- Email: [email protected]
Gastrointestinal absorption, dynamic tissue-specific accumulation, and isomer composition of dechlorane plus and related analogs in common carp by dietary exposure
Ecotoxicology and Environmental Safety 100 (2014) 32–38
Contents lists available at ScienceDirect
Ecotoxicology and Environmental Safety journal homepage: www.elsevier.com/locate/ecoenv
Gastrointestinal absorption, dynamic tissue-specific accumulation, and isomer composition of dechlorane plus and related analogs in common carp by dietary exposure Yan-Hong Zeng a,b, Xiao-Jun Luo a,n, Bin Tang a,b, Xiao-Bo Zheng a,b, Bi-Xian Mai a a b
State Key Laboratory of Organic Geochemistry, Guangzhou Institute of Geochemistry, Chinese Academy of Sciences, Guangzhou 510640, PR China University of Chinese Academy of Sciences, Beijing 100049, PR China
art ic l e i nf o
a b s t r a c t
Article history: Received 1 August 2013 Received in revised form 22 October 2013 Accepted 25 November 2013 Available online 20 December 2013
Dechlorane plus (DP) is a high-productive volume substance which had been identified as ubiquitous pollutant and has been widely detected in biota. In the present study, common carp (Cyprinus carpio) was exposed to known amounts of commercial DP-25 under laboratory conditions. The gastrointestinal absorption and tissue-specific bioaccumulation of DP and its dechlorinated analogs in common carp were investigated. The higher absorption efficiencies but lower assimilation efficiencies of anti-isomers indicated stereoselective metabolism of anti-isomers in fish. Linear uptake curves were seen in serum and muscle, but the depuration curves for all the four tissues (muscle, serum, liver and gonad) did not follow the first-order kinetics. The liver exhibited a high affinity for anti-isomers during the experiment. Other tissues, such as serum, muscle, and gonad, showed a selective accumulation of syn-DP in the early stages of the experiment, particularly the serum. However, the deviation of fanti between different tissues disappeared at late stages of the experiment, and the fanti values in all tissues were close to that in commercial mixtures. Our results suggest that the bioaccumulation of DP is a complex and multifactorial process. & 2013 Elsevier Inc. All rights reserved.
Keywords: DP Gastrointestinal absorption Tissue-specific accumulation Isomer composition
1. Introduction Dechlorane plus (DP) was introduced as a substitute for Dechlorane (an additive chlorinated flame retardant) by Hooker Chemical in the 1960s (Hoh et al., 2006). It has been used in wire and circuit boards, paint, and hard connectors in computers (Tomy et al., 2007). The commercial mixture consists primarily of 2 isomers (syn- and anti-DP) with a minor content of 2 dechlorinated isomers (syn- and anti-Cl11-DP) (Zheng et al., 2010). DP has been classified as a high-productive volume substance by the United States Environmental Protection Agency (Sverko et al., 2011). Through its long commercial history, it has not received attention until recently. DP was first identified in ambient air, fish, and sediment samples from the Great Lakes region in 2006 (Hoh et al., 2006). DP has since been measured in sediment (Qi et al., 2010; Qiu et al., 2007), surface soil (Yu et al., 2010), indoor dust (Zhu et al., 2007), biota (Gauthier and Letcher, 2009; Tomy et al., 2007), and even in air and seawater from the Arctic to Antarctica (Möller et al., 2010). Recent studies have also reported detection of DP in humans (Ben et al., 2013; Yan et al., 2012).
n
Corresponding author. Fax: +86 20 85290706. E-mail address: [email protected] (X.-J. Luo).
0147-6513/$ - see front matter & 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.ecoenv.2013.11.021
The detection of DP in biota suggests that the isomers are bioavailable and bioaccumulative. As reported in previous studies, the bioaccumulation tendencies of the isomers vary. Tomy et al., 2007 reported that the anti-isomer was dominant at higher trophic levels while the syn-isomer dominated in lower trophic level organisms in food web samples from Lake Winnipeg and Lake Ontario. Wu et al., 2010 also found that the fraction of anti-DP decreased upon moving up the trophic levels of aquatic organisms in a reservoir near an electronic waste recycling workshop in South China. Different extents of enrichment (anti-) and depletion (syn-) of the isomers were found in Lake Winnipeg biota by Tomy and colleagues (Tomy et al., 2007). The complexity of environmental DP isomer compositions often makes the fate studies of DP highly confusing. For example, different ratios of anti-DP to total DP (fanti) of a biota may be due to the accumulation of different DP isomer compositions from the environment (Xian et al., 2011) and perhaps due to isomer-specific bioavailability of DP in the biota. In Previous studies, accumulation of DPs in juvenile rainbow trout (Tomy et al., 2008), rat (Li et al., 2013a), and Quail (Li et al., 2013b) has been investigated in laboratory experiment. Different accumulation potential for anti- and syn-isomer was observed in rainbow trout, rat and quail. However, there is little understanding of the bioaccumulation differentiation of the isomers. Laboratory exposure experiments in controlled conditions are convenient
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
methods for investigating the bioaccumulation processes of contaminants. Therefore, the main objective of the present study was to improve the understanding of the bioaccumulation process of DP. To the best of our knowledge, only a single laboratory study has been conducted to explore this issue in fish (Tomy et al., 2008), but that study used juvenile rainbow trout (Oncorhynchus mykiss) exposed to syn- and anti-DP separately and no dechlorinated analogs were spiked in the food. Moreover, gastrointestinal absorption was not taken into consideration. The gastrointestinal tract is a critical segment which influences and limits the compound absorption as most compounds in fish are believed to be absorbed by the intestine. However, there is no information concerning about the gastrointestinal absorption of DP in fish. In the present study, common carp (Cyprinus carpio) was exposed to contaminated food pellets for 50 days. The contaminated food was made by known amounts of commercial DP-25 dissolved in cod liver and then mixed with fish food pellets. After the exposure period, a depuration period lasting a further 40 days was designed for the carp, during which the carp were fed with non-spiked food. The purposes of the present study were (1) to investigate gastrointestinal absorption by comparing the concentration and congener profile of DP between spiked food and feces and (2) to investigate the uptake and depuration behaviors of the DP congeners in different carp tissues after exposure to high doses of DP-25 mixtures.
2. Materials and methods 2.1. Fish exposure Common carp is a kind of freshwater and omnivorous fish. The carp reach sexual maturity in 1 to 3 years and lay eggs. In March, 2013, forty common carps, approximately 12 cm in length, were purchased from an aquarium market in Guangzhou, China. Four carps were randomly collected as the control group and kept in one glass tank (80 cm 35 cm 50 cm). The remaining carp (n¼ 36) were kept in another glass tank (120 cm 40 cm 60 cm) as the treated group. Air stones were placed in the tank to maintain oxygen saturation in the water. The water in each tank was heated to a constant temperature of 22 7 1 1C and circulated using a submersible pump at a rate of 1.5 L/min. Fish were acclimated to the non-spiked diet for a period of 7 days prior to exposure. Fish were fed food at a rate of 1 percent of their body weight/day based on their weight. Commercial DP-25 (Z 99 percent purity, particle diameter r 6 μm, Chlorine content of 65.1 percent and density of 1.8 g/cc) was acquired from Jiangsu Anpon Co., Ltd and cod liver oil was obtained from Peter Moller (Norway). DP-25 was delivered in artificially contaminated food. The DP dose of fish exposed was approximately 1.5 mg per day for each fish. DP-25 mixtures were first dissolved in isooctane and then diluted to the desired concentration with cod liver oil. Then, 1 mL of cod liver oil was mixed with 10 g of fish food pellets (Yangzhou Five-Star Aquatic Products Co., LTD, PR China). Non-spiked food was prepared by mixing food with solute-free cod liver oil. Food was homogenized by mixing in a shaking incubator (24 h, 25 1C). The spiked and non-spiked food was stored in amber containers in a cool, dark place throughout their use. The treated group (36 common carps) was exposed to spiked food for 50 days, followed by non-spiked food for 40 days. The control group was fed a non-spiked diet throughout the experiment. The food was collected on days 5 and 45 (spiked food in uptake period) and on days 55 and 90 (non-spiked food in depuration period). Feces were collected on days 5, 10, 15, 25, 35, 40, and 50 (uptake period), and on days 60, 70, 80, 85, and 90 (depuration period). During the entire experimental period (90 days), 2 fish were sampled from the tank every 5 days and were carefully dissected, resulting in 36 fish samples (internal tissues and gills were removed). The blood, liver, and gonad of fish sampled on the same day were pooled into 1 sample. Blood samples were drawn from the dorsal aorta by using syringes, transferred into 5-mL Teflon tubes, and centrifuged at 3000 rpm for 15 min to obtain serum. The fish carcasses (n ¼36) and pooled serum (n¼18), liver (n¼ 18), and gonad samples (n¼18) were obtained for analysis of DP (anti-DP and syn-DP) and dechlorinated products (anti-Cl10-DP, anti-Cl11-DP, and syn-Cl11-DP).
