Hepatic Ethoxyresorufin O-deethylase Activity Research Articles

Chromium accumulation and biomarker responses in the Neotropical fish Prochilodus lineatus caged in a river under the influence of tannery activities

In order to assess the effects of tannery effluents (TE) in organism health, juveniles of Prochilodus lineatus were submitted to in situ tests at four different river locations: site A – upstream of the tannery; site B – next to the tannery; and sites C and D – downstream of the tannery. After 96 h exposure in the river, samples of fish tissue, river water and sediment were collected in order to quantify chromium (Cr) concentrations. Tissue samples were used to assess the activity of ethoxyresorufin-O-deethylase (EROD) and glutathione S-transferase (GST), the content of glutathione (GSH) and metallothionein (MT) and the occurrence of lipid peroxidation (LPO) and DNA damage. Higher Cr concentrations were detected in the water and sediments from site B and in the liver of fish confined at site B, compared to the other sites. Fish caged at site B demonstrated higher levels of liver MT and hepatic EROD activity in relation to fish caged at the other sites. Moreover, fish from site B presented increased liver and branchial GST activities, as well as more GSH in the liver, than fish from site A. There were no significant variations in the occurrence of LPO and DNA damage among fish caged at the different sites. Thus, TE increased Cr levels in the water, sediments, and fish livers and stimulated the synthesis of MT and GSH and the activities of EROD and GST. In conclusion, TE affect the quality of the river and promote changes in biochemical biomarkers and Cr accumulation in P. lineatus.

Contaminant concentrations and biomarkers in 21-day old Herring Gulls (Larus argentatus) and Double-crested Cormorants (Phalacrocorax auritus) from eastern Lake Ontario, and from Hamilton Harbour in western Lake Ontario in 1989 and 1990

This study measured concentrations of polychlorinated biphenyls, polycyclic aromatic hydrocarbons, organochlorine pesticides, trace metals and biochemical markers of contaminant exposure and potential toxicity (plasma and hepatic retinol, hepatic retinyl palmitate, ethoxyresorufin O-deethylase, hepatic porphyrins) in 21 day-old Herring Gull (Larus argentatus) and Double-crested Cormorant (Phalacrocorax auritus) chicks. The chicks were collected from breeding colonies at Hamilton Harbour, an industrialized site in western Lake Ontario, and at Snake and Pigeon Islands in eastern Lake Ontario in 1989 and 1990. The polycyclic aromatic hydrocarbons, benzo[a]pyrene and naphthalene, were detected in bile and concentrations were higher in both species, but particularly in Cormorants, at Hamilton Harbour in comparison to eastern Lake Ontario. Ethoxyresorufin O-deethylase activity correlated positively with polyaromatic hydrocarbon exposure in Cormorants. At Hamilton Harbour, Cormorants had significantly greater liver to body weight ratios, increased hepatic ethoxyresorufin O-deethylase activities and elevated plasma retinol concentrations compared to those from eastern Lake Ontario. Liver concentrations of selected polychlorinated biphenyl congeners were generally low (<2 ug g−1 w.wt) in birds from both areas but Hamilton Harbour Herring Gulls had mean concentrations that were significantly higher than eastern Lake Ontario Gulls, while Cormorants had low and similar polychlorinated biphenyl concentrations between sites. Trace metal levels did not vary substantially between sites for either species.

Effects of in vivo exposure to polyfluorinated dibenzo-p-dioxins on organo-somatic indices and ethoxyresorufin-O-deethylase activity in mice (Mus musculus)

ABSTRACTIn this study, five different congeners of polyfluorinated dibenzo-P-dioxins (PFDDs) (1,8-di-FDD, 1,3,8-tri-FDD, 1,3,6,8-tetra-FDD, 2,3,7,8-tetra-FDD and 1,2,3,4,5,6,7,8-octa-FDD), representing different numbers and positions of fluorine substituents of all 75 PFDD congeners, were synthesized and purified to evaluate their potential environmental impact on living organisms. Their toxicity was evaluated by determining the impact on the organo-somatic indices (OSI) and ethoxyresorufin-O-deethylase (EROD) activity in mice (Mus musculus) after intragastric administration with different doses (0.5–100 μg/kg body weight) for 3 days. The results showed that these PFDDs significantly inhibited the growth and changed the OSI in mouse tissues. Notably, hepatic EROD activity was markedly induced in mice after exposure to these PFDDs, probably indicating a high affinity of binding to the aryl hydrocarbon receptor. Overall, these findings provided some preliminary but alarming toxicity data of PFDDs, and filled information gaps in the toxicological databases for living organisms.

