Rapid method for determination of dioxin-like polychlorinated biphenyls and other congeners in marine sediments using sonic extraction and photodiode array detection.

A rapid method has been developed to measure dioxin-like polychlorinated biphenyl (PCB) congeners as well as other selected PCBs in sediment. The analytes were extracted from sediment by sonication with dichloromethane, and the PCBs were separated from interfering compounds on a gravity-flow cleanup column packed with acidic, basic, and neutral silica gels eluted with 1:1 hexane:pentane (v/v). Subsequently, the dioxin-like PCB congeners were resolved from nonplanar PCBs and other chlorinated compounds by high-performance liquid chromatography (HPLC). Two important advantages of PDA over conventional UV detection are the ability to identify individual analytes by comparing their UV spectra with those of reference standards and the ability to establish the spectral homogeneity (purity) of the analytes by comparing spectra within a peak to the apex spectrum. The HPLC-PDA method was tested with reference and marine sediment samples. Concentrations of selected dioxin-like PCBs, selected nonplanar PCBs, and summed PCBs in sediments and National Institute of Standards and Technology standard reference materials determined by our rapid HPLC-PDA method were comparable with the levels in the same samples analyzed by alternative comprehensive methods (i.e., gas chromatography-electron capture detection or high-resolution gas chromatography-high-resolution mass spectrometry).

Global distribution of halogenated dimethyl bipyrroles in marine mammal blubber.

Four halogenated dimethyl bipyrroles (HDBPs), hypothesized to be naturally produced, were quantitated in marine mammal blubber from a number of species obtained from various locations worldwide. HDBPs were found in samples from all locations studied. Concentrations of total HDBPs (SigmaHDBPs) ranged from 0.4 ng/g lipid weight in ringed seals (Phoca hispida) from the White Sea to 2,540 ng/g lipid weight in Dall's porpoise (Phocoenoides dalli) from the northwestern North Pacific Ocean. At their highest levels, SigmaHDBPs made up 11% of the total quantitated organohalogen body burden of adult male Dall's porpoises. In two beluga (Delphinapterus leucas) data subsets, it was found that males contained significantly higher concentrations of SigmaHDBPs than females. No significant effects of age or sex on SigmaHDBPs were observed in harbor seal (Phoca vitulina) and bowhead whale (Balaena mysticetus) data subsets. The geographical distribution of concentrations did not resemble that of the ubiquitous anthropogenic organohalogen, polychlorinated biphenyl congener CB-153. Higher concentrations of HDBPs and different patterns of congeners were observed in samples from Pacific as opposed to non-Pacific Ocean influenced environments. Concentrations of HDBPs in beluga from the Arctic and St. Lawrence River were similar. Their high abundance in north Pacific Ocean biota and widespread occurrence suggest that HDBPs undergo extensive transport from a source located primarily in the Pacific Ocean. Evidence from HDBP congener patterns indicates that both ocean currents and atmospheric transport likely play a role in the movement of HDBPs. These results imply that HDBPs and anthropogenic organohalogens have different sources and support the natural production hypothesis.

An efficient clean-up method for the GC-MS determination of methylsulfonyl-PCBs/DDEs extracted from various marine mammal tissues.

Existing methods for analyzing methylsulfonyl-PCBs in animal tissues require multiple laborious clean-up steps prior to quantitation. In this paper, we report a new, efficient method for the isolation and determination of methylsulfonyl-PCB metabolites in the blubber, liver, kidney, lung, and milk from mammals of marine origin using: solvent extraction, GPC fractionation, and clean-up on an adsorption chromatography column prior to analysis by gas chromatography-mass spectrometry. The method is rapid and can be completed in about half the analysis time required by most other reported methods. The method exhibits excellent analyte recoveries (89-95%), and good reproducibilities with CV's ranging from 3-12% depending on analyte concentration. Detection limits are approximately 1 ng/g lipid. The method is further validated by comparing its quantitative results to that of an independent reference analytical method for several different marine mammal blubber samples. Finally, the method is applied to profile the distribution of methylsulfones among the various tissue types studied for a single bottle-nose dolphin (Tursiops truncatus) sample. In addition, the extent of their transplacental transfer from mother to unborn fetus for several pilot whale (Globicephala melaena) mother-fetus pairs is examined.

