Environ. Sci. Technol. 2003, 37, 1249-1255
Polychlorinated Biphenyls, Dibenzo-p-dioxins, Dibenzofurans, and p,p′-DDE in Livers of White-Tailed Sea Eagles from Eastern Germany, 1979-1998 K U R U N T H A C H A L A M K A N N A N , * ,† KURUNTHACHALAM SENTHIL KUMAR,‡ HARUHIKO NAKATA,† JERZY FALANDYSZ,§ GU ¨ NTER OEHME,| AND SHIGEKI MASUNAGA‡ Wadsworth Center, Empire State Plaza, P.O. Box 509, and Department of Environmental Toxicology and Health, State University of New York, Albany, New York 12202-0509, Graduate School of Environment and Information Sciences, Yokohama National University, Japan, Department of Environmental Chemistry and Ecotoxicology, University of Gdan ´ sk, Gdan ´ sk, Poland, and Martin Luther University of Halle-Wittenberg, Institute of Zoology, Halle, Germany
Concentrations of polychlorinated dibenzo-p-dioxins (PCDDs), dibenzofurans (PCDFs), polychlorinated biphenyls (including coplanar PCBs), and 1,1-dichloro-2,2-bis(pchlorophenyl)ethylene (p,p′-DDE) were measured in livers of white-tailed sea eagles collected during 1979 to 1998 from eastern Germany and western Poland. Concentrations of PCDDs, PCDFs, PCBs, and p,p′-DDE were, respectively, in the ranges of 4.2-341 pg/g, 3.3-651 pg/g, 64-89 400 ng/g, and 13-686 000 ng/g, wet wt. 23478-penta CDF was the predominant congener in the livers, accounting for, on average, 35% of the total PCDD/DF concentrations. The scatter plot of principal components analysis of the PCDD/DF congener profile in livers of white-tailed sea eagles showed a clustering of adult female sea eagles on the right side of the plot. The cluster position was driven by the positive loadings of 2378-TCDD, 23478-PeCDF, and 234678HxCDF and the negative loadings of 123789-HxCDD, 1234678HpCDD, OCDD, and 2378-TCDF on principal component 1, which suggested characteristic accumulation/elimination of PCDD/DF congeners in adult females. Concentrations of total PCBs were significantly correlated with those of PCDDs and/or PCDFs. Non-ortho coplanar PCBs accounted for 77% of the total TEQs. Chlorobiphenyl 126 (3,3′,4,4′,5pentaCB) accounted for 66% of the PCB-TEQs. PCDDs and PCDFs accounted for, on average, 19% of the total TEQs. Concentrations of PCDDs/DFs in livers of sea eagles collected prior to 1992 were not significantly different from those collected after 1992. * Corresponding author phone: (518)474-0015; fax: (518)473-2895; e-mail:
[email protected]. Corresponding address: Wadsworth Center, Empire State Plaza, P.O. Box 509, Albany, NY 122010509. † State University of New York. ‡ Yokohama National University. § University of Gdan ´ sk. || Martin Luther University of Halle-Wittenberg. 10.1021/es020942z CCC: $25.00 Published on Web 02/26/2003
2003 American Chemical Society
Introduction White-tailed sea eagles (Haliaeetus albicilla) are top predators in the aquatic food chain and feed mainly on fish, mammals, and other fish-eating birds. A reduction in the reproductive ability of white-tailed sea eagles was observed in countries surrounding the Baltic Sea between the early 1950s and the mid 1980s (1, 2). Studies have linked these reproductive problems to exposure to organochlorine pollutants such as DDT and its metabolite, p,p′-DDE, and to polychlorinated biphenyls (PCBs) (1, 3). Concentrations of DDT and PCBs in white-tailed sea eagles were among the highest reported for any organisms from the Baltic Sea (1, 4, 5). Notably high concentrations of PCBs and DDT have been reported in tissues of white-tailed sea eagle from the Swedish, Polish, Finnish, and East German coasts of the Baltic Sea (3, 6-9). Following the ban on the production and usage of DDT and PCBs in the late 1970s and the early 1980s, the reproductive success of white-tailed sea eagles increased, and 60-75% of the eggs produced hatchling (9). The population of sea eagles in Germany increased from a low of 120 pairs during the 1950s to a current population of 380 pairs (in 2001). Although several studies have reported PCB and DDT concentrations in white-tailed eagles from the Baltic countries (3, 6, 7, 9), prior to this study, no concentrations and trends of polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) in sea eagles from eastern Germany have been reported. PCDDs/DFs accounted for up to 50% of the TEQs in plasma, liver, and eggs of bald eagles collected from British Columbia, Canada (10-12). 2,3,7,8Tetrachlorodibenzo-p-dioxin equivalents (TEQs) contributed by PCDDs and PCDFs relative to PCBs in livers of fish-eating birds is less known. In this study, archived livers of white-tailed sea eagles collected from 1979 to 1998 in Brandenburg, MecklenburgVorpommern, and Sachsen (or Saxonia) in eastern Germany provided an opportunity to measure PCDDs, PCDFs, PCBs, and p,p′-DDE. The objectives of this study were (1) to determine concentrations and congener profiles of PCDDs and PCDFs and of total PCBs and p,p′-DDE in livers of whitetailed sea eagles, collected from 1979 to 1998, from inland and coastal regions of eastern Germany and western Poland; (2) to examine the trends in concentrations of these compounds in white-tailed sea eagles, collected over a period of 20 years; and (3) to estimate TEQs so as to elucidate the magnitude of dioxin-like toxicity contributed by PCBs, PCDDs, and PCDFs.
