THE ENFLUENCE OF A TRANQUSLEZER FED IN COMBINATION WTTH AN ESTROGE-NIC CQMPOUNQ 0N P'HYfiflO-LOGICAL ACTWITIES: GRQWTH, AND MARKET QUALETIES OF CHICKENS “tests for HM Degree 0‘ M. 5. MECHEGAN STATE UNWER’SETY Kenneth George Rood 1959 THESIS TL. LIBRA R Y E Michigan State § .- I: University {4 THE INFLUENCE OF A TRANQUILIZER FED IN COMBINATION WITH AN ESTROGENIC COMPOUND ON PHYSIOLOGICAL ACTIVITIES. GROWTH, AND MARKET QUALITIES OF CHICKENS By Kenneth George Rood AN ABSTRACT Submitted to the College of Agriculture Michigan State University of Agriculture and Applied Science in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Poultry Science 1959 Approved ' ' , ~¢quyp) Kenneth G. Rood l. The consequences of an estrogen and a tranquilizer. fed singly and in combination. to a known strain of White Plymouth Rocks was investigated. The tranquilizer (a perphenazine derivative) was incorporated in the feed at 4 grams per ton and a synthetic estrogenic compound (dienestrol diacetate) was incorporated in the feed at 21.14 grams per ton at two levels of protein intake. The positions of the eight (8) dietary treatments were randomly selected and replicated among sixteen (16) identical pens of thirty—two (32) males each and sixteen (16) identical pens of thirty—two (32) females each. Body weight. feed efficiency. feathering. fleshing. pigmentation and serum xanthophyil was scored on the live birds. Carcass grade. abdominal fat weight. comb. gonadal and oviducal weights were determined after slaughter. At nine weeks of age. one—half of the birds in each pen were weighed. scored and slaughtered. The remaining birds in each pen continued on to thirteen weeks of age. at which time they were weighed. scored and slaughtered. The lower protein recipients were from 5 to 8 percent lighter in body weight than those in corresponding groups on the higher protein ration. The oral administration of dienestrol diacetate brought body weight of the lower protein recipients up to the body weight of the higher protein con— trol fed birds. The addition of the perphenazine derivative to a low protein ration suggested a protein Sparing effect. ii Kenneth G. Hood 9 Q. I The addition of an estrogen or a tranquilizer to ei— ther a high or low protein ration did not increase fat deposition in the abdominal area of either sex. The males fed the lower protein ration exhibited a higher serum xanthophyll content than males fed a higher protein level and slightly higher than that found in the blood serum of the females. Dienestrol diacetate physiologically suppresses hypophy— seal gonadotrophin release resulting in decreased testicular weight and ultimately comb size in the male and decreased ovarian weight with increased oviducal weight in the female compared to that exhibited by the birds fed the control ration. Similar results were observed when the perphenazine deriva— tive was administered singly. suggesting an estrogen—like activity by this compound. When the estrogenic compound and the tranquilizer were fed in combination ovarian weight was very similar to that exhibited by birds fed the control ration. indicating that the suppression of hypophyseal secretions. when either compound was administered singly. was relieved. Oviducal weights indicated that when fed in combination. the two compounds counteracted each other eliminating their estro— genic activity and giving ovarian weights similar to the controls. In contrast testicular weight was increased sig— nificantly, when the combination was fed. compared to testi— cular weight of birds fed the control ration. iii Kenneth G. Hood 3. These data concerning an estrogen and a tranquilizer. fed singly and in combination. would offed excellent tools in the study of reproduction in the fowl. Since it is pos- sible to interrupt the reproductive cycle, apparently through endocrine interference, a more thorough investigation of endocrine inter—relationship can be made. iv THE INFLUENCE OF A TRANQUILIZER FED IN COMBINATION WITH AN ESTROGENIC COMPOUND ON PHYSIOLOGICAL ACTIVITIES. GROWTH, AND MARKET QUALITIES OF CHICKENS By Kenneth George Rood A THESIS Submitted to the College of Agriculture Michigan State University of Agriculture and Applied Science in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Poultry Science 1959 ACKNOWLEDGEMENTS The author wishes to express his most sincere apprecia— tion to Dr. R. K. Ringer of the Poultry Science Department. Michigan State University. for his guidance. assistance and constructive criticism of this manuscript. Appreciation is due Dr. H. C. Zindel, Head of the Poultry Science Department. for the facilities and the opportunity of pursuing this course of study and to Dr. T. Coleman for his criticism and corrections of the manuscript. The author also wishes to thank Mr. Elwood Speckmann and Mrs. Kenneth Poole for assistance in collecting and recording the data; to Dr. L. E. Dawson for his expert evaluation of the dressed carcasses. and to other members of the