Retention and Excretion of Polychlorinated Biphenyl Residues by Laying Hens G. F. FRIES, R. J. LILLIE, H. C. CECIL and J. BITMAN U.S. Department of Agriculture, Agricultural Research Service, Beltsville, Maryland 20705 (Received for publication December 7, 1976)
Poultry Science 56:1275-1280, 1977 INTRODUCTION Polychlorinated biphenyls (PCB) have been manufactured for more t h e n 4 0 years and primarily used in electrical applications. More recently, residues of these c o m p o u n d s were f o u n d widely spread in t h e e n v i r o n m e n t . Several severe incidents of PCB c o n t a m i n a t i o n occurred in which chickens exhibited serious toxicological effects and excessive PCB residues (Pichirallo, 1971). T h e source of PCBs, w h e n established, was c o n t a m i n a t e d feed caused by leakage of PCBs from heat exchange systems used in preparation of feedstuffs. This use of PCBs has s t o p p e d and t h e severe c o n t a m i n a t i o n incidents have n o t occurred for several years. However, t h e c o n t i n u e d m a n u f a c t u r e and use of PCBs present t h e possibility of future cont a m i n a t i o n . Even if t h e m a n u f a c t u r e of PCBs ceased, t h e r e w o u l d still be a threat because of a large fraction of t h e PCBs t h a t were manufact u r e d are still in use. T h e c o n t a m i n a t i o n incidents have led t o a n u m b e r of studies t o characterize toxicological effects a n d residue a c c u m u l a t i o n of PCBs in p o u l t r y . These studies have generally used only PCBs in a limited range of c h l o r i n a t i o n ; 4 2 %
1 Aroclors, Monsanto Chemical Co. Mention of a trade name does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture nor an endorsement by the Department over other products not mentioned.
(Harris and Rose, 1 9 7 2 ; Britton and Charles, 1 9 7 4 ) , 4 8 % ( S c o t t et al, 1 9 7 1 ; Rehfeld et al., 1972) and 54% (Platonow and F u n n e l , 1 9 7 2 ; P l a t o n o w et al, 1 9 7 3 ; T u m a s o n i s et al, 1 9 7 3 ; Teske et al., 1 9 7 4 ) . In contrast t o the n a r r o w percentage chlorine range of PCBs t h a t were studied, t h e r e are 12 d o m e s t i c commercial preparations in t h e U.S. t h a t vary in chlorine c o n t e n t f r o m 21 t o 6 8 % ' . Therefore, we carried out a comprehensive study t o d e t e r m i n e t h e effects of PCBs with widely varying chlorine c o n t e n t s o n hen laying performance, egg hatchability, progeny performance, and residue kinetics. I n f o r m a t i o n on hen p e r f o r m a n c e and e m b r y o n i c teratogenic effect have been presented previously, (Lillie et al, 1 9 7 4 , Cecil et al, 1 9 7 4 ) . T h e p u r p o s e of this paper is t o present data on PCB residue a c c u m u l a t i o n a n d elimination b y t h e hens used in t h e cited studies.
MATERIALS AND METHODS Twelve g r o u p s of 35 Single C o m b White Leghorn pullets, 4 1 weeks of age, were used. These birds were caged individually. A complete description of their feeding and managem e n t regime has been presented (Lillie et al, 1974). One of t h e g r o u p s served as a control receiving a s t a n d a r d breeder diet. Nine g r o u p s were fed various PCBs at either 2 or 20 p . p . m . as shown in Table 1. T h e o t h e r t w o groups were
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ABSTRACT Commercial polychlorinated biphenyl (PCB) mixtures with 21, 32, 42, 48, 54, and 68% chlorine were fed to caged White Leghorn hens at 20 p.p.m. for nine weeks. The 42, 48, and 54% mixtures were also fed at 2 p.p.m. Residues in egg at nine weeks were 0.7, 2.4, 14.4, 9.3, 11.4, and 21.5 p.p.m. for the 21, 32, 42, 48, 54, and 68% chlorine PCBs, respectively. Corresponding values in body fat were 24.0, 51.1, 89.3, 90.5, 124.5, and 52.4 p.p.m., respectively. Levels in eggs and body fat of groups fed 2 p.p.m. were approximately one-tenth the levels in the 20 p.p.m. group. After seven weeks of control feed, the values for the 20 p.p.m. groups were 0.7, 1.8, 2.8, 3.0, 7.2, and 1.8 for eggs and 8.1, 22.4, 26.4, 39.8, 91.7, and 43.6 for body fat. Estimated recovery of consumed PCBs increased from 8% to 55% as chlorine increased from 21 to 68%. Residues in excreta was about 10% of intake for all PCBs. Body fat retention was greater than egg elimination for 21 through 48% chlorine PCB, about the same for 54% and much less for the 68% chlorine PCB. The ratio of egg: fat residue in contaminated birds was .084:1.