Influence of life-history parameters on organochlorine concentrations in free-ranging killer whales (Orcinus orca) from Prince William Sound, AK.

Certain populations of killer whales (Orcinus orca) have been extensively studied over the past 30 years, including populations that use Puget Sound, WA, the inside waters of British Columbia, Southeastern Alaska and Kenai Fjords/Prince William Sound, Alaska. Two eco-types of killer whales, 'transient' and 'resident', occur in all of these regions. These eco-types are genetically distinct and differ in various aspects of morphology, vocalization patterns, diet and habitat use. Various genetic and photo-identification studies of eastern North Pacific killer whales have provided information on the male-female composition of most of these resident pods and transient groups, as well as the approximate ages, reproductive status and putative recruitment order (birth order) of the individual whales. Biopsy blubber samples of free-ranging resident and transient killer whales from the Kenai Fjords/Prince William Sound, AK region were acquired during the 1994-1999 field seasons and analyzed for selected organochlorines (OCs), including dioxin-like CB congeners and DDTs. Concentrations of OCs in transient killer whales (marine mammal-eating) were much higher than those found in resident animals (fish-eating) apparently due to differences in diets of these two killer whale eco-types. Certain life-history parameters such as sex, age and reproductive status also influenced the concentrations of OCs in the Alaskan killer whales. Reproductive female whales contained much lower levels of OCs than sexually immature whales or mature male animals in the same age class likely due to transfer of OCs from the female to her offspring during gestation and lactation. Recruitment order also influenced the concentrations of OCs in the Alaskan killer whales. In adult male residents, first-recruited whales contained much higher OC concentrations than those measured in non-first-recruited (e.g. second recruited, third recruited) resident animals in the same age group. This study provides baseline OC data for free ranging Alaskan killer whales for which there is little contaminant information.

Organochlorine contaminant levels in Eskimo harvested bowhead whales of arctic Alaska.

Organochlorine (OC) levels in liver and blubber of 20 bowhead whales (Balaena mysticetus) collected during the Eskimo subsistence harvest at Barrow (Alaska, USA) in 1992 and 1993 are presented. Liver sum DDT (lipid weight) was significantly greater in male whales than in females. Most of the organochlorines measured were at higher levels in longer (older) than in shorter (younger) males. For female bowhead whales, hexachlorobenzene and lipid levels decreased and other OC levels did not change significantly with increasing length. Most organochlorine contaminants have low concentrations in tissues of the bowhead whale compared to concentrations in tissues of other cetaceans, especially Odontocetes. Based on allowable daily intakes (ADI) levels established by the Canadian Northern Contaminants Program (Ottawa, Ontario, Canada) "safe" levels of blubber to consume were calculated. Chlordane levels in bowhead whale blubber results in the most restrictive consumption amount (50 g blubber/day). We expect no adverse effects related to these organochlorine contaminants to occur in bowhead whales or in consumers of their tissues. However, investigation of low level chronic exposure effects and a more rigorous assessment of histopathology, biomarkers, and immune status in the bowhead whale would be required to conclude "no effect" with more certainty.

Factors affecting organochlorine contaminant concentrations in milk and blood of northern fur seal (Callorhinus ursinus) dams and pups from St. George Island, Alaska.