Ploidy-, gender-, and dose-dependent alteration of selected biomarkers in Clarias gariepinus treated with benzo[a]pyrene

Naturally-occurring and artificially-induced polyploids have been documented in various fish species but to date no comparison has been reported of the impacts of ploidy on fish biomarker responses to organic pollutants. This study describes effects of ploidy, gender, and dose on biliary fluorescent aromatic compound (FAC) concentrations, hepatic ethoxyresorufin-O-deethylase (EROD) and glutathione S-transferase (GST) activities in one of the most commonly cultured warm-water species, the African catfish Clarias gariepinus. Recently matured male and female diploid and triploid fish were intraperitoneally (i.p.) injected with 0, 5 or 25mg/kg benzo[a]pyrene (BaP) and liver and gallbladder were sampled 48hr later. No significant differences were found between ploidies in bile concentrations of 7,8 dihydrodiolbenzo[a]pyrene (7,8D BaP), 1-hydroxybenzo[a]pyrene (1-OH BaP) or 3-hydroxybenzo[a]pyrene (3-OH BaP). However, concentrations of the biliary FACs did differ between males and females at different dose of injection with generally higher concentrations in females at the low dose of BaP and higher concentrations in males at the higher BaP concentration. Hepatic EROD activity did not exhibit gender-dependent difference, whereas it was significantly higher in triploids than diploids. GST activities were not significantly influenced by any of the tested factors. This work advanced our understanding of the role of ploidy, gender, and dose in biotransformation of pollutants in fish.

The influence of seasonality, fish size and reproductive status on EROD activity in Plagioscion squamosissimus: Implications for biomonitoring of tropical/subtropical reservoirs

Ethoxyresorufin-O-deethylase (EROD) activity is considered an important biomarker for aquatic environmental contamination. Although EROD activity has been widely used as a biomarker of exposure to polycyclic aromatic hydrocarbons in fish, this activity can be influenced in the field by spatial-, seasonal- or individual-related factors. We therefore performed a comparative study of hepatic EROD activity levels in the croaker Plagioscion squamosissimus to determine whether variations existed in enzyme activity levels, especially in relationship to reproductive status, fish size, age, and seasonality. For this purpose, we collected fish from three reservoirs with different pollution levels during the early-rainy (November 2012), rainy (March 2013), and dry (July–August 2013) seasons from the Tietê River, Brazil. We tested whether size, age and sex affected EROD activity among the localities and seasons. We found a marked effect of pollution during the dry season on variation among the localities in EROD activity in P. squamosissimus. An analysis of covariance indicated that sex had a significant negative effect on the seasonal variability of the EROD activity levels at the most polluted locality (near São Paulo). A possible explanation for the statistical association between EROD activity and sex is that the reproductive status of the females influenced the EROD activity levels. A possible explanation for the statistical association between EROD activity and sex is that the reproductive status of the females influenced the EROD activity levels, also largely reported in the literature. Our results suggest that reproductive status can be a significant confounding factor for determining EROD activity in female P. squamosissimus in freshwater ecosystems when compared to males. Moreover, our results suggest that the presence of phenanthrene during the dry season at Barra Bonita reservoir might explain the highest EROD activity responses in this period of study.