Materials and Methods Livers of 24 white-tailed sea eagles collected from 1979 to 1998 in Brandenburg, Mecklenburg-Vorpommern, and Sachsen in eastern Germany (Figure 1) were analyzed. All samples from Sachsen were collected east of the Elbe River. Similarly, livers of four white-tailed sea eagles collected during 1996 to 1999 from western Poland were analyzed for comparison. All of the sea eagles were found dead or dying. Liver samples from freshly dead birds were wrapped in aluminum foil or put in glass jars cleaned with solvents and were stored frozen at -20 °C until analysis. Body weight, age category, sex, and date and location of collection are shown in Table 1. The samples analyzed were from 18 females, 9 males, and 1 bird for which the sex was not determined. Liver tissues were analyzed for PCDDs, PCDFs, PCBs, and p,p′-DDE following the methods described earlier (13-15). Approximately 3-5 g of liver tissue were homogenized with anhydrous sodium sulfate and extracted in a Soxhlet apVOL. 37, NO. 7, 2003 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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TABLE 1. Concentrations of 2,3,7,8-Substituted PCDDs, PCDFs (pg/g, wet wt), Total PCBs, and p,p′-DDE (ng/g, wet wt) in White-Tailed Sea Eagle Livers from Eastern Germany and Western Poland, 1979-1998a sample i.d.
date
location
biometry
fat %
ΣPCDD pg/g
ΣPCDF pg/g
ΣPCBs ng/g
p,p′-DDE ng/g
Brandenburg 6640g,im,F 5700g,ad,F 4200g,ad,F 4374g,juv,M 4425g,ad,M 5640g,ad,F 2970g,ad,M 4750g,Ju,F 4300g,ad,ND
5.79 5.95 5.01 5.95 4.82 4.9 4.41 6.82 4.83
63.6 91 341 8.4 7.4 60 90.7 4.8 103
113 120 651 5.3 12 93.1 130 7.8 153
4830 10100 89400 305 2820 12200 29400 202 41500
3760 15700 686000 98 4430 15300 88400 241 61100
Mecklenburg-Vorpommern 4325g,ad,F 5.69 5517g,ad,F 4.41 4830g,juv,F 6.82 4105g,ad,M 4.4 juv,M 7.92 3805g,juv,F 5.5 4280g,ad,M 7.03 4505g,ad,F 4.17 3620g,ad,F 4.75 4225g,ad,M 4.83 6500g,ad,F 5.37
55.3 89.5 6 66.2 5.9 25 11.1 72.7 171 8.3 26.6
60.2 69.9 8.7 52.4 10.9 38.9 19.3 101 186 11.6 40.6
7000 4360 313 10300 340 7800 6000 11500 25800 340 4040
19100 8370 325 22500 353 1090 2170 28600 44350 330 866
136 107 23 7.4
179 100 47.7 5.2
37400 10000 1810 220
23600 11400 2790 184
217 64 197 3240
277 13 219 993
M79/5 M79/6 M79/8 M87/18 M90/3 M90/4 M90/7 BB93/19 BB94/2
17/4/1979 2/5/1979 12/6/1979 4/11/1987 28/3/1990 12/4/1990 15/8/1990 1/10/1993 15/1/1994
Waldlewitz Muritz Stegelitz Dassow Babke Kotzow Melzower Forst Wilmersdorf Thomsdorf
MV87/21 MV91/10 MV91/12 MV91/8 MV92/2 MV92/3 MV92/9 MV93/2 MV94/11 MV94/19 MV95/5
29/3/1987 6/5/1991 25/6/1991 6/4/1991 5/1/1992 6/2/1992 28/4/1992 15/2/1993 10/4/1994 17/5/1994 21/2/1995
Dabelow Julchendorf Bossow Bergfeld Lahnvitz Lewitz Torgelow Muritz Dobbin Karow Kolpinsee
SA93/17 S96/7 S97/22 S98/18
4/9/1993 4/3/1996 28/8/1997 28/7/1998
Stackelitz Niederspree Ebersbach-Rodern Klein Krauscha
