Polycyclic Aromatic Hydrocarbons Metabolites In Bile Research Articles

Spatiotemporal patterns in the prevalence of microscopic hepatic changes in Gulf of Mexico Tilefish (Lopholatilus chamaeleonticeps) and associations with hepatic PAHs

Following the 2010 Deepwater Horizon blowout, demersal longline surveys were conducted across the Gulf of Mexico (GoM) continental shelf to evaluate polycyclic aromatic hydrocarbon (PAH) exposure, tissue accumulation, and health indices in demersal fishes. Tilefish (Lopholatilus chamaeleonticeps), a target species due to Gulf-wide distribution with documented high exposure to PAHs, were collected in the north central GoM at repeat stations 2012 to 2015, and from the northwest GoM, Bay of Campeche, and Yucatán Shelf in 2015 and 2016. Liver samples (n = 239) were analyzed for microscopic hepatic changes (MHCs) by a board-certified veterinary pathologist. Histological analyzes identified 14 MHCs. Prevalence of MHCs was generally uniform throughout the GoM, except for low prevalence on the Yucatán Shelf. Inflammatory and vacuolar changes were most prevalent, while pre-neoplasia and neoplasia were rare. Tilefish sampled annually in the north central GoM showed increases in inflammatory MHCs and glycogen-type vacuolar change over time, while lipid-type vacuolar change decreased over time. Short-term exposure to PAHs was assessed by measuring PAH metabolites in bile (n = 100) using high performance liquid chromatography with fluorescence detection. Longer-term accumulation of PAHs in tissue was assessed by analyzing liver (n = 111) for PAHs and alkylated homologs using QuEChERS extractions and gas chromatography tandem mass spectrometry. Six MHCs including glycogen-type vacuolar change, biliary fibrosis, foci of cellular alteration, parasites, hepatocellular atrophy, and necrosis were significantly associated with hepatic PAH accumulation in Tilefish from the northern central GoM; however, no MHCs were associated with biliary PAH metabolites. Combined with previous studies of PAH exposure and health indices in north central GoM Tilefish post-Deepwater Horizon, which also identified decreases in hepatic lipid storage and Fulton's condition factor that were correlated to increasing PAH exposure, these data indicate concerning temporal trends and changes in hepatic energy storage.

Exposure to pollutants present in Iguaçu River Southern Brazil affect the health of Oreochromis niloticus (Linnaeus, 1758): Assessment histological, genotoxic and biochemical

Urban sewage is a source of major contamination in aquatic systems and contributes to environmental and human health disturbances. This study investigates the effects of sewage-polluted waters from Iguaçu River on the health of juvenile Oreochromis niloticus. Two hundred four specimens were exposed to riverine water in four groups: no diluted, 25 and 50 % diluted water and a control group without tested water for 72 days. Biological samples were obtained for histopathological, neurotoxicity, antioxidant defenses, genotoxicity, metallothionines expression and polycyclic aromatic hydrocarbons (PAHs) metabolites. The results showed histopathological alterations in liver and gills, genotoxic alteration in erythrocytes, reduction of acetylcholinesterase activity in brain and muscle, activation of antioxidant defenses in the liver, recruitment of metals by metallothionein and the detection of PAHs metabolites in bile. These results demonstrate that juveniles of O. niloticus are susceptible to Iguaçu River exposure water and they can be used as indicator of water quality.

Histological Effects of Light Crude Oil on Sciaenops ocellatus Under Experimental Conditions.

The health status of Sciaenops ocellatus was studied by qualitative and quantitative histopathological analysis through histological damage prevalence and degree of tissue change (DTC) in response to 96h and 9days of exposure to 0.1, 0.8, and 8g/L of light crude oil in seawater. The histology of the liver, spleen, kidney, and gills of the fish were analyzed and compared between treatments. Our results showed that the exposed fish developed lesions associated with degenerative and necrotic changes. The highest frequency of damage and DTC scores were observed in the liver and kidney from 96h to 9-days post-exposure. Generalized additive models of location, scale, and shape, showed that the DTC was strongly associated with crude oil compounds such as napthalene, Cd, Ni, Pb, and bile polycyclic aromatic hydrocarbon metabolites. Our findings suggest that exposure to crude oil affects fish health, producing irreversible histological damage.