2.2. Sample preparation The extraction and analysis procedures used for the fish carcass (approximately 2 g), liver, and gonad were similar to those described in previous studies (Hoh et al., 2006). Briefly, freeze-dried samples were ground into powder and spiked with surrogate standards (BDE 181) prior to Soxhlet extraction with 50 percent acetone in
33
hexane for 24 h. The extract was concentrated and then solvent exchanged to hexane (10 mL). The lipid content was determined by gravimetric measurement from an aliquot of extract. The remaining extract was treated with 3 mL of concentrated sulfuric acid (H2SO4) (analytical reagent grade, Guangzhou Chemical Reagent Factory, China) twice to remove any lipids present in the samples. After centrifugation for 15 min, the hexane layer was recovered and combined with 6 mL of hexane to wash the pooled H2SO4 residue. The lipid-free extract was washed with sodium sulfate (5 percent, w/w) to remove the residual H2SO4 and concentrated to 1 mL, further cleaned up on a multilayer column (i.d.¼ 1 cm) packed with 8-cm neutral silica, 8-cm acidified silica, and a 2-cm layer of anhydrous sodium sulfate. The DP isomers and their dechlorinated congeners were obtained by eluting with 30 mL hexane/ dichloromethane (v/v¼ 1:1). After solvent exchange to isooctane (1 mL), internal standards (BDE 128) were added before gas chromatography/mass spectrometry analysis (GC/MS). A sample preparation method similar to the fish tissue preparation method was used for the food and feces samples. After Soxhlet extraction, the extract was concentrated, subjected to solvent exchange with hexane (10 mL), treated with H2SO4, and cleaned as described above. Serum pretreatment was performed as previously described with a minor modification (Malmberg et al., 2005). Briefly, after spiking with the surrogate standards (BDE 181), approximately 0.5 g serum was denatured using hydrochloric acid (6 N, 1 mL) and 2-propanol (6 mL), and the mixtures were shaken vigorously. Mixtures were then extracted 3 times with methyl tert-butyl ether: hexane (1:1, v/v, 6 mL). The combined organic extracts were washed with a potassium chloride solution (1 percent, w/w, 4 mL) and blown to near dryness under N2 for gravimetric lipid weight determination. The extracts were redissolved with 9 mL of hexane, then treated with H2SO4, and cleaned as described above. 2.3. Instrumental analysis DP and its dechlorinated congeners were analyzed by GC/MS (Agilent 7890A/5975B MSD; Agilent Technology, CA) with an electron capture negative ionization source in a selective ion-monitoring mode and separation performed using a DB-XLB capillary column, 30 m 0.25 mm ID 0.25-mm film (J&W Scientific). The carrier gas was helium with a constant column flow rate of 1.3 mL/min. The column temperature program was as follows: maintained at 110 1C for 1 min, heated to 200 1C at a 1st rate of 8 1C/min and then maintained at 200 1C for 1 min, heated to 240 1C at a 2nd rate of 3 1C/min and maintained at 240 1C for 2 min, heated to 280 1C at a 3rd rate of 5 1C/min and maintained at 280 1C for 10 min, and finally heated to 310 1C at a 4th rate of 10 1C/min and maintained at 310 1C for 10 min. We automatically injected 1 mL of sample into the pulsed splitless mode. The temperature of the injection port was set at 290 1C. Ion fragments m/z 653.8 and 651.8 for DP isomers (syn-DP and anti-DP), m/z 618 and 620 for Cl11-DP isomers (anti-Cl11-DP and syn-Cl11-DP), m/z 584 and 586 for antiCl10-DP, and m/z 79 and 81 for BDE 181 and BDE 128 were monitored. syn-Cl11-DP was previously identified as a dechlorinated product of syn-DP (Wang et al., 2011). Semiquantitation was achieved for syn-Cl11-DP in reference to the response of anti-Cl11-DP on GC/MS. The compounds were quantified using the internal calibration method based on a 9-point calibration curve.
2.4. Data analysis The net assimilation efficiency of each DP compound was calculated using the following equation: α ¼ ðbody burden in common carp at time tÞ=ðcumulative mass exposed to common carp up to time tÞ
where α is net assimilation efficiency. The net assimilation efficiency calculations mostly involve the determination of ingestion minus egestion and excretion. In the present study we have explored net assimilation efficiency given that our target was primarily to examine the differences in assimilation and depuration between syn-DP congeners and anti-DP congeners. All statistical analyses were conducted using PASW Statistics 18.0 to test for the inter-tissues variability of DP levels and isomer compositions. DP concentrations determined in fish tissues were not normalized to total lipid content, due to higher variations of these concentrations when normalized to lipid content.
3. Results and discussion 3.1. Background levels and quality control After the 90-d experiment, the length of common carp was 12.4 7 0.8 cm, which did not change significantly compared to days 1 (12.1 70.6 cm) (p o0.05). No mortality was observed during the experiment. Procedural blanks covering the whole procedure were performed in parallel with the samples at each batch extraction. Anti-DP was detected in the blanks, but syn-DP,
34
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
anti-Cl10-DP, syn-Cl11-DP, and anti-Cl11-DP were not detected in the blanks. The method detection limit (MDL) was defined as the mean value plus 3-fold standard deviation for anti-DP detected in the procedural blanks (n¼ 7). For syn-DP, anti-Cl10-DP, syn-Cl11-DP, and anti-Cl11-DP, which were not detected in blanks, a signal-to-noise ratio of 10 was set as the MDL. The MDLs for syn-DP, anti-DP, antiCl10-DP, syn-Cl11-DP, and anti-Cl11-DP were between 0.02 and 0.11 ng/g wet weight (ww) in fish tissues, and between 0.18 and 0.47 ng/g dry weight (dw) in food and feces. The recoveries of BDE 181 for all samples (n ¼112) ranged from 80 percent to 128 percent. During the 90-day experiment, no anti-Cl10-DP, syn-Cl11DP, or anti-Cl11-DP were detected in the control fish tissues or the non-spiked food. Both anti-DP and syn-DP were detected in control fish tissues and the non-spiked food. The average concentrations of syn-DP and anti-DP in control fish tissues were 197 12 pg/g and 34 726 pg/g (ww) and 0.52 70.38 ng/g and 1.5 71.1 ng/g (dw) in non-spiked food, respectively. These levels were several orders of magnitude lower than those of the dosed fish and the spiked food. 3.2. Gastrointestinal absorption and fecal excretion of DP congeners Syn-DP, anti-DP, syn-Cl11-DP, and anti-Cl11-DP were detected in 100 percent of the food and fecal samples while no anti-Cl10-DP was detected. The average concentrations of syn-DP, anti-DP, synCl11-DP, and anti-Cl11-DP in the spiked food were 2071 724, 7826 71031, 4.0 70.3, and 33.570.4 ng/g (dw), respectively. The concentrations of DP congeners in feces collected in the uptake period were in the range of 1.5–8.2-fold compared to that in the administered food. A ratio of the chemical concentration in the feces to that in the food is directly related to the chemical absorption and excretion in the gut if the chemicals are not metabolized by endogenous enzymes in the gastrointestinal system. The higher the ratio, the lower the absorption efficiency or the higher excretion efficiency is. The concentration ratios of feces to food in the uptake period were in the range of 1.55–4.98 for synDP, 1.53–4.31 for anti-DP, 1.86–8.26 for syn-Cl11-DP and 1.53–6.31 for anti-Cl11-DP. The ratios of feces to food for syn-DP were slightly higher than those for anti-DP in each sampling in the uptake period except for the 50-day sampling (p o0.05). Similar results were also seen between syn-Cl11-DP and anti-Cl11-DP (po 0.05). These phenomena imply that the absorption efficiencies of antiisomers in the gastrointestinal system were higher than those of the syn-isomers. In the study of Tomy et al., 2008, accumulation rate of syn-DP was found to be faster than that of anti-DP in juvenile rainbow trout. Obviously, the difference in gastrointestinal absorption rate between the two isomers cannot be responsible for stereoselective accumulation of syn-DP in fish given that anti-DP rather than syn-DP were absorbed more efficiently through the gut. The concentrations of four chemicals in the feces collected in the 60-day (the first feces sample for the depuration period) sharply decreased by 1 order of magnitude compared to the feces in the uptake period. An exponential decrease trend of concentration of target chemicals over the depuration period was observed (Fig. 1b). To evaluate the differences in excretion rate between synDP and anti-DP, a ratio of feces to food (non-spiked food) was calculated (Fig. 1b). The calculated result indicated that there were no systematic differences in the ratio of feces to food between synDP and anti-DP (p o0.05), indicating no stereoselective excretion for DP in common carp. The congener profile of DP can be expressed by the fractions of the anti-isomer (fanti: ratio of anti-DP concentration to total concentration of anti-DP and syn-DP; fanti-Cl11: ratio of anti-Cl11DP concentration to total concentration of anti-Cl11-DP and synCl11-DP). fanti in the spiked food was 0.791 70.02, which was
slightly higher than those in feces collected in the uptake period (range: 0.765–0.787; mean, 0.775) except for at 50 days (0.850). The fanti-Cl11 in the spiked food was 0.893, which was also higher than that in feces in the uptake period (range: 0.853–0.873; mean, 0.866) except for at 50 days (0.920) (Fig. S1). These results further confirm that syn-isomers have lower absorption efficiencies or higher elimination efficiencies than anti-isomers in the gut. fanti and fanti-Cl11 were significantly lower in feces in the depuration period than in feces in the uptake period and in the spiked food (p o0.05) (Fig. S1), indicating anti-isomers have been selectively metabolized in common carps. 3.3. Uptake and depuration kinetics of DP in fish tissues The uptake and depuration kinetics of DP in the serum, muscle, liver, and gonad were investigated. The four chemicals (anti-DP, syn-DP, anti-Cl11-DP and syn-Cl11-DP), except for syn-Cl11-DP in the serum, were detected in all target tissues. All chemicals exhibited linear accumulation kinetics in serum (r2 ¼0.88, r2 ¼0.84 and r2 ¼ 0.69 for anti-DP, syn-DP and anti-Cl11-DP, respectively) and muscle (r2 ¼ 0.84, r2 ¼0.81, r2 ¼0.84 and r2 ¼ 0.81 for anti-DP, synDP anti-Cl11-DP and syn-Cl11-DP, respectively) samples during the uptake period (Fig. 2a and b) and reached the highest concentrations at the end of the uptake period (50th day). No steady state was observed in the serum or muscle for any chemical during the 50-day exposure. Depuration of the chemicals in both muscle and serum showed an initial rapid depuration for the first 10 days followed by a fluctuating depuration over the remainder of the experiment. No obvious trend towards a decrease was found in the last 30 days of the depuration. There are no previous studies regarding DP uptake and clearance curves in the fish serum, but the DP uptake and clearance curves in the muscle observed in the present study were similar to results reported by other researchers during bioaccumulation studies of hydroponic contaminants in fish (Fisk et al., 1998; Tomy et al., 2004). The uptake kinetics of DP and their dechlorinated analogs in the liver and gonad were not linear (Fig. 2c and d). The concentrations in the gonad showed co-variation with those in the liver with the 5-day hysteresis during the uptake period. This hysteresis in the gonad might be attributed to maternal transfer of DP in fish. The depuration curves of DP in the liver showed 2-stage elimination kinetics, with an initial decrease to day 70, followed by a slight increase. The concentrations of DP and their analogs fluctuate and follow a zigzag pattern in the depuration. This unique accumulation pattern of DP in the gonad of carp could be because gonads collected in different individuals may be in different stages of embryonic development as some of the DP was determined in fish ovaries and some was determined in fish eggs. It remains unclear as to why no linear uptake occurred in the liver in the uptake period and accumulation in the liver was still occurring at the end of the experiment. Enterohepatic circulation and redistribution of lipophilic contaminants can affect liver concentrations and ultimately give rise to complex uptake and depuration kinetics (Roberts et al., 2002). Additionally, the potential metabolism of DP and analogs in the liver and the high variation in the concentrations between individual fish could also contribute to the complex uptake and depuration kinetics. Moreover, during the experimental period, the undulating tissue concentrations of DP isomers and Cl11-DP isomers could be partly due to the individual variation. Meanwhile, based on the research of Tomy et al., 2008, which reported half-lives of DP in the 40 to 50 day length, so the depuration phase (40 d) of the present experiment was not long enough. This is probably a major contributor to the inability for us to find a significant elimination rate. The mean assimilation efficiencies of syn-DP, anti-DP, syn-C11DP, and anti-C11-DP calculated from days 5, 10, 15, 20, 25, 30, 35,
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
35
Conc. of DP isomers (ng/g dry weight)
Conc. ratios in feces to spiked food
Syn-DP Anti-DP Syn-Cl11-DP
8 7
Anti-Cl11-DP
6 5 4 3 2 1 0
5
10
15
20
25
30
35
40
45
50
55
Time (days)
700
700
Ratio for syn-DP Ratio for anti-DP
600
600
500
500
400
400
300
300
200
200
100
100
Syn-DP concentration Anti-DP concentration
0
55
60
65
70
0 75
80
85
90
Conc. ratios of feces to non-spiked food
Depuration period
Uptake period 9
95
Time (days)
Fig. 1. (a) Concentration ratios of DP congeners and dechlorinated congeners in feces to spiked food during the uptake period of common carp; (b) concentrations of DP congeners and the concentration ratios of DP congeners in feces to non-spiked food during the depuration period of common carp. The solid line indicates an exponential decrease trend of the concentrations of DP isomers in the feces along with depurated time.
