenic and anti-androgenic effects.

Alkylphenols, such as nonylphenol and 4-tert-octylphenol (OP), are byproducts of the biodegradation of alkylphenol ethoxylates and present substantial ecological and health risks in aquatic environments and higher life forms. In this context, our study aimed to explore the effect of OP on reproductive endocrine function in both female and male zebrafish. Over a period of 21 days, the zebrafish were subjected to varying concentrations of OP (0, 0.02, 0.1, and 0.5 µg/L), based on the lowest effective concentration (EC10 = 0.48 µg/L) identified for zebrafish embryos. OP exposure led to a pronounced increase in hepatic vitellogenin (vtg) mRNA expression and 17ß-estradiol biosynthesis in both sexes. Conversely, OP exhibits anti-androgenic properties, significantly diminishes gonadal androgen receptor (ar) mRNA expression, and reduces endogenous androgen (testosterone and 11-ketotestosterone) levels in male zebrafish. Notably, cortisol and thyroid hormone (TH) levels demonstrated concentration-dependent elevations in zebrafish, influencing the regulation of gonadal steroid hormones (GSHs). These findings suggest that prolonged OP exposure may result in sustained reproductive dysfunction in adult zebrafish, which is largely attributable to the intricate reciprocal relationship between hormone levels and the associated gene expression. Our comprehensive biological response analysis of adult zebrafish offers vital insights into the reproductive toxicological effects of OP, thereby enriching future ecological studies on aquatic systems.

Sex differences in 4-tert-octylphenol toxicokinetics: Exploration of sex as an effective covariate through an in vivo modeling approach.

4-tert-octylphenol (4-tert-OP) is a potentially harmful substance, which is found widely in the environment. Nevertheless, information on the in vivo toxicokinetics of 4-tert-OP is lacking, and quantitative risk assessment studies are urgently needed. Therefore, we aimed to quantitatively identify differences in the toxicokinetics of 4-tert-OP and its distribution among tissues between sexes. To this end, following exposure of male and female rats to 10 or 50 mg/kg 4-tert-OP orally and 4 or 8 mg/kg 4-tert-OP intravenously, we conducted a quantitative analysis of samples using ultra-high performance liquid chromatography-tandem mass spectrometry. The results revealed that the 4-tert-OP plasma concentration profiles differed between sexes; however, systemic absorption of 4-tert-OP through the gastrointestinal tract occurred within 0.5 h of exposure in both sexes. Although small, the excretion percentage of 4-tert-OP in urine and feces was lower in males than females (0.06-0.08% vs. 0.82-1.11% of exposure). Significant sex differences were also confirmed in the tissue distribution patterns of 4-tert-OP, and overall, the average tissue distribution in males was lower than that in females. The distribution of 4-tert-OP to liver, adipose, spleen, kidney, brain, and lung in both sexes was predominant. A covariate exploration modeling approach revealed that sex explained the differences in 4-tert-OP toxicokinetics between sexes. These significant differences in the toxicokinetics and tissue distribution of 4-tert-OP between sexes will be important for the scientific precision human risk assessment of 4-tert-OP.

Development of an HPLC-DAD method for the simultaneous analysis of four xenoestrogens in water and its application for monitoring photocatalytic degradation.

A high-performance liquid chromatography with diode array detector (HPLC-DAD) method was developed and validated for the simultaneous quantification of 4 xenoestrogens in water for monitoring their photocatalytic degradation in synthetic water. The analytical parameters evaluated were linearity, limits of detection, and quantification (LODs and LOQs), selectivity, and accuracy, according to the US Food and Drug Administration (FDA) and Eurachem guidelines. The developed method shows good linearity (R2 > 0.995 for all compounds), and LODs ranged from 0.02 to 0.04 mg L-1, while LOQs ranged from 0.05 to 0.11 mg L-1. Moreover, accuracy expressed as recovery and precision were within the required limits. Therefore, the developed method was considered accurate, and reliable. In addition, it was successfully applied for monitoring a mixture of 4 xenoestrogens in water during the photocatalytic treatment.•An HPLC-DAD method was developed to quantify 4 xenoestrogens in water simultaneously.•The developed HPLC-DAD method shows excellent linearity, selectivity, and accuracy.•A mixture of 4 xenoestrogens was reliably monitored during their photocatalytic degradation.