Proteomics and lipidomics analyses reveal modulation of lipid metabolism by perfluoroalkyl substances in liver of Atlantic cod (Gadus morhua)

The aim of the present study was to investigate effects of defined mixtures of polycyclic aromatic hydrocarbons (PAHs) and perfluoroalkyl substances (PFASs), at low, environmentally relevant (1× = L), or high (20× = H) doses, on biological responses in Atlantic cod (Gadus morhua). To this end, farmed juvenile cod were exposed at day 0 and day 7 via intraperitoneal (i.p.) injections, in a two-week in vivo experiment. In total, there were 10 groups of fish (n = 21-22): two control groups, four separate exposure groups of PAH and PFAS mixtures (L, H), and four groups combining PAH and PFAS mixtures (L/L, H/L, L/H, H/H). Body burden analyses confirmed a dose-dependent accumulation of PFASs in cod liver and PAH metabolites in bile. The hepatosomatic index (HSI) was significantly reduced for three of the combined PAH/PFAS exposure groups (L-PAH/H-PFAS, H-PAH/L-PFAS, H-PAH/H-PFAS). Analysis of the hepatic proteome identified that pathways related to lipid degradation were significantly affected by PFAS exposure, including upregulation of enzymes in fatty acid degradation pathways, such as fatty acid β-oxidation. The increased abundances of enzymes in lipid catabolic pathways paralleled with decreasing levels of triacylglycerols (TGs) in the H-PFAS exposure group, suggest that PFAS increase lipid catabolism in Atlantic cod. Markers of oxidative stress, including catalase and glutathione S-transferase activities were also induced by PFAS exposure. Only minor and non-significant differences between exposure groups and control were found for cyp1a and acox1 gene expressions, vitellogenin concentrations in plasma, Cyp1a protein synthesis and DNA fragmentation. In summary, our combined proteomics and lipidomics analyses indicate that PFAS may disrupt lipid homeostasis in Atlantic cod.

Fluctuating asymmetry of sclerotized structures of Haliotrematoides spp. (Monogenea: Dactylogyridae) as bioindicators of aquatic contamination

Abstract Fluctuating asymmetry (FA) and increased variation in bilateral symmetry in a sample of monogeneans can indicate disturbance in developmental stability caused by environmental and/or genomic stress. This study was conducted to determine the presence of FA in the sclerotized structures of Haliotrematoides spp. (Monogenea) collected along the continental shelf of the Yucatan Peninsula. We hypothesize the potential effect of chemical contamination on FA. Geometric morphometrics with Procrustes ANOVA was used to test FA in the ventral and dorsal anchors in Haliotrematoides spp. Then, Generalized Additive Model for Location, Scale and Shape (GAMLSS) were used to assess the statistical associations of heavy metal concentrations, Polycyclic Aromatic Hydrocarbons (PAHs), gene expression, PAHs metabolites in bile, fish morphometric measurements, physicochemical and spatial variables on FA patterns. Principal Component Analysis showed no clear group formation according to shape variability by sampling site. Evidence of FA for size and shape in ventral anchors of Haliotrematoides striatohamus was found, in contrast to dorsal anchors, which only displayed shape FA. For Haliotrematoides sp. only evidence for shape FA of ventral and dorsal anchors was found. The GAMLSS models revealed a significant positive relationship between the FA of Haliotrematoides spp. and PAHs, Pb, Naphthalene, Ni, pH, Hydroxy-Phenanthrene, Depth, DNA Methylation, Glutathione peroxydase and Condition factor (K). These results partially support our hypothesis that FA in the anchors would be produced by aquatic contamination but in association to other environmental variables and host related factors.