Poultry Science Department for the assistance in proces— sing the chickens used in this study. Special thanks is expressed by the author to his wife. Myrtle. and children Bill, Bob. and Lynn for their patience. understanding and encouragement. vi TABLE OF CONTENTS Introduction Review of Literature Objectives Experimental Procedure Randomization of Experimental Chickens Dietary Treatments Feather Score Fleshing Score Pigmentation Score Serum Xanthophyil Determination Dressed Carcass Grading Criteria Abdominal Fat Weight. Gonad and Oviduct Weight Determinations Statistics Results Body Weight and Feed Conversion Summary of Live Scores Dressed Carcass Grade Abdominal Fat Weight Serum Xanthophyil Gonadal. Oviducal and Comb Weights Discussion Summary and Conclusions Bibliography vii 14 15 17 19 19 19 23 24 36 37 37 38 38 42 53 66 LIST OF TABLES Table Page 1. High energy pigment starter ration (basal from 1 to 5 weeks . . . . . . . . . . . . . . . . 16 2. Grower ration (basal from 5 to 9 weeks). . . . 18 3. Finisher ration (basal from 9 to 13 weeks) . . 21 4. Analysis of variance for 9—week body weights as affected by sex. treatment. replication and interaction . . . . . . . . . . . . . . 26 5. The effect of protein level on body weight of chickens at 9 weeks of age . . . . . . . . . 27 6. The effects of treatment on body weight of males and females at 9 weeks of age . . . . . . . 28 7. Analysis of variance for 13—week body weights as affected by sex. treatment. replication and interaction. . . . . . . . . . . . . . . . . 29 8. The effects of protein level on body weights of males and females at 13 weeks of age . . . . 30 9. The effects of treatment on body weight of males and females at 13 weeks of age . . . . . . . 31 10. Effect of treatment on the mean body weight of males and females at 9 and 13 weeks of age . 32 1.1. The effect of treatment on weight gain and feed efficiency of males and females during the growing period (5 to 9 weeks of age) . . . . 33 122. The effect of treatment on weight gain and feed efficiency of males and females during the finishing period (9 to 13 weeks of age). . . 34 -13. Effect of treatment on dressed grade (expressed in percentage) of males and females ( 9 and 13 weeks of age combined). . . . . . . . . . 57 14- The effect of treatment on abdominal fat weight of males and females at 13 weeks of age . . 58 viii 15. 16. 17. 18. 19. 20. 21. The effect of treatment and sex on serum xanthophyll expressed as optical density at 9 and 13 weeks of age . . . . . . . . . . 59 Effect of the oral administration of Lipamone or Quietrol. fed singly or in combination. on the testicular weights of 9-week old cockerels . . . . . . . . . . . . . . . . . 60 The effects of protein level of the ration and Lipamone or Quietrol, fed singly or in combi— nation. on testes weight of 13—week old cockerels . . . . . . . . . . . . . . . . . 61 Effect of the oral administration of Lipamone or Quietrol. fed singly or in combination. on the ovarian weight of 9—week old pullets . . . . 62 Effect of the oral administration of Lipamone or Quietrol. fed singly or in combination. on the mean oviducal weight of 9-week old pullets . 63 The effects of protein level of the ration and Lipamone or Quietrol. fed singly or in combi- nation. on oviducal weights of 13—week 01d pullets . . . . . . . . . . . . . . . . . 64 The effect of protein level and treatment on comb weights of males and females at 9 weeks of age . . . . . . . . . . . . . . . . . 65 ix LIST OF FIGURES Figure Page 1 Testis Sections at 13 weeks of age 56 INTRODUCTION The search for products. either natural or synthetic. that will stimulate growth in chickens and improve appearance of the finished product is important to the grower and pro— cessor. Estrogens are examples of synthetic products that will improve carcass quality by promoting fat distribution and will render the edible portion more tender. Broilers. roasters and adult chickens fed an estrogenic compound are slightly heavier in body weight than birds not receiving the estrogen. This phenomena is caused primarily by increased fat deposition and is not considered by many investigators to be true growth. Tranquilizers allay or diminish anxiety and motivation when administered to some mammals. In these mammals, increased weight gains and improved feed conversions were observed when a tranquilizer was fed singly or in combination with other compounds. A reduction in motor activity with a subsequent improvement in feed conversion without deleterious side effects would be an economic advantage to the poultry industry. The consequences of a combination of an estrogen and a trElnquilizer have not been adequately investigated; there— fore , the main objective of this study was to determine the influence of a tranquilizer, administered singly and in com— bination with dienestrol diacetate at two levels of protein intake_ on growing chickens. REVIEW OF LITERATURE Body weight gains and feed conversion: When diets