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RESULTS AND DISCUSSION The PCBs eliminated in the excreta has about the same analog composition as material fed. Thus, as a general conclusion, it can be assumed that there were no differences in absorption of the various analogs. A few possible exceptions to this conclusion will be discussed later. Excreta levels of residue were erratic during the feeding period as illustrated by three representative PCBs in Fig. 1. In spite of the variability, it is reasonable to conclude that a "steady state" concentration was reached within one week. When feeding PCB stopped, residue concentrations immediately dropped to
20
40
60
80
100
DAYS
FIG. 1. Residue concentrations of representative PCBs in wet excreta of hens fed 20 p.p.m. PCB. Each point is a one-day composite of 10 hens.
negligible levels. The average level of excreta residues for all groups during the feeding period is presented in Table 1. The percent of intake that was eliminated ranged from 3.8% for Arochor 1221 to 10.8% for Arochor 1254. There was a trend towards higher levels of residues with increasing chlorination. However, the differences were not statistically significant because of the great week to week variation. It can be concluded that more than 90% of the PCBs are absorbed by the chickens as is common with other animal species (Albro and Fishbein, 1972). The two exceptions at the 2 p.p.m. intake of 1242 and 1248 are more apparent than real because of the difficultires in quantitating the low levels. The concentrations of PCB residue in eggs of all groups followed the same qualitative trends over time while the quantitative differences were related to degree of chlorination. Initially, the concentration rose rapidly and approached a "steady state" after 40 days. When PCB feeding stopped, there was rapid initial decline in concentration followed by a much slower decline after two weeks on clean feed. Concentration curves for two PCBs representing high and low chlorination are shown in Fig. 2. The quantitative differences among the PCBs are best illustrated by the concentrations reached at steady state and at the end of the study (Table 2). These values based on the individual samples collected at 9 and 16 weeks. Residues in the eggs of the hens fed 20 p.p.m. ranged from less than 1 p.p.m. with Aroclor 1221 to over 20 p.p.m. with Aroclor 1268 at 9 weeks (Table 2). The residue concen-
Downloaded from http://ps.oxfordjournals.org/ at UCSF Library on April 11, 2015
fed a polychlorinated terphenyl and a polybrominated biphenyl. Residues of the terphenyl were not determined and results of the polybrominated biphenyl group were presented separately (Fries et al., 1976). The groups were fed their respective diets for 9 weeks and all were given the control diets for the next 7 weeks. Samples of excreta, eggs, and body fat were obtained at various times from each group during the study. The excreta samples were composites of a one-day total collection from ten hens per group each week of the study. Composite egg samples were obtained weekly and consisted of 10 randomly selected eggs from each group. At the end of the PCB feeding period (9 weeks) and at the end of the complete experiment (16 weeks) five randomly selected hens from each group were sacrificed and body fat samples were obtained. The fat samples and the last laid eggs of these hens were analyzed individually. The samples were analyzed by routine methods in use in our laboratory (Fries et al., 1973). Florosil columns were used for cleanup. Identification and quantitation were by electron capture gas chromatography. The PCB fed to each group was used as a standard for samples from that group. The quantities were calculated by comparing the total area under all the peaks in the samples with the total area under all peaks in the standard. This method of quantitation is subject to some error because all chlorobiphenyls do not have the same electron capture response (Zitko, 1971). However, any error introduced would be systematic and would not change the major conclusions of the paper.
POLYCHLORINATED BIPHENYL RESIDUES
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TABLE 1. — Poly chlorinated biphenyl residues in the excreta of hens during the nine weeks PCBs were fed Excreta residues
Aroclora
Fed, p.p.m.
Level, p.p.m.