Potential of Periophthalmus sobrinus and Siganus sutor as bioindicator fish species for PAH pollution in tropical waters

Polycyclic Aromatic Hydrocarbons (PAHs) are ubiquitous class of toxic organic compounds in coastal waters. The aim of this study was to assess the potential of goby fish (Periophthalmus sobrinus) and rabbit fish (Siganus sutor) as bioindicators for PAH pollution in Zanzibar coastal waters. The fish samples were collected from four sites along the coasts of Zanzibar between February to July, 2016. The analysis of PAH metabolites from bile was conducted using Synchronous Fluorescence Scanning (SFS) spectroscopy. The parent PAH compounds (naphthalene; fluorene; phenanthrene; pyrene; fluoranthene; anthracene; benzo (a) pyrene) in the fish muscle tissues were analyzed using Gas Chromatography with Mass Spectrometric detection (GC/MS). The levels of PAH metabolites in bile of goby fish (P. sobrinus) ranged from 100 ng mL−1 to 690 ng mL−1. The PAH contents in goby tissues fish ranged from 0.028 ± 0.008 ng g−1 ww to 0.037 ± 0.005 ng g−1 ww with the average of 0.032 ± 0.005 ng g−1 ww. PAH metabolites were not detected in the bile samples from rabbit fish (S. sutor) in any sampled areas. The results suggest that the goby fish (P. sobrinus) could be promising PAH biondicator in coastal waters of tropical regions including Zanzibar islands. This is the first study applying biliary PAH metabolites in bioindicator fish species to trace PAH pollution conducted in West Indian Ocean region that can serve as a baseline information of biliary PAH metabolites as biomarkers of fish PAH pollution in the regional waters.

Biological effects of polycyclic aromatic hydrocarbons (PAH) and their first metabolic products in in vivo exposed Atlantic cod (Gadus morhua)

ABSTRACTThe monitoring of the presence of polycyclic aromatic hydrocarbons (PAH) in the aquatic environment is a worldwide activity since some of these compounds are well-established carcinogens and mutagens. Contaminants in this class are in fact regarded as priority hazardous substances for environmental pollution (Water Framework Directive 2000/60/EC). In this study, Atlantic cod (Gadus morhua) was selected to assess in vivo effects of two PAH and their first metabolic products, namely, the corresponding trans-dihydrodiols, using biological markers. Fish were exposed for 1 wk to a single PAH (naphthalene or chrysene) and its synthetic metabolites ((1R,2R)-1,2-dihydronaphthalene-1,2-diol and (1R,2R)-1,2-dihydrochrysene-1,2-diol) by intraperitoneal injection in a continuous seawater flow system. After exposure, PAH metabolism including PAH metabolites in bile and ethoxyresorufin O-deethylase (EROD) activity, oxidative stress glutathione S-transferases (GST) and catalase (CAT) activities, and genotoxicity such as DNA adducts were evaluated, as well as general health conditions including condition index (CI), hepatosomatic index (HSI), and gonadosomatic index (GSI). PAH metabolite values were low and not significantly different when measured with the fixed-wavelength fluorescence screening method, while the gas chromatography–mass spectroscopy (GC-MS) method showed an apparent dose response in fish exposed to naphthalene. DNA adduct levels ≥0.16 × 10−8 relative adduct level (RAL) were detected. It should be noted that 0.16 × 10−8 RAL is considered the maximal acceptable background level for this species. The other biomarkers activities of catalase, GST, and EROD did not display a particular compound- or dose-related response. The GSI values were significantly lower in some chrysene- and in both naphthalene- and naphthalene diol-exposed groups compared to control.

PAH Metabolites in Bile of European Eel (Anguilla anguilla) from Morocco.

Environmental pollution of fish with organic contaminants is a topic of rising attention in Morocco. Polycyclic aromatic hydrocarbons (PAH) are prominent organic contaminants which are rapidly metabolized in fish. Their metabolites are accumulated in the bile fluid and can be used to assess PAH exposure. The two PAH metabolites 1-hydroxypyrene and 1-hydroxyphenanthrene were quantified in European eels (Anguilla anguilla) from two Moroccan river systems by high-performance liquid chromatography with fluorescence detection. Mean values ranged from 52 to 210ng/mL 1-hydroxypyrene and from 61 to 73ng/mL 1-hydroxyphenanthrene. The overall concentrations of PAH metabolites in eel from Morocco appeared moderate compared to eel from European rivers and coastal sites. The present study provides first information on concentrations of PAH metabolites in fish from Morocco.