containing 15 percent and 18 percent protein tvere fed to 12~week old cockerels. Bird (1948) observed that ‘the birds fed the high—protein diet gained more weight and sshowed an improved feed conversion when compared to those fed 21 lower protein ration. but were "lean. blue and lacking fflinish". Experimenting with various high protein broiler nuashes. Gassner and Wilgus (1948) demonstrated improved rate C)f'gain. improved feed efficiency and improved grade of the (ilressed carcasses over that obtained on the standard or low Ixrotein mash. The average grades obtained on these high pnrotein rations were as good as those observed in birds treated ‘wzith.estrogens. Lorenz (1954) used rations containing vary— irlg amounts of fiber in a study of the effect of protein intake on the response of broilers (killed at 12 weeks of Eigue) to different estrogen treatments. The low—fiber diet puexunitted the greatest response to the various treatments. bc>t11 in additional gains and amount of fattening. Feed BfIKiciency was decreased. as compared with control birds. by all 'treatments on the high—fiber ration. but dienestrol diacuetate actually improved feed conversion on the low—fiber diet. Lorenz and Bachman (1947) reported slight stimulation 0f total growth in 8 to 12—week old cockerels fed dienestrol or abdominal fat weight. indicating that no advantage was derived from the tranquilizer in these respects. Wolterink _t _l. (1958) found that birds fed a combination of dienestrol diacetate and Reserpine in a 12 percent protein ration fed to White Rock cockerels from the first to the tenth week of zage improved weight gain and resulted in greater fat depo— sition when compared to similar birds not receiving the zadditives. I3igmentation: In some geographical areas. yellow skinned. highly [)1gmented poultry meats are more desirable than the paler or Vvllite skinned varieties. This is especially true in the (baited States. where the yellow skinned breeds predominate. cmantrasted to England. where the consumer prefers poultry Inerving white skin. white shanks. and white fat. Since Palmer (.1915) first demonstrated that xanthophyll is the yellow pigment of chicken body fat, numerous investigators have 0(1nfirmed this finding and demonstrated a relationship be- ‘twueen xanthophyll content of feeds and the intensity of yellow Piggmentation in shanks and body fat. Hammond and Harshaw (1E141) demonstrated that yellow pigment was absent in the shanks and skin of Light Sussex chicks. regardless of the diet fed. White Wyandotte. White Leghorn and Rhode Island Red (zhicks were found to have approximately the same shank 5"“1 Sliin color on any given diet. Collins gt a1. (1955) found. the shank color of New Hampshires to be significantly darkexr than that of White Plymouth Rocks. A significant diacetate at 0.01 percent of the ration. Quisenberry and Kruger (1948) also obtained an increased rate of gain. as well as greater feed consumption and increased net efficiency of feed conversion. by feeding dienestrol diacetate. The administration of estrogens to chickens tends to decrease metabolism and activity and the fattening effects may be attributed to these factors (Bird. 1946). The current impetus for tranquilizers in animal feeding is directed toward these same effects. The quiting effect of tranquilizers on animal metabolism is believed to be obtained by the concen— tration of a greater share of the nutrients toward weight gains and production and less toward motor activity. In cattle feeding. tranquilizers in combination with an anti— biotic and stilbestrol resulted in greater feed efficiency than either of these ingredients fed alone or in single combination with the tranquilizer (Anonymous, 1958). Very little research has been conducted in the use of tranquilizers as an additive to poultry feeds. The results of Hewitt and Reynolds (1957) have shown Reserpine to be successful in reducing cannibalism in pheasants but the rate (of body weight was also reduced. Meprobemate and promazine. Ivhen fed to White Leghorns, did not lessen characteristic 11ervousness but did retard body weight gains (Babcock and Tfiaylor. 1957). Rood £3.2l- (1958), feeding Reserpine to fiemale broiler chicks from the fourth to the eleventh week (Df age with phenobarbital as a comparative compound. found IND significant differences in weight gain. feed conversion, sex difference in shank color was observed and some of the strains within each breed differed significantly. A further study of the same five strains of New Hampshires on each of three consecutive trials showed that environment markedly influenced shank color. Culton and Bird (1941) presented evidence indicating that certain common poultry feedstuffs such as meat scraps. fish meal and soybean oil meal contain a factor. or factors. which inhibit the deposition of yellow pigment in the shanks of growing chicks. It would appear that some lots of these common protein supplements must be free of inhibiting effects, otherwise a high degree of pigmentation could not be produced under normal practical conditions. Certain brands of cod liver oil were found to be potent sources of a pigment-suppressing factor (Hammond and Harshaw. 