Levels of organochlorine contaminants in blood of northern fur seal (Callorhinus ursinus) pups and the blood and milk of their dams early in the lactation period are reported here. The contaminants included 15 selected individual polychlorinated biphenyl (PCB) congeners and DDT metabolites identified through high-performance liquid chromatography. Congeners CB-77, -81, -126, -169 and -189 were below the limits of detection in milk and blood samples analyzed. Congener-specific concentrations of PCBs in the blood of pups were compared based on the age of their dam (< or = 5 years or > 7 years). Pups of young (presumably primiparous) dams had significantly elevated levels of CBs-101, -118, -128, -138, -153/87, -170/194, and -180 than pups of older (multiparous) dams. Congeners CB-128 and -170/194 were detected in the blood of pups of young dams but not in the blood of pups of older dams nor in any of the dams blood. Additionally, pups had higher blood levels in seven of 10 detected PCB congeners as compared to the levels measured in milk when adjusted for lipid content. Levels of DDT metabolites and toxic equivalency quotients of dioxin-like congeners followed similar trends. Lipid-normalized concentrations of CB-101 and total PCBs were significantly higher in the blood of dams than in their milk. CB-128, -156, -157, -170/194, p,p'-DDT, o,p'-DDT, p,p'-DDD and o,p'-DDD were not detected in dam blood samples, but were detected in milk samples. Calculation of 'biomagnification factors' from milk to pup blood indicated a biomagnification of CB-101, -105, -118, -138, -153/87, and -180. Significant mean accumulation factors ranged from 1.5 to 7.5. Inter-annual differences in exposure levels and specific congener concentrations in both milk and blood were apparent. Northern fur seal pups, especially first-born, have a substantial exposure to organochlorine contaminants at a critical developmental stage.

Analyses of tissues of eight marine species from Atlantic and Pacific coasts for dioxin-like chlorobiphenyls (CBs) and total CBs.

Eight commercially and recreationally important marine species were collected in 1993 and 1994 from several Atlantic and Pacific coastal regions of the contiguous United States. Approximately 700 edible tissue samples (e.g., whole body of mussel, crustacean muscle and hepatopancreas, and fish muscle) were analyzed for dioxin-like chlorobiphenyls (CBs) and other selected CB congeners using a rapid high-performance liquid chromatography photodiode array detection method (HPLC/PDA). Total CBs and toxic equivalents (TEQs) of dioxin-like CBs were also determined. The most abundant congeners measured in these tissues were the moderately chlorinated CBs (e.g., CB 138, 153), with mean concentrations ranging from below the limits of detection (approximately 0.2 ng/g) to 1,500 ng/g, wet weight. Certain dioxin-like CBs (e.g., CBs 77, 105, 118, 126) were also found in several of these samples (mean concentrations ranging from below the limits of detection [approximately 0.4 ng/g] to 680 ng/g). Similar to previous studies, the majority of seafood tissues contained total CB concentrations that were below the U.S. Food and Drug Administration's (FDA) tolerance limit for CBs of 2,000 ng/g, wet weight (2.0 ppm). Furthermore, the majority of samples that contained CB levels below the FDA CB tolerance limit also had CB TEQs that were lower than the FDA's advisory level for TCDD (25 ppt or 25 pg/g, wet weight) in fish from the Great Lakes, which is used in evaluating CB TEQs. Several crustacean hepatopancreas samples collected from certain Atlantic and Pacific urban sites (e.g., Dungeness crab from Elliott Bay in Puget Sound, WA, American lobster from Deer Island in Boston Harbor, MA), however, did contain total CB and CB TEQs that exceeded the FDA CB tolerance and TCDD advisory limits. Mono-ortho- (e.g., CBs 118, 105) and non-ortho-substituted congeners (e.g., CBs 77 and 126) were the largest contributors to the CB TEQs of the hepatopancreas samples that exceeded the action limit.http://link.springer-ny. com/link/service/journals/00244/bibs/37n2p205.html

Organochlorine contaminants in blubber of four seal species: integrating biomonitoring and specimen banking.