Amphibian Metamorphosis Assay: Investigation of the potential effects of five chemicals on the hypothalamic-pituitary thyroid axis of Xenopus laevis.

2-Ethylhexyl 4-hydroxybenzoate (2-EHHB), 4-tert-octylphenol (4-OP), 4-nonylphenol-branched (4-NP), benzyl butyl phthalate (BBP) and dibutyl phthalate (DBP) were evaluated using a 21-day Amphibian Metamorphosis Assay (AMA). Xenopus laevis larvae were exposed nominally to each chemical at 3.6, 10.9, 33.0, and 100 µg/L, except 4-NP concentrations were 1.8, 5.5, 16.5 and 50 µg/L. Endpoints included mortality, developmental stage, hind limb length (HLL), snout-vent length (SVL), body weight (BW), and thyroid histopathology. BBP and 4-OP accelerated development compared to controls at the mean measured concentration of 3.5 and 39.8 µg/L, respectively. An increase in developmental stage frequency distribution was observed for 4-OP at 39.8 and 103 µg/L, BBP at all concentrations and DBP at 143 µg/L. Normalized HLL was increased on study day (SD) 21 for all tested substances except 4-NP. Histopathology revealed accelerated development and mild thyroid follicular cell hypertrophy at all BBP concentrations, but moderate severity at 105 µg/L. Increased BW occurred for all chemicals except 4-OP. Increased SVL was observed for 4-NP, BBP and DBP on SD 21. There was insufficient evidence that 4-NP and 2-EHHB affected the hypothalamic-pituitary thyroid axis, however, BBP, DBP and 4-OP showed potential effects on amphibian metamorphosis and thyroid activity, albeit through different lines of evidence.

Simultaneous Quantification of Bisphenol-A and 4-Tert-Octylphenol in the Live Aquaculture Feed Artemia franciscana and in Its Culture Medium Using HPLC-DAD.

Aquaculture, a mass supplier of seafood, relies on plastic materials that may contain the endocrine disruptors bisphenol-A (BPA) and tert-octylphenol (t-OCT). These pollutants present toxicity to Artemia, the live aquaculture feed, and are transferred through it to the larval stages of the cultured organisms. The purpose of this work is the development and validation of an analytical method to determine BPA and t-OCT in Artemia and their culture medium, using n-octylphenol as the internal standard. Extraction of the samples was performed with H2O/TFA (0.08%)-methanol (3:1), followed by SPE. Analysis was performed in a Nucleosil column with mobile phases A (95:5, v/v, 0.1% TFA in H2O:CH3CN) and B (5:95, v/v, 0.08% TFA in H2O:CH3CN). Calibration curves were constructed in the range of concentrations expected following a 24 h administration of BPA (10 µg/mL) or t-OCT (0.5 µg/mL), below their respective LC50. At the end of exposure to the pollutants, their total levels appeared reduced by about 32% for BPA and 35% for t-OCT, and this reduction could not be accounted for by photodegradation (9-19%). The developed method was validated in terms of linearity, accuracy, and precision, demonstrating the uptake of BPA and t-OCT in Artemia.

17α-ethinylestradiol and 4-tert-octylphenol concurrently disrupt the immune response of common carp.