40, 45, and 50 were estimated to be 3.8 percent, 3.2 percent, 20.4 percent, and 14.8 percent, respectively. These assimilation efficiencies of DP isomers were comparable to those (6.0 percent for syn-DP and 3.9 percent for anti-DP) determined in juvenile rainbow trout (Tomy et al., 2008). Meanwhile, the assimilation efficiencies of syn-isomers were significantly higher than those of anti-isomers (p o0.05). However, the ratios of feces to food indicate that the uptake efficiencies of syn-isomers were lower than those of the anti-isomers. Therefore, the relatively low assimilation efficiencies of anti-isomers could be attributed to stereoselective metabolism of anti-isomers in carp. This is also consistent with the structural conformation of the isomers as it has been suggested that because of the configuration of the pendant chlorocyclopentane moieties, the anti-isomer would be more susceptible to biological attack (Tomy et al., 2008). 3.4. Dynamic tissue distribution and isomer composition The concentrations of DP and the analogs in the liver were higher than those in the muscle, serum, and gonad throughout the experiment (po0.1) (Fig. 3). Lipid content is not a cause for this tissue distribution because the lipid contents in the liver (1.7071.56 percent) were significantly lower than those in the muscle (3.3971.69 percent) and the gonad (3.7171.96 percent). This result indicates that DP and analogs prefer to accumulate in the liver than in other tissues, which is consistent with that reported by Li et al., 2013a and is also in line with reports on other highly halogenated flame retardants (Guvenius et al., 2002; Iwata et al., 2004). In Chinese sturgeon, liver also showed higher dechlorane concentrations than muscle and adipose tissue (Peng et al., 2012). Sequestration of DP isomers by hepatic proteins could be a cause for their disposition in the liver although this was not identified (Li et al., 2013b). Exposure to xenobiotic pollutants such as polychlorinated dibenzo-p-diosins, polychlorinated dibenzo-furans, and coplanar PCBs may induce hepatic binding proteins leading to hepatic sequestration of the compounds (Kubota et al., 2004). A dynamic tissue distribution for DP and its analogs among tissues was observed. At the start of exposure, the concentration ratios of the liver to other tissues were the highest, and then the ratio exponentially decreased over time to close to or slightly lower than 1 at the end of the experiment (Fig. 3). This result indicated that the liver is the first organ to which contaminants deposit after absorption from the gastrointestinal tract in fish. This result was consistent with a study on the tissue distribution of
PBDEs in 2 predatory fish species (Zeng et al., 2013). In that study, the liver was found to readily achieve equilibrium with the serum while the muscle did not until the end of the experiment, using a tissue fugacity comparison. The muscle is the main organ for contaminant deposition. The percentage of DP and analogs in the muscle was greater than 60 percent during the entire experimental period (Fig. S2). This is because the muscle is the largest tissue in volume in fish, despite its low concentration. The percentage in the muscle increased over time and reached the maximum at the 75th day (95 percent), and then a decreasing trend was seen until the end of the experiment. A concomitant decrease in the liver from 23 percent at the 5th day to 3 percent at the 75th day was observed. The gradual accumulation of contaminants occurring in the muscle was the main reason for the above observation. The decreased proportion in the muscle along with a corresponding increase in the liver in the last 15 days of the experiment implies a redistribution of DP and analogs among tissues. A non-regular trend was found for the gonadal proportion during the whole experiment, and this may be due to the variations in the number of gonads between individual fish. DP and analogs in the gonad accounted for 2 percent to 20 percent of the total amount in fish with most cases being greater than 5 percent (Fig. S2). Meanwhile, the concentration of DP and analogs in the gonad was higher than that in the muscle (p o0.05), which is consistent with the report of Peng et al. (2012), where the ratio of the chemical concentration in the eggs to those in the maternal tissues was greater than 1. These results indicate that DP and analogs readily underwent maternal transfer. Considering the fact that a high content of the pollutant usually exhibits a significant development toxicity to naturally hatched larvae (Abdelouahab et al., 2009), such a high accumulation of DP and Cl11-DP in the gonad suggests that the maternal transfer of DP congeners and the corresponding adverse effects for larvae should be elucidated further. The isomer composition of DP and analogs also showed tissuespecific and dynamic changes over time (Fig. 4). In the early stage of the uptake experiment (from day 0 to the 20th day), fanti and fanti-Cl11 values in tissues were lower than those in the commercial DP mixtures, indicating a preference for syn-isomers. fanti and fantiCl11 values then increased over time until they were close to those in commercial DP mixtures, especially fanti in the serum. Serum accumulated more syn-DP than anti-DP compared with other tissues while the liver showed a preference for anti-DP to syn-DP
36
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
Depuration
Uptake
Uptake
300 250
Depuration
80
Serum
70
Muscle
200 150
y = -18.77+4.85x r2 = 0.88; p < 0.001
Concentration of congener (ng/g ww)
Concentration of congener (ng/g ww)
60
100 50 y = -1.32+1.48x r2 = 0.84; p < 0.001
3.0 2.5 2.0 1.5 1.0
y = 0.05+0.022x r2 = 0.69; p = 0.002
0.0
y = -6.34+1.30x r2 = 0.84; p < 0.001
30 20 10
1.0 0.8
y = -1.80+0.37x 2 r = 0.81; p < 0.001 y = 0.026+0.018x 2 r = 0.84; p < 0.001
0.6 0.4 y = 0.012+0.013x 2 r = 0.81; p < 0.001
0.0 0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95
0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95
Time (days)
Time (days)
600
Liver
Gonad
500
300 200 100
syn-DP anti-DP syn-Cl11-DP
5 4
anti-Cl11-DP
3 2
Concentration of congener (ng/g ww)
400
Concentration of congener (ng/g ww)
40
0.2
0.5
500
50
400 300 200 100 0 8 7 6 5 4 3 2 1
1
0
0
-1 0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95
0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95
Time (days)
Time (days)
Fig. 2. Uptake and depuration curves of syn-DP, anti-DP, syn-Cl11-DP, and anti-Cl11-DP throughout the experimental period in (a) serum, (b) muscle, (c) liver, and (d) gonad of common carp.
compared with other tissues in the early stage of the experiment. The fact that the liver preferred anti-DP to syn-DP was in line with the results of a rat exposure experiment (Li et al., 2013a). The greatest deviation of fanti between the serum and liver reached 0.09 (0.70 vs. 0.79). However, in the late stage of the experiment, the larger deviation of fanti between different tissues disappeared and no significant differences were found between different tissues (p4 0.1). These results indicate that the isomer-specific accumulation of DP is a very complex and multi-factorial process.
In the current state, it was not possible to provide a detailed mechanism for this accumulation. Using rats exposed to different doses of DP, Li et al., 2013a found that no stereoselective accumulation was seen in the low dose exposure group, whereas a selective accumulation of syn-DP was observed in the high dose exposure group. This result indicates that the fanti value in rat was dose-dependent. Tissuespecific fanti has been reported in several previous studies. Zhang et al., 2011 reported that the brain of mud carp has a high affinity for
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
37
35
30
30
25
25 20
Concentration ratios of liver to other tissues
20 15
15
10
10
5
5
0
0
40
30
35
25
30 20
25 20
15
15
Liver/Muscle Liver/Serum Liver/Gonad
10
10 5
5
0
0 0
10
20
30
40
50
60
70
80
90
0
10
20
30
40
50
60
70
80
90
Time (days)
fanti (anti-DP/(anti-DP+sym-DP))
Muscle
Liver
Serum
Gonad
0.80
0.75
0.70
0.65
fanti-Cl11(anti-Cl11-DP/(anti-Cl11-DP + syn-Cl11-DP))
Fig. 3. Concentration ratios of (a) syn-DP, (b) anti-DP, (c) anti-Cl11-DP, (d) syn-Cl11-DP in the liver compared to other tissues of common carp throughout the experimental period.