Sachsen 5420g,im,F 3400g,im,F 3800g,ad,M 4920g,im,F
7.72 4.44 4.82 9.9
PON4 PON7 PON1 PON2
1996 1997 1998-1 1998-2
western Poland western Poland western Poland western Poland
Western Poland F ad,M ad,F F
4.84 5.94 2.86 9.26
a
8.25 4.19 5.46 24.7
8.73 3.29 13.6 32.7
ju ) juvenile (1 year), im ) immature (2-4 yr), ad ) adult (5 yrs and above), F ) female, M ) male, ND ) not determined.
FIGURE 1. Map of Germany showing sampling areas of whitetailed sea eagles. paratus for 16 h using dichloromethane and hexane (400 mL; 3:1). An aliquot of the concentrated extract was used for a gravimetric estimation of fat content. Seventeen 2,3,7,8substituted 13C-labeled tetra-CDD through octa-CDD and CDF congeners, and 12 dioxin-like PCBs (IUPAC Nos. 81, 77, 126, 169, 105, 114, 118, 123, 156, 157, 167, 189), were spiked as surrogate standards (Cambridge Isotope Laboratories, Andover, MA). Aliquots were then treated with sulfuric acid in a separatory funnel. The solution was concentrated to 2 mL and was sequentially subjected to silica gel, alumina, and silica gel-impregnated-active carbon chromatography. Details of the column chromatography procedures have been described elsewhere (13). PCB congeners other than nonortho congeners were separated from PCDDs/DFs and were analyzed by high-resolution gas chromatograph interfaced with a high-resolution mass spectrometer (HRGC-HRMS). 1250
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Identification and Quantification. Identification and quantification of 2,3,7,8-substituted PCDD/DF congeners and dioxin-like PCBs (non-ortho and mono-ortho-substituted congeners) were performed with a Hewlett-Packard 6890 Series gas chromatograph interfaced with a Micromass Autospec - Ultima high-resolution mass spectrometer (HRMS). The HRMS was operated in an electron impact (EI) and selected ion monitoring (SIM) mode at a resolution R >10 000 (10% valley). Separation was achieved with DB-5 (0.25 mm i.d. × 60 m; J&W Scientific, Folsom, CA) and DB-17 capillary (0.25 mm i.d. × 60 m; J&W Scientific) columns. The column oven temperature was programmed from an initial temperature of 160 °C to a final temperature of 310 °C (total run time, 60 min). Recoveries of 17 2,3,7,8-substituted 13Clabeled PCDDs/DFs ranged from 57 to 95% (mean: 78%). Reported concentrations were not corrected for the recoveries of surrogate standards. Total PCBs refers to the sum of all the identified PCB congeners, whereas total PCDD and PCDF refers to the sum of 2,3,7,8-substituted congeners. Data were analyzed statistically by Student’s t-test and principal components analysis (PCA). PCA transforms original variables into their linear combinations, which are uncorrelated to one another and which account for decreasing proportions of the data variance. PCA was performed using a statistical software supplied by Esumi Co. Ltd. (Tokyo, Japan).
