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Research Note: Effects of Dietary Butylated Hydroxyanisole and Cysteine on Toxicity of Lathyrus odoratus to Broiler and Japanese Quail Chicks1
Research Note: Effects of Dietary Butylated Hydroxyanisole and Cysteine on Toxicity of Lathyrus odoratus to Broiler and Japanese Quail Chicks 1 Y. C. RAHARJO,2 P. R. CHEEKE,2 and G. H. ARSCOTT3 Departments of Animal and Poultry Science, Oregon State University, Corvalis, Oregon 97331 (Received for publication January 15, 1987) ABSTRACT The effect of .75% dietary butylated hydroxyanisole (BHA) and 1% cysteine on the toxicity of Lathyrus odoratus seed to Japanese quail and broiler chicks was examined. In both species, the feeding of Lathyrus seed as a component of a complete diet depressed (P<.05) body weight gain and feed intake. Typical signs of lathyrism, including ruffled feathers, enlarged hocks, curled toes, ataxia, leg paralysis, and mortality, were observed. Neither BHA nor cysteine exerted protective effects against the lathyrogenic effects. As assessed by mortality, these additives appeared instead to potentiate Lathyrus toxicity. (Key words: lathyrism, butylated hydroxyanisole, toxicity, cysteine) 1988 Poultry Science 67:153-155 INTRODUCTION
Osteolathyrism, characterized by spastic paralysis of the legs and skeletal deformities in humans and animals, is caused by the consumption of seeds of various Lathyrus species (Padmanaban, 1980). The toxic compounds are (3amino propionitrile (BAPN) and its homologues. Lathyrism has been an important public health problem in India, where the crop has been extensively grown because of its ability to thrive under adverse conditions when other crops may fail. In previous work in our laboratory, dietary cysteine and synthetic antioxidants such as ethoxyquin and butylated hydroxyanisole (BHA) have been found to reduce the toxicity of pyrrolizidine alkaloids (Miranda et al., 1981a,b; 1982). The objective of this study was to determine if these additives have similar protective activity against the toxicity of lathyrus odoratus seeds. MATERIALS AND METHODS
One experiment with Japanese quail chicks and two with broiler chicks (Hubbard strain)
'Oregon Agricultural Experiment Technical Paper No. 8089. 2 Animal Science Department. 3 Poultry Science Department.
153
were conducted. Composition of diets is given for quail (Table 1) and broilers (Table 2). In Experiment 1,11 Japanese quail chicks (1 day of age) were randomly assigned to each of the treatment groups, consisting of control, 46.3% Lathyrus seed (LS), LS + .75% BHA, and LS + 1% cysteine. The LS seed replaced dietary soybean meal. Birds on each treatment were penned as a group in one cage and fed the diets ad libitum for 14 days. In Experiment 2, three replicates of five birds/replicate were assigned to each treatment. One-day-old broiler chicks were used. Treatments included: 1) 100% basal diet (Table 2); 2) 80% basal diet, 20% LS; 3) 79.25% basal diet, 20% LS, .75% BHA; and 4) 79% basal diet, 20% LS and 1% cysteine. The test period was 21 days. In Experiment 3, there were three replicates, each with five broiler chicks (14 days of age). Treatments were as in Experiment 2, except that 10% LS was used. The test period was 21 days. The true metabolizable energy (TME) value of LS, determined according to the method of Sibbald (1982) using three adult roosters, was 3,321 kcal/kg on a dry weight basis. Crude protein content was 25.31% on a dry weight basis. Diets were formulated to be isonitrogenous and isocaloric. Feed and water were provided ad libitum. Body weights and feed intake were measured weekly. Data were statistically analyzed using one-way analysis of variance and least significant difference (Steel and Tome, 1980).
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TABLE 1. Composition of Japanese quail diets (Experiment 1) Diet (%) Lathyrus Basal
Ingredients Yellow corn Soybean meal (44% CP 2 ) Herring meal (60% CP) Ground LS Safflower oil Dicalcium phosphate Vitamin-mineral premix 3 DL-Methionine BHA
Cysteine Total Calculated nutrient composition: Metabolizable energy, kcal/kg CP, % Sulfur amino acids, % 1
BHA = butylated hydroxyanisole.