Muscle
Liver
Gonad
0.90
0.85
0.80
0.75 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90
5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90
Time (days)
Time (days)
Fig. 4. fanti and fanti-Cl11 values calculated in the muscle, liver, serum, and gonad of common carp throughout the experimental period. Dotted lines indicate fanti and fanti-Cl11 values of DP in the spiked food.
anti-DP. Peng et al., 2012 recently reported the tissue distribution of DP in Chinese sturgeon. They found that the fanti in maternal tissues was significantly higher than that in eggs, while fanti-Cl11 in maternal tissue was significantly lower than that in eggs. Meanwhile, fanti in heart, intestine, and gonad were lower than those in the muscle and liver. Combining these results with our finding, the factors determining the isomer composition of DP in biota were DP concentration, different affinity for isomers in different tissues, and reaching equilibrium between the biota and the environment. To evaluate whether dechlorination occurred in fish, the concentration ratios of dechlorinated analogs to their parent compound were calculated for muscle (the predominant tissue for DP deposition) (Fig. S3). This result indicated that the ratio for syn-isomer and anti-isomer showed a decreasing trend in the depuration period and suggested that dechlorinated analogs are
eliminated more rapidly than their corresponding parent chemicals. This result did not support the dechlorination metabolism of the 2 DP isomers in fish. However, fanti and fanti-Cl11 in fish were lower than those in the commercial DP mixture, and the uptake efficiency of anti-isomer was higher than those of the syn-isomer, thereby strongly suggesting the selective degradation of antiisomers in fish. Thus, a metabolic pathway other than dechlorination, such as hydroxylated metabolism, may occur and more studies are needed to investigate this issue.
4. Conclusion In summary, dynamic tissue-specific accumulation of DP isomers was observed in common carp under laboratory conditions.
38
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
The higher absorption efficiencies but the lower assimilation efficiencies of anti-isomers indicated a stereoselective metabolism of antiisomer in common carp. A dynamic tissue distribution with an increasing proportion in the muscle along with a decreasing proportion in the liver over time was observed. The isomer composition of DPs and their dechlorinated analogs also exhibited tissue specificity and dynamic changes over the experimental period. Our result suggests that isomer-specific accumulation of DPs and their dechlorinated analogs in fish is a complex and multi-factorial process. Capsule Gastrointestinal absorption and tissue-specific accumulation of dechlorane plus in common carp under laboratory conditions were demonstrated. Acknowledgments This work was financially supported by the Chinese Academy of Sciences (No. KZCX2-EW-QN 105) and the National Natural Science Foundation of China (No. 41273118, 81273118, 41230639). This is a contribution of IS-1787 from GIGACS. Appendix A. Supporting information Supplementary data associated with this article can be found in the online version at http://dx.doi.org/10.1016/j.ecoenv.2013.11.021. References Abdelouahab, N., Suvorov, A., Pasquier, J.C., Langlois, M.F., Praud, J.P., Takser, L., 2009. Thyroid disruption by low-dose bde-47 in prenatally exposed lambs. Neonatology 96, 120–124. Ben, Y.-J., Li, X.-H., Yang, Y.-L., Li, L., Di, J.-P., Wang, W.-Y., Zhou, R.-F., Xiao, K., Zheng, M.-Y., Tian, Y., 2013. Dechlorane Plus and its dechlorinated analogs from an ewaste recycling center in maternal serum and breast milk of women in Wenling, China. Environ. Pollut. 173, 176–181. Fisk, A.T., Cymbalisty, C.D., Tomy, G.T., Muir, D.C., 1998. Dietary accumulation and depuration of individual C10-, C11-and C14-polychlorinated alkanes by juvenile rainbow trout (Oncorhynchus mykiss). Aquat. Toxicol. 43, 209–221. Gauthier, L.T., Letcher, R.J., 2009. Isomers of Dechlorane Plus flame retardant in the eggs of herring gulls (Larus argentatus) from the Laurentian Great Lakes of North America: temporal changes and spatial distribution. Chemosphere 75, 115–120. Guvenius, D.M., Hassanzadeh, P., Bergman, A., Norent, K., 2002. Metabolites of polychlorinated biphenyls in human liver and adipose tissue. Environ. Toxicol. Chem. 21, 2264–2269. Hoh, E., Zhu, L.Y., Hites, R.A., 2006. Dechlorane plus, a chlorinated flame retardant, in the Great Lakes. Environ. Sci. Technol. 40, 1184–1189. Iwata, H., Watanabe, M., Okajima, Y., Tanabe, S., Amano, M., Miyazaki, N., Petrov, E.A., 2004. Toxicokinetics of PCDD, PCDF, and coplanar PCB congeners in Baikal seals, Pusa sibirica: age-related accumulation, maternal transfer, and hepatic sequestration. Environ. Sci. Technol. 38, 3505–3513. Kubota, A., Iwata, H., Tanabe, S., Yoneda, K., Tobata, S., 2004. Levels and toxicokinetic behaviors of PCDD, PCDF, and coplanar PCB congeners in common cormorants from Lake Biwa, Japan. Environ. Sci. Technol. 38, 3853–3859.
Li, Y., Yu, L., Wang, J., Wu, J., Mai, B., Dai, J., 2013a. Accumulation pattern of Dechlorane Plus and associated biological effects on rats after 90d of exposure. Chemosphere 90, 2149–2156. Li, Y., Yu, L., Zhu, Z., Dai, J., Mai, B., Wu, J., Wang, J., 2013b. Accumulation and effects of 90‐day oral exposure to Dechlorane Plus in quail (Coturnix coturnix). Environ. Toxicol. Chem. 32, 1649–1654. Möller, A., Xie, Z.Y., Sturm, R., Ebinghaus, R., 2010. Large-scale distribution of dechlorane plus in air and seawater from the Arctic to Antarctica. Environ. Sci. Technol. 44, 8977–8982. Malmberg, T., Athanasiadou, M., Marsh, G., Brandt, I., Bergmant, Å., 2005. Identification of hydroxylated polybrominated diphenyl ether metabolites in blood plasma from polybrominated diphenyl ether exposed rats. Environ. Sci. Technol. 39, 5342–5348. Peng, H., Zhang, K., Wan, Y., Hu, J., 2012. Tissue distribution, maternal transfer, and age-related accumulation of dechloranes in Chinese sturgeon. Environ. Sci. Technol. 46, 9907–9913. Qi, H., Liu, L.Y., Jia, H.L., Li, Y.F., Ren, N.Q., You, H., Shi, X.Y., Fan, L.L., Ding, Y.S., 2010. Dechlorane plus in surficial water and sediment in a northeastern Chinese river. Environ. Sci. Technol. 44, 2305–2308. Qiu, X.H., Marvin, C.H., Hites, R.A., 2007. Dechlorane plus and other flame retardants in a sediment core from Lake Ontario. Environ. Sci. Technol. 41, 6014–6019. Roberts, M.S., Magnusson, B.M., Burczynski, F.J., Weiss, M., 2002. Enterohepatic circulation. Clin. Pharmacokinet. 41, 751–790. Sverko, E., Tomy, G.T., Reiner, E.J., Li, Y.F., McCarry, B.E., Arnot, J.A., Law, R.J., Hites, R.A., 2011. Dechlorane plus and related compounds in the environment: a review. Environ. Sci. Technol. 45, 5088–5098. Tomy, G.T., Palace, V.P., Halldorson, T., Braekevelt, E., Danell, R., Wautier, K., Evans, B., Brinkworth, L., Fisk, A.T., 2004. Bioaccumulation, biotransformation, and biochemical effects of brominated diphenyl ethers in juvenile lake trout (Salvelinus namaycush). Environ. Sci. Technol. 38, 1496–1504. Tomy, G.T., Pleskach, K., Ismail, N., Whittle, D.M., Helm, P.A., Sverko, E., Zaruk, D., Marvin, C.H., 2007. Isomers of dechlorane plus in Lake Winnipeg and Lake Ontario food webs. Environ. Sci. Technol. 41, 2249–2254. Tomy, G.T., Thomas, C.R., Zidane, T.M., Murison, K.E., Pleskach, K., Hare, J., Arsenault, G., Marvin, C.H., Sverko, E., 2008. Examination of isomer specific bioaccumulation parameters and potential in vivo hepatic metabolites of syn- and antiDechlorane Plus isomers in juvenile rainbow trout (Oncorhynchus mykiss). Environ. Sci. Technol. 42, 5562–5567. Wang, J., Tian, M., Chen, S.J., Zheng, J., Luo, X.J., An, T.C., Mai, B.X., 2011. Dechlorane plus in house dust from e-waste recycling and urban areas in south china: sources, degradation, and human exposure. Environ. Toxicol. Chem. 30, 1965–1972. Wu, J.P., Zhang, Y., Luo, X.J., Wang, J., Chen, S.J., Guan, Y.T., Mai, B.X., 2010. Isomerspecific bioaccumulation and trophic transfer of dechlorane plus in the freshwater food web from a highly contaminated site, south china. Environ. Sci. Technol. 44, 606–611. Xian, Q., Siddique, S., Li, T., Feng, Y.-l., Takser, L., Zhu, J., 2011. Sources and environmental behavior of Dechlorane Plus—a review. Environ. Int. 37, 1273–1284. Yan, X., Zheng, J., Chen, K.H., Yang, J., Luo, X.J., Yu, L.H., Chen, S.J., Mai, B.X., Yang, Z.Y., 2012. Dechlorane Plus in serum from e-waste recycling workers: influence of gender and potential isomer-specific metabolism. Environ. Int. 49, 31–37. Yu, Z.Q., Lu, S.Y., Gao, S.T., Wang, J.Z., Li, H.R., Zeng, X.Y., Sheng, G.Y., Fu, J.M., 2010. Levels and isomer profiles of Dechlorane Plus in the surface soils from e-waste recycling areas and industrial areas in South China. Environ. Pollut. 158, 2920–2925. Zeng, Y.H., Yu, L.H., Luo, X.J., Chen, S.J., Wu, J.P., Mai, B.X., 2013. Tissue accumulation and species‐specific metabolism of technical pentabrominated diphenyl ether (DE‐71) in two predator fish. Environ. Toxicol. Chem. 32, 757–763. Zhang, Y., Wu, J.P., Luo, X.J., Wang, J., Chen, S.J., Mai, B.X., 2011. Tissue distribution of Dechlorane Plus and its dechlorinated analogs in contaminated fish: high affinity to the brain for anti-DP. Environ. Pollut. 159, 3647–3652. Zheng, J., Wang, J., Luo, X.J., Tian, M., He, L.Y., Yuan, J.G., Mai, B.X., Yang, Z.Y., 2010. Dechlorane Plus in human hair from an E-waste recycling area in South China: comparison with dust. Environ. Sci. Technol. 44, 9298–9303. Zhu, J., Feng, Y.L., Shoeib, M., 2007. Detection of dechlorane plus in residential indoor dust in the city of Ottawa, Canada. Environ. Sci. Technol. 41, 7694–7698.