Bioconcentration of phenanthrene and metabolites in bile and behavioral alterations in the tropical estuarine guppy Poecilia vivipara

Quantification of polycyclic aromatic hydrocarbon (PAH) metabolites in fish bile is widely used to evaluate levels of internal PAH contamination in fish, whereas behavioral effects are deemed important to address potential risks to fish populations. The estuarine guppy Poecilia vivipara was exposed for 96h to waterborne phenanthrene at concentrations of 10, 50, 200 and 500μgL−1. Phenanthrene and metabolites in bile were analyzed by fixed fluorescence at 260/380nm (excitation/emission) wavelengths. Phenanthrene increased in the bile of exposed fish in a dose-dependent pattern, and log bile bioconcentration factors ranged from 4.3 to 3.9 at 10 and 500μgL−1 phenanthrene, respectively, values that are similar to predicted bioconcentration factors based on phenanthrene Kow. Swimming resistance index was reduced to 81% of control values at 500μgL−1. Alteration of swimming speed was non monotonic, with a significant speed increase relative to control fish in treatments 50 and 200μgL−1 phenanthrene, respectively, followed by a speed decrease in fish exposed to 500μgL−1. However, swimming trajectories of fish exposed to 50, 200 and 500μgL−1 was altered by the development of a repetitive circular swimming behavior, in contrast to the controls that explored the entire experimental arena. This change in swimming patterns apparently explains the reduction in prey capture rates at 200μgL−1 phenanthrene. This study provides important information enabling the use of the estuarine guppy P. vivipara to monitor PAH metabolites in bile and its bioconcentration, linking internal exposure with ecologically relevant behavioral effects in the species.

Effects of water accommodated fractions of crude oils and diesel on a suite of biomarkers in Atlantic cod (Gadus morhua)

The aim of this study was to characterize concentration- and time-dependent responses in juvenile Atlantic cod (Gadus morhua) following exposure for one and three weeks to the water-soluble fraction (WAF) of three weathered oils: Arabian Light crude oil (ALC), North Sea crude oil (NSC) and ship-diesel. The sum of polycyclic aromatic hydrocarbons (PAH) in water was highest after one week of exposure and within environmentally relevant concentrations. PAH metabolites in bile confirmed exposure to and uptake of PAHs. Hepatic cytochrome P450 1A (CYP1A) gene expression (mRNA quantification) increased dramatically following exposure to all three oil types (fold-change up to 165) and there was a time lag between gene and protein expression. Hepatic CYP1A protein concentration and ethoxyresorufin-O-deethylase (EROD) activity were more variable among individuals and treatments than gene expression. EROD activity in liver and gills increased in fish exposed to WAF from the two crude oils, but not in fish exposed to WAF from diesel. Exposure to diesel appeared to induce oxidative stress to a greater extent than exposure to crude oils. Other biomarkers (glutathione S-transferases, acetylcholine esterase, vitellogenin) did not appear to respond to the exposure and hence did not discriminate among oils. Biomarker responses in cod after exposure to weathered crude oils and diesel suggested that the CYP1A system and oxidative stress markers have the highest potential for discriminating among different oil types and to monitor the environmental consequences of spills.

Comparative study of different exposure routes on the biotransformation and genotoxicity of PAHs in the flatfish species, Scophthalmus maximus