The aquatic environment is massively polluted with endocrine-disrupting compounds (EDCs) including synthetic estrogens (e.g. 17α-ethinylestradiol, EE2) and alkylphenols (e.g. 4-tert-octylphenol, 4t-OP). A major mechanism of action for estrogenic EDCs is their interaction with estrogen receptors and consequently their modulation of the action of enzymes involved in steroid conversion e.g. aromatase CYP19. We now studied the effects of EE2 and 4t-OP on the anti-bacterial immune response of common carp. We investigated effects on the number/composition of inflammatory leukocytes and on the gene expression of mediators that regulate inflammation and EDC binding. In vitro we found that high concentrations of both EE2 and 4t-OP down-regulated IFN-γ2 and IFN-γ-dependent immune responses in LPS-stimulated monocytes/macrophages. Similarly, during bacterial infection in fish, in vivo treated with EE2 and 4t-OP, decreased gene expression of il-12p35 and of ifn-γ2 was found in the focus of inflammation. Moreover, during A. salmonicida-induced infection in EE2-treated carp, but not in fish fed with 4t-OP-treated food, we found an enhanced inflammatory reaction manifested by high number of inflammatory peritoneal leukocytes, including phagocytes and higher expression of pro-inflammatory mediators (inos, il-1ß, cxcl8_l2). Furthermore, in the liver, EE2 down-regulated the expression of acute phase proteins: CRPs and C3. Importantly, both in vitro and in vivo, EDCs altered the expression of estrogen receptors: nuclear (erα and erß) and membrane (gpr30). EDCs also induced up-regulation of the cyp19b gene. Our findings reveal that contamination of the aquatic milieu with estrogenic EDCs, may considerably violate the subtle and particular allostatic interactions between the immune response and endogenous estrogens and this may have negative consequences for fish health.

Molecular interactions of thyroxine binding globulin and thyroid hormone receptor with estrogenic compounds 4-nonylphenol, 4-tert-octylphenol and bisphenol A metabolite (MBP).

AIM: Endocrine disruption due to environmental chemical contaminants is a global human health issue. The aim of present study was to investigate the structural binding aspects of possible interference of commonly detected environmental contaminants on thyroid function. MATERIAL AND METHODS: Three compounds, 4-tert-octylphenol (4-tert-OP), 4-nonylphenol (4-NP), and 4-methyl-2,4-bis(4-hydroxypentyl)pent-1-ene (MBP) were subjected to induced fit docking (IFD) against thyroxine binding globulin (TBG) and thyroid hormone receptor (THR). Structural analysis included molecular interactions of the amino acid residues and binding energy estimation between the ligands and the target proteins. KEY RESULTS: All the ligands were successfully placed in the ligand binding pocket of TBG and THR using induced fit docking (IFD). The IFD results revealed high percentage of commonality in interacting amino acid residues between the aforementioned compounds and the native ligand for both TBG and THR. The results of our study further revealed that all the compounds have the potential to interfere with thyroid transport and signaling. However, MBP showed higher binding affinity for both TBG and THR, suggesting higher thyroid disruptive potential as compared to 4-t-OP and 4-NP. Furthermore, our results also suggest that the reported disruptive effects of BPA could actually be exerted through its metabolite; MBP. SIGNIFICANCE: This work implies that all the three compounds 4-NP, 4-t-OP and especially MBP have the potential to interfere with thyroid hormone transport and signaling. This potentially leads to disruption of thyroid hormone function.

4-Nonylphenol and 4-tert-octylphenol induce anxiety-related behaviors through alternation of 5-HT receptors and transporters in the prefrontal cortex.

Environmental endocrine disruptors 4-nonylphenol (NP) and 4-tert-octylphenol (OP) may cast huge harm to human health. We used a rat model to observe the influence of NP or/and OP exposure on anxiety-related behaviors and the underlying mechanisms. Eighty male Sprague-Dawley (SD) rats were randomly divided into 10 groups: control group (corn oil), NP groups [30, 90, 270 mg/kg], OP groups [40, 120, 360 mg/kg] and NO groups [(mixed with the corresponding NP, OP alone exposed low, medium and high dose according to the natural environment exists NP:OP = 4:1]. The rats were orally administered every other day for 30 days. The neurobehaviors of rats were evaluated by open-field test (OFT) and elevated plus-maze test (EPM), and the concentrations of 5-HT, monoamine oxidase (MAOA), serotonin transporter (SERT), vesicular monoamine transporter 2 (VAMT2), 5-hydroxytryptamine 1A (5-HT1A), 5-hydroxytryptamine 2A (5-HT2A)，and 5-hydroxytryptamine 2C (5-HT2C) in the rat prefrontal cortex were analyzed by ELISA. OFT and EPM tests showed that NP or/and OP exposure induced anxiety-related behaviors in rats. 5-HT levels were significantly increased compared with the control group. The levels of MAOA, SERT, VAMT2, 5-HT1A, 5-HT2A, and 5-HT2C in the prefrontal cortex reduced in different degrees by high-doses NP or/and OP exposure. In summary, NP or/and OP exposure might cause anxiety-related behaviors in rats through regulating neurotransmitter 5-HT levels by altering the expression of 5-HT decomposition enzyme MAOA, transporters SERT and VMAT2, and 5-HT receptors 5-HT1A, 5-HT2A, and 5-HT2C.