Contents lists available at ScienceDirect
Ecotoxicology and Environmental Safety journal homepage: www.elsevier.com/locate/ecoenv
Gastrointestinal absorption, dynamic tissue-specific accumulation, and isomer composition of dechlorane plus and related analogs in common carp by dietary exposure Yan-Hong Zeng a,b, Xiao-Jun Luo a,n, Bin Tang a,b, Xiao-Bo Zheng a,b, Bi-Xian Mai a a b
State Key Laboratory of Organic Geochemistry, Guangzhou Institute of Geochemistry, Chinese Academy of Sciences, Guangzhou 510640, PR China University of Chinese Academy of Sciences, Beijing 100049, PR China
art ic l e i nf o
a b s t r a c t
Article history: Received 1 August 2013 Received in revised form 22 October 2013 Accepted 25 November 2013 Available online 20 December 2013
Dechlorane plus (DP) is a high-productive volume substance which had been identified as ubiquitous pollutant and has been widely detected in biota. In the present study, common carp (Cyprinus carpio) was exposed to known amounts of commercial DP-25 under laboratory conditions. The gastrointestinal absorption and tissue-specific bioaccumulation of DP and its dechlorinated analogs in common carp were investigated. The higher absorption efficiencies but lower assimilation efficiencies of anti-isomers indicated stereoselective metabolism of anti-isomers in fish. Linear uptake curves were seen in serum and muscle, but the depuration curves for all the four tissues (muscle, serum, liver and gonad) did not follow the first-order kinetics. The liver exhibited a high affinity for anti-isomers during the experiment. Other tissues, such as serum, muscle, and gonad, showed a selective accumulation of syn-DP in the early stages of the experiment, particularly the serum. However, the deviation of fanti between different tissues disappeared at late stages of the experiment, and the fanti values in all tissues were close to that in commercial mixtures. Our results suggest that the bioaccumulation of DP is a complex and multifactorial process. & 2013 Elsevier Inc. All rights reserved.
Keywords: DP Gastrointestinal absorption Tissue-specific accumulation Isomer composition
1. Introduction Dechlorane plus (DP) was introduced as a substitute for Dechlorane (an additive chlorinated flame retardant) by Hooker Chemical in the 1960s (Hoh et al., 2006). It has been used in wire and circuit boards, paint, and hard connectors in computers (Tomy et al., 2007). The commercial mixture consists primarily of 2 isomers (syn- and anti-DP) with a minor content of 2 dechlorinated isomers (syn- and anti-Cl11-DP) (Zheng et al., 2010). DP has been classified as a high-productive volume substance by the United States Environmental Protection Agency (Sverko et al., 2011). Through its long commercial history, it has not received attention until recently. DP was first identified in ambient air, fish, and sediment samples from the Great Lakes region in 2006 (Hoh et al., 2006). DP has since been measured in sediment (Qi et al., 2010; Qiu et al., 2007), surface soil (Yu et al., 2010), indoor dust (Zhu et al., 2007), biota (Gauthier and Letcher, 2009; Tomy et al., 2007), and even in air and seawater from the Arctic to Antarctica (Möller et al., 2010). Recent studies have also reported detection of DP in humans (Ben et al., 2013; Yan et al., 2012).
n
Corresponding author. Fax: +86 20 85290706. E-mail address: [email protected] (X.-J. Luo).
0147-6513/$ - see front matter & 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.ecoenv.2013.11.021
The detection of DP in biota suggests that the isomers are bioavailable and bioaccumulative. As reported in previous studies, the bioaccumulation tendencies of the isomers vary. Tomy et al., 2007 reported that the anti-isomer was dominant at higher trophic levels while the syn-isomer dominated in lower trophic level organisms in food web samples from Lake Winnipeg and Lake Ontario. Wu et al., 2010 also found that the fraction of anti-DP decreased upon moving up the trophic levels of aquatic organisms in a reservoir near an electronic waste recycling workshop in South China. Different extents of enrichment (anti-) and depletion (syn-) of the isomers were found in Lake Winnipeg biota by Tomy and colleagues (Tomy et al., 2007). The complexity of environmental DP isomer compositions often makes the fate studies of DP highly confusing. For example, different ratios of anti-DP to total DP (fanti) of a biota may be due to the accumulation of different DP isomer compositions from the environment (Xian et al., 2011) and perhaps due to isomer-specific bioavailability of DP in the biota. In Previous studies, accumulation of DPs in juvenile rainbow trout (Tomy et al., 2008), rat (Li et al., 2013a), and Quail (Li et al., 2013b) has been investigated in laboratory experiment. Different accumulation potential for anti- and syn-isomer was observed in rainbow trout, rat and quail. However, there is little understanding of the bioaccumulation differentiation of the isomers. Laboratory exposure experiments in controlled conditions are convenient
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
methods for investigating the bioaccumulation processes of contaminants. Therefore, the main objective of the present study was to improve the understanding of the bioaccumulation process of DP. To the best of our knowledge, only a single laboratory study has been conducted to explore this issue in fish (Tomy et al., 2008), but that study used juvenile rainbow trout (Oncorhynchus mykiss) exposed to syn- and anti-DP separately and no dechlorinated analogs were spiked in the food. Moreover, gastrointestinal absorption was not taken into consideration. The gastrointestinal tract is a critical segment which influences and limits the compound absorption as most compounds in fish are believed to be absorbed by the intestine. However, there is no information concerning about the gastrointestinal absorption of DP in fish. In the present study, common carp (Cyprinus carpio) was exposed to contaminated food pellets for 50 days. The contaminated food was made by known amounts of commercial DP-25 dissolved in cod liver and then mixed with fish food pellets. After the exposure period, a depuration period lasting a further 40 days was designed for the carp, during which the carp were fed with non-spiked food. The purposes of the present study were (1) to investigate gastrointestinal absorption by comparing the concentration and congener profile of DP between spiked food and feces and (2) to investigate the uptake and depuration behaviors of the DP congeners in different carp tissues after exposure to high doses of DP-25 mixtures.
2. Materials and methods 2.1. Fish exposure Common carp is a kind of freshwater and omnivorous fish. The carp reach sexual maturity in 1 to 3 years and lay eggs. In March, 2013, forty common carps, approximately 12 cm in length, were purchased from an aquarium market in Guangzhou, China. Four carps were randomly collected as the control group and kept in one glass tank (80 cm 35 cm 50 cm). The remaining carp (n¼ 36) were kept in another glass tank (120 cm 40 cm 60 cm) as the treated group. Air stones were placed in the tank to maintain oxygen saturation in the water. The water in each tank was heated to a constant temperature of 22 7 1 1C and circulated using a submersible pump at a rate of 1.5 L/min. Fish were acclimated to the non-spiked diet for a period of 7 days prior to exposure. Fish were fed food at a rate of 1 percent of their body weight/day based on their weight. Commercial DP-25 (Z 99 percent purity, particle diameter r 6 μm, Chlorine content of 65.1 percent and density of 1.8 g/cc) was acquired from Jiangsu Anpon Co., Ltd and cod liver oil was obtained from Peter Moller (Norway). DP-25 was delivered in artificially contaminated food. The DP dose of fish exposed was approximately 1.5 mg per day for each fish. DP-25 mixtures were first dissolved in isooctane and then diluted to the desired concentration with cod liver oil. Then, 1 mL of cod liver oil was mixed with 10 g of fish food pellets (Yangzhou Five-Star Aquatic Products Co., LTD, PR China). Non-spiked food was prepared by mixing food with solute-free cod liver oil. Food was homogenized by mixing in a shaking incubator (24 h, 25 1C). The spiked and non-spiked food was stored in amber containers in a cool, dark place throughout their use. The treated group (36 common carps) was exposed to spiked food for 50 days, followed by non-spiked food for 40 days. The control group was fed a non-spiked diet throughout the experiment. The food was collected on days 5 and 45 (spiked food in uptake period) and on days 55 and 90 (non-spiked food in depuration period). Feces were collected on days 5, 10, 15, 25, 35, 40, and 50 (uptake period), and on days 60, 70, 80, 85, and 90 (depuration period). During the entire experimental period (90 days), 2 fish were sampled from the tank every 5 days and were carefully dissected, resulting in 36 fish samples (internal tissues and gills were removed). The blood, liver, and gonad of fish sampled on the same day were pooled into 1 sample. Blood samples were drawn from the dorsal aorta by using syringes, transferred into 5-mL Teflon tubes, and centrifuged at 3000 rpm for 15 min to obtain serum. The fish carcasses (n ¼36) and pooled serum (n¼18), liver (n¼ 18), and gonad samples (n¼18) were obtained for analysis of DP (anti-DP and syn-DP) and dechlorinated products (anti-Cl10-DP, anti-Cl11-DP, and syn-Cl11-DP).