Vertical distribution of AhR-activating compounds in sediments contaminated by modernized pulp and paper industry

Increased ethoxyresorufin-O-deethylase (EROD) activity is a sensitive biomarker of exposure to the chemicals which activate the aryl hydrocarbon receptor (AhR) and induce the cytochrome P450 system, such as many polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs) and polychlorinated biphenyls (PCBs). Pulp bleaching was one of the main sources of PCDDs and PCDFs until elemental chlorine free (ECF) and total chlorine free bleaching processes since 1990s have remarkably decreased but not completely eliminate discharges of these chemicals. In addition, historically contaminated sediments may act as a source of these persistent contaminants. In this study, the contamination history and recovery of a watercourse heavily loaded by the chemical wood industry were studied by analyzing PCDDs, PCDFs and PCBs from vertical sediment samples and by measuring hepatic EROD activity from rainbow trout intraperitoneally dosed with the sediment extracts. No PCDDs or PCDFs were found above the chromatographic limit of detection from the study area and only small amounts of PCB congeners 101, 138, 153, and 180 were present. No increased EROD activity was observed in fish indicating the absence of any AhR-activating compounds in the surface sediment, to about 15 cm depth, representing about the last 20 years when kraft pulping and ECF bleaching with activated wastewater treatment have been used. It can be concluded that nowadays organochlorines and other AhR-ligands do not harm the previously heavily polluted watercourse.

Recovery of Historically Contaminated Watercourse Polluted by the Chemical Wood Industry: EROD Activity in Fish as Biomarker

Despite outstanding process alterations over decades, pulp- and paper-mill-contaminated sediments and continuing exposure by the effluents may still have effects on biota. In this study, ecotoxicological impacts in the boreal watercourse were analyzed by measuring ethoxyresorufin-O-deethylase (EROD) induction from wild fish populations and from experimentally exposed fish. In order to assess the role of sediment-borne chemicals, juvenile rainbow trout (Oncorhynchus mykiss) were exposed in the laboratory to the surface sediments of Lake Vatianjärvi and Southern Lake Saimaa, both watercourses impacted by the chemical wood industry for approximately a century. Hepatic EROD activity was also measured from roach (Rutilus rutilus) and perch (Perca fluviatilis) caught in Lake Vatianjärvi. Increased EROD activity was not observed in wild fish caught in Lake Vatianjärvi nor in rainbow trout exposed to the sediment of Lake Vatianjärvi, but it was observed in rainbow trout exposed to the sediment of Southern Lake Saimaa, probably explained by critically high concentrations of retene. The results of both the laboratory and field study indicate the absence of EROD-inducing compounds of the previously heavily contaminated Lake Vatianjärvi.

Variation of Basal EROD Activities in Ten Passerine Bird Species – Relationships with Diet and Migration Status

Inter-specific differences in animal defence mechanisms against toxic substances are currently poorly understood. The ethoxyresorufin-O-deethylase (EROD) enzyme plays an important role in defence against toxic chemicals in a wide variety of animals, and it is an important biomarker for environmental contamination. We compared basal hepatic EROD activity levels among ten passerine species to see if there is inter-specific variation in enzyme activity, especially in relation to their diet and migration status. Migratory insectivores showed higher EROD activity compared to granivores. We hypothesize that the variable invertebrate diet of migratory insectivores contains a wider range of natural toxins than the narrower diet of granivores. This may have affected the evolution of mixed function oxidases (MFO) system and enzyme activities. We further tested whether metabolic rates or relative liver size were associated with the variation in detoxification capacity. We found no association between EROD activity and relative (per mass unit) basal metabolic rate (BMR). Instead, EROD activity and relative liver mass (% of body mass) correlated positively, suggesting that a proportionally large liver also functions efficiently. Our results suggest that granivores and non-migratory birds may be more vulnerable to environmental contaminants than insectivores and migratory birds. The diet and migration status, however, are phylogenetically strongly connected to each other, and their roles cannot be fully separated in our analysis with only ten passerine species.

