Biotransformation enzymes and thyroid axis disruption in juvenile rainbow trout (Oncorhynchus mykiss) exposed to hexabromocyclododecane diastereoisomers.

Juvenile rainbow trout (Oncorhynchus mykiss) were fed either a reference diet or one of three diets enriched with alpha, beta, or gamma diastereoisomers of hexabromocyclododecane (HBCD, C12H18Br6) for 56 days. This exposure period was followed by 112 days during which all fish were fed the reference diet. Potential effects of HBCD on phase I and II biotransformation enzyme activities and thyroid axis disruption were examined. Disruption of the thyroid axis was most evident in the gamma-HBCD exposed group, as indicated by lower circular FT4 and higher FT3 as well as an increase in thyroid epithelial cell height. However, fish fed the alpha-HBCD enriched diet also exhibited altered glucuronyltransferase activity and thyroid epithelial cell heights and the beta-HBCD group had altered FT4 and FT3 and glucuronyltransferase activity. T4ORD activity was not affected after 14 days, but was significantly lower among all HBCD exposed fish compared to the reference fish after 56 days. Results from these experiments indicate that all isomers have the potential to disrupt thyroid homeostasis.

Dietary exposure of juvenile rainbow trout (Oncorhynchus mykiss) to 1,2-bis(2,4,6-tribromophenoxy)ethane: bioaccumulation parameters, biochemical effects, and metabolism.

Juvenile rainbow trout (Oncorhynchus mykiss) were exposed in the laboratory to an environmentally relevant dose of 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE) via their diet for 49 days, followed by 154 days of untreated food to examine bioaccumulation parameters, potential biochemical effects, and metabolic products. There was a linear increase in the amount of BTBPE in fish during the uptake phase of the experiment, and an uptake rate constant of 0.0069 +/- 0.0012 (arithmetic mean +/- 1 x standard error) nmoles per day was calculated. The elimination of BTBPE from the fish obeyed first-order depuration kinetics (r2 = 0.6427, p < 0.001) with a calculated half-life of 54.1 +/- 8.5 days. The derived biomagnification factor of 2.3 +/- 0.9 suggests that this chemical has a high potential for biomagnification in aquatic food webs. Debrominated and hydroxylated metabolites were not detected in liver extracts and suggest that either biotransformation or storage of BTBPE-metabolites in the hepatic system of fish is minor or that our exposure time frame was too short. Similar concentrations of circulating thyroid hormones, liver deiodinase enzyme activity, and thyroid glandular histology suggest that BTBPE is not a potent thyroid axis disruptor.

Bioaccumulation and trophic transfer of some brominated flame retardants in a Lake Winnipeg (Canada) food web.

The extent of bioaccumulation and trophic transfer of brominated diphenyl ether (BDE) congeners, hexabromocyclododecane (HBCD) diastereoisomers (alpha, beta, and gamma), decabromodiphenylethane (DBDPE), and bis(2,4,6-tribromophenoxy)ethane (BTBPE) was examined in a Lake Winnipeg (Canada) food web. Six species of fish, zooplankton, mussels, sediment, and water from the south basin of the lake were selected for study. Significant positive correlations were found between concentrations of total (sigma) polybrominated diphenylethers (PBDEs; p < 0.005), sigmaHBCDs (p < 0.0001), BTBPE (p < 0.0001), and lipid content in fish. Strong positive linear relationships also were observed from individual plots of BDE 47, BDE 209, and DBDPE concentrations (lipid wt) and trophic level (based on delta15N), suggesting that these compounds biomagnify in the Lake Winnipeg food web. Biomagnification factors varied for the chemicals studied. Plots of log bioaccumulation factors for mussel and zooplankton versus log octanol-water partition coefficient (Kow) were similar and suggest that neither mussels nor zooplankton are in equilibrium with the water. Fifteen BDE congeners were consistently detected in water (dissolved phase, n = 3), with BDE 47 having the greatest concentration (17 pg/L). The rank order of compounds in water (arithmetic mean +/- standard error) were sigmaPBDEs (49 +/- 12 pg/ L) > alpha-HBCD (11 +/- 2 pg/L) > BTBPE (1.9 +/- 0.6 pg/L). Concentrations of DPDPE, BDE 209, and beta- and -gamma-HBCD isomers were below their respective method detection limits (MDLs) in water. Total PBDE concentrations in sediment (n = 4) were greater than any other brominated flame retardant examined in the present study and ranged from 1,160 to 1,610 ng/g (dry wt), with BDE 209 contributing roughly 50% of the total. The gamma-HBCD isomer was detected at concentrations of 50 +/- 20 pg/g (dry wt) in sediment, whereas BTBPE and DBDPE were consistently below their respective MDLs in sediment.

Dietary accumulation of hexabromocyclododecane diastereoisomers in juvenile rainbow trout (Oncorhynchus mykiss) I: bioaccumulation parameters and evidence of bioisomerization.