In this study, laboratory experiments were carried out in order to come to a better understanding of the fate of polycyclic aromatic hydrocarbons (PAHs) in the marine environment and especially on their bioaccumulation, biotransformation and genotoxic effects in fish. Juveniles of turbot (Scophthalmus maximus) were exposed to PAHs through different routes via (1) a mixture of dissolved PAHs, (2) a PAH-polluted sediment and (3) an oil fuel elutriate. Fish were exposed 4 days followed by a 6-day depuration period. In each experiment, PAH concentrations in the seawater of the tanks were analysed regularly by gas chromatography coupled with mass spectrometry. Muscle and liver samples were also analysed for parent PAH levels and PAH bioconcentration factors were calculated. Biotransformation was evaluated by measuring the levels of PAH metabolites in fish bile. Genotoxicity was assessed by the alkaline comet assay. Regardless of exposure route, the parent PAH concentrations in the liver and muscle showed a peak level 1 day after the beginning of the exposure, followed by a decrease up to the background level towards the end of the experiment, except for the exposure to dissolved PAHs for which levels were relatively low throughout the study. As a consequence, no bioaccumulation was observed in fish tissues at the end of the experiment. In contrast, regardless of exposure routes, a rapid production of biliary metabolites was observed throughout the whole exposure experiment. This was especially true for 1-hydroxypyrene, the major metabolite of pyrene. After 6 days of recovery in clean water, a significant decrease in the total metabolite concentrations occurred in bile. Fish exposed through either route displayed a significant increase in DNA strand breaks after 4 days of exposure, and significant correlations were observed between the level of biliary PAH metabolites and the level of DNA lesions in fish erythrocytes. Overall results indicate that exposure to either a mixture of dissolved PAHs, a PAH-contaminated sediment or a dispersed oil fuel elutriate leads to biotransformation and increase in DNA damage in fish. The quantification of PAH metabolites in bile and DNA damage in erythrocytes appear to be suitable for environmental monitoring of marine pollution either in the case of accidental oil spills or sediment contamination.

How flood events affect rainbow trout: Evidence of a biomarker cascade in rainbow trout after exposure to PAH contaminated sediment suspensions

Increasing frequency and intensity of flood events are major concerns in the context of climate change. In addition to the direct hydrological implications of such events, potential ecotoxicological impacts are of increasing interest. It is vital to understand mechanisms of contaminant uptake from suspended particulate matter (SPM) and related effects in aquatic biota under realistic conditions. However, little is known about these processes. Due to recent changes in climate, during summer temperatures of German rivers frequently exceed 25°C. Effects of re-suspension of sediments on biota under elevated temperature regimes are likely to differ from those under lower temperature regimes. To elucidate this differential response of aquatic vertebrates, rainbow trout were exposed to suspensions of sediment from the Rhine River that was spiked with a mixture of polycyclic aromatic hydrocarbons (PAH). The experiments were conducted under two different temperature regimes (24°C or 12°C). Physicochemical parameters, including concentration of PAHs in SPM, and biomarkers in fish (biliary PAH metabolites, 7-ethoxyresorufin O-deethylase activity, lipid peroxidation (LPO), mRNA expression of some genes and micronuclei) were measured over the course of a 12d study. Concentrations of pyrene and phenanthrene decreased over time, while no decrease was observed for chrysene and benzo[a]pyrene. The biomarker cascades, more specifically the temporal dynamics of biomarker reactions, did not only show quantitative differences (i.e. different induction intensity or rate of biomarker responses) at the two temperatures but also qualitative differences, i.e. different biomarker responses were observed. A slight significant increase of biliary metabolites in fish was observed in un-spiked sediment at 24°C. In bile of fish exposed to PAH spiked sediment concentrations of 1-hydroxypyrene and 1-hydroxyphenanthrene increased significantly during the first two days, and then decreased. At 12°C uptake of PAHs was slower and maximum metabolite concentrations in bile were less than in fish exposed at 24°C. Following a latency of two days, concentrations of PAH metabolites in bile of fish exposed at 24°C were followed by a peak in LPO. PAHs spiked into sediments under laboratory conditions were significantly more bioavailable than the PAHs that were already present in un-spiked field-collected sediments.