Biotransformation modulation and genotoxicity in white seabream upon exposure to paralytic shellfish toxins produced by Gymnodinium catenatum

Fish are recurrently exposed to paralytic shellfish toxins (PSTs) produced by Gymnodinium catenatum. Nevertheless, the knowledge regarding metabolism of PSTs and their toxic effects in fish is scarce. Consequently, the current study aims to investigate the role of phase I and II detoxification enzymes on PST metabolism in the liver of white seabream ( Diplodus sargus), assessing ethoxyresorufin- O-deethylase (EROD) and glutathione S-transferase (GST) activities. Moreover, the genotoxic potential of PSTs was examined through the erythrocytic nuclear abnormality (ENA) assay. Fish were intracoelomically (IC) injected with a nominal dose (expressed as saxitoxin equivalents) of 1.60 μg STXeq kg −1 semipurified from a G. catenatum cell culture with previously determined toxin profile. Fish were sacrificed 2 and 6 days after IC injection. PST levels determined in fish liver were15.2 and 12.2 μg STXeq kg −1, respectively, at 2 and 6 days after the injection. Though several PSTs were administered, only dcSTX was detected in the liver after 2 and 6 days. This was regarded as an evidence that most of the N-sulfocarbamoyl and decarbamoyl toxins were rapidly biotransformed in D. sargus liver and/or eliminated. This was corroborated by a hepatic GST activity induction at 2 days after injection. Hepatic EROD activity was unresponsive to PSTs, suggesting that these toxins enter phase II of biotransformation directly. The genotoxic potential of PSTs was also demonstrated; these toxins were able to induce cytogenetic damage, such as chromosome (or chromatid) breaks or loss and segregational anomalies, measured by the ENA assay. Overall, this study pointed out the ecological risk associated with the contamination of fish with PSTs generated by G. catenatum blooms, providing the necessary first data for a proper interpretation of biomonitoring programs aiming to assess the impact of phytoplankton blooms in fish.

Polychlorinated biphenyl accumulation and hepatic EROD activity in golden grey mullet Liza aurata from Tunis Bay, southern Mediterranean

Liza aurata was used as a bioindicator species to monitor contamination by organic xenobiotics of estuarine water at the outlet of Khélij channel [canal], Tunisia, from November 2002 to December 2003. Polychlorinated biphenyl (PCB) concentrations, including coplanar congeners, were measured in mullet muscle tissue. Mean PCB concentrations ranged between 77 and 180 ng g−1 wet weight. The PCB profiles were dominated by hexachlorobiphenyl congeners accounting for 25–52% of the total profiles. Total 2,3,7,8-tetrachlorodibenzo-p-dioxin toxic equivalents of coplanar PCBs, including mono- and non-ortho congeners, was 4 pg g−1 wet weight in December 2003 vs 82 pg g−1 wet weight in April 2003. Biological effects in mullet were evaluated by measuring ethoxyresorufin-o-deethylase (EROD) activity in fish liver. EROD activity was significantly elevated, compared to baseline values in control fish (43 ± 23 pmol min−1 mg−1 protein). The highest enzymatic activity of 1 202 ± 813 pmol min−1 mg−1 protein was recorded in July 2003.

Reduced cytochrome P4501A activity and recovery from oxidative stress during subchronic benzo[a]pyrene and benzo[e]pyrene treatment of rainbow trout

This study assessed the role of aryl hydrocarbon receptor (AHR) affinity, and cytochrome P4501A (CYP1A) protein and activity in polyaromatic hydrocarbon (PAH)-induced oxidative stress. In the 1–100 nM concentration range benzo[a]pyrene (BaP) but not benzo[e]pyrene (BeP) competitively displaced 2 nM [ 3H]2, 3, 7, 8-tetrachloro-dibenzo-p-dioxin from rainbow trout AHR2α. Based on appearance of fluorescent aromatic compounds in bile over 3, 7, 14, 28 or 50 days of feeding 3 μg of BaP or BeP/g fish/day, rainbow trout liver readily excreted these polyaromatic hydrocarbons (PAHs) and their metabolites at near steady state rates. CYP1A proteins catalyzed more than 98% of ethoxyresorufin-O-deethylase (EROD) activity in rainbow trout hepatic microsomes. EROD activity of hepatic microsomes initially increased and then decreased to control activities after 50 days of feeding both PAHs. Immunohistochemistry of liver confirmed CYP1A protein increased in fish fed both PAHs after 3 days and remained elevated for up to 28 days. Neither BaP nor BeP increased hepatic DNA adduct concentrations at any time up to 50 days of feeding these PAHs. Comet assays of blood cells demonstrated marked DNA damage after 14 days of feeding both PAHs that was not significant after 50 days. There was a strong positive correlation between hepatic EROD activity and DNA damage in blood cells over time for both PAHs. Neither CYP1A protein nor 3-nitrotyrosine (a biomarker for oxidative stress) immunostaining in trunk kidney were significantly altered by BaP or BeP after 3, 7, 14, or 28 days. There was no clear association between AHR2α affinity and BaP and BeP-induced oxidative stress.