Ecological risks of phenolic endocrine disrupting compounds in an urban tropical river.

The distribution of emerging organic contaminants in drinking water sources in Africa is a subject with very scanty data and information. In order to fill knowledge gaps, we report here the distribution and potential ecological risks of three phenolic compounds (bisphenol A (BPA), 4-nonylphenol (NP), and 4-tert-octylphenol (OP)), which have been previously identified to have the potential of endocrine disrupting activity, in surface water and sediment of the New Calabar River. The compounds were quantified using GC-MS. At all sampling sites, a similar concentration pattern of BPA > NP > OP was recorded, with the exception of Choba sampling station in which the levels of these endocrine disrupting compounds were low or undetectable. The levels of BPA in surface water ranged from 1.20 to 63.64 µg/L, whereas those of NP and OP ranged from < 0.20 to 2.15 µg/L and from < 0.10 to 0.68 µg/L, respectively. For sediments, measured levels were from 1.20 to 66.57 µg/kg for BPA, from < 0.35 to 3.37 µg/kg for NP, and from < 0.13 to 0.90 µg/kg for OP. Risk quotients (RQs) assessed for some sensitive organisms (algae, Daphnia magna, and fish) were above 1 for BPA and NP, whereas RQs for OP were below 1. This implies that BPA and NP at the levels detected could have potential risks to the sensitive organisms considered, but low risk for OP.

Metabolomics Reveals Metabolic Changes Caused by Low-Dose 4-Tert-Octylphenol in Mice Liver.

Background: Humans are constantly exposed to low concentrations of 4-tert-octylphenol (OP). However, studies investigating the effects of low-dose OP on the liver are scarce, and the mechanism of these effects has not been thoroughly elucidated to date. Methods: Adult male institute of cancer research (ICR) mice were exposed to low-dose OP (0, 0.01 and 1 µg/kg/day) for 7 consecutive days. Weights of mice were recorded daily during the experiment. Blood serum levels of OP, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were determined, and haematoxylin-eosin (HE) staining of the liver was performed. We applied an integrated metabolomic and enzyme gene expression analysis to investigate liver metabolic changes, and the gene expression of related metabolic enzymes was determined by real-time PCR and ELISA. Results: OP in blood serum was increased after OP exposure, while body weights of mice were unchanged. Liver weight and its organ coefficient were decreased significantly in the OP (1 µg/kg/day) group, but ALT and AST, as well as the HE staining results, were unchanged after OP treatment. The levels of cytidine, uridine, purine and N-acetylglutamine were increased significantly, and the level of vitamin B6 was decreased significantly in mice treated with OP (1 µg/kg/day). The mRNA and protein levels of Cda and Shmt1 were both increased significantly in OP (1 µg/kg/day)-treated mice. Conclusions: Through metabolomic analysis, our study firstly found that pyrimidine and purine synthesis were promoted and that N-acetylglutamine was upregulated after low-dose OP treatment, indicating that the treatment disturbed nucleic acid and amino acid metabolism in mice liver.

Analytical methods for determination of bisphenol A, 4-tert-octylphenol and 4-nonylphenol in herrings and physiological fluids of the grey seal.