2.2. Sample preparation The extraction and analysis procedures used for the fish carcass (approximately 2 g), liver, and gonad were similar to those described in previous studies (Hoh et al., 2006). Briefly, freeze-dried samples were ground into powder and spiked with surrogate standards (BDE 181) prior to Soxhlet extraction with 50 percent acetone in
33
hexane for 24 h. The extract was concentrated and then solvent exchanged to hexane (10 mL). The lipid content was determined by gravimetric measurement from an aliquot of extract. The remaining extract was treated with 3 mL of concentrated sulfuric acid (H2SO4) (analytical reagent grade, Guangzhou Chemical Reagent Factory, China) twice to remove any lipids present in the samples. After centrifugation for 15 min, the hexane layer was recovered and combined with 6 mL of hexane to wash the pooled H2SO4 residue. The lipid-free extract was washed with sodium sulfate (5 percent, w/w) to remove the residual H2SO4 and concentrated to 1 mL, further cleaned up on a multilayer column (i.d.¼ 1 cm) packed with 8-cm neutral silica, 8-cm acidified silica, and a 2-cm layer of anhydrous sodium sulfate. The DP isomers and their dechlorinated congeners were obtained by eluting with 30 mL hexane/ dichloromethane (v/v¼ 1:1). After solvent exchange to isooctane (1 mL), internal standards (BDE 128) were added before gas chromatography/mass spectrometry analysis (GC/MS). A sample preparation method similar to the fish tissue preparation method was used for the food and feces samples. After Soxhlet extraction, the extract was concentrated, subjected to solvent exchange with hexane (10 mL), treated with H2SO4, and cleaned as described above. Serum pretreatment was performed as previously described with a minor modification (Malmberg et al., 2005). Briefly, after spiking with the surrogate standards (BDE 181), approximately 0.5 g serum was denatured using hydrochloric acid (6 N, 1 mL) and 2-propanol (6 mL), and the mixtures were shaken vigorously. Mixtures were then extracted 3 times with methyl tert-butyl ether: hexane (1:1, v/v, 6 mL). The combined organic extracts were washed with a potassium chloride solution (1 percent, w/w, 4 mL) and blown to near dryness under N2 for gravimetric lipid weight determination. The extracts were redissolved with 9 mL of hexane, then treated with H2SO4, and cleaned as described above. 2.3. Instrumental analysis DP and its dechlorinated congeners were analyzed by GC/MS (Agilent 7890A/5975B MSD; Agilent Technology, CA) with an electron capture negative ionization source in a selective ion-monitoring mode and separation performed using a DB-XLB capillary column, 30 m 0.25 mm ID 0.25-mm film (J&W Scientific). The carrier gas was helium with a constant column flow rate of 1.3 mL/min. The column temperature program was as follows: maintained at 110 1C for 1 min, heated to 200 1C at a 1st rate of 8 1C/min and then maintained at 200 1C for 1 min, heated to 240 1C at a 2nd rate of 3 1C/min and maintained at 240 1C for 2 min, heated to 280 1C at a 3rd rate of 5 1C/min and maintained at 280 1C for 10 min, and finally heated to 310 1C at a 4th rate of 10 1C/min and maintained at 310 1C for 10 min. We automatically injected 1 mL of sample into the pulsed splitless mode. The temperature of the injection port was set at 290 1C. Ion fragments m/z 653.8 and 651.8 for DP isomers (syn-DP and anti-DP), m/z 618 and 620 for Cl11-DP isomers (anti-Cl11-DP and syn-Cl11-DP), m/z 584 and 586 for antiCl10-DP, and m/z 79 and 81 for BDE 181 and BDE 128 were monitored. syn-Cl11-DP was previously identified as a dechlorinated product of syn-DP (Wang et al., 2011). Semiquantitation was achieved for syn-Cl11-DP in reference to the response of anti-Cl11-DP on GC/MS. The compounds were quantified using the internal calibration method based on a 9-point calibration curve.
2.4. Data analysis The net assimilation efficiency of each DP compound was calculated using the following equation: α ¼ ðbody burden in common carp at time tÞ=ðcumulative mass exposed to common carp up to time tÞ
where α is net assimilation efficiency. The net assimilation efficiency calculations mostly involve the determination of ingestion minus egestion and excretion. In the present study we have explored net assimilation efficiency given that our target was primarily to examine the differences in assimilation and depuration between syn-DP congeners and anti-DP congeners. All statistical analyses were conducted using PASW Statistics 18.0 to test for the inter-tissues variability of DP levels and isomer compositions. DP concentrations determined in fish tissues were not normalized to total lipid content, due to higher variations of these concentrations when normalized to lipid content.
3. Results and discussion 3.1. Background levels and quality control After the 90-d experiment, the length of common carp was 12.4 7 0.8 cm, which did not change significantly compared to days 1 (12.1 70.6 cm) (p o0.05). No mortality was observed during the experiment. Procedural blanks covering the whole procedure were performed in parallel with the samples at each batch extraction. Anti-DP was detected in the blanks, but syn-DP,
34
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
anti-Cl10-DP, syn-Cl11-DP, and anti-Cl11-DP were not detected in the blanks. The method detection limit (MDL) was defined as the mean value plus 3-fold standard deviation for anti-DP detected in the procedural blanks (n¼ 7). For syn-DP, anti-Cl10-DP, syn-Cl11-DP, and anti-Cl11-DP, which were not detected in blanks, a signal-to-noise ratio of 10 was set as the MDL. The MDLs for syn-DP, anti-DP, antiCl10-DP, syn-Cl11-DP, and anti-Cl11-DP were between 0.02 and 0.11 ng/g wet weight (ww) in fish tissues, and between 0.18 and 0.47 ng/g dry weight (dw) in food and feces. The recoveries of BDE 181 for all samples (n ¼112) ranged from 80 percent to 128 percent. During the 90-day experiment, no anti-Cl10-DP, syn-Cl11DP, or anti-Cl11-DP were detected in the control fish tissues or the non-spiked food. Both anti-DP and syn-DP were detected in control fish tissues and the non-spiked food. The average concentrations of syn-DP and anti-DP in control fish tissues were 197 12 pg/g and 34 726 pg/g (ww) and 0.52 70.38 ng/g and 1.5 71.1 ng/g (dw) in non-spiked food, respectively. These levels were several orders of magnitude lower than those of the dosed fish and the spiked food. 3.2. Gastrointestinal absorption and fecal excretion of DP congeners Syn-DP, anti-DP, syn-Cl11-DP, and anti-Cl11-DP were detected in 100 percent of the food and fecal samples while no anti-Cl10-DP was detected. The average concentrations of syn-DP, anti-DP, synCl11-DP, and anti-Cl11-DP in the spiked food were 2071 724, 7826 71031, 4.0 70.3, and 33.570.4 ng/g (dw), respectively. The concentrations of DP congeners in feces collected in the uptake period were in the range of 1.5–8.2-fold compared to that in the administered food. A ratio of the chemical concentration in the feces to that in the food is directly related to the chemical absorption and excretion in the gut if the chemicals are not metabolized by endogenous enzymes in the gastrointestinal system. The higher the ratio, the lower the absorption efficiency or the higher excretion efficiency is. The concentration ratios of feces to food in the uptake period were in the range of 1.55–4.98 for synDP, 1.53–4.31 for anti-DP, 1.86–8.26 for syn-Cl11-DP and 1.53–6.31 for anti-Cl11-DP. The ratios of feces to food for syn-DP were slightly higher than those for anti-DP in each sampling in the uptake period except for the 50-day sampling (p o0.05). Similar results were also seen between syn-Cl11-DP and anti-Cl11-DP (po 0.05). These phenomena imply that the absorption efficiencies of antiisomers in the gastrointestinal system were higher than those of the syn-isomers. In the study of Tomy et al., 2008, accumulation rate of syn-DP was found to be faster than that of anti-DP in juvenile rainbow trout. Obviously, the difference in gastrointestinal absorption rate between the two isomers cannot be responsible for stereoselective accumulation of syn-DP in fish given that anti-DP rather than syn-DP were absorbed more efficiently through the gut. The concentrations of four chemicals in the feces collected in the 60-day (the first feces sample for the depuration period) sharply decreased by 1 order of magnitude compared to the feces in the uptake period. An exponential decrease trend of concentration of target chemicals over the depuration period was observed (Fig. 1b). To evaluate the differences in excretion rate between synDP and anti-DP, a ratio of feces to food (non-spiked food) was calculated (Fig. 1b). The calculated result indicated that there were no systematic differences in the ratio of feces to food between synDP and anti-DP (p o0.05), indicating no stereoselective excretion for DP in common carp. The congener profile of DP can be expressed by the fractions of the anti-isomer (fanti: ratio of anti-DP concentration to total concentration of anti-DP and syn-DP; fanti-Cl11: ratio of anti-Cl11DP concentration to total concentration of anti-Cl11-DP and synCl11-DP). fanti in the spiked food was 0.791 70.02, which was
slightly higher than those in feces collected in the uptake period (range: 0.765–0.787; mean, 0.775) except for at 50 days (0.850). The fanti-Cl11 in the spiked food was 0.893, which was also higher than that in feces in the uptake period (range: 0.853–0.873; mean, 0.866) except for at 50 days (0.920) (Fig. S1). These results further confirm that syn-isomers have lower absorption efficiencies or higher elimination efficiencies than anti-isomers in the gut. fanti and fanti-Cl11 were significantly lower in feces in the depuration period than in feces in the uptake period and in the spiked food (p o0.05) (Fig. S1), indicating anti-isomers have been selectively metabolized in common carps. 3.3. Uptake and depuration kinetics of DP in fish tissues The uptake and depuration kinetics of DP in the serum, muscle, liver, and gonad were investigated. The four chemicals (anti-DP, syn-DP, anti-Cl11-DP and syn-Cl11-DP), except for syn-Cl11-DP in the serum, were detected in all target tissues. All chemicals exhibited linear accumulation kinetics in serum (r2 ¼0.88, r2 ¼0.84 and r2 ¼ 0.69 for anti-DP, syn-DP and anti-Cl11-DP, respectively) and muscle (r2 ¼ 0.84, r2 ¼0.81, r2 ¼0.84 and r2 ¼ 0.81 for anti-DP, synDP anti-Cl11-DP and syn-Cl11-DP, respectively) samples during the uptake period (Fig. 2a and b) and reached the highest concentrations at the end of the uptake period (50th day). No steady state was observed in the serum or muscle for any chemical during the 50-day exposure. Depuration of the chemicals in both muscle and serum showed an initial rapid depuration for the first 10 days followed by a fluctuating depuration over the remainder of the experiment. No obvious trend towards a decrease was found in the last 30 days of the depuration. There are no previous studies regarding DP uptake and clearance curves in the fish serum, but the DP uptake and clearance curves in the muscle observed in the present study were similar to results reported by other researchers during bioaccumulation studies of hydroponic contaminants in fish (Fisk et al., 1998; Tomy et al., 2004). The uptake kinetics of DP and their dechlorinated analogs in the liver and gonad were not linear (Fig. 2c and d). The concentrations in the gonad showed co-variation with those in the liver with the 5-day hysteresis during the uptake period. This hysteresis in the gonad might be attributed to maternal transfer of DP in fish. The depuration curves of DP in the liver showed 2-stage elimination kinetics, with an initial decrease to day 70, followed by a slight increase. The concentrations of DP and their analogs fluctuate and follow a zigzag pattern in the depuration. This unique accumulation pattern of DP in the gonad of carp could be because gonads collected in different individuals may be in different stages of embryonic development as some of the DP was determined in fish ovaries and some was determined in fish eggs. It remains unclear as to why no linear uptake occurred in the liver in the uptake period and accumulation in the liver was still occurring at the end of the experiment. Enterohepatic circulation and redistribution of lipophilic contaminants can affect liver concentrations and ultimately give rise to complex uptake and depuration kinetics (Roberts et al., 2002). Additionally, the potential metabolism of DP and analogs in the liver and the high variation in the concentrations between individual fish could also contribute to the complex uptake and depuration kinetics. Moreover, during the experimental period, the undulating tissue concentrations of DP isomers and Cl11-DP isomers could be partly due to the individual variation. Meanwhile, based on the research of Tomy et al., 2008, which reported half-lives of DP in the 40 to 50 day length, so the depuration phase (40 d) of the present experiment was not long enough. This is probably a major contributor to the inability for us to find a significant elimination rate. The mean assimilation efficiencies of syn-DP, anti-DP, syn-C11DP, and anti-C11-DP calculated from days 5, 10, 15, 20, 25, 30, 35,
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
35
Conc. of DP isomers (ng/g dry weight)
Conc. ratios in feces to spiked food
Syn-DP Anti-DP Syn-Cl11-DP
8 7
Anti-Cl11-DP
6 5 4 3 2 1 0
5
10
15
20
25
30
35
40
45
50
55
Time (days)
700
700
Ratio for syn-DP Ratio for anti-DP
600
600
500
500
400
400
300
300
200
200
100
100
Syn-DP concentration Anti-DP concentration
0
55
60
65
70
0 75
80
85
90
Conc. ratios of feces to non-spiked food
Depuration period
Uptake period 9
95
Time (days)
Fig. 1. (a) Concentration ratios of DP congeners and dechlorinated congeners in feces to spiked food during the uptake period of common carp; (b) concentrations of DP congeners and the concentration ratios of DP congeners in feces to non-spiked food during the depuration period of common carp. The solid line indicates an exponential decrease trend of the concentrations of DP isomers in the feces along with depurated time.