A chemometrical approach to study interactions between ethynylestradiol and an AhR‐agonist in stickleback (Gasterosteus aculeatus)

AbstractQuantifiable responses in fish, such as induction of certain proteins, can be used as indicators of chemical contamination of waterways. In order to evaluate differences in ethoxyresorufin‐O‐deethylase (EROD) induction capacity of the gill and the liver and effects on organs and biomarker proteins, e.g. gill and liver EROD, hepatosomatic index (HSI), nephrosomatic index (NSI), gonadosomatic index (GSI), spiggin, vitellogenin and sperm motility were analysed in male three‐spined sticklebacks (Gasterosteus aculeatus) exposed for 21 days to β‐naphthoflavone (βNF) alone (Exp 1) or in combination with 17α‐ethynylestradiol (EE2) (Exp 2). The sperm motility variables were studied using computer‐assisted sperm analysis (CASA).Exp 1: Gill EROD activity was significantly induced in fish exposed to ≥1.2 µg/l and hepatic EROD activity in fish exposed to ≥6 µg/l. No significant effect of ßNF on the production of spiggin or vitellogenin or on sperm variables was found.Exp 2: A significant additative effect of EE2 + βNF was shown for gill EROD. A significant antagonistic effect of the two compounds was found on NSI where an increased EE2 concentration led to an increase in NSI while an increased concentration of βNF led to a decreased NSI. Interestingly, the results showed that exposure to intermediate concentrations of EE2 and ßNF led to a significant increase in the sperm variables. In the aquatic environment mixtures of numerous chemicals with oestrogenic activity are present, so if the capacity to induce gill EROD activity is a general property of oestrogen‐acting chemicals, our findings are important. Copyright © 2010 John Wiley & Sons, Ltd.

Ethoxyresorufin- O-deethylase enzyme activities and accumulation of secondary/tertiary lysosomes in rabbitfish Siganus oramin as biomarkers for xenobiotic exposures

The sensitivities of using hepatic and intestinal ethoxyresorufin- O-deethylase (EROD) activities and hepatic accumulation of secondary/tertiary (2°/3°) lysosomes to detect xenobiotic exposures were assessed in the rabbitfish Siganus oramin in a metropolitan harbour, subtropical Hong Kong, over a complete seasonal cycle of one year. Additional information on the body-burden pollutants and physiological indices in S. oramin, and seasonal variables in seawater quality, were extracted from published data and re-analyzed. Under the influences of pollutant cocktail and seasonal factors, neither the hepatic nor intestinal EROD activity was indicative of total polycyclic aromatic hydrocarbons (∑ PAH), total polychlorinated biphenyls, condition factor and hepatosomatic index (HSI) in S. oramin. However, the relative ratio of hepatic to intestinal EROD activities provided an indication to differentiate the xenobiotic intake route in the fish through diffusion via gills/skin or consumption of contaminated food. In addition, the elevated hepatic accumulation of 2°/3° lysosomes was closely associated with the dominant temporal trends of zinc and ∑ PAH, as well as reduced HSI, in S. oramin. Being minimally influenced by any investigated seasonal factors, the hepatic 2°/3° lysosomes in S. oramin was recommended as an effective biomarker of xenobiotic exposures and toxic effects for use in coastal pollution monitoring programmes.