1941). Although the reason for the inhibition of pigmentation \vhen these oils were fed was not determined. a large portion of the inhibiting factor could be eliminated by heating the cod liver oil to 230°C. in a vacuum for three hours. It was ‘theorized that the pigment might be destroyed by oxidation induced by the unsaturated compounds in the cod liver 011. 'P0 test this theory. various oils and other so—called con— ‘taminants were fed. These supplements had no appreciable (affect on pigmentation, resulting in the conclusion that the (Halor of the shanks and skin in the yellow skin breeds was (hapendent on the interaction of the quantity of the pigment— 1nducing ingredient and the amount of yellow pigment-suppressing factor present in the diet. Day and Williams (1958) significantly increased the pigmentation exhibited in broilers by increasing the calorie: protein ratio the last two weeks of the feeding period. Palmer's pioneer work proved that. in hens. dietary xanthophylls (but not carotene) occur in the blood plasma and in the fat and skin. especially in the shanks. Xanthophyll (lutein) is esterified and stored in the liver. skin. fat. face and beak of the hen. Hollander and Owen (1939) noted xanthophyll in the iridial carotenoids and demonstrated their lability by showing that alteration of the diet can alter the eye color of hens. The contribution which carotenoids make to the pigmen— tation of the plumage of birds is considerable. and feather carotenoids can be considered analogous to the carotenoids stored in the external structures of sea animals. In both (eases the carotenoids are xanthophyllic. produced by the zrnimals from the alimentary carotenoids (Goodwin. 1952). Xanthophyils are laid down in the skin and shanks as essters in a non—laying chicken. During the laying period tilese are mobilized in the free state and deposited into the Euggs. In pullets. carotenoids are mobilized into the blood b)’ heavy doses of estrogens and this is probably how the Piggments are transferred to the eggs when the pullet comes ifrto lay (Common and Bolton. 1946). The basic findings that Stlfilking increases in blood lipid levels occur during periods 0f (Ivarian activity have been confirmed. Further. Lorenz _t _t. (1938) demonstrated that liver lipids are also increased in hens during periods of ovarian activity. Contributions to the understanding of the lipogenic mechanism were made by Entenman gt gt. (1938). who demon— strated lipemia in intact immature female birds following administration of crude gonadotrophin. and by Lorenz gt gt. (1938). who produced lipemia in immature birds of both sexes by the administration of crude estrin. The blood lipids of the laying female are considerably higher than in the non—laying female or in the male. and most of the surplus fat is deposited in the egg in the form of yolk. Since the blood lipids of males receiving estrogen are also increased. the surplus blood fat in males must be stored in the tissue. thus making the bird fatter and more desirable as a meat bird. Feathering: In birds with six—dimorphic feathers. the administra— t;ion of an estrogen can alter the shape of the feather and tile nature and distribution of feather pigment from the Sfaxually-indifferent pattern typical of the male into the di rection of female plumage normal for the species and breed. 0f" additional practical importance to broiler growers is the Biifect of estrogens on rate of feather growth. By experi— merrtally plucking small areas, Lorenz (1954) observed that esixrogen treatment tends to inhibit the formation of new ifirrfeathers but does stimulate the growth of those already (HHIpted. The number of pinfeathers present when the birds are marketed is thus reduced. Carcass Quality: The degree of external fatness designates finish, while the degree of internal fat deposited in the muscle tissue determines quality and tenderness of the flesh. An even distribution of fat in all parts of the body is essential to optimum quality in the carcass. Maw (1935) found that the different cereals cause the deposition of fats in entirely different manners; for instance. cornmeal causes a high percent of the total body fat to be deposited in the flesh and much less fat in the abdominal cavity and in the skin of the bird. whereas the cereals. oats. barley. and wheat, show the reverse in varying degrees. Wheat gave an excellent ex— ternal appearance. a uniform layer of fat deposited over the entire carcass. but very little deposition in the actual flesh. Several investigators have shown that fat deposited in body tissues replaces moisture. thus increasing the edible portion Of the carcass. Harms gt gt. (1958) demonstrated that as the amount of Yellow corn was increased in the diet. finish of birds