Blubber samples from four Alaska seal species (bearded seal, Erignathus barbatus, harbor seal, Phoca vitulina, northern fur seal, Callorhinus ursinus, ringed seal, P. hispida) were collected for inclusion in the US National Biomonitoring Specimen Bank, as well as for immediate analysis as part of the contaminant monitoring component of the US National Marine Fisheries Service's Marine Mammal Health and Stranding Response Program. The blubber samples were analyzed for organochlorine (OC) contaminants (e.g., PCB congeners, pesticides, DDTs). Results for bearded and ringed seals from the Alaska Arctic revealed low blubber concentrations of OC contaminants. Harbor seals from Prince William Sound. Gulf of Alaska, had somewhat higher blubber concentrations of OC contaminants. In contrast, northern fur seals sampled from the Pribilof Islands had blubber concentrations of certain OC contaminants that were about an order of magnitude higher than those found in the other seal species. Differences in contaminant concentrations among the Alaska seals may be explained by differences in feeding habits and migratory patterns, age or gender did not appear to account for the differences observed. The highest concentrations of OCs were found in harbor seals stranded along the northwestern US mainland, which is consistent with higher concentrations of anthropogenic contaminants being found in urban coastal areas than in more remote Arctic environments. The integration of real-time contaminant monitoring with specimen banking provides important baseline data that can be used to plan and manage banking activities. This includes identifying appropriate specimens that are useful in assessing temporal trends and increasing the utility of the banked samples in assessing chemical contaminant accumulation and relationships to biological effects.

Screening for planar chlorobiphenyl congeners in tissues of marine biota by high-performance liquid chromatography with photodiode array detection.

A rapid method has been developed to screen for planar chlorobiphenyl (CB) congeners, as well as certain other CBs and DDTs, in tissue samples from marine biota. The analytes were extracted from tissue matrices with 1:1 hexane/pentane (v/v) and interfering compounds were separated from the CBs on a gravity-flow column packed with acidic, basic and neutral silica gel eluted with 1:1 hexane/methylene chloride (v/v). Subsequently, the planar CB congeners were resolved from the DDTs and other CBs by HPLC on Cosmosil PYE analytical columns cooled to 9 degrees C and were measured by an ultraviolet (UV) photodiode array (PDA) detector. Two important advantages of PDA over conventional UV detection were the ability to identify individual analytes by comparing their UV spectra to those of reference standards and the ability to establish the spectral homogeneity (purity) of the analytes by comparing spectra within a peak to the apex spectrum. The HPLC/PDA method was tested with tissue samples from fish, shellfish and marine mammals; concentrations of certain CBs and DDTs in samples determined by screening compared favorably with those in the same samples analyzed by a comprehensive method (e.g., gas chromatography/high resolution mass spectrometry). However, the HPLC/PDA method was about an order of magnitude less sensitive than determinations by comprehensive methods.

Relationships between hepatic neoplasms and related lesions and exposure to toxic chemicals in marine fish from the U.S. West Coast.

English sole (Parophrys vetulus) inhabiting polluted waterways and embayments of Puget Sound, Washington, are affected with a variety of multiple, co-occurring idiopathic hepatic lesions, including unique degenerative conditions, putatively preneoplastic foci of cellular alteration, and neoplasms. Results of a statistical analysis of the patterns of co-occurrence of these lesions in wild English sole are consistent with the concept that these lesions represent morphologically identifiable steps forming a sequence of progression ultimately leading to the development of hepatic neoplasms. This progressive sequence parallels the pattern identified in experimental models of chemically induced hepatocarcinogenesis in rodents. The rationale for the hypothesis that these lesions in wild English sole can be caused by exposure to certain hepatoxic and hepatocarcinogenic xenobiotic compounds in the marine environment is based on the demonstration of significant and consistent statistical associations between levels of aromatic hydrocarbons (AHs) in sediment and prevalences of these idiopathic liver lesions; a significant contribution by sediment AHs to the variability in hepatic neoplasm prevalence in a logistic regression model; significantly increased probabilities for several idiopathic lesions in sole from chemically contaminated sites in Puget Sound; significant correlations between uptake of polycyclic aromatic hydrocarbons, as measured by levels of fluorescent metabolites of aromatic compounds in bile of sole, and prevalences of several hepatic lesion types; and experimental induction of unique degenerative, proliferative, and putatively preneoplastic focal lesions in English sole injected with either benzo(a)pyrene or a polycyclic aromatic hydrocarbons (PAH) enriched fraction of an extract from a contaminated urban sediment from Puget Sound.(ABSTRACT TRUNCATED AT 250 WORDS)

Overview of studies on liver carcinogenesis in English sole from Puget Sound; evidence for a xenobiotic chemical etiology. I: Pathology and epizootiology.