40, 45, and 50 were estimated to be 3.8 percent, 3.2 percent, 20.4 percent, and 14.8 percent, respectively. These assimilation efficiencies of DP isomers were comparable to those (6.0 percent for syn-DP and 3.9 percent for anti-DP) determined in juvenile rainbow trout (Tomy et al., 2008). Meanwhile, the assimilation efficiencies of syn-isomers were significantly higher than those of anti-isomers (p o0.05). However, the ratios of feces to food indicate that the uptake efficiencies of syn-isomers were lower than those of the anti-isomers. Therefore, the relatively low assimilation efficiencies of anti-isomers could be attributed to stereoselective metabolism of anti-isomers in carp. This is also consistent with the structural conformation of the isomers as it has been suggested that because of the configuration of the pendant chlorocyclopentane moieties, the anti-isomer would be more susceptible to biological attack (Tomy et al., 2008). 3.4. Dynamic tissue distribution and isomer composition The concentrations of DP and the analogs in the liver were higher than those in the muscle, serum, and gonad throughout the experiment (po0.1) (Fig. 3). Lipid content is not a cause for this tissue distribution because the lipid contents in the liver (1.7071.56 percent) were significantly lower than those in the muscle (3.3971.69 percent) and the gonad (3.7171.96 percent). This result indicates that DP and analogs prefer to accumulate in the liver than in other tissues, which is consistent with that reported by Li et al., 2013a and is also in line with reports on other highly halogenated flame retardants (Guvenius et al., 2002; Iwata et al., 2004). In Chinese sturgeon, liver also showed higher dechlorane concentrations than muscle and adipose tissue (Peng et al., 2012). Sequestration of DP isomers by hepatic proteins could be a cause for their disposition in the liver although this was not identified (Li et al., 2013b). Exposure to xenobiotic pollutants such as polychlorinated dibenzo-p-diosins, polychlorinated dibenzo-furans, and coplanar PCBs may induce hepatic binding proteins leading to hepatic sequestration of the compounds (Kubota et al., 2004). A dynamic tissue distribution for DP and its analogs among tissues was observed. At the start of exposure, the concentration ratios of the liver to other tissues were the highest, and then the ratio exponentially decreased over time to close to or slightly lower than 1 at the end of the experiment (Fig. 3). This result indicated that the liver is the first organ to which contaminants deposit after absorption from the gastrointestinal tract in fish. This result was consistent with a study on the tissue distribution of
PBDEs in 2 predatory fish species (Zeng et al., 2013). In that study, the liver was found to readily achieve equilibrium with the serum while the muscle did not until the end of the experiment, using a tissue fugacity comparison. The muscle is the main organ for contaminant deposition. The percentage of DP and analogs in the muscle was greater than 60 percent during the entire experimental period (Fig. S2). This is because the muscle is the largest tissue in volume in fish, despite its low concentration. The percentage in the muscle increased over time and reached the maximum at the 75th day (95 percent), and then a decreasing trend was seen until the end of the experiment. A concomitant decrease in the liver from 23 percent at the 5th day to 3 percent at the 75th day was observed. The gradual accumulation of contaminants occurring in the muscle was the main reason for the above observation. The decreased proportion in the muscle along with a corresponding increase in the liver in the last 15 days of the experiment implies a redistribution of DP and analogs among tissues. A non-regular trend was found for the gonadal proportion during the whole experiment, and this may be due to the variations in the number of gonads between individual fish. DP and analogs in the gonad accounted for 2 percent to 20 percent of the total amount in fish with most cases being greater than 5 percent (Fig. S2). Meanwhile, the concentration of DP and analogs in the gonad was higher than that in the muscle (p o0.05), which is consistent with the report of Peng et al. (2012), where the ratio of the chemical concentration in the eggs to those in the maternal tissues was greater than 1. These results indicate that DP and analogs readily underwent maternal transfer. Considering the fact that a high content of the pollutant usually exhibits a significant development toxicity to naturally hatched larvae (Abdelouahab et al., 2009), such a high accumulation of DP and Cl11-DP in the gonad suggests that the maternal transfer of DP congeners and the corresponding adverse effects for larvae should be elucidated further. The isomer composition of DP and analogs also showed tissuespecific and dynamic changes over time (Fig. 4). In the early stage of the uptake experiment (from day 0 to the 20th day), fanti and fanti-Cl11 values in tissues were lower than those in the commercial DP mixtures, indicating a preference for syn-isomers. fanti and fantiCl11 values then increased over time until they were close to those in commercial DP mixtures, especially fanti in the serum. Serum accumulated more syn-DP than anti-DP compared with other tissues while the liver showed a preference for anti-DP to syn-DP
36
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
Depuration
Uptake
Uptake
300 250
Depuration
80
Serum
70
Muscle
200 150
y = -18.77+4.85x r2 = 0.88; p < 0.001
Concentration of congener (ng/g ww)
Concentration of congener (ng/g ww)
60
100 50 y = -1.32+1.48x r2 = 0.84; p < 0.001
3.0 2.5 2.0 1.5 1.0
y = 0.05+0.022x r2 = 0.69; p = 0.002
0.0
y = -6.34+1.30x r2 = 0.84; p < 0.001
30 20 10
1.0 0.8
y = -1.80+0.37x 2 r = 0.81; p < 0.001 y = 0.026+0.018x 2 r = 0.84; p < 0.001
0.6 0.4 y = 0.012+0.013x 2 r = 0.81; p < 0.001
0.0 0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95
0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95
Time (days)
Time (days)
600
Liver
Gonad
500
300 200 100
syn-DP anti-DP syn-Cl11-DP
5 4
anti-Cl11-DP
3 2
Concentration of congener (ng/g ww)
400
Concentration of congener (ng/g ww)
40
0.2
0.5
500
50
400 300 200 100 0 8 7 6 5 4 3 2 1
1
0
0
-1 0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95
0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95
Time (days)
Time (days)
Fig. 2. Uptake and depuration curves of syn-DP, anti-DP, syn-Cl11-DP, and anti-Cl11-DP throughout the experimental period in (a) serum, (b) muscle, (c) liver, and (d) gonad of common carp.
compared with other tissues in the early stage of the experiment. The fact that the liver preferred anti-DP to syn-DP was in line with the results of a rat exposure experiment (Li et al., 2013a). The greatest deviation of fanti between the serum and liver reached 0.09 (0.70 vs. 0.79). However, in the late stage of the experiment, the larger deviation of fanti between different tissues disappeared and no significant differences were found between different tissues (p4 0.1). These results indicate that the isomer-specific accumulation of DP is a very complex and multi-factorial process.
In the current state, it was not possible to provide a detailed mechanism for this accumulation. Using rats exposed to different doses of DP, Li et al., 2013a found that no stereoselective accumulation was seen in the low dose exposure group, whereas a selective accumulation of syn-DP was observed in the high dose exposure group. This result indicates that the fanti value in rat was dose-dependent. Tissuespecific fanti has been reported in several previous studies. Zhang et al., 2011 reported that the brain of mud carp has a high affinity for
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
37
35
30
30
25
25 20
Concentration ratios of liver to other tissues
20 15
15
10
10
5
5
0
0
40
30
35
25
30 20
25 20
15
15
Liver/Muscle Liver/Serum Liver/Gonad
10
10 5
5
0
0 0
10
20
30
40
50
60
70
80
90
0
10
20
30
40
50
60
70
80
90
Time (days)
fanti (anti-DP/(anti-DP+sym-DP))
Muscle
Liver
Serum
Gonad
0.80
0.75
0.70
0.65
fanti-Cl11(anti-Cl11-DP/(anti-Cl11-DP + syn-Cl11-DP))
Fig. 3. Concentration ratios of (a) syn-DP, (b) anti-DP, (c) anti-Cl11-DP, (d) syn-Cl11-DP in the liver compared to other tissues of common carp throughout the experimental period.