re— ceiving the diet was improved. The first suggestion that birds might be fattened for market by estrogen administration was made by Lorenz (1943) Who sued subcutaneous implantation of diethylstilbestrol (DES) pellets on cockerels. A single 12 to 15 mg. pellet PrOduced a noticeable effect on growing birds within a week. DES has a relatively low oral estrogenic potency on chickens arui large doses are required to produce fattening. Jaap (1944), Munro and Kosin (1946). Thayer gt a. (194.5) ob— s<2ryed that a slight improvement in finish was obtained when EB-wveek old Barred Plymouth Rock cockerels consumed 75 mg. of tIIeeestrogen during a three week feeding period and 100 mg. iri a four week period. Birds that consumed 50 mg. of the (esrtrogenic compound in a two week period did not show an improvement in grade compared to the group receiving no (esrtrogen. Feeding DES to 6— and 11—week old White Leghorn (Ecyckerels for 4 weeks produced only questionable increases .ir1 abdominal fat deposition when over 200 mg. of estrogen \Neare consumed. and none when the total dose was lower (7 to 144 mg.)(Lorenz. 1945). Glazener and Jull (1946) obtained 'ezccellent fattening in 6—week old Barred Plymouth Rocks x New Hampshire cross—bred cockerels fed DES for 3 weeks; but 'tlley used massive doses (630 mg. per bird). Some'improvement irl grade has been reported on relatively low doses by Sykes S:E_ El, (1945). who fed DES at the rate of 1 mg. per day. Rhode Island Red and Barred Plymouth Rock cockerels 5— to lil—Ndeeks of age were used. and various treatment periods {Harunitted total doses of 30 to 130 mg. of the estrogenic Compound per bird. Black (1946) also reported improvement in. 17—week old Light Sussex cockerels fed 68 mg. of DES over aPeriod of 7 weeks. The reasons for these discrepancies are Iuyt obvious but may be related to different criteria of car- Cass quality or the effect of breed on responsiveness to eStrogens. 10. Dianisylhexene. fed to chickens. is much more potent as an estrogen by the oral route than is DES and has produced better fattening responses at lower doses. Lorenz (1945) obtained good fat deposition by feeding 144 and 236 mg. of the compound over a period of 4 weeks to 6—week old White Leghorn cockerels. Thayer gt gt. (1945) fed levels varying from 44 to 220 mg. of dianisylhexene per kilo of feed and repor— ted improvement in grade over their entire range of experi— Inental conditions. Birds receiving the higher levels exhibited asymptoms of overdosage. with loss of appetite. leg weakness. 21nd occasional death. Due to its relatively high estrogenic potency when fed tC) cockerels. dienestrol was the most promising of the compounds teasted. In preliminary trials it appeared to be an excellent farttening agent. Its solubility characteristics made the pusssibility of practical application seem remote. The dizacetate of dienestrol. however. is more easily handled arui the lability of the ester linkages would have the same pfnysiological action as the free phenol. Dienestrol is elfifective when fed at the level of 0.01 percent or less of thf? feed for 3 to 6 weeks. Lorenz and Bachman (1947) found thert this estrogen gave satisfactory results when fed at leviels of 0.0033 percent. but more time was required to PITHiuce results than that at higher levels. At the 0.01 P91W3ent level or above. lipemia was produced. but not at the O-(H333 percent level. 1]. The primary objective of estrogen treatment of poultry is improvement of carcass quality. Estrogen treatment has at least some influence on each component of carcass quality. and for the most part the influence is favorable. Fatness is the best—known advantage gained; extra fat is deposited in all parts of the bird. Muscle fat. important for "juiciness" of the meat. is consistently increased. and. in the leg tissue especially. may be more than doubled (Lorenz. 1943; Sykes gt _gt. 1945). Increases in subcutaneous fat improve the appear— ance of the dressed carcass. Visceral fat may be increased inany fold (Lorenz, 1943 and Bird. 1946). Warden gt gt. (1958) demonstrated that the administra— tion of dienestrol diacetate at 1/3 pound per ton. signifi— (2antly reduced the deposition of abdominal fat in the males IJut not as markedly in the females compared to that exhi— t)ited by similar birds receiving an implant of DES. They cnoncluded that this effect may be due to a decreased calorie: Ixrotein ratio of the fattener rather than to the influence (3f dienestrol diacetate. Finish is improved and a smooth "silky" texture is ilnparted to the dressed carcass by the increased subcutaneous fkit and by the effects of estrogen on skin structure and feuither growth. Improved skin texture is a definite con— Se‘Tluence of estrogen treatment. presumably due to hypertrophy 0f"the epidermal strata (Bird. 1948). Dienestrol diacetate Wtua more effective in producing this hypertrophy than dian— isb’lhexene when both were administered orally and in the same 12. dosage. Endocrine