Results and Discussion PCDDs and PCDFs. Concentrations of seven 2,3,7,8substituted PCDD and 10 PCDF congeners in 28 white-tailed sea eagles, collected over a period of 20 years, ranged from 4.2 to 341 and from 3.3 to 651 pg/g, wet wt, respectively (Table 1). The greatest concentrations, 341 pg/g, wet wt, for
FIGURE 2. Percentage composition of PCDD and PCDF congeners in white-tailed sea eagle livers collected from eastern Germany and western Poland between 1979 and 1998. PCDDs and 651 pg/g, wet wt, for PCDFs were both found in the liver of an adult female eagle collected in 1979 from Stegelitz, Brandenburg. This individual was found in emaciated condition. However, lipid content in the liver of this bird (5.01%) was comparable to content for other birds in relatively good body condition. The same individual contained the greatest liver concentrations of total PCBs and p,p′-DDE in liver. Five of the 28 individuals analyzedsM79/ 8, M90/7, MV91/8, MV92/3, and MV94/11swere in emaciated condition. Mean ((SD) lipid content in livers of sea eagles analyzed in this study was 5.7 ( 1.4%. The lipid content in livers of the emaciated individuals were (4.4-5.5%) similar to those found at good condition. However, concentrations of PCDDs, PCDFs, PCBs, and p,p′-DDE were generally elevated in emaciated individuals. Mean lipid-normalized concentration (2450 pg/g, lipid wt) of total PCDDs/DFs in livers of white-tailed sea eagles from eastern Germany were approximately 10-20-fold greater than the lipid-normalized concentrations in muscle of white-tailed sea eagles from Finland (16). Concentrations of PCDDs/DFs in white-tailed sea eagle livers were similar to those observed in livers of dead bald eagles (23-4500 pg/ g, wet wt) from Michigan, U.S.A. (17) and British Columbia, Canada (39-6000 pg/g, wet wt) (11). The mean concentration of PCDFs was approximately 1.4-fold greater than the concentrations of PCDDs in most of the sea eagle livers (Table 3, Supporting Information). Among PCDFs, 23478-penta CDF was the predominant congener in almost all livers, accounting for, on average, 35% of the total PCDD/DF concentrations (Figure 2). This pattern was similar to what was measured in breast muscle of white-tailed sea eagles from Finland (6, 16). 23478-PeCDF was the predominant congener in livers of bald eagles from the North American Great Lakes (17). This congener has been shown to preferentially accumulate in liver tissues of several fish-eating water birds (17-20). High concentrations of 23478PeCDF in bird tissues have been attributed to exposure to technical PCB mixtures, which contain this isomer as a major impurity (21). Technical PCB mixtures such as Aroclors and Kanechlors also contain 2,3,7,8-TCDF as a major impurity. However, concentrations of 2,3,7,8-TCDF were relatively lower than those of 23478-PeCDF. Among PCDDs, 2378TCDD, 12378-pentaCDD, and 123678-HexaCDD were the major congeners found in the sea eagle livers (Table 3, Supporting Information). High concentrations of these three PCDD congeners in bald eagles have been associated with exposure to pentachlorophenol originating from pulp mills (22). The proportions of individual PCDD/DF congeners relative to total PCDD/DF concentrations in white-tailed sea eagle livers were examined using PCA to delineate potential patterns in the data. Since the samples analyzed in this study
included several variables, PCA was used to transform a number of correlated variables into a smaller number of uncorrelated variables. The first principal component accounts for as much of the variability in the data as possible, and each succeeding component accounts for as much of the remaining variability as possible. For PCA, PCDD/DF congener concentrations were normalized to percentages of the total PCDD/DF concentrations. The contributions of PC1, PC2, PC3, and PC4 to the total variance were 26.3, 22, 11.6, and 9.3%, respectively (Table 4, Supporting Information). The scores for the first and second principal components for each sample have been plotted (Figure 3). The first principal component, PC1, had high eigenvalues for congeners 2378TCDD, 23478-PeCDF, and 234678-HxCDF, whereas eigenvalues were low (negative) for 