2
CP = Crude protein.
seed (LS)
57.00 30.00 8.00
LS + Cysteine
LS + BHA 1
37.00
36.00
.00
.00
.00
1.75 3.00
10.20 46.30 3.00 3.00
10.30 46.30 3.15 3.00
8.80 46.30 3.40 3.00
.25 .00 .00 .00
.25 .25 .00 .00
.25 .25 .75 .00
.25 .25 .00
1.00
100.00
100.00
100.00
100.00
3,015 24.00
3,025 24.05
3,007 24.07
.83
.83
.83
3,018 24.05 1.79
.00
37.00
'Supplied per kilogram of diet: Ca, 97.5 mg; Mn, 60 mg; Fe, 20 mg; Cu, 2 mg; I, 1.2 mg; Zn, 27.5 mg; vitamin A, 3,304 IU; vitamin D 3 , 1,111 ICU; riboflavin, 3.3 mg; d-pantothenic acid, 5.51 mg; niacin, 20 mg; choline, 191 mg; vitamin B 1 2 , 5.51 /ug; vitamin E, 1.1 IU; vitamin K, .55 mg; folacin, .22 mg.
RESULTS AND DISCUSSION
All birds fed diets containing LS developed typical signs of lathyrism, including ruffled feathers, enlarged hock joints, curled toes, ataxia, leg paralysis, and death (Padmanaban, 1980). Signs of lathyrism were evident within 7 days. Feed intake and body weight gain of both quail and broilers were adversely affected by inclusion of LS (Table 3). In the quail, fed 46% LS, feed intake and weight gain were depressed by 70% and 58% compared with values for birds fed the basal diet, and mortality was 27% (Table 3). Inclusion of 20% LS decreased feed intake and weight gain of chickens by approximately 70% and 82%, respectively, and resulted in 57% mortality (Experiment 2). In Experiment 3, with 10% LS and older chicks, growth depression, severity of clinical signs, and mortality were lower than in Experiment 2. Cysteine and BHA did not have protective activity against LS toxicity; both additives actually increased mortality (Table 3). We have no explanation for why cysteine produced an adverse effect. Thornton et al. (1962) found that the antioxidant ascorbic acid increased the
TABLE 2. Composition of the basal chick diet (Experiments 2 and 3) Ingredients Yellow corn Soybean meal (44% CP 1 ) Herring meal (60% CP) Corn oil Limestone Defluorinated rock phosphate Salt, iodized Vitamin-mineral premix 2 DL-Methionine
(%) 58.00 30.00 5.00 4.20 .60
1.50 .25 .30 .15
Calculated composition: Metabolizable energy, kcal/kg CP, % Crude fiber, % Calcium, % Phosphorus, %
3,223 23.00 2.33 1.02 .72
'CP = Crude Protein. 2
Supplied per kilogram of diet: Ca, 97.5 mg; Mn, 60 mg; Fe, 20 mg; Cu, 2 mg; I, 1.2 mg;Zn, 27.5 mg; vitamin A, 3,304 IU; vitamin D 3 , 1,111 ICU; riboflavin, 3.3 mg; d-pantothenic acid, 5.51 mg; niacin, 20 mg; choline, 191 mg; vitamin B u , 5.51 Mg; vitamin E, 1.1 IU; vitamin K, .55 mg; folacin, .22 mg.
RESEARCH NOTE
155
TABLE 3. Performance of birds fed diets containing Lathyrus odoratus-seeds (LS) with and without butylated hydroxyanisole (BHA) and cysteine1
Experiment
Treatment
1
Basal 46.3% LS 46.3% LS + BHA 46.3% LS + cysteine SE 2
2
Basal 28% LS 20% LS + BHA 20% LS + cysteine SE
3
Basal 10% LS 10% LS + BHA 10% LS + cysteine SE
Body weight gain
Total feed intake
Feed: Gain
No. survivors/ no. started
87 40 65 59
(g/g) 2.57 4.36 4.52 4.33
10/11 8/11 6/11 4/11
437 129 102 120 7.5
1.58 a 2.27° 2.41 b 1.96 c .104
15/15 8/15 6/15 7/15
1,525 1,077 1,080 1,123 40.9
1.93 a 2.30 b 2.57 b 2.37 b .178
15/15 15/15 13/15 12/15
-(g/bird) 41.8 a 17.4 b 22.5 b 21.7 b 2.96 277 a 5 7 b,c 41c 62b 6.3 794a 469 b 436 b 468b 42.7
Means within columns and experiments followed by different superscripts are significantly different (P<.05). 1
Japanese quail chicks were used in Experiment 1 and broiler chicks in Experiments 2 and 3.
