Catalytic and immunochemical properties of hepatic cytochrome P450 1A in three avian species treated with beta-naphthoflavone or isosafrole.

Induction of cytochrome P450 1A (CYP1A) can be used as a biomarker of exposure to planar halogenated aromatic hydrocarbons (PHAHs). Our objective was to characterize the induction of CYP1A activity and protein in three avian species following in vivo treatment with beta-naphthoflavone (BNF) and/or isosafrole. Alkoxyresorufin-O-dealkylase (alk-ROD) activities of hepatic microsomes from Herring Gulls (Larus argentatus) (HGs), Double-crested Cormorants (Phalacrocorax auritus) (DCCs) and chickens (Gallus domesticus) were measured using ethoxy-, methoxy-, pentoxy- and benzyloxy-resorufin, in the presence and absence of the inhibitors ellipticine or furafylline. Immunoreactivity of microsomal proteins with antibodies to several CYP1A proteins was investigated. CYP1A protein and alk-ROD activities of HGs and DCCs, but not chickens, were induced by isosafrole. Ellipticine was a potent and non-selective inhibitor of alk-ROD activity in all three species, while furafylline inhibition of alk-ROD activities varied among species and treatments. In all three species, BNF induced a protein immunoreactive with monoclonal antibody to CYP1A1 from the marine fish Stenotomus chrysops (scup), but a CYP1A2-like protein was not detected in avian microsomes probed with polyclonal antibodies to mouse CYP1A2. Variations in responses among avian species indicate that CYP1A proteins and substrate specificities should be characterized for each species used in PHAH biomonitoring programs.

Multigenerational study of the effects of consumption of PCB-contaminated carp from Saginaw Bay, Lake Huron, on mink. 1. Effects on mink reproduction, kit growth and survival, and selected biological parameters.

This study was conducted to determine the multigenerational effects of consumption of PCB-contaminated carp (Cyprinus carpio) from Saginaw Bay (Lake Huron) on mink (Mustela vison) reproduction and health and to examine selected biomarkers as potential indicators of polyhalogenated hydrocarbon toxicity in mink. The mink were fed diets formulated to provide 0 (control), 0.25, 0.5, or 1.0 ppm polychlorinated biphenyls (PCBs) through substitution of Saginaw Bay carp for ocean fish in the diets. To determine whether the effects of PCB exposure were permanent, half of the parental (P1) animals were switched from their respective treatment diets to the control diet after whelping the first of two F1 generations. Effects of in utero and lactational exposure to PCBs on subsequent reproductive performance of the F1 animals were examined by switching half of the first-year F1 offspring (kits) to the control diet at weaning, while the other half was continued on their parental diet (continuous exposure). Continuous exposure to 0.25 ppm, or more, of PCBs delayed the onset of estrus (as determined by vulvar swelling and time of mating) and lessened the whelping rate. Litters whelped by females continually exposed to 0.5 ppm, or more, of PCBs had greater mortality and lesser body weights than controls. Continuous exposure to 1.0 ppm PCBs had a variable effect on serum T4 and T3 concentrations. Compared to the controls, there were significant differences in kidney, liver, brain, spleen, heart, and thyroid gland weights of the mink continually exposed to 1.0 ppm PCBs. There was an increase in the incidence of periportal and diffuse vacuolar hepatocellular lipidosis in the P1 mink with continuous exposure to increasing concentrations of PCBs. Plasma and liver PCB concentrations of the adult and kit mink were, in general, directly related to the dietary concentration of PCBs and the duration and time of exposure. Short-term parental exposure to PCBs had detrimental effects on survival of subsequent generations of mink conceived months after the parents were placed on "clean" feed. The lowest observed adverse effect level (LOAEL) for dietary PCBs in this study was 0.25 ppm.

Effects induced by feeding organochlorine-contaminated carp from Saginaw Bay, Lake Huron, to laying White Leghorn hens. I. Effects on health of adult hens, egg production, and fertility.