Amount, mg./day
%of intake
1221 1232 1242 1242 1248 1248 1254 1254 1268
20 20 2 20 2 20 2 20 20
0.76 1.12 0.42 1.69 0.61 1.68 0.16 2.12 1.67
0.10 0.14 0.05 0.20 0.07 0.22 0.02 0.26 0.22
3.8 5.4 20.8 8.5 28.4 9.4 8.1 10.8 8.7
trations increased with increased chlorine content of the PCB except for the hens receiving 20 p.p.m. of Arochor 1242. The cause of this anomalous residue level is unknown but could be related to reduced egg production without decreased feed intake by this group. When the hens were placed on clean feed, there was a rapid initial decline in PCB concentrations in eggs followed by a slower rate of decline (Fig. 2). When the values are plotted, the curves take the form of a sum of two exponentials. Rate constants were not estimated because of the small number of observations for each PCB and the short observation
-PCB10
/ "
-+-
Poultry Science 56:1275-1280, 1977 INTRODUCTION Polychlorinated biphenyls (PCB) have been manufactured for more t h e n 4 0 years and primarily used in electrical applications. More recently, residues of these c o m p o u n d s were f o u n d widely spread in t h e e n v i r o n m e n t . Several severe incidents of PCB c o n t a m i n a t i o n occurred in which chickens exhibited serious toxicological effects and excessive PCB residues (Pichirallo, 1971). T h e source of PCBs, w h e n established, was c o n t a m i n a t e d feed caused by leakage of PCBs from heat exchange systems used in preparation of feedstuffs. This use of PCBs has s t o p p e d and t h e severe c o n t a m i n a t i o n incidents have n o t occurred for several years. However, t h e c o n t i n u e d m a n u f a c t u r e and use of PCBs present t h e possibility of future cont a m i n a t i o n . Even if t h e m a n u f a c t u r e of PCBs ceased, t h e r e w o u l d still be a threat because of a large fraction of t h e PCBs t h a t were manufact u r e d are still in use. T h e c o n t a m i n a t i o n incidents have led t o a n u m b e r of studies t o characterize toxicological effects a n d residue a c c u m u l a t i o n of PCBs in p o u l t r y . These studies have generally used only PCBs in a limited range of c h l o r i n a t i o n ; 4 2 %
1 Aroclors, Monsanto Chemical Co. Mention of a trade name does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture nor an endorsement by the Department over other products not mentioned.
(Harris and Rose, 1 9 7 2 ; Britton and Charles, 1 9 7 4 ) , 4 8 % ( S c o t t et al, 1 9 7 1 ; Rehfeld et al., 1972) and 54% (Platonow and F u n n e l , 1 9 7 2 ; P l a t o n o w et al, 1 9 7 3 ; T u m a s o n i s et al, 1 9 7 3 ; Teske et al., 1 9 7 4 ) . In contrast t o the n a r r o w percentage chlorine range of PCBs t h a t were studied, t h e r e are 12 d o m e s t i c commercial preparations in t h e U.S. t h a t vary in chlorine c o n t e n t f r o m 21 t o 6 8 % ' . Therefore, we carried out a comprehensive study t o d e t e r m i n e t h e effects of PCBs with widely varying chlorine c o n t e n t s o n hen laying performance, egg hatchability, progeny performance, and residue kinetics. I n f o r m a t i o n on hen p e r f o r m a n c e and e m b r y o n i c teratogenic effect have been presented previously, (Lillie et al, 1 9 7 4 , Cecil et al, 1 9 7 4 ) . T h e p u r p o s e of this paper is t o present data on PCB residue a c c u m u l a t i o n a n d elimination b y t h e hens used in t h e cited studies.
MATERIALS AND METHODS Twelve g r o u p s of 35 Single C o m b White Leghorn pullets, 4 1 weeks of age, were used. These birds were caged individually. A complete description of their feeding and managem e n t regime has been presented (Lillie et al, 1974). One of t h e g r o u p s served as a control receiving a s t a n d a r d breeder diet. Nine g r o u p s were fed various PCBs at either 2 or 20 p . p . m . as shown in Table 1. T h e o t h e r t w o groups were
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ABSTRACT Commercial polychlorinated biphenyl (PCB) mixtures with 21, 32, 42, 48, 54, and 68% chlorine were fed to caged White Leghorn hens at 20 p.p.m. for nine weeks. The 42, 48, and 54% mixtures were also fed at 2 p.p.m. Residues in egg at nine weeks were 0.7, 2.4, 14.4, 9.3, 11.4, and 21.5 p.p.m. for the 21, 32, 42, 48, 54, and 68% chlorine PCBs, respectively. Corresponding values in body fat were 24.0, 51.1, 89.3, 90.5, 124.5, and 52.4 p.p.m., respectively. Levels in eggs and body fat of groups fed 2 p.p.m. were approximately one-tenth the levels in the 20 p.p.m. group. After seven weeks of control feed, the values for the 20 p.p.m. groups were 0.7, 1.8, 2.8, 3.0, 7.2, and 1.8 for eggs and 8.1, 22.4, 26.4, 39.8, 91.7, and 43.6 for body fat. Estimated recovery of consumed PCBs increased from 8% to 55% as chlorine increased from 21 to 68%. Residues in excreta was about 10% of intake for all PCBs. Body fat retention was greater than egg elimination for 21 through 48% chlorine PCB, about the same for 54% and much less for the 68% chlorine PCB. The ratio of egg: fat residue in contaminated birds was .084:1.