2
Standard error.
mortality of chickens fed a diet containing BAPN. They suggested that ascorbic acid may inhibit the activity of monoamine oxidase, an enzyme that detoxifies BAPN to cyanoamine acetate (Roy, 1981). It is possible that BHA could have a similar action, and thus intensify the toxicity of lathyrogens. In summary, these results indicate that cysteine and BHA, which have protective effects against alkaloid toxicity, do not have a similar effect with lathyrogens, and may even potentiate the toxicity of Lathyrus to avian species. REFERENCES Miranda, C. L., D. R. Buhler, H. S. Ramsdell, P. R. Cheeke and J. A. Schmitz. 1982. Modification of chronic hepatotoxicity of pyrrolizidine (Senecio) alkaloids by BHA and cysteine. Toxicol. Lett. 10:177182. Miranda, C. L., H. M. Carpenter, P. R. Cheeke, and D. R. Buhler. 1981a. Effect of ethoxyquin on the toxicity
of the pyrrolizidine alkaloid monocrotaline and on hepatic drug metabolism in mice. Chem. Biol. Interact. 37:95-107. Miranda, C. L., R. L. Reed, P. R. Cheeke, and D. R. Buhler. 1981b. Protective effects of butylated hydroxyanisole against the acute toxicity of monocrotaline in mice. Toxicol. Appl. Pharmacol. 59:424430. Padmanaban, G. 1980. Lathyrogens. Pages 239-263 in: Toxic Constituents of Plant Foodstuffs. I.E. Liener, ed. Academic Press, New York, NY. Roy, D. N. 1981. Toxic amino acids and proteins from Lathyrus plants and other leguminous species. A literature review. Nutr. Abstr. Rev. 51:691-707. Sibbald, I. R. 1982. Measurement of bioavailable energy in poultry feeding stuffs: a review. Can. J. Anim. Sci. 62:983-1048. Steel, R.G.D. and J. H. Torrie. 1980. Principles and Procedures of Statistics. A Biomedical Approach. 2nd ed. McGraw Hill Book Co., New York, NY. Thornton, P. A., D. Brownrigg, and F. B. Mather. 1962. The influence of dietary ascorbic acid and the severity of p-amino propionitrile (BAPN) toxicity in turkeys. Poultry Sci. 41:1586-1590.
Osteolathyrism, characterized by spastic paralysis of the legs and skeletal deformities in humans and animals, is caused by the consumption of seeds of various Lathyrus species (Padmanaban, 1980). The toxic compounds are (3amino propionitrile (BAPN) and its homologues. Lathyrism has been an important public health problem in India, where the crop has been extensively grown because of its ability to thrive under adverse conditions when other crops may fail. In previous work in our laboratory, dietary cysteine and synthetic antioxidants such as ethoxyquin and butylated hydroxyanisole (BHA) have been found to reduce the toxicity of pyrrolizidine alkaloids (Miranda et al., 1981a,b; 1982). The objective of this study was to determine if these additives have similar protective activity against the toxicity of lathyrus odoratus seeds. MATERIALS AND METHODS
One experiment with Japanese quail chicks and two with broiler chicks (Hubbard strain)
'Oregon Agricultural Experiment Technical Paper No. 8089. 2 Animal Science Department. 3 Poultry Science Department.