Biomarkers of environmental contaminants in the coastal waters of Estonia (Baltic Sea): effects on eelpouts (Zoarces viviparus)

The eastern Baltic Sea near the Estonian coast is heavily navigated by numerous cargo ships and oil tankers. Hundreds of accidents and oil spills happen yearly in this area. Yet, there is a lack of data concerning the distribution and effects of the environmental contaminants, especially polycyclic aromatic hydrocarbons (PAHs). Different parts of the Baltic Sea have different levels of contamination; therefore a wide range of monitoring stations in coastal areas in the Gulf of Finland and Gulf of Riga were chosen. The aim of the present research was to document the responses of chosen biomarkers of environmental contaminants in different unstudied areas of the Estonian coastal sea. During 2009 and 2010 we measured PAH metabolites, EROD activities, geno- and cytotoxicity, histology, parasites and other biomarkers from the eelpout (Zoarces viviparus), a resident benthic fish species. The results showed that fish from the Gulf of Riga emitted lower levels of fluorescence in fixed wavelength analyses (representing equivalents of PAH metabolites in bile and urine), and consistently, showed less geno- and cytotoxicity and parasite infection, higher liver somatic index (LSI) and a higher condition factor (CF) than fish inhabiting areas close to the Baltic proper and in the Gulf of Finland. The results point to the effect of long-range contaminant transportation, whether atmospheric or hydrodynamic, and also to the intensive shipping activity in international routes. This study fills the gap of knowledge in this area that has persisted until now. Nevertheless, more studies in this area on the different groups of contaminants are necessary, to specify the factors that are responsible for observed biological effects.

The silvering process of European eel (Anguilla anguilla) influences PAH metabolite concentrations in bile fluid: Consequences for monitoring

The stock of the catadromous European eel (Anguilla anguilla L.) continues to decline and there is growing evidence that poor health status due to contaminants might be a key element in this decrease. Organic contaminants such as polycyclic aromatic hydrocarbons (PAHs) belong to the major threats to yellow eel in their growth habitat and their metabolites are detectable in the bile. Starting the silvering process eels undergo physiological and morphological changes including cessation of feeding and downstream migration back to their spawning grounds. Reduced feed intake results in a diminishment of bile production and induces accumulation of e.g. PAH-metabolites in bile. Therefore, the aim of the present study was to demonstrate the impact of silvering on biliary PAH metabolite concentrations and to utilize normalization procedures to overcome silvering related accumulation effects of PAH-metabolites in eel bile. We investigated the hydroxyl-metabolites of pyrene (1-OH Pyr) and phenantrene (1-OH Phen) in the bile of different maturation stages of eels (silvering index I–V) from nine German rivers. We detected increasing absolute PAH metabolite levels in bile during the silvering process. The highest rise could be observed at the transition from pre migration stage III to the migrating stage IV, suggesting the onset of cessation of feeding at this stage. A cessation bias in PAH metabolite measurement could be diminished by normalization of absolute values against bile pigments (A380, biliverdin). In conclusion, we demonstrated the impact of silvering on PAH metabolite concentrations in eel bile and present suitable normalization procedures to overcome silvering related accumulation effects. Thus, for a future eel monitoring we recommend (1) to regularly monitor PAH metabolites in bile, (2) to determine silvering index of eel and (3) to normalize PAH metabolite values in bile based on maturation/silvering stages. The knowledge of the silvering stage is mandatory for an unbiased evaluation of PAH contamination of European eel towards an international harmonized eel monitoring program.

PAH biomarkers in common eelpout (Zoarces viviparus) from Danish waters

Eelpouts (Zoarces viviparus) sampled at surveillance stations during the fall of 2007 and spring 2008 in different Danish coastal areas, were studied for biomarkers of polycyclic aromatic hydrocarbons (PAHs) exposure and effects. Two analytical techniques, synchronous fluorescence spectrometry (SFS) and high-performance liquid chromatography with fluorescence detection (HPLC/F), were applied for detecting PAH metabolites in bile and urine. CYP1A activity, in this study regarded as potential biomarker of effect, was measured as 7-ethoxyresorufin-O-deethylase (EROD) activity in liver of eelpouts from different stations. Biliary PAH metabolite measurements were used for monitoring the environmental PAH load at the surveillance stations. There was found significant difference in biliary PAH metabolite content between sexes with male fish containing higher concentrations of PAH metabolites than females. The urinary PAH metabolite content did not show the same spatial trends as biliary PAH metabolites. However, fish from Aarhus Bight and Vejle Fjord had significantly higher levels of PAH metabolites in both urine and bile compared to the reference station Agersø. Normalisation methods applied for bile and urine matrices did not have any effect or only slightly reduced the coefficients of variation in data sets. The CYP1A activity in eelpout liver did not show the same spatial distribution trends between sampling sites as did biliary or urinary PAH metabolite contents. Male eelpouts showed significantly higher CYP1A activity than females in fall sampling period but there were no differences found in the spring period. General comparison between both seasons showed that eelpouts sampled in the fall had significantly higher CYP1A activity than fish sampled during spring season. Overall, the results of this study describe selected biomarker responses in eelpouts to environmental PAH load at the different areas along Danish coasts.