This study was conducted to determine the effects of consumption of halogenated hydrocarbon compounds, primarily polychlorinated biphenyls (PCBs), contained in Great Lakes fish by the domestic chicken (Gallus domesticus). In this article we report the results of feeding White Leghorn hens for a period of 8 wk diets that contained 31-35% ocean fish and/or carp (Cyprinus carpio) from Saginaw Bay, Lake Huron, MI, which provided 0.3 (control), 0.8 (low-dose group), or 6.6 (high-dose group) mg PCB/kg, wet weight (ww). These concentrations were analogous to 3.3, 26, or 59 pg 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) equivalents (TEQs)/g diet, ww, respectively. There were no significant effects on feed consumption among the groups. An unexpectedly high incidence of fatty liver hemorrhagic syndrome (FLHS) was observed in hens from the control (78% FLHS) and low-dose (75% FLHS) groups when compared to the high-dose group (15% FLHS). Birds in the control and low-dose groups had a significant increase in liver and body weights. Significant decreases in egg production, weight, and fertility were immediate in all dose groups, with the effect being permanent in the control and low-dose groups. Although the incidence of FLHS was an unexpected complication, the fact that there were no significant effects on egg production, egg weights, or fertility in the high-dose group suggests that the no-observable-adverse-effect concentration (NOAEC) for these parameters is in excess of 26 mg total weathered PCBs/kg egg, ww. This value was the average concentration of PCBs in the high-dose group eggs during the last week of the study.

Effects induced by feeding organochlorine-contaminated carp from Saginaw Bay, Lake Huron, to laying White Leghorn hens. II. Embryotoxic and teratogenic effects.

Carp from Saginaw Bay, Lake Huron, MI, was fed to White Leghorn chickens for a period of 8 wk. The diets contained 0.3 (control; 0% carp), 0.8 (3.4% carp), and 6.6 (35% carp) mg polychlorinated biphenyls (PCBs)/kg diet, by wet weight (ww). These concentrations corresponded to 3.3, 26, and 59 pg 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) equivalents/g diet ww, respectively. Though the diets were not acutely toxic to the adult laying hens, dose- and time-dependent responses were observed in the embryos and chicks. Toxicity was manifested as a dose-dependent increase in embryo mortality and decreased hatching rates. Furthermore, embryos and chicks displayed various deformities, including (1) head and neck edema and hemorrhage, (2) abdominal edema and hemorrhage, (3) foot and leg deformities, (4) skull and brain deformities, (5) yolk-sac deformities, and (6) miscellaneous deformities. The types of deformities observed were similar to those reported for embryos and chicks of colonial waterbirds in Saginaw Bay, as well as in controlled studies where technical mixtures or individual congeners of polychlorinated diaromatic hydrocarbons (PCDAHs) were fed to chickens. Increasing concentrations of carp also significantly affected the various organ weights in 18-d embryos and hatched chicks. At 18 d of incubation, weights of the embryos' livers were directly proportional to the concentration of PCBs in the diets. The weights of the spleens and bursae were inversely proportional to the dietary PCB concentration. After 3 additional days of incubation, significant effects in body, brain, liver, heart, and bursa weights were observed in hatched chicks. The concentrations of total PCBs, as well as 2,3,7,8-TCDD equivalents (TEQs) in the diets, were in the range of those that have been shown to cause similar adverse effects in other species. This study has shown that fish, the primary food source of colonial waterbirds in Saginaw Bay, are capable of causing adverse reproductive effects in a model avian species, the chicken. However, due to differences in the relative potency to cause effects on different endpoints in different species, the results of this study should not be used to predict the threshold for effects in other species.

Contaminants of fishes from Great Lakes-influenced sections and above dams of three Michigan rivers: III. Implications for health of bald eagles.

Recently, there have been discussions of the relative merits of passage of fishes around hydroelectric dams on three rivers (Au Sable, Manistee, and Muskegon) in Michigan. A hazard assessment was conducted to determine the potential for adverse effects on bald eagles that could consume such fishes from above and below dams on the three primary rivers. The hazard assessments were verified by comparing the reproductive productivities of eagles nesting in areas where they ate primarily fish from either above or below dams on the three primary rivers, as well as on two additional rivers in Michigan, the Menominee and Thunder Bay. Concentrations of organochlorine insecticides (OCI), polychlorinated biphenyls (total PCBs), 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents (TCDD-EQ), and total mercury (Hg) were measured in composite samples of fishes from above and below hydroelectric dams on the Manistee and Muskegon Rivers, which flow into Lake Michigan, and the Au Sable River, which flows into Lake Huron. Mean concentrations of OCI, total PCBs, and TCDD-EQ were all greater in fishes from below the dams than in those from above. The hazard assessment indicated that current concentrations of Hg and OCI other than DDT (DDT+DDE+DDD) in fish from neither above nor below dams would present a significant hazard to bald eagles (Haliaeetus leucocephalus). Both total PCBs and TCDD-EQ in fishes from below the dams currently present a significant hazard to bald eagles, since their mean hazard quotients (HQ) were all greater than one.

