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The transfer of dietary butylated hydroxytoluene (BHT) into the body and egg fat of laying hens
Fd Cosmet. Toxicol. Vol. 3, pp. 475--477. Pergamon Press 1965. Printed in Great Britain
SHORT PAPERS The Transfer of Dietary Butylated Hydroxytoluene (BHT) into the Body and Egg Fat of Laying Hens P. G. C. VAN STRATUM and H. J. Vos Central Institute for Nutrition and Food Research T.N.O., Zeist, Netherlands (Received 27 May 1965) Abstract--Laying hens fed 500 ppm of butylated hydroxytoluene (BHT; 3,5-di-tert-butyl-4-
hydroxytoluene) in the diet, produced eggs with 20 ppm of the antioxidant in the fat fraction. Fifty-five ppm of BHT was found after 21 wk in the body fat. A dietary level of 100 ppm BHT resulted in less than 5 ppm in the egg fat and in body fat values of 5 ppm or less. At lower dietary concentrations of BHT neither the egg fat nor the body fat tissues contained detectable amounts of unchanged BHT. It is concluded that normal levels of BHT in feedstuffs will not cause retention of unchanged antioxidant in egg or body tissues of laying hens. Since butylated hydroxytoluene (BHT; 3,5-di-tert-butyl-4-hydroxytoluene), which is widely used as an antioxidant for the stabilization of lipids in animal feedstuffs, accumulates in the tissue fat of chickens fed on B H T in the diet (Francois & Pihet, 1960), the possible retention of unchanged antioxidant in the body fat and eggs of laying hens that had been treated orally has now been investigated. Individually housed laying birds were fed on a standard diet with the following percentage composition: maize meal, 32.8; ground wheat, 8.0; ground oats, I0.0; soya-bean meal, 20.0; herring meal, 6.0; meat scraps, 3.0; lucerne meal, 4.0; dried whey, 3-0; mineral mixture, 4.0; degummed soya-bean oil, 5.0; vitamin additives, 4.2. The hens were allowed free access to water and ground oyster shell. After adaptation, 3 groups of 4 birds were fed on the standard diet which was supplemented respectively with 0 (controls), 5 and 50 p p m of BHT, the antioxidant being dissolved in the soya-bean oil component of the ration. After 8 wk, the dietary concentrations of BHT were changed respectively to 0 (controls), 100 and 500 ppm. The birds remained in good health throughout the 29 wk on test. Egg weights and egg production were not significantly influenced by treatment with BHT, and this finding is in agreement with the work of Shellenberger, Parrish & Sanford (1957) who showed that the health, production and reproductive performance of laying hens were not affected by the daily intake of 1250 p p m of BHT in the diet during 8 months. At 0, 4, 8, 10, 12, 16 and 20 wk, one egg was collected f r o m each hen, and the mixed yolks from the 4 eggs of each group were analyzed for BHT. The residual lipids f r o m the boiling ethanol-chloroform extracts were fractionated by column chromatography on silicic acid, using isooctane as the eluent, and ultraviolet absorption spectral measurements were made of the eluate between 270 and 340 m~ using an appropriate blank prepared from the egg yolks of untreated hens. The B H T content was estimated by the method of Vos (1959). At 26 wk, B H T was determined in the lipids of 4 eggs from each hen. The results are summarized in Table 1. 475
TRANSFER OF BHT INTO BODY AND EGGS
476
Table 1. BHT content of egg fat of laying hens fed various levels of BH T BHT in egg fat (ppm) at wk Dietary level o f B H T (ppm) 5 ppm for 8 wk then 100 ppm thereafter 50 ppm for 8 wk then 500 ppm thereafter
0
4
8
10
12
16
20
26
0
0
0
0
0
0
0
T
0
0
0
24
20
18
18
24"