153
were conducted. Composition of diets is given for quail (Table 1) and broilers (Table 2). In Experiment 1,11 Japanese quail chicks (1 day of age) were randomly assigned to each of the treatment groups, consisting of control, 46.3% Lathyrus seed (LS), LS + .75% BHA, and LS + 1% cysteine. The LS seed replaced dietary soybean meal. Birds on each treatment were penned as a group in one cage and fed the diets ad libitum for 14 days. In Experiment 2, three replicates of five birds/replicate were assigned to each treatment. One-day-old broiler chicks were used. Treatments included: 1) 100% basal diet (Table 2); 2) 80% basal diet, 20% LS; 3) 79.25% basal diet, 20% LS, .75% BHA; and 4) 79% basal diet, 20% LS and 1% cysteine. The test period was 21 days. In Experiment 3, there were three replicates, each with five broiler chicks (14 days of age). Treatments were as in Experiment 2, except that 10% LS was used. The test period was 21 days. The true metabolizable energy (TME) value of LS, determined according to the method of Sibbald (1982) using three adult roosters, was 3,321 kcal/kg on a dry weight basis. Crude protein content was 25.31% on a dry weight basis. Diets were formulated to be isonitrogenous and isocaloric. Feed and water were provided ad libitum. Body weights and feed intake were measured weekly. Data were statistically analyzed using one-way analysis of variance and least significant difference (Steel and Tome, 1980).
RAHARJO ET AL.
154
TABLE 1. Composition of Japanese quail diets (Experiment 1) Diet (%) Lathyrus Basal
Ingredients Yellow corn Soybean meal (44% CP 2 ) Herring meal (60% CP) Ground LS Safflower oil Dicalcium phosphate Vitamin-mineral premix 3 DL-Methionine BHA
Cysteine Total Calculated nutrient composition: Metabolizable energy, kcal/kg CP, % Sulfur amino acids, % 1
BHA = butylated hydroxyanisole.
2
CP = Crude protein.
seed (LS)
57.00 30.00 8.00
LS + Cysteine
LS + BHA 1
37.00
36.00
.00
.00
.00
1.75 3.00
10.20 46.30 3.00 3.00
10.30 46.30 3.15 3.00
8.80 46.30 3.40 3.00
.25 .00 .00 .00
.25 .25 .00 .00
.25 .25 .75 .00
.25 .25 .00
1.00
100.00
100.00
100.00
100.00
3,015 24.00
3,025 24.05
3,007 24.07
.83
.83
.83
3,018 24.05 1.79
.00
37.00
'Supplied per kilogram of diet: Ca, 97.5 mg; Mn, 60 mg; Fe, 20 mg; Cu, 2 mg; I, 1.2 mg; Zn, 27.5 mg; vitamin A, 3,304 IU; vitamin D 3 , 1,111 ICU; riboflavin, 3.3 mg; d-pantothenic acid, 5.51 mg; niacin, 20 mg; choline, 191 mg; vitamin B 1 2 , 5.51 /ug; vitamin E, 1.1 IU; vitamin K, .55 mg; folacin, .22 mg.
RESULTS AND DISCUSSION
All birds fed diets containing LS developed typical signs of lathyrism, including ruffled feathers, enlarged hock joints, curled toes, ataxia, leg paralysis, and death (Padmanaban, 1980). Signs of lathyrism were evident within 7 days. Feed intake and body weight gain of both quail and broilers were adversely affected by inclusion of LS (Table 3). In the quail, fed 46% LS, feed intake and weight gain were depressed by 70% and 58% compared with values for birds fed the basal diet, and mortality was 27% (Table 3). Inclusion of 20% LS decreased feed intake and weight gain of chickens by approximately 70% and 82%, respectively, and resulted in 57% mortality (Experiment 2). In Experiment 3, with 10% LS and older chicks, growth depression, severity of clinical signs, and mortality were lower than in Experiment 2. Cysteine and BHA did not have protective activity against LS toxicity; both additives actually increased mortality (Table 3). We have no explanation for why cysteine produced an adverse effect. Thornton et al. (1962) found that the antioxidant ascorbic acid increased the
TABLE 2. Composition of the basal chick diet (Experiments 2 and 3) Ingredients Yellow corn Soybean meal (44% CP 1 ) Herring meal (60% CP) Corn oil Limestone Defluorinated rock phosphate Salt, iodized Vitamin-mineral premix 2 DL-Methionine
(%) 58.00 30.00 5.00 4.20 .60
1.50 .25 .30 .15
Calculated composition: Metabolizable energy, kcal/kg CP, % Crude fiber, % Calcium, % Phosphorus, %
3,223 23.00 2.33 1.02 .72
'CP = Crude Protein. 2
Supplied per kilogram of diet: Ca, 97.5 mg; Mn, 60 mg; Fe, 20 mg; Cu, 2 mg; I, 1.2 mg;Zn, 27.5 mg; vitamin A, 3,304 IU; vitamin D 3 , 1,111 ICU; riboflavin, 3.3 mg; d-pantothenic acid, 5.51 mg; niacin, 20 mg; choline, 191 mg; vitamin B u , 5.51 Mg; vitamin E, 1.1 IU; vitamin K, .55 mg; folacin, .22 mg.