system and secondary sex characteristics: The experiments of Berthold (1949) on transplantation of fowl testes are regarded as the first demonstration of endocrine secretion. The relation of the gonads and their endocrine products to the development of secondary sexual characteristics. to the functions of primary and accessory sex organs, and to overt sexual behavior have been the cen— ter of interest to numerous researchers. Surgical castration .has been practiced for many years; however. since the isola- tion of the hormonally active material by Dodds gt 21.0937) 21nd the synthesis of DES and other potent estrogenic stilbene (ierdvatives. the older method is rapidly being replaced by tile administration of the synthetic compounds. Behavioral responses to administered estrogens include (iiIninished aggressiveness and cessation of fighting and crowing. [hivis and Domm (1943) presented evidence to show that estro— géin.is a direct though weak stimulator of masculine sex beihaviour in the male. but not in the female. Estrogens Hu)st striking direct effect is stimulation of sexual recep— tiveness. Chickens treated with adequate estrogen dosage reeuiily assume the typical feminine receptive position. 59luitting with slightly spread wings when approached by a nOt‘mal cock or by some other stimulus such as the human caretaker. 13. Retention of estrogen in edible tissue: The significance to human health of tissue retention of estrogens was examined by Bird gt a1. (1947). Menopausal women were fed diets containing livers or rendered fat from treated birds daily for 6 days. Some vaginal cornification was obtained when tissues of dianisylhexene—treated birds were fed. but no demonstrable effect was produced by tissues of birds treated with dienestrol diacetate. Lowe (1949) made biological assays to determine the quantity of residual estrogen remaining in the flesh and skin. liver. gizzard. heart. and abdominal adipose tissues of cockerels which had been fed dienestrol diacetate. All the tissues assayed con— tained negligible amounts of estrogen and the conclusion drawn was that a person would have to consume at least 12 to 120 pounds of chicken skin and flesh every other day to obtain a clinical dose of estrogen of the magnitude that is used in practice (1 mg. to 10 mg.) (Selye. 1947; Robson. 1947). The edible tissue of chickens fed dienestrol diacetate were examined by Umberger and Goss (1959) for estrogenic residues. Within limits of the assay method. no estrogenic residues could be detected in muscle, abdominal fat. skin fat, heart. gizzard, or spleen. The results suggest that dienestrol diacetate is not stored in the edible tissues but that a period of 24 hours off the estrogen feed before slaugh- ter is not sufficiently long to insure complete removal of the estrogen from the usual pathways of excretion. OBJECTIVES The poultry industry is an extremely competitive business, continually searching for new products and methods to improve the efficiency of production and the acceptance of the finished product. This study was designed to furnish information for the eventual practical application of a tranquilizer as an additive to poultry feeds. The tranquil— izer. administered singly and in combination with an estro— genic compound at various levels of protein intake. was studied to test the following objectives: a. To measure the physiological activity of the tranquilizer administered alone and in combin- ation with an estrogenic compound. b. To determine the influence of the dietary treatments on weight gain and feed efficiency. 0. To establish the effect of protein intake and the addition of a tranquilizer and an estrogenic compound on pigmentation. de- position of abdominal fat. gonadal and oviducal weight. d. To determine the influence of the dietary treatments on live market quality and dressed carcass grade. EXPERIMENTAL PROCEDURE The chicks used in this study were a known broiler strain of White Plymouth Rocks purchased from a local hat- chery. Thirty-seven (37) wing—banded chicks of one sex were placed in each individual pen. Sixteen (16) identical pens of males were housed on one side of a brooder house with sixteen identical pens of females on the opposite side (see appendix for experimental design). The sexes were reared separately to eliminate sexual differences in behavior and feed conversion. During the first five weeks of the ex- perimental period. a high energy pigment starter (Table l) was fed to all pens (rations formulated by Poultry Nutrition Department. Michigan State University). This was a control period to facilitate selection of birds that exhibited rela— tively uniform growth rate before initiation of the dietary treatments. .Randomlzation of experimental chickens: Just prior to the thirty—fifth day of age. the birds lVere weighed and segregated by weight into 30 gram increment lcrts, All birds in the middle or average weight division Were uniformly distributed into the pens. The