T~Trace *S~4-3. Results for separate hens at 26 wk in ppm: 100-ppm group: trace ~5, 0, 0, trace; 500-ppm group: 33, 21. 18 and 23. No BHT was found in the egg fat of hens with a dietary intake of 5 or 50 ppm BHT over 8 wk. Feeding at 500 ppm resulted in the accumulation of 20 ppm of the antioxidant in the egg lipids after 2 wk, and this concentration remained roughly constant during the following 16 wk. The lowest dietary concentration of BHT, which caused detectable amounts of the antioxidant to become apparent in the egg fat, was 100 ppm. The lipid component of this ration (5.0 ~o of soya-bean oil) contained 2000 ppm of BHT, i.e. 20 times the concentration usually recommended for stabilizing fats and oils. If lipids, stabilized with normal concentrations of BHT, are incorporated into the rations of laying hens, it is unlikely that detectable amounts of unchanged antioxidant would be found in the fat of the eggs. After 29 wk, the birds were killed, and the solvent extracts of the fatty tissues were analyzed for the antioxidant; the fats of the control hens being used for the preparation of blanks. In the group fed on 500 ppm BHT, 2 samples of body fat contained 56 and 52 p p m respectively of unchanged antioxidant; these concentrations are approximately one-half of that recommended for the stabilization of food fats. These results agree with the value of 60 ppm of BHT which was found in the body fat of broiler chickens, fed 1000 ppm BHT for 8 wk (Francois & Pihet, 1960). In the group of laying hens fed 100 ppm BHT, 2 samples of body fat contained < 5 and 5 ppm of BHT; these values approach thelowest concentrations detectable by our analytical methods. Again, the concentration of BHT in the lipid component of this mixed diet was 20 times that recommended for practical usage, and it might be argued that normal stabilization with BHT of the lipid components of feedstuffs would not cause retention of unchanged antioxidant in the tissue of laying hens. Acknowledgements--The sample of BHT ("Ionol") was kindly supplied by Shell Nederland Chemic N.V. at
Rotterdam.
REFERENCES Francois, A. C. & Pihet, A. (1960). Influence de l'ingestion d'antioxyg~nes sur la composition de certains tissus et sur la stabilit6 des graisses de r~serve du porc et du poulet. Annls Zootech. 9, 195. Shellenberger, T. E., Parrish, D. B. & Sanford, P. E. (1957). Effects of antioxidants, DPPD and BHT, on health, production and reproduction of laying hens. Poult. Sci. 36, 1313. Vos, H. J. (1959). D6tection et dosage des antioxyg~nes dans les mati~re~ grasses et dans les produits alimentaire~. Techq. lait. 14, 45.
SHORT PAPERS
Passage du butylate d'hydroxytolu/bne alimentaire (BHT) dans le corps et le jaune des oeufs de poules pondeuses R~sum6----Des poules pondeuses auxquelles on a fait absorber 500 ppm de butylate d'hydroxytoluene (BHT; 3,5-di-tert-butyl-4-hydroxytolu~.ne) m~lang6s aux aliments, ont pondu des oeufs dont les jaunes contenaient 20 ppm du produit antioxydant. On a retrouv6 55 ppm de BHT dans les graisses du corps de la poule au bout de 21 semaines. A la dose de 100 ppm de BHT, on a retrouv6 moins de 5 ppm dans le jaune d'oeuf et, dans les graisses du corps, 5 ppm ou moins. Pour des concentrations alimentaires inf6rieures de BHT, n i l e jaune d'oeuf ni les tissus adipeux de la poule ne contenaient de quantit6s d6celables de BHT non transform6. On en conclut que des quantit6s normales de BHT dans la nourriture ne provoqueront pas de r6tention d'antioxydant non transform6 dans I'oeuf ou darts les tissus des poules pondeuses.
Der lJbergang yon verfiittertem butyliertem Hydroxytoluol (BHT) in K/~rper und Eifett von Legehennen Zusammenfassung--Legehennen, die mit dem Futter 500 ppm butyliertes Hydroxytoluol (BHT; 3,5-Di-tert-butyl-4-hydroxytoluol) erhielten, legten Eier mit 20 ppm des Antioxydationsmittels in der Fettfraktion. Nach 21 Wochen wurden 55 ppm BHT im K/Srperfett festgestellt. Bei Verfiitterung yon 100 ppm BHT wurden unter 5 ppm im Eifett und 5 ppm oder darunter im K6rperfett gefunden. Bei noch geringerem BHT-Anteil im Futter enthielten das Eifett und die Korperfettgewebe keine feststellbaren Mengen unver~.ndertes BHT. Daraus lasst sich schliessen, dass normale Mengen BHT im Futter keine Retention unver~.nderten Antioxydationsmittels in den Eiern und den K6rpergeweben von Legehennen verursachen.