•The aim of this work was to develop the methods of determination for phenol derivatives: bisphenol A (BPA), 4-tert-octylphenol (OP) and 4-nonylphenol (NP), in the whole body of herring Clupea harengus and in physiological fluids of the Baltic grey seal Halichoerus grypus grypus (blood and milk).•Methods were based on liquid chromatography coupled with a fluorescence detector (HPLC-FL).•These methods were satisfactorily validated, each showing good recovery (>80%) and precision (RSD < 15%). Regarding the limit of quantification (LOQ), this was established at <2 ng g-1 for herring, <0.07 ng cm-3 for blood and <0.1 ng cm-3 for milk.

Effects of Bisphenol A and 4-tert-Octylphenol on Embryo Implantation Failure in Mouse.

Miscarriage due to blastocyst implantation failure occurs in up to two-thirds of all human miscarriage cases. Calcium ion has been shown to be involved in many cellular signal transduction pathways as well as in the regulation of cell adhesion, which is necessary for the embryo implantation process. Exposure to endocrine-disrupting chemicals (EDs) during early gestation results in disruption of intrauterine implantation and uterine reception, leading to implantation failure. In this study, ovarian estrogen (E2), bisphenol A (BPA), or 4-tert-octylphenol (OP), with or without ICI 182,780 (ICI) were injected subcutaneously from gestation day 1 to gestation day 3 post-coitus. The expression levels of the calcium transport genes were assessed in maternal uteri and implantation sites. The number of implantation sites was significantly low in the OP group, and implantation sites were absent in the E2, ICI and EDs + ICI groups. There were different calcium transient transport channel expression levels in uterus and implantation site samples. The levels of TRPV5 and TRPV6 gene expression were significantly increased by EDs with/without ICI treatment in utero. Meanwhile, TRPV5 and TRPV6 gene expression were significantly lower in implantation sites samples. NCX1 and PMCA1 mRNA levels were significantly decreased by OP and BPA in the implantation site samples. Compared to vehicle treatment in the uterus, both the MUC1 mRNA and protein levels were markedly high in all but the BPA group. Taken together, these results suggest that both BPA and OP can impair embryo implantation through alteration of calcium transport gene expressions and by affecting uterine receptivity.

4-tert-octylphenol injures motility and viability of human sperm by affecting cAMP-PKA/PKC-tyrosine phosphorylation signals.

4-tert-octylphenol (4t-OP) is a well-known xenoestrogen. Our objective was to explore the effects and molecular mechanisms of 4t-OP on human sperm. Sperm samples were exposed to 0, 0.1, or 0.3 mM 4t-OP for two hours. Results showed that both sperm viability and motility were significantly injured by 0.3 mM 4t-OP. We applied comparative proteomics to explore the molecular targets affected by 4t-OP. 81 differentially expressed (DE) proteins were identified. Bioinformatic analysis showed that these proteins were highly associated with motility and apoptosis, and were mostly enriched in cAMP-PKA/PKC-phosphorylation-associated pathway. We further verified that 0.1 mM and 0.3 mM 4t-OP significantly decreased cAMP activity of sperm. Expression of RACK1 and PRDX6 were detected by western blot (WB) to verify their tendencies in gels; antiapoptotic factor BCL2 was also detected by WB. The data indicated that 4-tert-octylphenol injures the motility and viability of human sperm probably by affecting cAMP-PKA/PKC-tyrosine phosphorylation signals.

Ferrofluid-based liquid-phase microextraction: Analysis of four phenolic compounds in milks and fruit juices.

A novel liquid-phase microextraction based on the ferrofluid was successfully applied for the extraction and determination of four phenolic compounds (bisphenol-A, bisphenol-AF, tetrabromobisphenol-A and 4-tert-octylphenol) in milks and fruit juices. In this study, a range of alkyl (C4-C10) alcohols as the carrier liquid were used for the preparation of ferrofluids. The study showed that an appropriate chain length of alkyl alcohol may improve the extraction efficiency and maintain the integrity of ferrofluids during extraction, so 1-heptanol (C7) was selected as the optimal carrier liquid. Specifically, the effects of various parameters on the extraction of phenolic compounds were investigated and optimized. Under the optimized conditions, the limits of detection were between 0.35 and 0.66 µg L-1. The intra-day and inter-day precision (RSD %) for the analytes at 10 and 50 µg L-1 were in the range of 3.2-6.7 % and 2.9-7.1%, respectively.