Polychlorinated biphenyls and 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents in eggs of double-crested cormorants from a colony near Green Bay, Wisconsin, USA.

Great Lakes colonial waterbirds have experienced poor reproduction and a greater incidence of birth defects than those in remote areas. An egg was collected from each of 1,000 marked cormorant nests at Spider Island (Lake Michigan). Nine pools comprised of three eggs were randomly selected for instrumental quantification of polychlorinated biphenyls (PCB) congeners, calculation of 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents (TEq) and measurement of equivalents by bioassay (TCDD-EQ). PCB analysis of the nine samples was semi-automated with high performance liquid chromatography (HPLC) columns including a porous graphitic carbon column. TEqs were calculated from concentrations of PCB congeners and bioassay-derived toxic equivalency factors (TEfs), and TCDD-EQ were measured directly with an H4IIE bioassay. Total PCB concentrations ranged from 9.7 to 38 micrograms/g, wet weight (ww). Mean concentrations of PCB 77, 126, and 169 were 2, 7, and 1 ng/g, ww. The mean TEqs and TCDD-EQ were 150 and 350 pg/g, ww, respectively. Thus, PCB congeners contributed less than 50% of the total TCDD-EQs as measured by the bioassay.

Dietary exposure of mink to carp from Saginaw Bay, Michigan: 2. Hematology and liver pathology.

The effects of consumption of environmental contaminants contained in carp (Cyprinus carpio) from Saginaw Bay, Michigan on various hematological parameters and liver integrity of adult female mink (Mustela vision) were determined. Mink were fed diets that contained 0 (control), 10, 20, or 40% carp prior to and throughout the reproductive period (182 days). The diets contained 0.015, 0.72, 1.53, and 2.56 mg polychlorinated biphenyls (PCBs)/kg diet and 1.0, 19, 40, and 81 pg 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents (TEQs)/g diet, respectively. Mink fed the diets containing carp showed a general dose-dependent occurrence of clinical signs commonly associated with chlorinated hydrocarbon toxicity, including listlessness, nervousness when approached, anorexia, and melena. Erythrocyte counts were less in mink exposed to Saginaw Bay carp than in controls, while the number of white blood cells was greater than in controls. Significant differences (p < 0.05) in the concentrations of neutrophils, lymphocytes, monocytes, and eosinophils were also found between the control and carp-fed groups, but are considered to be of limited clinical or biological importance. Hematocrit values for the mink fed the 20 and 40% carp diets were significantly less than those of mink in the control and 10% carp groups. There were no significant differences in hemoglobin concentrations among the groups. Necropsies revealed enlarged yellowish livers in many of the carp-fed mink, especially those fed the 40% carp diet. Liver, spleen, and lung weights of carp-fed mink were significantly greater than those of control mink.(ABSTRACT TRUNCATED AT 250 WORDS)

Dietary exposure of mink to carp from Saginaw Bay, Michigan. 1. Effects on reproduction and survival, and the potential risks to wild mink populations.

Carp (Cyprinus carpio) collected from Saginaw Bay, Michigan, containing 8.4 mg total polychlorinated biphenyls (PCBs)/kg and 194 ng of 2,3,7,8-tetrachloro-dibenzo-p-dioxin equivalents (TEQs)/kg, were substituted for marine fish at levels of 0, 10, 20, or 40% in the diets of adult ranch mink (Mustela vison). The diets, containing 0.015, 0.72, 1.53, and 2.56 mg PCBs/kg diet, or 1.03, 19.41, 40.02, and 80.76 ng TEQs/kg diet, respectively, were fed to mink prior to and throughout the reproductive period to evaluate the effects of a naturally-contaminated prey species on their survival and reproductive performance. The total quantities of PCBs ingested by the mink fed 0, 10, 20, or 40% carp over the 85-day treatment period were 0.34, 13.2, 25.3, and 32.3 mg PCBs/mink. respectively. The corresponding quantities of TEQs ingested by the mink over the same treatment period were 23, 356, 661, and 1,019 ng TEQs/mink, respectively. Consumption of feed by mink was inversely proportional to the PCB and TEQ content of the diet. The diet containing Saginaw Bay carp caused impaired reproduction and/or reduced survival of the kits. Compared to controls, body weights of kits at birth were significantly reduced in the 20 and 40% carp groups, and kit body weights and survival in the 10 and 20% carp groups were significantly reduced at three and six weeks of age. The females fed 40% carp whelped the fewest number of kits, all of which were stillborn or died within 24 hours. Lowest observable adverse effect levels (LOAEL) of 0.134 mg PCBs/kg body weight/day or 3.6 ng TEQs/kg body weight/day for adult female mink were determined. The potential effects of exposure of wild mink to contaminated Great Lakes fish were assessed by calculating "maximum allowable daily intakes" and "hazard indices" based on total concentrations of PCB residues in several species of Great Lakes fish and mink toxicity data derived from the study.