Polycyclic Aromatic Hydrocarbons and Risk to Threatened and Endangered Chinook Salmon in the Lower Columbia River Estuary

Polycyclic aromatic hydrocarbons (PAHs), derived from oil and fuel combustion, are ubiquitous nonpoint source pollutants that can have a number of detrimental effects on fish and wildlife. In this study, we monitored PAH exposure in outmigrant juvenile Chinook salmon from the Lower Columbia River to evaluate the risk that these contaminants might pose to the health and recovery of threatened and endangered salmonids. Juvenile Chinook salmon (Oncorhynchus tshawytscha) were collected by beach seine from five sites in the Lower Columbia River from Bonneville Dam to the mouth of the estuary (Warrendale, the Willamette-Columbia Confluence, Columbia City, Beaver Army Terminal, and Point Adams) and from a site in the Lower Willamette near downtown Portland (Morrison Street Bridge). Sediment samples were also collected at the same sites. Concentrations of PAHs in sediment samples were relatively low at all sites with average total PAH concentrations <1000 ng/g dry weight (wt.). However, we found PAHs in stomach contents of salmon from all sites at concentrations ranging from <100 to >10,000 ng/g wet wt. Metabolites of low and high molecular-weight PAHs were also detected in bile of salmon from all sites; for metabolites fluorescing at phenanthrene (PHN) wavelengths, concentrations ranged from 1.1 to 6.0 μg/mg bile protein. Levels of PAHs in stomach contents and PAH metabolites in bile were highest in salmon from the Morrison Street Bridge site in Portland and the Willamette-Columbia Confluence, Columbia City, and Beaver Army Terminal sites. Mean PAH concentrations measured in some stomach content samples from the Columbia City, Beaver Army Terminal, and Morrison Street Bridge sites were near the threshold concentration (approximately 7200-7600 ng/g wet wt.) associated with variability and immune dysfunction in juvenile salmonids (Meador et al., Can J Fish Aquat Sci 63:2364-2376, 2006; Bravo et al., Environ Toxicol Chem 30:704-714, 2011). Mean levels of biliary fluorescent aromatic compounds (FACs)-PHN in juvenile Chinook collected at the Morrison Street Bridge site in Portland, at the Confluence and Columbia City sites, and at the Beaver Army Terminal site were at or above a threshold effect concentration of 2 μg/mg protein for FACs-PHN linked to growth impairment, altered energetics, and reproductive effects (Meador et al., Environ Toxicol Chem 27(4):845-853, 2008). These findings suggest that PAHs in the food chain are a potential source of injury to juvenile salmon in the Lower Columbia and Lower Willamette rivers.

High matrix interference affecting detection of PAH metabolites in bile of Atlantic hagfish ( Myxine glutinosa) used for biomonitoring of deep-water oil production

The characteristic biology and wide distribution of hagfish species makes them relevant for use in pollution biomonitoring at great water depths, particularly in regions where deep-water oil production may take place. The exposure of fish to petrogenic contaminants can normally be detected from the level of polycyclic aromatic hydrocarbon (PAH) metabolites in bile fluid. Some of these metabolites are strong fluorophores, allowing analytical detection by means of simple fluorometric techniques such as fixed wavelength fluorescence (FF) and synchronous fluorescence scanning (SFS). In the present study bile from Atlantic hagfish ( Myxine glutinosa) collected in pristine areas (Barents Sea and southwestern Norway) displayed strong bile fluorescence levels, suggesting the presence of PAH contaminants. However, gas-chromatography–mass spectrometry (GC–MS) analyses ruled out PAHs as the origin for this fluorescence signal. Rather, the bile of Myxine contains components resulting in unusually strong background fluorescence interfering at the wavelength pairs used for detection of PAH metabolites. Possible background for the observed matrix interference and implications for detection of PAH metabolites in hagfish is discussed.