Assessment of biological effects of pollutants in a hyper eutrophic tropical water body, Lake Beira, Sri Lanka using multiple biomarker responses of resident fish, Nile tilapia (Oreochromis niloticus)

Biomarkers measured at the molecular and cellular level in fish have been proposed as sensitive "early warning" tools for biological effect measurements in environmental quality assessments. Lake Beira is a hypertrophic urban water body with a complex mixture of pollutants including polycyclic aromatic hydrocarbons (PAHs) and Microcystins. In this study, a suite of biomarker responses viz. biliary fluorescent aromatic compounds (FACs), hepatic ethoxyresorufin O-deethylase (EROD) and glutathione S-transferase (GST), brain and muscle cholinesterases (ChE), serum sorbitol dehydrogenase (SDH), and liver histology of Oreochromis niloticus, the dominant fish inhabiting this tropical Lake were evaluated to assess the pollution exposure and biological effects. Some fish sampled in the dry periods demonstrated prominent structural abnormalities in the liver and concomitant increase in serum SDH and reduction in hepatic GST activities in comparison to the control fish and the fish sampled in the rainy periods. The resident fish with apparently normal liver demonstrated induction of hepatic EROD and GST activities and increase in biliary FACs irrespective of the sampling period indicating bioavailability of PAHs. Muscle ChE activities of the resident fish were depressed significantly indicating exposure to anticholinesterase substances. The results revealed that fish populations residing in this Lake is under threat due to the pollution stress. Hepatic abnormalities in the fish may be mainly associated with the pollution stress due to recurrent exposure to PAHs and toxigenic Microcystis blooms in the Lake.

Cytochrome P4501A biomarker indication of oil exposure in harlequin ducks up to 20 years after the Exxon Valdez oil spill

Hydrocarbon-inducible cytochrome P4501A (CYP1A) expression was measured, as ethoxyresorufin-O-deethylase (EROD) activity, in livers of wintering harlequin ducks (Histrionicus histrionicus) captured in areas of Prince William Sound, Alaska, USA, oiled by the 1989 Exxon Valdez spill and in birds from nearby unoiled areas, during 2005 to 2009 (up to 20 years following the spill). The present work repeated studies conducted in 1998 that demonstrated that in harlequin ducks using areas that received Exxon Valdez oil, EROD activity was elevated nearly a decade after the spill. The present findings strongly supported the conclusion that average levels of hepatic EROD activity were higher in ducks from oiled areas than those from unoiled areas during 2005 to 2009. This result was consistent across four sampling periods; furthermore, results generated from two independent laboratories using paired liver samples from one of the sampling periods were similar. The EROD activity did not vary in relation to age, sex, or body mass of individuals, nor did it vary strongly by season in birds collected early and late in the winter of 2006 to 2007, indicating that these factors did not confound inferences about observed differences between oiled and unoiled areas. We interpret these results to indicate that harlequin ducks continued to be exposed to residual Exxon Valdez oil up to 20 years after the original spill. This adds to a growing body of literature suggesting that oil spills have the potential to affect wildlife for much longer time frames than previously assumed.

Response of CYP1A Gene expression in fish liver of catfish (Ariopsis felis) from Gulf of Mexico and their relationship with the genetic variability

We determined the hepatic Cytochrome P4501A (CYP1A) mRNA and Ethoxyresorufin-O-deethylase (EROD) activities in the fish, Ariopsis felis, from highly polluted to relatively pristine regions in the southwest Gulf of Mexico and their relationship with the genetic polymorphisms of the same fish. We hypothesized that a high genetic variation reflects interindividual variability in levels of CYP1A mRNA underlying the pathway culminating in EROD induction caused by the environmental contaminants. Catfish from Laguna de Mecoacan exhibited marked induction of CYP1A mRNA and high levels of hepatic EROD activities, whereas fish from Laguna de Celestun showed no induction of CYP1A mRNA and moderately low levels of EROD activities. In contrast, the similarity index considering all samples varied from 0.4 to 0.87, showing a wide range of variation. A dendrogram showed a clear grouping of fish collected from the Laguna de Terminos, Rio Coatzacoalcos and Laguna de Celestun, with discrete subgroups according to region. In contrast, fish from Laguna de Mecoacan were grouped together completely separate from the rest of the fish. Despite the low number of fish from Mecoacan (a high bootstrap support was observed in this group), the results indicated a significant genetic variability in comparison with the other ecosystems included. The differential level of expression of CYP1A and the EROD activity observed among the ecosystems analyzed could be due to the high range of genetic variation, with special emphasis on fish collected in Mecoacan where it is possible to find a subspecies of Ariopsis felis.