The Role of Chlorogenic Acid Supplementation in Anemia and Mineral Disturbances Induced by 4-Tert-Octylphenol Toxicity.

4-tert-octylphenol (OP) is an endocrine-disrupting chemical that causes harmful effects to human health. Chlorogenic acid is the major dietary polyphenol present in various foods and beverages. The aim of the present study was to evaluate the protective role of chlorogenic acid in anemia and mineral disturbance occurring in OP toxicity in rats. Thirty-two male albino rats were divided into four equal groups (8 rats/group) as follows. The first (control) group was treated daily with an oral dose of 1 ml saline for two weeks. The second group was treated daily with an oral dose of 60 mg chlorogenic acid/kg body weight for two weeks. The third and fourth groups received daily intraperitoneal (ip) injections with 100 mg OP/kg body weight for two weeks; the fourth group was treated daily with an oral dose of 60 mg chlorogenic acid/kg body weight for three weeks starting one week before OP injections. The results revealed that OP induced significant decreases in hemoglobin, hematocrit, red blood cells, mean cell volume, mean cell hemoglobin, mean cell hemoglobin concentration, platelet count, white blood cells, lymphocyte and neutrophil percent, transferrin receptor, serum calcium, phosphorous, sodium, potassium, chloride, glutathione-S-transferase, glutathione peroxidase, catalase, glutathione reductase, and superoxide dismutase. Moreover, significant increases in serum hepcidin, ferritin, transferrin, erythropoietin, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, urea, creatinine, selenium, zinc, manganese, copper, iron, malondialdehyde, and protein carbonyl levels were found in OP groups. OP exposure also induced cell apoptosis. Chlorogenic acid pretreatment in OP-treated groups restored all the mentioned parameters to approach the normal values. In conclusion, chlorogenic acid protects from anemia and mineral disturbances in 4-tert-octylphenol toxicity by ameliorating oxidative stress and apoptosis.

Supramolecular solvents combined with layered double hydroxide-coated magnetic nanoparticles for extraction of bisphenols and 4-tert-octylphenol from fruit juices.

A magnesium-aluminum layered double hydroxides coated on magnetic nanoparticles (Fe3O4@MgAl-LDHs NPs) combined with the supramolecular solvents (SUPRASs) as the magnetic supramolecular fluids were successfully applied to the extraction and determination of four phenolic compounds (Bisphenol-A (BPA), bisphenol-AF (BPAF), tetrabromobisphenol-A (TBBPA) and 4-tert-octylphenol (4-TOP)). Magnetic supramolecular fluids (ferrofluids) composites were prepared through adding the mixture of n-octanol and tetrahydrofuran in distilled water as the supramolecular solvents into the solution containing magnetic Fe3O4@MgAl-LDHs NPs. Under the optimized conditions, the calibration curve was obtained in range of 5-1000µgL-1. The limits of detection (S/N=3) for BPA, BPAF, TBBPA and 4-TOP, were obtained for 0.54, 0.48, 0.37 and 0.63µgL-1, respectively. The intra-day and inter-day precision (RSDs %) for bisphenols (BPs) and 4-TOP at 100 and 300µgL-1 were in the range of 4.0-7.8. The enrichment factors were between 16 and 24.

Glutathione biosynthesis plays an important role against 4-tert-octylphenol-induced oxidative stress in Ceratophyllum demersum.

4-tert-octylphenol (OP) is a persistent environmental pollutant with an endocrine-disrupting property. In the present study, we examined the effect of various concentrations of OP (0, 0.5, 1, 1.5, 2 and 3 mg L-1) applied to an aquatic plant, the submersed macrophyte Ceratophyllum demersum. The toxic effect caused by OP inhibited the plant's growth rate, reduced total chlorophyll content and increased levels of the reactive oxygen species (ROS) O2•- and H2O2. OP treatment significantly increased the activities of antioxidant enzymes including superoxide dismutase, guaiacol peroxidase, glutathione reductase and ascorbate peroxidase. The contents of the non-enzymatic antioxidant glutathione (GSH) and ratio of GSH to glutathione disulfide were markedly increased with OP treatment. Pretreatment with buthionine sulfoximine, a specific and potent inhibitor of GSH biosynthesis, significantly reduced total GSH content and conferred a more severe toxic phenotype on OP exposure. Thus, with OP-induced oxidative stress, C. demersum might actively regulate the antioxidant machinery, especially the biosynthesis and redox state of GSH.