Livers of wild English sole (Parophrys vetulus) from polluted waterways and embayments of Puget Sound, Washington, are affected by a spectrum of multiple, co-occurring idiopathic hepatic lesions, including neoplasms, putative preneoplastic foci of cellular alteration, and unique degeneration conditions. Results from a statistical analysis of the patterns of co-occurrence of these lesions in wild English sole indicate that these lesions represent morphologically identifiable steps leading to the development of hepatic neoplasms. This sequence parallels the lesion progression in experimental models of chemically induced liver carcinogenesis in rodents. The hypothesis that these lesions in wild English sole can be caused by exposure to certain xenobiotic hepatotoxic and hepatocarcinogenic compounds in Puget Sound is based on: a) statistical associations between levels of aromatic hydrocarbons (sigma AHs) in sediment and prevalences of these idiopathic liver lesions, b) the contribution of sigma AHs in accounting for the variability in hepatic neoplasm prevalence in a logistic regression model, c) elevated odds ratios for several idiopathic hepatic lesion types in sole from polluted sites in Puget Sound, d) significant correlations between prevalences of idiopathic hepatic lesions and levels of fluorescent metabolites of aromatic compounds (FACs) in bile of English sole, and e) experimental induction of putatively preneoplastic focal lesions in English sole injected with a PAH-enriched fraction of an extract from a contaminated urban sediment from Puget Sound, that were morphologically identical to lesions found in wild English sole from the same site.

Uptake of aromatic and chlorinated hydrocarbons by juvenile chinook salmon (Oncorhynchus tshawytscha) in an urban estuary.

A study was conducted to assess the potential for uptake of toxic chemicals by down-stream migrant salmon in an urban estuary. Juvenile chinook salmon (Oncorhynchus tshawytscha) were collected from the Duwamish Waterway (located in Seattle, Washington) and from the Nisqually River (a reference site). The mean concentrations of aromatic hydrocarbons and polychlorinated biphenyls (PCBs) in the stomach contents (food organisms) of salmon from the Duwamish Waterway were approximately 650 times and 4 times, respectively, higher than those in salmon from the Nisqually River. Similarly, the mean concentration of bile metabolites of aromatic compounds which fluoresce at benzo[a]pyrene wavelengths was 24 times higher in the urban salmon compared to the reference salmon, whereas the mean concentration of PCBs in liver of urban salmon was 3 times higher than that in reference salmon. The study clearly demonstrated that, during their residency in this urban estuary, juvenile chinook salmon bioaccumulate substantial levels of toxic chemicals. The possible effects of these chemical exposures on the health and survival of this species are not presently known.

High-performance liquid chromatographic method for isolating coprostanol from sediment extracts.

We report a rapid, largely automated high-performance liquid chromatographic (HPLC) method, which uses an HPLC column packed with alkylnitrile-substituted secondary alkylamine (aminocyano) bonded phase, to isolate coprostanol from interfering compounds in sediment extracts. Coprostanol is then quantitated, as the trimethylsilyl ether, by gas chromatography with flame ionization detection. Results from using the HPLC method to analyze a sediment reference material for coprostanol were statistically comparable to a previously used gravity-flow column method. We also report the coprostanol concentrations in several sediment samples from the California coast which reflect a range of sewage contamination (62-15,000 ng/g).

High-performance liquid chromatographic method for isolating organic contaminants from tissue and sediment extracts.

Interest in the assessment of the anthropogenic contamination of the marine environment has accelerated in recent years. Existing methods to analyze environmental samples (e.g., sediments or tissues) for trace amounts of organic contaminants such as aromatic hydrocarbons and chlorinated compounds are tedious and costly. We report a rapid, efficient high-performance liquid chromatography (HPLC) procedure which uses a size-exclusion column to separate the analytes of interest from interfering compounds in the sample matrix. Analytical results from the HPLC method were, in general, comparable to a gravity-column method which had been used for several years. The HPLC method had several other advantages: improved precision; the ability to monitor chromatographic conditions; the potential for automating analyses; and reduced consumption of solvents and other materials.

Field and laboratory studies of the etiology of liver neoplasms in marine fish from Puget Sound.