Catalytic and immunochemical detection of hepatic and extrahepatic microsomal cytochrome P450 1A1 (CYP1A1) in white-sided dolphin ( Lagenorhynchus acutus)

We have characterized microsomal systems and measured the levels of microsomal cytochrome P450 1A1 (CYP1A1) and ethoxyresorufin-O-deethylase (EROD) activity in multiple internal organs of male and female white-sided dolphin ( Lagenorhynchus acutus) from the northwest Atlantic Ocean. Internal organs were sampled within 24 h of death, sometimes in a period of hours, collection times which are significantly less than usually seen for marine mammals. Tissue autolysis, as assessed by histological analysis of liver, was minimal to none in all individuals. Total P420 did not correlate with time from death to sampling, suggesting that it is a poor indicator of P450 degradation in cetacean tissues where perfusion is not practical. The total hepatic microsomal P450 content, cytochrome b5 content, and NADPH-cytochrome c (P450) reductase (CPR) activity averaged 0.29 nmol mg −1, 0.12 nmol mg −1, and 238 nmol mg −1 min −1, respectively. Microsomal CPR activity in liver was higher than that in lung and kidney, and was higher than that reported in liver of most other cetacean species. Immunodetected CYP1A1 content was low in all organs, less than 3 pmoles CYP1A equivalents mg −1. EROD activity ranged from 9 to 376 pmoles mg −1 min −1 and was greater in liver than in other tissues. Hepatic microsomal EROD activity and CYP1A1 content did not correlate. However, hepatic EROD activity, but not CYP1A1 protein content, was well correlated with both total PCB and Σmono- ortho PCB concentrations in blubber. Length, as a proxy for age, did not correlate with hepatic EROD activity or CYP1A1 protein levels, and sex did not influence the relationship between EROD and contaminant concentrations. We cannot easily control for the extent of tissue degradation in cetacean studies nor do we have a complete history of these animals. Therefore, other factors such as degradation or hormonal state may have a role in the observed relationships. Yet, as in other mammals, hepatic tissues appear to be a major site of CYP1A1 expression and probably of biotransformation of CYP1A substrates in white-sided dolphin. The expression of an EROD catalyst in liver likely reflects induction by PCBs, but the P450 enzyme catalyzing hepatic EROD activity in these whales may not be CYP1A1.

Weighing the evidence of ecological risk from PAHs contamination in the estuarine environment of Salina Cruz Bay, México

Results of bulk-phase chemical measurements, toxicological tests combined with bioaccumulation measures in fishes, were used to evaluate the toxicity of the 16 USEPA priority polycyclic aromatic hydrocarbons from the sediment collected from eight stations of the Ventosa Estuarine System, located close to the main center of processing oil in the Mexican Pacific coast. Levels of the sum of polycyclic aromatic hydrocarbons varied from 22 to 6,850 microg kg(-1) dry weight. Based on sediment quality guidelines, the compounds with high environmental priority were acenaphtylene, acenaphtene, and phenanthrene. Acute toxicity tests with Vibrio fischeri and Daphnia magna as well as chronic toxicity with Panagrellus redivivus were performed. The quantification of hepatic ethoxyresorufin O-deethylase activity was used to assess the induction of the mixed function oxygenase system of brown trout. However, because it is often difficult to blend the results from such very different assays into a unified decision about the potential for impacts, a weight-of-evidence (WOE) approach to sediment quality investigations was followed. These assays provided measurement endpoints that could be used to develop an overall evaluation of the potential for environmental impacts from the oil processing operations. WOE provides a valuable tool for assessing the results of environmental investigations because it provides a framework for considering the strengths and weaknesses of environmental measurements, an approach for addressing uncertainty in the measurements, and documentation of the evaluation and its assumptions.