Contaminants in fishes from Great Lakes-influenced sections and above dams of three Michigan rivers. I: Concentrations of organo chlorine insecticides, polychlorinated biphenyls, dioxin equivalents, and mercury.

Fishes of the Great Lakes contain hazardous chemicals such as synthetic halogenated hydrocarbons and metals. These fish can move from the lakes into the Great Lakes tributaries of Michigan. In doing so, they transport concentrations of contaminants which may represent a risk to wildlife. Concentrations of mercury (Hg), total polychlorinated biphenyls (PCBs), 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents (TCDD-EQ), total DDT complex, aldrin, endrin, dieldrin, heptachlor, heptachlor epoxide, lindane, hexachlorobenzene, cis-chlordane, oxychlordane, endosulfan-I, methoxychlor, trans-chlordane, and trans-nonachlor were determined in composite samples of fishes from above and below Michigan hydroelectric dams, which separate the fishes which have access to the Great Lakes from fishes that do not. Mean concentrations of total PCBs, TCDD-EQ, DDT, and most of the other pesticides were greater in composite samples of six species of fishes from below than above the dams on the Au Sable, Manistee, and Muskegon Rivers. Concentrations of mercury, were the same or greater above the dams than below. However, this difference was statistically significant only on the Au Sable. Mercury concentrations ranged from less than 0.05 mg/kg to 0.73 mg Hg/kg, ww. Total concentrations of PCBs ranged from 0.02 to 1.7 mg/kg, ww. Concentrations of 2,3,7,8-tetrachlordibenzo-p-dioxin equivalents varied among fishes and locations. The concentrations of TCDD-EQ ranged from 2.4 to 71 micrograms/kg, ww, with concentrations in carp being the greatest. Concentrations of TCDD-EQ were greater than the concentrations which would be expected to occur, due solely to the presence of polychlorinated dibenzo-p-dioxins (PCDD), polychlorinated dibenzofurans (PCDF), and technical mixtures of PCBs.

Contaminants in fishes from Great Lakes-influenced sections and above dams of three Michigan rivers. II: Implications for health of mink.

Populations of mink (Mustela vison) have declined in many areas of the world. Such declines have been linked to exposures to synthetic, halogenated hydrocarbons. In the Great Lakes region, mink are fewer in areas along the shore of the Great Lakes and their tributaries where mink have access to fish from the Great Lakes. Recently, there has been discussion of the relative merits of passage of fishes around hydroelectric dams on rivers in Michigan. A hazard assessment was conducted to determine the potential for adverse effects on mink, which could consume such fishes from above or below dams on the rivers. Concentrations of organochlorine insecticides, polychlorinated biphenyls (PCBs), 2,3,7,8-tetrachlordibenzo-p-dioxin equivalents (TCDD-EQ), and total mercury were measured in composite samples of fishes from above or below hydroelectric dams on the Manistee and Muskegon Rivers, which flow into Lake Michigan, and the Au Sable River, which flows into Lake Huron. Concentrations of organochlorine insecticides, PCBs, and TCDD-EQ were all greater in fishes from below the dams than those from above. Concentrations of neither organochlorine insecticides nor mercury in fishes are currently a risk to mink above or below the dams. All of the species of fishes collected from downstream of the dams contained concentrations of PCBs and TCDD-EQ, which represent a hazard to mink. The hazard index for PCBs was less than one for the average of all species from the upstream reaches of the Manistee and Au Sable Rivers, but not the Muskegon. The hazard index (concentration in fish/NOAEC) was greater than 1 for all of the species collected from below the dams, in all three rivers. The greatest hazard index was observed for carp (Cyprinus carpio) downstream on the Muskegon River. Because the concentrations of PCBs used in the hazard assessment were corrected for relative toxic potencies, the hazard ratios based on PCBs should be similar to those based on TCDD-EQ. This was found to be true. Thus, either total PCBs or TCDD-EQ could be used as the critical toxicant in the hazard assessment. However, if uncorrected concentrations of PCBs, expressed as Aroclors, were used in the hazard assessment, the toxicity of the weathered mixture would have been underestimated by approximately five-fold, and, in that instance, TCDD-EQ would be the critical contaminant for the hazard assessment.(ABSTRACT TRUNCATED AT 400 WORDS)