RESEARCH NOTE
155
TABLE 3. Performance of birds fed diets containing Lathyrus odoratus-seeds (LS) with and without butylated hydroxyanisole (BHA) and cysteine1
Experiment
Treatment
1
Basal 46.3% LS 46.3% LS + BHA 46.3% LS + cysteine SE 2
2
Basal 28% LS 20% LS + BHA 20% LS + cysteine SE
3
Basal 10% LS 10% LS + BHA 10% LS + cysteine SE
Body weight gain
Total feed intake
Feed: Gain
No. survivors/ no. started
87 40 65 59
(g/g) 2.57 4.36 4.52 4.33
10/11 8/11 6/11 4/11
437 129 102 120 7.5
1.58 a 2.27° 2.41 b 1.96 c .104
15/15 8/15 6/15 7/15
1,525 1,077 1,080 1,123 40.9
1.93 a 2.30 b 2.57 b 2.37 b .178
15/15 15/15 13/15 12/15
-(g/bird) 41.8 a 17.4 b 22.5 b 21.7 b 2.96 277 a 5 7 b,c 41c 62b 6.3 794a 469 b 436 b 468b 42.7
Means within columns and experiments followed by different superscripts are significantly different (P<.05). 1
Japanese quail chicks were used in Experiment 1 and broiler chicks in Experiments 2 and 3.
2
Standard error.
mortality of chickens fed a diet containing BAPN. They suggested that ascorbic acid may inhibit the activity of monoamine oxidase, an enzyme that detoxifies BAPN to cyanoamine acetate (Roy, 1981). It is possible that BHA could have a similar action, and thus intensify the toxicity of lathyrogens. In summary, these results indicate that cysteine and BHA, which have protective effects against alkaloid toxicity, do not have a similar effect with lathyrogens, and may even potentiate the toxicity of Lathyrus to avian species. REFERENCES Miranda, C. L., D. R. Buhler, H. S. Ramsdell, P. R. Cheeke and J. A. Schmitz. 1982. Modification of chronic hepatotoxicity of pyrrolizidine (Senecio) alkaloids by BHA and cysteine. Toxicol. Lett. 10:177182. Miranda, C. L., H. M. Carpenter, P. R. Cheeke, and D. R. Buhler. 1981a. Effect of ethoxyquin on the toxicity
of the pyrrolizidine alkaloid monocrotaline and on hepatic drug metabolism in mice. Chem. Biol. Interact. 37:95-107. Miranda, C. L., R. L. Reed, P. R. Cheeke, and D. R. Buhler. 1981b. Protective effects of butylated hydroxyanisole against the acute toxicity of monocrotaline in mice. Toxicol. Appl. Pharmacol. 59:424430. Padmanaban, G. 1980. Lathyrogens. Pages 239-263 in: Toxic Constituents of Plant Foodstuffs. I.E. Liener, ed. Academic Press, New York, NY. Roy, D. N. 1981. Toxic amino acids and proteins from Lathyrus plants and other leguminous species. A literature review. Nutr. Abstr. Rev. 51:691-707. Sibbald, I. R. 1982. Measurement of bioavailable energy in poultry feeding stuffs: a review. Can. J. Anim. Sci. 62:983-1048. Steel, R.G.D. and J. H. Torrie. 1980. Principles and Procedures of Statistics. A Biomedical Approach. 2nd ed. McGraw Hill Book Co., New York, NY. Thornton, P. A., D. Brownrigg, and F. B. Mather. 1962. The influence of dietary ascorbic acid and the severity of p-amino propionitrile (BAPN) toxicity in turkeys. Poultry Sci. 41:1586-1590.