Influence of Dietary Coexposure to Benzo(a)pyrene on the Biotransformation and Distribution of 14C-Methoxychlor in the Channel Catfish (Ictalurus punctatus)

Methoxychlor (MXC) is an organochlorine pesticide whose mono- and bis-demethylated metabolites, 2-(4-hydroxyphenyl)-2-(4-methoxyphenyl)-1,1,1-trichloroethane (OH-MXC) and 2,2-bis(4-hydroxyphenyl)-1,1,1-trichloroethane (HPTE), respectively, are estrogenic and antiandrogenic. Studies in vitro showed that treatment of channel catfish with a polycyclic aromatic hydrocarbon increased phase I and phase II metabolism of MXC. To determine the in vivo significance, groups of four channel catfish were treated by gavage for 6 days with 2 mg/kg (14)C-MXC alone or 2 mg/kg (14)C-MXC and 2 mg/kg benzo(a)pyrene (BaP). On day 7, blood and tissue samples were taken for analysis. Hepatic ethoxyresorufin O-deethylase activity was 10-fold higher in the BaP-treated catfish, indicating CYP1A induction. More MXC-derived radioactivity remained in control (42.8 +/- 4.1%) than BaP-induced catfish (28.5 +/- 3.2%), mean percent total dose +/- SE. Bile, muscle and fat contained approximately 90% of the radioactivity remaining in control and induced catfish. Extraction and chromatographic analysis showed that liver contained MXC, OH-MXC, HPTE, and glucuronide but not sulfate conjugates of OH-MXC and HPTE. Liver mitochondria contained more MXC, OH-MXC, and HPTE than other subcellular fractions. Bile contained glucuronides of OH-MXC and HPTE, and hydrolysis of bile gave HPTE and both enantiomers of OH-MXC. The muscle, visceral fat, brain and gonads contained MXC, OH-MXC, and HPTE in varying proportions, but no conjugates. This study showed that catfish coexposed to BaP and MXC retained less MXC and metabolites in tissues than those exposed to MXC alone, suggesting that induction enhanced the elimination of MXC, and further showed that potentially toxic metabolites of MXC were present in the edible tissues.

The zebrafish gill model: Induction of CYP1A, EROD and PAH adduct formation

Zebrafish ( Danio rerio) is a common model species in fish toxicology, and the zebrafish gill is potentially useful in screening waterborne pollutants. We have previously developed a gill-based ethoxyresorufin- O-deethylase (EROD) assay, and proposed gill EROD activity as a biomarker for exposure to waterborne aryl hydrocarbon receptor (AHR) agonists. In this study we modified the gill EROD assay for use in zebrafish. We used immunohistochemistry to localize CYP1A induction, and microautoradiography to localize irreversible binding of the prototypic polycyclic aromatic hydrocarbon, 7,12-dimethylbenz[a]anthracene (DMBA) in zebrafish gills. Gill filament and liver microsomal EROD activities were measured after waterborne exposure of zebrafish and rainbow trout to benzo[a]pyrene (BaP) or β-naphthoflavone (βNF). The results showed considerably lower relative EROD induction by βNF (1 μM) in zebrafish than in rainbow trout, both in gills (13-fold versus 230-fold compared to control) and in liver (5-fold versus 320-fold compared to control). The induced hepatic EROD activity was similar in the two species, whereas the basal activity was considerably higher in zebrafish than in rainbow trout. In zebrafish gills, βNF enhanced DMBA adduct formation and CYP1A immunostaining. Ellipticine blocked DMBA adduct formation and EROD activity following βNF exposure but had no effect on CYP1A immunostaining. A notable finding was that the localization of DMBA adducts differed from that of CYP1A protein in βNF-induced fish; CYP1A immunoreactivity was evenly distributed in the gills whereas DMBA adduction was confined to the leading edges of the filaments and the gill rakers, i.e., structures being highly exposed to DMBA-containing inhaled water. The results show that the modified method is suitable for determination of gill EROD activity in zebrafish, although rainbow trout seems more sensitive. They also imply that the sites of DMBA adduct formation in zebrafish gills are markedly influenced by kinetic factors.