Accumulation of 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents by double-crested cormorant (Phalacrocorax auritus, Pelicaniformes) chicks in the North American Great Lakes.

Concentrations of polychlorinated biphenyls (PCBs), and 2,3,7,8-tetrachlordibenzo-p-dioxin equivalents (TCDD-EQ) were determined in eggs and chicks of double-crested cormorants (DCC) which were collected in 1989 from eight locations in the Laurentian Great Lakes. The mean biomagnification factor (BMF) from forage fish to eggs was found to be 31.3. Absolute and relative concentrations as well as rates of accumulation of total concentrations of PCBs and TCDD-EQ were measurable in all of the samples. The concentrations of both PCBs and TCDD-EQs decreased immediately upon hatching of chicks, due to growth dilution. Initial decreases in absolute masses of TCDD-EQ in chicks were also observed, which indicates that there can be significant elimination of these compounds during early development. The initial rates of accumulation by chicks were dependent only on the mass of fish consumed. After the chicks began thermoregulating, the rates of accumulation, expressed as a concentration, normalized to body weight, became greater. Rates of accumulation of both PCBs and TCDD-EQ were correlated with their respective concentrations in forage fish consumed by the chicks. The relative potency, expressed as the ratio of the concentration of TCDD-EQ to that of total PCBs was calculated to determine if there was significant trophic-level enrichment of the TCDD-EQs, relative to total concentrations of PCBs. A significant enrichment was observed at the more and less contaminated locations, but the degree of enrichment was greater at the less contaminated locations (26 vs 72 micrograms/g).

H4IIE rat hepatoma cell bioassay-derived 2,3,7,8-tetrachlorodibenzo-p-dioxin equivalents in colonial fish-eating waterbird eggs from the Great Lakes.

Fish-eating waterbirds from the Great Lakes of North America have shown symptoms of poisoning similar to those observed in laboratory exposures of various avian species to planar halogenated hydrocarbons (PHHs). PHHs, include among others, polychlorinated biphenyls (PCBs), polychlorinated dibenzo-p-dioxins (PCDDs), and polychlorinated dibenzofurans (PCDFs), and have been implicated in some of the reproductive problems of Great Lakes water-birds. The objectives of this study were to assess the overall potencies of PCB-containing extracts from colonial water-bird eggs taken from the Great Lakes and to compare the potencies with the location and spatial distribution of the colonies. The potencies of the extracts were assessed by their ability to induce cytochrome P450IA1-associated ethoxyresorufin O-deethylase (EROD) activity in H4IIE rat hepatoma cells as compared to the standard, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The H4IIE bioassay-derived TCDD-equivalents (TCCD-EQs) in the waterbird eggs concur with residue analyses and biological data from other studies. The greatest concentrations of TCDD-EQs were found in waterbird eggs from historically polluted, industrialized or urbanized areas in which the reproductive impairment of colonial waterbirds was most severe. However, significant concentrations of TCDD-EQs were detected at all sites tested; with a range of 49 to 415 pg TCDD-EQ/g egg, uncorrected for extraction efficiencies. The H4IIE bioassay proved to be a useful biomonitoring tool to assess the overall potency of complex PHH mixtures in environmental samples.

Development of a potentially wearable glucose sensor for patients with diabetes mellitus: design and in-vitro evaluation.

A potentially wearable glucose sensor was developed, consisting of an oxygen electrode as detector and a dynamic enzyme perfusion system as selector. The selector is a hollow fibre, which can be placed subcutaneously and dialyses glucose from tissue fluid. In this design the problems of enzyme instability and oxygen limitation might be circumvented. The sensor measures glucose reliably for over two weeks, provided a new 10 ml syringe containing a glucose oxidase solution is connected to the system each day.