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RESULTS AND DISCUSSION The PCBs eliminated in the excreta has about the same analog composition as material fed. Thus, as a general conclusion, it can be assumed that there were no differences in absorption of the various analogs. A few possible exceptions to this conclusion will be discussed later. Excreta levels of residue were erratic during the feeding period as illustrated by three representative PCBs in Fig. 1. In spite of the variability, it is reasonable to conclude that a "steady state" concentration was reached within one week. When feeding PCB stopped, residue concentrations immediately dropped to
20
40
60
80
100
DAYS
FIG. 1. Residue concentrations of representative PCBs in wet excreta of hens fed 20 p.p.m. PCB. Each point is a one-day composite of 10 hens.
negligible levels. The average level of excreta residues for all groups during the feeding period is presented in Table 1. The percent of intake that was eliminated ranged from 3.8% for Arochor 1221 to 10.8% for Arochor 1254. There was a trend towards higher levels of residues with increasing chlorination. However, the differences were not statistically significant because of the great week to week variation. It can be concluded that more than 90% of the PCBs are absorbed by the chickens as is common with other animal species (Albro and Fishbein, 1972). The two exceptions at the 2 p.p.m. intake of 1242 and 1248 are more apparent than real because of the difficultires in quantitating the low levels. The concentrations of PCB residue in eggs of all groups followed the same qualitative trends over time while the quantitative differences were related to degree of chlorination. Initially, the concentration rose rapidly and approached a "steady state" after 40 days. When PCB feeding stopped, there was rapid initial decline in concentration followed by a much slower decline after two weeks on clean feed. Concentration curves for two PCBs representing high and low chlorination are shown in Fig. 2. The quantitative differences among the PCBs are best illustrated by the concentrations reached at steady state and at the end of the study (Table 2). These values based on the individual samples collected at 9 and 16 weeks. Residues in the eggs of the hens fed 20 p.p.m. ranged from less than 1 p.p.m. with Aroclor 1221 to over 20 p.p.m. with Aroclor 1268 at 9 weeks (Table 2). The residue concen-
Downloaded from http://ps.oxfordjournals.org/ at UCSF Library on April 11, 2015
fed a polychlorinated terphenyl and a polybrominated biphenyl. Residues of the terphenyl were not determined and results of the polybrominated biphenyl group were presented separately (Fries et al., 1976). The groups were fed their respective diets for 9 weeks and all were given the control diets for the next 7 weeks. Samples of excreta, eggs, and body fat were obtained at various times from each group during the study. The excreta samples were composites of a one-day total collection from ten hens per group each week of the study. Composite egg samples were obtained weekly and consisted of 10 randomly selected eggs from each group. At the end of the PCB feeding period (9 weeks) and at the end of the complete experiment (16 weeks) five randomly selected hens from each group were sacrificed and body fat samples were obtained. The fat samples and the last laid eggs of these hens were analyzed individually. The samples were analyzed by routine methods in use in our laboratory (Fries et al., 1973). Florosil columns were used for cleanup. Identification and quantitation were by electron capture gas chromatography. The PCB fed to each group was used as a standard for samples from that group. The quantities were calculated by comparing the total area under all the peaks in the samples with the total area under all peaks in the standard. This method of quantitation is subject to some error because all chlorobiphenyls do not have the same electron capture response (Zitko, 1971). However, any error introduced would be systematic and would not change the major conclusions of the paper.
POLYCHLORINATED BIPHENYL RESIDUES
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TABLE 1. — Poly chlorinated biphenyl residues in the excreta of hens during the nine weeks PCBs were fed Excreta residues
Aroclora
Fed, p.p.m.
Level, p.p.m.
Amount, mg./day
%of intake
1221 1232 1242 1242 1248 1248 1254 1254 1268
20 20 2 20 2 20 2 20 20
0.76 1.12 0.42 1.69 0.61 1.68 0.16 2.12 1.67
0.10 0.14 0.05 0.20 0.07 0.22 0.02 0.26 0.22
3.8 5.4 20.8 8.5 28.4 9.4 8.1 10.8 8.7
trations increased with increased chlorine content of the PCB except for the hens receiving 20 p.p.m. of Arochor 1242. The cause of this anomalous residue level is unknown but could be related to reduced egg production without decreased feed intake by this group. When the hens were placed on clean feed, there was a rapid initial decline in PCB concentrations in eggs followed by a slower rate of decline (Fig. 2). When the values are plotted, the curves take the form of a sum of two exponentials. Rate constants were not estimated because of the small number of observations for each PCB and the short observation
-PCB10
/ "
-+-