same procedure Wafii followed in the weight increment above the average and t1lenthe group below the average. then the group that weighed tWCI increments (60 grams) above the average. then two incre— merrts below and so on until 32 birds were placed in each of 16. the experimental pens. The same procedure was followed for both males and females assuring a uniform distribution of weight throughout the experimental pens. This selection aided in the elimination of any significant differences in weight at the initiation of dietary treatment. Table l - High energy pigment starter ration (basal from 1 to 5 weeks) Lbs. or gmsT7 Ingredient 100 lbs. Corn, grd. No. 2 yellow 45.00 Alfalfa leaf meal. 20 percent protein 2.50 Soybean oil meal. 50 percent protein 36.00 Menhaden fish meal. 60 percent protein 2.00 Meat and bone scraps. 50 percent protein 2.50 Ground limestone 0.75 Dicalcium phOSphate 1.25 Salt. iodized 0.50 Whey, delactose (50 percent lactose) 0.50 Brewers dried yeast 0.50 No. 2 yellow grease 8.00 Vitamin supplement No. l 34 grams Vitamin 1) (3.000 ICU/gm.) 10 grams (Iholine caloride (25 percent supplement) 30 grams \fiitamin 812 supplement (6 mg./lb.) 0.10 Methionine 0.10 Delamix 0.10 Niacin 0.10 1 gram 100 lbs. CEilculated analysis Crude protein percent 25 Crude fat percent 10.5 CI'ude fiber percent 2.5 Productive energy (Cal./lbs.) 1025 17. Dietary treatments: At the end of the fifth week and after the experimental pens were formed as previously described. the following dietary treatments were initiated. goge Treatment Description HPC High protein basal control HPL High protein basal plus Lipamonea HPLQ High progein basal plus Lipamone plus Quietrol HPQ High protein basal plus Quietrol “LPC Low protein basal control LPL Low protein basal plus Lipamone LPLQ Low protein basal plus Lipamone plus Quietrol LPQ ‘ Low protein basal plus Quietrol The positions of the various treatments were randomly selected to eliminate positional variables. The estrogenic (Bompound (Lipamone) was administered orally at one-third [Maund per ton (21.14 gms./ton). The tranquilizer (Quietrol) lvas administered orally at 2 mg. per pound of feed (4 grams [Her ton). During the growing period. 5 to 9 weeks. the two llavels of protein intake were calculated at 20 and 16 percent ITHspectively as shown in Table 2. At the end of the ninth 3- Lipamone. 14 percent dienestrol diacetate in corn distil- lers dried grains. manufactured and distributed by White Laboratories. Inc.. Kenilworth. N.J. (Now. American Scien— tific Laboratories, Inc.. Madison. Wis.). 'b- Quietrol, containing 50 grams of perphenazine. l—(2—hr— (iroxyethyl)-4Z§—(2—chloro—10—phenothiazinyl)—propyl ?— jpiperazine, per pound_ manufactured and distributed by \Vhite Laboratories, Inc.. Kenilworth, N.J. 18. week individual body weights, and feather. fleshing and pig- ment scores were recorded. Table 2 — Grower ration (basal from 5 to 9 weeks) High protein Low protein ’ lbs. or gms./ lbs. or gms./ Ingredient 100 lbs. 100 lbs. Corn, grd. No. 2 yellow 54.88 72.78 Soybean oil meal, Solv. 44 percent 30.00 16.00 Fat. N0. 2 yellow grease 5.00 1.00 Alfalfa leaf meal. 20 percent protein 2.50 2.50 Meat and bone scraps, 50 percent " 2.50 2.50 Fish meal (Menhaden 60 percent protein .50 .50 Whey, delactosed product .50 .50 Yeast. dried brewers .50 .50 Salt, iodized .50 .50 Dicalcium phosphate 2.00 2.00 Limestone. grd. (98 percent CaCO3) 1.00 1.00 Delamix .10 .10 'Vitamin suppl. 2490 .05 .05 (Iholine Chloride (25 percent dry mix) — —— .10 Nicarbazin (25 percent dry mix) .05 .05 \litamin D3 (3.000 ICU/gm.) 10 grams 10 grams Vfiitamin A (10.000 IU/gm.) 10 grams 10 grams (halculated analysis: Crwzde protein percent 20.2 15.6 Crude fat percent 7.7 4.3 Crude fiber percent 3.5 3 3 Productive energy (Cal/lb.) 998 984 19. Feathering was scored by the author. as suggested by Arscott (1958). as follows: Feather score: Description: 0 Back completely bare 1 Large bare areas 2 Numerous pin feathers and small bare areas 3 Numerous pin feathers 4 Few pin feathers 5 No pin feathers Fleshing score: Fleshing was scored by the author on an arbitrary ba— sis. The bird was suspended by the legs so as to determine 'the degree of fleshing on the legs. thigh and breast. A ascore of five was highest. zero the lowest (indicating poor :fleshing). I’igmentation score: Pigmentation was determined visually by the author by ccnnparing the shanks to discs on the Heiman-Carver Yolk Color Rotor. One—half of the birds from each pen were sacrificed at 9 “Hacks of age for dressed carcass grade, abdominal fat weigghts and serum xanthophyll determinations. The remaining birds in each pen continued on experiment until. the thirteenth week of age with the following changes: the lligh protein basal was reduced from 20 percent to 16 per— cent lxrotein; the low protein basal was reduced from 16 per— Cent‘to 13 percent protein. both rations remaining isocaloric 20. (1000 % calories per pound). The finisher formula is given in Table 3. At