Evaluation of PAH Metabolites in Bile of Common Carp, Cyprinus carpio L. with Fixed Wavelength Fluorescence in a Field and Laboratory Study

The seasonal and inter-gender variability of polycyclic aromatic hydrocarbon (PAH) metabolites in the bile of common carp (Cyprinus carpio) was investigated with a field study in Karakaya Dam Lake (KDL) and a laboratory-conducted study with the fish samples of KDL and Sultansuyu Dam Lake (Turkey). The KDL has been polluted by urban, industrial, and agricultural activities for many years. A seasonal trend was seen in PAH metabolite levels with higher levels detected in May 2005 samples in KDL. Their levels in the bile of fish were not found to be related to sex which showed that gender is not a confounding factor on PAH uptake. The results showed significant decreases in bile metabolite levels in the KDL fish by approximately 75% versus increases in the SDL fish after acclimation at laboratory conditions. These results supported that PAH metabolites in bile were not stable and short-term differences in living conditions may lead to great differences in their levels. Moreover, these rapid changes in bile metabolites with acclimation may also indicate that dam lakes were not badly polluted with PAHs and the feeding status of fish should be taken into consideration when using bile metabolites as biomarkers.

Origin and distribution of polycyclic aromatic hydrocarbon pollution in sediment and fish from the biosphere reserve of Urdaibai (Bay of Biscay, Basque country, Spain)

The Urdaibai estuary is a UNESCO biosphere reserve impacted by recreational, agricultural and industrial activities. Polycyclic aromatic hydrocarbons (PAHs) are major pollutants in Urdaibai, and their control and the identification of their sources is central on the preservation of the area. Chemical analysis by GC–MS showed a significant pollution by PAHs in Urdaibai sediment samples, mainly from pyrolytic sources, with minor contributions from oil spills. Measurement of the dioxin-like activity using a yeast-based bioassay showed an excess of biological activity in sediment samples from the inner part of the estuary. Analysis of PAH metabolites in bile of thicklip grey mullet ( Chelon labrosus) showed a large excess of 2-naphthol over 1-naphthol or 1-pyrenol, suggesting a specific contamination in the inner Urdaibai estuary by some industrial process. Therefore, the combination of these three techniques defined different PAH pollution sources in Urdaibai: a major pyrolytic origin, occasional oil spills, and specific industrial activities.

Genotoxicity of Environmentally Relevant Concentrations of Water-Soluble Oil Components in Cod (Gadus morhua)

Large discharges from oil and gas production platforms (produced water) have led to concerns for adverse biological effects in marine areas. The aim of this study was to investigate the development of DNA adductformation and related biomarkers in fish after chronic exposure to water-soluble components of oil. Atlantic cod (Gadus morhua) were exposed for up to 44 weeks to three treatments (low, pulsed, high) containing environmentally relevant concentrations of low-molecular-weight polycyclic aromatic hydrocarbons (PAHs) and short-chained alkylphenols (APs). A time- and dose-related pattern of DNA adduct formation (measured using 32P-postlabeling) was observed. The results suggested that an extended exposure period (more than 16 weeks) would be required for the formation of DNA adduct levels above background. Interestingly, fish receiving pulsed high exposure did not develop elevated concentrations of DNA adducts, possibly due to DNA repair processes. No obvious relationship between DNA adduct concentration and cytochrome P4501A activity (EROD) was observed. This study has demonstrated the genotoxic potential of water-soluble oil components, relevant for operational discharges (produced water) and chronic oil spills. The quantification of PAH metabolites in bile and hepatic DNA adduct formation appear to be suitable for environmental monitoring of chronic oil pollution.