Muscle
Liver
Gonad
0.90
0.85
0.80
0.75 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90
5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90
Time (days)
Time (days)
Fig. 4. fanti and fanti-Cl11 values calculated in the muscle, liver, serum, and gonad of common carp throughout the experimental period. Dotted lines indicate fanti and fanti-Cl11 values of DP in the spiked food.
anti-DP. Peng et al., 2012 recently reported the tissue distribution of DP in Chinese sturgeon. They found that the fanti in maternal tissues was significantly higher than that in eggs, while fanti-Cl11 in maternal tissue was significantly lower than that in eggs. Meanwhile, fanti in heart, intestine, and gonad were lower than those in the muscle and liver. Combining these results with our finding, the factors determining the isomer composition of DP in biota were DP concentration, different affinity for isomers in different tissues, and reaching equilibrium between the biota and the environment. To evaluate whether dechlorination occurred in fish, the concentration ratios of dechlorinated analogs to their parent compound were calculated for muscle (the predominant tissue for DP deposition) (Fig. S3). This result indicated that the ratio for syn-isomer and anti-isomer showed a decreasing trend in the depuration period and suggested that dechlorinated analogs are
eliminated more rapidly than their corresponding parent chemicals. This result did not support the dechlorination metabolism of the 2 DP isomers in fish. However, fanti and fanti-Cl11 in fish were lower than those in the commercial DP mixture, and the uptake efficiency of anti-isomer was higher than those of the syn-isomer, thereby strongly suggesting the selective degradation of antiisomers in fish. Thus, a metabolic pathway other than dechlorination, such as hydroxylated metabolism, may occur and more studies are needed to investigate this issue.
4. Conclusion In summary, dynamic tissue-specific accumulation of DP isomers was observed in common carp under laboratory conditions.
38
Y.-H. Zeng et al. / Ecotoxicology and Environmental Safety 100 (2014) 32–38
The higher absorption efficiencies but the lower assimilation efficiencies of anti-isomers indicated a stereoselective metabolism of antiisomer in common carp. A dynamic tissue distribution with an increasing proportion in the muscle along with a decreasing proportion in the liver over time was observed. The isomer composition of DPs and their dechlorinated analogs also exhibited tissue specificity and dynamic changes over the experimental period. Our result suggests that isomer-specific accumulation of DPs and their dechlorinated analogs in fish is a complex and multi-factorial process. Capsule Gastrointestinal absorption and tissue-specific accumulation of dechlorane plus in common carp under laboratory conditions were demonstrated. Acknowledgments This work was financially supported by the Chinese Academy of Sciences (No. KZCX2-EW-QN 105) and the National Natural Science Foundation of China (No. 41273118, 81273118, 41230639). This is a contribution of IS-1787 from GIGACS. Appendix A. Supporting information Supplementary data associated with this article can be found in the online version at http://dx.doi.org/10.1016/j.ecoenv.2013.11.021. References Abdelouahab, N., Suvorov, A., Pasquier, J.C., Langlois, M.F., Praud, J.P., Takser, L., 2009. Thyroid disruption by low-dose bde-47 in prenatally exposed lambs. Neonatology 96, 120–124. Ben, Y.-J., Li, X.-H., Yang, Y.-L., Li, L., Di, J.-P., Wang, W.-Y., Zhou, R.-F., Xiao, K., Zheng, M.-Y., Tian, Y., 2013. Dechlorane Plus and its dechlorinated analogs from an ewaste recycling center in maternal serum and breast milk of women in Wenling, China. Environ. Pollut. 173, 176–181. Fisk, A.T., Cymbalisty, C.D., Tomy, G.T., Muir, D.C., 1998. Dietary accumulation and depuration of individual C10-, C11-and C14-polychlorinated alkanes by juvenile rainbow trout (Oncorhynchus mykiss). Aquat. Toxicol. 43, 209–221. Gauthier, L.T., Letcher, R.J., 2009. Isomers of Dechlorane Plus flame retardant in the eggs of herring gulls (Larus argentatus) from the Laurentian Great Lakes of North America: temporal changes and spatial distribution. Chemosphere 75, 115–120. Guvenius, D.M., Hassanzadeh, P., Bergman, A., Norent, K., 2002. Metabolites of polychlorinated biphenyls in human liver and adipose tissue. Environ. Toxicol. Chem. 21, 2264–2269. Hoh, E., Zhu, L.Y., Hites, R.A., 2006. Dechlorane plus, a chlorinated flame retardant, in the Great Lakes. Environ. Sci. Technol. 40, 1184–1189. Iwata, H., Watanabe, M., Okajima, Y., Tanabe, S., Amano, M., Miyazaki, N., Petrov, E.A., 2004. Toxicokinetics of PCDD, PCDF, and coplanar PCB congeners in Baikal seals, Pusa sibirica: age-related accumulation, maternal transfer, and hepatic sequestration. Environ. Sci. Technol. 38, 3505–3513. Kubota, A., Iwata, H., Tanabe, S., Yoneda, K., Tobata, S., 2004. Levels and toxicokinetic behaviors of PCDD, PCDF, and coplanar PCB congeners in common cormorants from Lake Biwa, Japan. Environ. Sci. Technol. 38, 3853–3859.
Li, Y., Yu, L., Wang, J., Wu, J., Mai, B., Dai, J., 2013a. Accumulation pattern of Dechlorane Plus and associated biological effects on rats after 90d of exposure. Chemosphere 90, 2149–2156. Li, Y., Yu, L., Zhu, Z., Dai, J., Mai, B., Wu, J., Wang, J., 2013b. Accumulation and effects of 90‐day oral exposure to Dechlorane Plus in quail (Coturnix coturnix). Environ. Toxicol. Chem. 32, 1649–1654. Möller, A., Xie, Z.Y., Sturm, R., Ebinghaus, R., 2010. Large-scale distribution of dechlorane plus in air and seawater from the Arctic to Antarctica. Environ. Sci. Technol. 44, 8977–8982. Malmberg, T., Athanasiadou, M., Marsh, G., Brandt, I., Bergmant, Å., 2005. Identification of hydroxylated polybrominated diphenyl ether metabolites in blood plasma from polybrominated diphenyl ether exposed rats. Environ. Sci. Technol. 39, 5342–5348. Peng, H., Zhang, K., Wan, Y., Hu, J., 2012. Tissue distribution, maternal transfer, and age-related accumulation of dechloranes in Chinese sturgeon. Environ. Sci. Technol. 46, 9907–9913. Qi, H., Liu, L.Y., Jia, H.L., Li, Y.F., Ren, N.Q., You, H., Shi, X.Y., Fan, L.L., Ding, Y.S., 2010. Dechlorane plus in surficial water and sediment in a northeastern Chinese river. Environ. Sci. Technol. 44, 2305–2308. Qiu, X.H., Marvin, C.H., Hites, R.A., 2007. Dechlorane plus and other flame retardants in a sediment core from Lake Ontario. Environ. Sci. Technol. 41, 6014–6019. Roberts, M.S., Magnusson, B.M., Burczynski, F.J., Weiss, M., 2002. Enterohepatic circulation. Clin. Pharmacokinet. 41, 751–790. Sverko, E., Tomy, G.T., Reiner, E.J., Li, Y.F., McCarry, B.E., Arnot, J.A., Law, R.J., Hites, R.A., 2011. Dechlorane plus and related compounds in the environment: a review. Environ. Sci. Technol. 45, 5088–5098. Tomy, G.T., Palace, V.P., Halldorson, T., Braekevelt, E., Danell, R., Wautier, K., Evans, B., Brinkworth, L., Fisk, A.T., 2004. Bioaccumulation, biotransformation, and biochemical effects of brominated diphenyl ethers in juvenile lake trout (Salvelinus namaycush). Environ. Sci. Technol. 38, 1496–1504. Tomy, G.T., Pleskach, K., Ismail, N., Whittle, D.M., Helm, P.A., Sverko, E., Zaruk, D., Marvin, C.H., 2007. Isomers of dechlorane plus in Lake Winnipeg and Lake Ontario food webs. Environ. Sci. Technol. 41, 2249–2254. Tomy, G.T., Thomas, C.R., Zidane, T.M., Murison, K.E., Pleskach, K., Hare, J., Arsenault, G., Marvin, C.H., Sverko, E., 2008. Examination of isomer specific bioaccumulation parameters and potential in vivo hepatic metabolites of syn- and antiDechlorane Plus isomers in juvenile rainbow trout (Oncorhynchus mykiss). Environ. Sci. Technol. 42, 5562–5567. Wang, J., Tian, M., Chen, S.J., Zheng, J., Luo, X.J., An, T.C., Mai, B.X., 2011. Dechlorane plus in house dust from e-waste recycling and urban areas in south china: sources, degradation, and human exposure. Environ. Toxicol. Chem. 30, 1965–1972. Wu, J.P., Zhang, Y., Luo, X.J., Wang, J., Chen, S.J., Guan, Y.T., Mai, B.X., 2010. Isomerspecific bioaccumulation and trophic transfer of dechlorane plus in the freshwater food web from a highly contaminated site, south china. Environ. Sci. Technol. 44, 606–611. Xian, Q., Siddique, S., Li, T., Feng, Y.-l., Takser, L., Zhu, J., 2011. Sources and environmental behavior of Dechlorane Plus—a review. Environ. Int. 37, 1273–1284. Yan, X., Zheng, J., Chen, K.H., Yang, J., Luo, X.J., Yu, L.H., Chen, S.J., Mai, B.X., Yang, Z.Y., 2012. Dechlorane Plus in serum from e-waste recycling workers: influence of gender and potential isomer-specific metabolism. Environ. Int. 49, 31–37. Yu, Z.Q., Lu, S.Y., Gao, S.T., Wang, J.Z., Li, H.R., Zeng, X.Y., Sheng, G.Y., Fu, J.M., 2010. Levels and isomer profiles of Dechlorane Plus in the surface soils from e-waste recycling areas and industrial areas in South China. Environ. Pollut. 158, 2920–2925. Zeng, Y.H., Yu, L.H., Luo, X.J., Chen, S.J., Wu, J.P., Mai, B.X., 2013. Tissue accumulation and species‐specific metabolism of technical pentabrominated diphenyl ether (DE‐71) in two predator fish. Environ. Toxicol. Chem. 32, 757–763. Zhang, Y., Wu, J.P., Luo, X.J., Wang, J., Chen, S.J., Mai, B.X., 2011. Tissue distribution of Dechlorane Plus and its dechlorinated analogs in contaminated fish: high affinity to the brain for anti-DP. Environ. Pollut. 159, 3647–3652. Zheng, J., Wang, J., Luo, X.J., Tian, M., He, L.Y., Yuan, J.G., Mai, B.X., Yang, Z.Y., 2010. Dechlorane Plus in human hair from an E-waste recycling area in South China: comparison with dust. Environ. Sci. Technol. 44, 9298–9303. Zhu, J., Feng, Y.L., Shoeib, M., 2007. Detection of dechlorane plus in residential indoor dust in the city of Ottawa, Canada. Environ. Sci. Technol. 41, 7694–7698.