Juvenile rainbow trout (Oncorhynchus mykiss) were exposed to three diastereoisomers (alpha, beta, gamma) of hexabromocyclododecane (C12H18Br6) via their diet for 56 d followed by 112 d of untreated food to examine bioaccumulation parameters and test the hypothesis of in vivo bioisomerization. Four groups of 70 fish were used in the study. Three groups were exposed to food fortified with known concentrations of an individual diastereoisomer, while a fourth group were fed unfortified food. Bioaccumulation of the gamma-diastereoisomer was linear during the uptake phase, while the alpha- and beta-diastereoisomers were found to increase exponentially with respective doubling times of 8.2 and 17.1 d. Both the beta- and the gamma-diastereoisomers followed a first-order depuration kinetics with calculated half-lives of 157 +/- 71 and 144 +/- 60 d (+/-1 x standard error), respectively. The biomagnification factor (BMF) for the alpha-diastereoisomer (BMF = 9.2) was two times greater than the beta-diastereoisomer (BMF = 4.3); the large BMF for the beta-diastereoisomer is consistent with this diastereoisomer dominating higher-trophic-level organisms. Although the BMF of the beta-diastereoisomer suggests that it will biomagnify, it is rarely detected in environmental samples because it is present in small quantities in commercial mixtures. Results from these studies also provide evidence of bioisomerization of the beta- and gamma-diastereoisomers. Most importantly, the alpha-diastereoisomer that was recalcitrant to bioisomerization by juvenile rainbow trout in this study and known to be the dominant diastereosiomer in fish was bioformed from both the beta- and the gamma-diastereoisomers. To our knowledge, this is the first report of bioisomerization of a halogenated organic pollutant in biota.

Refinements to the diastereoisomer-specific method for the analysis of hexabromocyclododecane.

The emergence of hexabromocyclododecane (HBCD) as a bromine-based flame retardant of concern is partly attributable to recent measurements on the environmental occurrence of the individual diastereoisomers (alpha, beta and gamma). These measurements were fuelled by a newly developed liquid chromatography/tandem mass spectrometric (LC/MS/MS)-based analytical method. However, in the course of our recent studies on the environmental fate and behaviour of the diastereoisomers of HBCD, some interesting features of the LC/MS/MS method became apparent. For example, the ion signal of the native ions was found to be dependent on the final extract volume. This was true for both biotic and sediment samples and was found to arise from the suppression of the ion signal due to endogenous material in the extracts that escape clean-up. We have also found differences in the stability of the diastereoisomers in different solvents. If left unaccounted for, both factors can compromise analytical measurement data. By way of a series of controlled experiments conducted at our two laboratories [Department of Fisheries & Oceans Canada (DFO) and Environment Canada (EC)], we illustrate these features and demonstrate that use of newly synthesized labelled HBCD isomers [(13-carbon (13C) and deuterium (d18)] can minimize and often circumvent matrix-related effects.

Bioaccumulation, biotransformation, and biochemical effects of brominated diphenyl ethers in juvenile lake trout (Salvelinus namaycush).

Juvenile lake trout (Salvelinus namaycush) were exposed to three dietary concentrations (0, approximately 2.5, and approximately 25 ng/g per BDE congener) of 13 BDE congeners (3-10 Br atoms) in the laboratory for 56 days, followed by 112 days of clean food, to examine bioaccumulation parameters and potential biochemical effects. The bioaccumulation of BDEs by the trout was highly influenced by biotransformation, via debromination, which resulted in bioaccumulation parameters that were much different than would be expected based on studies of chlorinated organic compounds (e.g., PCBs). Half-lives (t1/2's) for some BDE congeners (e.g., BDE-85 and -190) were much lower than expected based on their Kow, which was likely due to biotransformation, whereas t1/2's of other BDE congeners (e.g., BDE-66, -77, -153, and -154) were much longer than anticipated based on Kow. This was likely because the metabolites of BDE formed via debromination had the same chemical structure of these BDE congeners, which supplemented measured concentrations. The detection of three BDE congeners (an unknown penta, BDE-140, and an unknown hexa) in the fish that were not present in the food or in the control fish provide further evidence forthe debromination of BDEs. Half-lives of BDEs ranged from 38 +/- 9 to 346 +/- 173 days and biomagnification factors ranged from 1.6 (BDE-190) to 45.9 (BDE-100), but these bioaccumulation parameters need to be viewed with caution because they were highly influenced by debromination and relative abundance of individual BDEs that the fish were exposed to. CYP1A enzyme activity, measured as EROD, and free tri-iodothyronine (T3) concentrations in the plasma of lake trout varied significantly throughout the experiment but were not related to BDE exposure. In contrast, plasma levels of thyroxine levels (T4) were lower in both groups of PBDE-exposed fish compared with control fish after 56 days of exposure, and after 168 days in the high dose, suggesting that PBDEs may influence thyroid homeostasis at levels that are higher than what is normally found in the environment.