477
SHORT PAPERS The Transfer of Dietary Butylated Hydroxytoluene (BHT) into the Body and Egg Fat of Laying Hens P. G. C. VAN STRATUM and H. J. Vos Central Institute for Nutrition and Food Research T.N.O., Zeist, Netherlands (Received 27 May 1965) Abstract--Laying hens fed 500 ppm of butylated hydroxytoluene (BHT; 3,5-di-tert-butyl-4-
hydroxytoluene) in the diet, produced eggs with 20 ppm of the antioxidant in the fat fraction. Fifty-five ppm of BHT was found after 21 wk in the body fat. A dietary level of 100 ppm BHT resulted in less than 5 ppm in the egg fat and in body fat values of 5 ppm or less. At lower dietary concentrations of BHT neither the egg fat nor the body fat tissues contained detectable amounts of unchanged BHT. It is concluded that normal levels of BHT in feedstuffs will not cause retention of unchanged antioxidant in egg or body tissues of laying hens. Since butylated hydroxytoluene (BHT; 3,5-di-tert-butyl-4-hydroxytoluene), which is widely used as an antioxidant for the stabilization of lipids in animal feedstuffs, accumulates in the tissue fat of chickens fed on B H T in the diet (Francois & Pihet, 1960), the possible retention of unchanged antioxidant in the body fat and eggs of laying hens that had been treated orally has now been investigated. Individually housed laying birds were fed on a standard diet with the following percentage composition: maize meal, 32.8; ground wheat, 8.0; ground oats, I0.0; soya-bean meal, 20.0; herring meal, 6.0; meat scraps, 3.0; lucerne meal, 4.0; dried whey, 3-0; mineral mixture, 4.0; degummed soya-bean oil, 5.0; vitamin additives, 4.2. The hens were allowed free access to water and ground oyster shell. After adaptation, 3 groups of 4 birds were fed on the standard diet which was supplemented respectively with 0 (controls), 5 and 50 p p m of BHT, the antioxidant being dissolved in the soya-bean oil component of the ration. After 8 wk, the dietary concentrations of BHT were changed respectively to 0 (controls), 100 and 500 ppm. The birds remained in good health throughout the 29 wk on test. Egg weights and egg production were not significantly influenced by treatment with BHT, and this finding is in agreement with the work of Shellenberger, Parrish & Sanford (1957) who showed that the health, production and reproductive performance of laying hens were not affected by the daily intake of 1250 p p m of BHT in the diet during 8 months. At 0, 4, 8, 10, 12, 16 and 20 wk, one egg was collected f r o m each hen, and the mixed yolks from the 4 eggs of each group were analyzed for BHT. The residual lipids f r o m the boiling ethanol-chloroform extracts were fractionated by column chromatography on silicic acid, using isooctane as the eluent, and ultraviolet absorption spectral measurements were made of the eluate between 270 and 340 m~ using an appropriate blank prepared from the egg yolks of untreated hens. The B H T content was estimated by the method of Vos (1959). At 26 wk, B H T was determined in the lipids of 4 eggs from each hen. The results are summarized in Table 1. 475
TRANSFER OF BHT INTO BODY AND EGGS
476
Table 1. BHT content of egg fat of laying hens fed various levels of BH T BHT in egg fat (ppm) at wk Dietary level o f B H T (ppm) 5 ppm for 8 wk then 100 ppm thereafter 50 ppm for 8 wk then 500 ppm thereafter
0
4
8
10
12
16
20
26
0
0
0
0
0
0
0
T
0
0
0
24
20
18
18
24"