2378-TCDF and for the more highly chlorinated dibenzo-p-dioxins such as 123789-HxCDD, 1234678-HpCDD, and OCDD. The second principal component, PC2, had positive eigenvalues for congeners 123678HxCDD and for highly chlorinated PCDFs such as 123678HxCDF, 1234678- and 1234789-HpCDFs, and OCDF, but a negative value for 2378-TCDF (Table 4, Supporting Information). The scatter plot of the component scores showed a clustering of adult female sea eagles on the right side of the plot. The cluster position was driven by the positive loadings of 2378-TCDD, 23478-PeCDF, and 234678-HxCDF and the negative loadings of 123789-HxCDD, 1234678-HpCDD, OCDD, and 2378-TCDF on PC1. Eight of 10 individuals in this cluster were adult females. This cluster suggests a specific pattern of accumulation of PCDD/DF congeners in adult females; i.e., 2378-TCDD, 23478-PeCDF, and 234678-HxCDF were enriched in livers of adult female livers, whereas highly chlorinated PCDD congeners and 2378-TCDF were depleted relative to those in males and/or juveniles. Tissue-specific distribution of PCDD/DF congeners in cormorants has been shown (18, 19). In general, several of the PCDD congeners tended to be prevalent in blood, while PCDF congeners were relatively enriched in liver (18). Greater proportions of more highly chlorinated PCDDs were reported in cormorant eggs than in livers (19, 23). Highly chlorinated PCDD congeners, such as 123789-HxCDD, 1234678-HpCDD, and OCDD, may be transferred to eggs during oogenesis in females. 2378TCDF had negative loadings both for PC1 and PC2, suggesting faster elimination due to metabolism/transfer of this congener by adult females. 2,3,7,8-TCDF has been shown to be metabolized and excreted more rapidly than 2,3,7,8-TCDD (24). Since the number of samples analyzed for individual years was small, samples were grouped to represent different periods (e.g., pre-1992 versus post-1992) for statistical analysis. Concentrations were normalized to lipid content for comparisons. Concentrations of total PCDDs/DFs in livers of sea eagles collected prior to 1992 were not significantly different from those collected after 1992 (p >0.05) (Figure 4). No significant difference in total PCDD/DF concentrations was observed in adult females collected from 1987 to 1995. However, such differences could be confounded by biological and spatial variations. Small sample size and associated biological variations preclude a more extensive analysis of the data. It is also worth mentioning that adult white-tailed sea eagles on the Baltic coast are largely stationary in terms of breeding area; therefore, their contamination should reflect that of the local area. PCBs and DDE. Concentrations of PCBs and p,p′-DDE in livers of white-tailed sea eagles ranged from 64 to 89 400 and from 13 to 686 000 ng/g, wet wt, respectively (Table 1). PCB concentrations greater than 2000 ng/g, wet wt, were found in 18 of the 28 individuals analyzed. Concentrations of total PCBs in sea eagle livers collected from Poland in 1987 (1900 to 540 000 ng/g, wet wt) (3, 25) were similar to those found in sea eagles from eastern Germany in our study. A p,p′-DDE VOL. 37, NO. 7, 2003 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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1251
1252
TABLE 2. Concentrations of 2,3,7,8-TCDD Equivalents (TEQs; pg/g, wet wt; based on WHO-Bird TEFs) in White-Tailed Sea Eagle Liversa
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Brandenberg congener
WHO-TEF
M79/5
M79/6
M79/8
M87/18
M90/3
Mecklenburg-Vorpommern M90/4
M90/7
BB93/19
BB94/2
MV87/21
MV91/10
MV91/12
MV91/8
62 284 527 2.1
0.11 3.72 4.42 0.01
67 230 623 2.7
34.8 65.2 310 0.6
4.3 54.2 135 0.32
0.2 4.6 10.2 0.02
5.3 29.6 217 0.73
21 14.4 13.6
0.16 0.07 0.17
13 15 3.2
8.4 4.7 3.7
6.8 3.4 2
0.59 0.21 0.41
9.9 3.8 2.1
81 77 126 169
0.1 0.05 0.1 0.001
27 157 131 0.3
44 67 373 1
450 531 1120 9.5
0.7 5.6 3.5 0.01
Non-ortho PCBs 6 23.9 25.2 75.9 62.2 249 0.11 0.7
105 118 156
0.0001 0.00001 0.0001
6 2.5 1.2
7.2 6.3 3.9
49 51.2 23
0.41 0.12 0.12
2.6 1.3 0.56
2,3,7,8-D 1,2,3,7,8-D 1,2,3,4,7,8-D 1,2,3,6,7,8-D 1,2,3,7,8,9-D 1,2,3,4,6,7,8-D OCDD
1 1 0.05 0.01 0.1 0.001 0.0001
13.2 20.1 0.259 0.171 0.02 0.004