A series of field studies was conducted between 1979 and 1985 in Puget Sound, Washington State, to investigate etiological relationships between prevalences of hepatic neoplasms in bottom-dwelling marine fish species, with emphasis on English sole (Parophrys vetulus), and concentrations of toxic chemicals in sediments and affected fish. Statistically significant (p less than or equal to 0.05) correlations have been found between the prevalences of hepatic neoplasms in English sole and the following parameters: sediment concentrations of aromatic hydrocarbons, and concentrations of the metabolites of aromatic compounds in the bile of affected sole. A significant difference (p less than 0.001) was also found between the relative concentrations of aromatic free radicals in the liver microsomes of English sole with liver lesions compared to sole without liver lesions. Laboratory studies designed to evaluate the etiology of the liver neoplasms in English sole have also yielded evidence that is consistent with the view that high molecular weight aromatic hydrocarbons, e.g., benzo[a]pyrene (BaP), are hepatocarcinogens in English sole. The current status of a series of long-term (up to 18 months) exposures of English sole and rainbow trout (Salmo gairdneri) to selected fractions of Puget Sound sediment extracts, enriched with aromatic hydrocarbons and nitrogen-containing aromatic compounds, and to individual carcinogens (e.g., BaP) is discussed.

Toxic chemicals in sediments and biota from a creosote-polluted harbor: relationships with hepatic neoplasms and other hepatic lesions in English sole (Parophrys vetulus).

High prevalences of idiopathic hepatic lesions, including neoplasms (e.g., hepatocellular carcinomas, cholangiocellular carcinomas) (27%, 20 of 75 fish) and foci of cellular alteration (putative 'preneoplastic' lesions) (44%, 33 of 75 fish), were found in English sole (Parophrys vetulus) exposed to creosote-contaminated sediments in Eagle Harbor, Puget Sound, WA. Sediments from the contaminated region of the harbor contained particularly high concentrations of aromatic hydrocarbons (e.g., benzo[a]pyrene and benz[a]anthracene), and a variety of nitrogen-containing aromatic compounds (e.g., carbazole and acridine). The composition of the aromatic compounds was characteristic of creosote. Dramatically lower concentrations of aromatic compounds were found in sediments from a reference site in which the bottom-dwelling fish examined were free of detectable neoplastic or 'preneoplastic' hepatic lesions. Food organisms in the stomachs of the English sole from Eagle Harbor contained substantially higher concentrations of aromatic hydrocarbons than comparable organisms from the reference site. The concentrations of individual aromatic hydrocarbons in muscle and liver from the Eagle Harbor fish were low; however, high concentrations of metabolites of aromatic compounds were present in the bile. The findings strongly suggest an association between exposure to creosote and the prevalence of hepatic lesions, including neoplasms, in the bottom-dwelling fish, and furthermore support the putative role of aromatic hydrocarbons in liver carcinogenesis in fish.

Toxic chemicals in marine sediment and biota from Mukilteo, Washington: relationships with hepatic neoplasms and other hepatic lesions in English sole (Parophrys vetulus).

High prevalences of idiopathic liver lesions, including 7.5% hepatic neoplasms (e.g., hepatocellular and cholangiocellular carcinomas) and 16.7% foci of cellular alteration (putative preneoplastic hepatic lesions), were found in English sole (Parophrys vetulus) from waters near Mukilteo, a small community on Puget Sound in Washington State. Sediments from the sampling sites contained particularly high concentrations of aromatic hydrocarbons. Substantially lower concentrations of these compounds were found in sediments from another Puget Sound (reference) site in which fish showed no evidence of hepatic neoplasms or foci of cellular alteration. Stomach contents from the fish at Mukilteo contained substantially higher concentrations of the chemicals than did stomach contents of fish from the relatively uncontaminated site. High concentrations of metabolites of aromatic compounds were measured in the bile of fish from Mukilteo. These findings support previously observed relationships between sediment chemicals (e.g., aromatic hydrocarbons) and high prevalences of liver lesions in English sole from Puget Sound. In addition, a dietary route of uptake by English sole of environmental chemicals, including known carcinogens, was documented.