Hepatic glucuronidation of 4-tert-octylphenol in humans: inter-individual variability and responsible UDP-glucuronosyltransferase isoforms.

4-tert-Octylphenol (4-tOP) is an endocrine-disrupting chemical. It is mainly metabolized into glucuronide by UDP-glucuronosyltransferase (UGT) enzymes in humans. The purpose of this study was to assess inter-individual variability in and the possible roles of UGT isoforms in hepatic 4-tOP glucuronidation in the humans. 4-tOP glucuronidation activities in the liver microsomes and recombinant UGTs of humans were assessed at broad substrate concentrations, and kinetics were analyzed. Correlation analyses between 4-tOP and diclofenac or 4-hydroxybiphenyl activities in pooled and individual human liver microsomes were also performed. Typical CLint values were 17.8 mL/min/mg protein for the low type, 25.2 mL/min/mg protein for the medium type, and 47.7 mL/min/mg protein for the high type. Among the recombinant UGTs (13 isoforms) examined, UGT2B7 and UGT2B15 were the most active of catalyzing 4-tOP glucuronidation. Although the K m values of UGT2B7 and UGT2B15 were similar (0.36 and 0.42 µM, respectively), the CLint value of UGT2B7 (6.83 mL/min/mg protein) >UGT2B15 (2.35 mL/min/mg protein). Strong correlations were observed between the glucuronidation activities of 4-tOP and diclofenac (a probe for UGT2B7) or 4-hydroxybiphenyl (a probe for UGT2B15) with 0.79-0.88 of Spearman correlation coefficient (r s) values. These findings demonstrate that 4-tOP glucuronidation in humans is mainly catalyzed by hepatic UGT2B7 and UGT2B15, and suggest that these UGT isoforms play important and characteristic roles in the detoxification of 4-tOP.

Glucuronidation of 4-tert-octylphenol in humans, monkeys, rats, and mice: an in vitro analysis using liver and intestine microsomes.

4-tert-Octylphenol (4-tOP) is an endocrine-disrupting chemical. It is mainly metabolized into glucuronide by UDP-glucuronosyltransferase (UGT) enzymes in mammals. In the present study, the glucuronidation of 4-tOP in humans, monkeys, rats, and mice was examined in an in vitro system using microsomal fractions. The kinetics of 4-tOP glucuronidation by liver microsomes followed the Michaelis-Menten model for humans and monkeys, and the biphasic model for rats and mice. The K m, V max, and CL int values of human liver microsomes were 0.343 µM, 11.6 nmol/min/mg protein, and 33.8 mL/min/mg protein, respectively. The kinetics of intestine microsomes followed the Michaelis-Menten model for humans, monkeys, and rats, and the biphasic model for mice. The K m, V max, and CL int values of human intestine microsomes were 0.743 µM, 0.571 nmol/min/mg protein, and 0.770 mL/min/mg protein, respectively. The CL int values estimated by Eadie-Hofstee plots were in the order of mice (high-affinity phase) (3.0) > humans (1.0) ≥ monkeys (0.9) > rats (high-affinity phase) (0.4) for liver microsomes, and monkeys (10) > mice (high-affinity phase) (5.6) > rats (1.4) > humans (1.0) for intestine microsomes. The percentages of the CL int values of intestine microsomes to liver microsomes were in the order of monkeys (27 %) > rats (high-affinity phase in liver microsomes) (7.9 %) > mice (high-affinity phase in liver and intestine microsomes) (4.2 %) > humans (2.3 %). These results suggest that the metabolic abilities of UGT enzymes expressed in the liver and intestine toward 4-tOP markedly differ among species and imply that species differences are strongly associated with the toxicities of alkylphenols.