T~Trace *S~4-3. Results for separate hens at 26 wk in ppm: 100-ppm group: trace ~5, 0, 0, trace; 500-ppm group: 33, 21. 18 and 23. No BHT was found in the egg fat of hens with a dietary intake of 5 or 50 ppm BHT over 8 wk. Feeding at 500 ppm resulted in the accumulation of 20 ppm of the antioxidant in the egg lipids after 2 wk, and this concentration remained roughly constant during the following 16 wk. The lowest dietary concentration of BHT, which caused detectable amounts of the antioxidant to become apparent in the egg fat, was 100 ppm. The lipid component of this ration (5.0 ~o of soya-bean oil) contained 2000 ppm of BHT, i.e. 20 times the concentration usually recommended for stabilizing fats and oils. If lipids, stabilized with normal concentrations of BHT, are incorporated into the rations of laying hens, it is unlikely that detectable amounts of unchanged antioxidant would be found in the fat of the eggs. After 29 wk, the birds were killed, and the solvent extracts of the fatty tissues were analyzed for the antioxidant; the fats of the control hens being used for the preparation of blanks. In the group fed on 500 ppm BHT, 2 samples of body fat contained 56 and 52 p p m respectively of unchanged antioxidant; these concentrations are approximately one-half of that recommended for the stabilization of food fats. These results agree with the value of 60 ppm of BHT which was found in the body fat of broiler chickens, fed 1000 ppm BHT for 8 wk (Francois & Pihet, 1960). In the group of laying hens fed 100 ppm BHT, 2 samples of body fat contained < 5 and 5 ppm of BHT; these values approach thelowest concentrations detectable by our analytical methods. Again, the concentration of BHT in the lipid component of this mixed diet was 20 times that recommended for practical usage, and it might be argued that normal stabilization with BHT of the lipid components of feedstuffs would not cause retention of unchanged antioxidant in the tissue of laying hens. Acknowledgements--The sample of BHT ("Ionol") was kindly supplied by Shell Nederland Chemic N.V. at
Rotterdam.
REFERENCES Francois, A. C. & Pihet, A. (1960). Influence de l'ingestion d'antioxyg~nes sur la composition de certains tissus et sur la stabilit6 des graisses de r~serve du porc et du poulet. Annls Zootech. 9, 195. Shellenberger, T. E., Parrish, D. B. & Sanford, P. E. (1957). Effects of antioxidants, DPPD and BHT, on health, production and reproduction of laying hens. Poult. Sci. 36, 1313. Vos, H. J. (1959). D6tection et dosage des antioxyg~nes dans les mati~re~ grasses et dans les produits alimentaire~. Techq. lait. 14, 45.
SHORT PAPERS
Passage du butylate d'hydroxytolu/bne alimentaire (BHT) dans le corps et le jaune des oeufs de poules pondeuses R~sum6----Des poules pondeuses auxquelles on a fait absorber 500 ppm de butylate d'hydroxytoluene (BHT; 3,5-di-tert-butyl-4-hydroxytolu~.ne) m~lang6s aux aliments, ont pondu des oeufs dont les jaunes contenaient 20 ppm du produit antioxydant. On a retrouv6 55 ppm de BHT dans les graisses du corps de la poule au bout de 21 semaines. A la dose de 100 ppm de BHT, on a retrouv6 moins de 5 ppm dans le jaune d'oeuf et, dans les graisses du corps, 5 ppm ou moins. Pour des concentrations alimentaires inf6rieures de BHT, n i l e jaune d'oeuf ni les tissus adipeux de la poule ne contenaient de quantit6s d6celables de BHT non transform6. On en conclut que des quantit6s normales de BHT dans la nourriture ne provoqueront pas de r6tention d'antioxydant non transform6 dans I'oeuf ou darts les tissus des poules pondeuses.
Der lJbergang yon verfiittertem butyliertem Hydroxytoluol (BHT) in K/~rper und Eifett von Legehennen Zusammenfassung--Legehennen, die mit dem Futter 500 ppm butyliertes Hydroxytoluol (BHT; 3,5-Di-tert-butyl-4-hydroxytoluol) erhielten, legten Eier mit 20 ppm des Antioxydationsmittels in der Fettfraktion. Nach 21 Wochen wurden 55 ppm BHT im K/Srperfett festgestellt. Bei Verfiitterung yon 100 ppm BHT wurden unter 5 ppm im Eifett und 5 ppm oder darunter im K6rperfett gefunden. Bei noch geringerem BHT-Anteil im Futter enthielten das Eifett und die Korperfettgewebe keine feststellbaren Mengen unver~.ndertes BHT. Daraus lasst sich schliessen, dass normale Mengen BHT im Futter keine Retention unver~.nderten Antioxydationsmittels in den Eiern und den K6rpergeweben von Legehennen verursachen.
477