the end of the thirteenth week the individual weights and scores were recorded and the birds sacrificed for serum xanthophyll. carcass grade and abdominal fat weights. Serum xanthophyll determination: In this experiment. the procedure for determination of serum xanthophyll outlined by Davis and Kratzer (1958) was modified. Approximately 10 ml. of whole blood was collected: without an anti—coagulent, from each bird at the time of slaughter. The blood samples were placed in a 35°F. cooler until all birds were slaughtered (approximately 7 hours). Each tube was ringed to allow the clot to withdraw and free 'the serum. Some samples were centrifuged to facilitate serum <3ollection. The next day the serum was poured off into small trials and frozen. 0n the day of serum xanthophyll determination, the sanmples were separated by treatment and sex and allowed to inmaw; A sufficient volume of serum was needed to run dup— liczate samples for comparison of technique. This necessita— terl pooling. 2, 3. and in some cases. 4 serum samples from diiiferent birds on the same treatment. These pooled samples Werwa then centrifuged to separate any cells that may have been Present. and the serum decanted. To each 1 ml. of serum was addeui 10 m1. of acetone. This solution was mixed by bubbling with Elir and then centrifuged to remove the precipitate. The SuperTuatant of each mixture was poured into a clean dry SPBCtINDphotometer cuvette and the optical density compared to a lalank (a cuvette containing acetone). Table 3 - Finisher ration (basal from 9 to 13 weeks) 21. High Protein lbs. or gms./ Low Protein lbs. or gms./ Ingredients 100 lbs. 100 lbs. Corn. No. 2 grd. yellow 68.59 78.63 Soybean oil meal, dehulled (50% protein) 13.60 6.00 No. 2 yellow grease 3.50 1.00 Wheat. middling flour 5.00 5.00 Meat and bone scraps 2.50 2.50 Alfalfa leaf meal (20% protein) 2.50 2.50 Whey, delactosed prod. .50 .50 Yeast. dried brewers .50 .50 Fish meal, Menhaden .50 .50 Limestone, grd. 1.00 1.00 Dicalcium phosphate 1.00 1.00 Salt, iodized .50 .50 Delamix .10 .10 Vitamin Suppl. 249C .05 .05 ‘Vitamin 312 suppl. (6mg/lb) .05 .05 Nicarbazin (25 percent mix) .05 .05 Choline chloride (25 percent dry mix) 10 gram 40 gram \Litamin A suppl. 10,000 USP/gm.) 10 gram 10 gram 'Vitamin D3 Suppl. (3.000 ICU/gm.) 10 gram 10 gram 100 lbs. 100 lbs. Calcnilated analysis: Crude protein percent 15.96 13.02 CTUG£B fat percent 6.64 4.47 Crudta fiber percent 2.66 2.69 prod. energy (Cal/lb.) 1041.30 1038.30 [\3 [O Dressed carcass grading criteria: The New York dressed birds were graded according to U.S.D.A. Standards for fleshing, conformation. pigmentation and defects. The following scores were then allocated to allow the determination of a percentage figure. Score: Description: 10 Grade A 9 Grade A with a defect 8 Grade A with pigmentation or fleshing deficiency - 7 Grade B 6 Grade B with a defect 5 Grade B with pigmentation or fleshing deficiency 4 Grade C O Rejected gggdominal fat weight, gonad and oxriduct weight determinations: The carcasses were placed in a freezer until further meiasurements could be made. A limited number of birds were thzuded each day to allow the determination of abdominal fat Weight, oviduct weight and gonad weight. The fat surrounding thee;proventriculus, gizzard and the pad of fat found in the abdcnninal area was removed and weighed in grams. The testes were :removed, trimmed of extraneous material and the weight recoxwied in milligrams. With the females. a similar procedure Was fxallowed for determining the weight of the oviduct. 23. Statistics: All data were statistically analyzed using the Duncan Multiple Range and Multiple F Test. Duncan (1955) and the Analysis of Variance and "t" Method. Snedecor (1946). RESULTS Body weight and feed conversion: Statistical analysis of the body weight groupings of the 1,024 birds selected at the end of the 35 day control period showed no significant differences except that of sex. Accordingly, weight differences at the end of the ninth week can be interpreted without reference to differences in the pre-treatment period. At the ninth week, the body weights were still highly uniform. No differences between replicates had appeared nor 1vas there any interaction between sex and treatment. The initial sex difference was somewhat larger. Now, however, :1 highly significant effect of treatment had appeared, Table 4. The birds on the 16 percent protein ration were from 5 -— 8 percent lighter than those in corresponding groups on thus 20 percent protein ration, Table 5. These differences welre not always significant, however, when the body weights are; separated according to protein level. as in Table 5. no (effect of the hormone or tranquilizer supplements on body Weight can be demonstrated. However, when the body weights are éieparated by sex, Table 6, it appears that the oral admitlistration of Lipamone either to the males or females brouéfilt body weights of the low protein recipients up to the bOdY'lfieaights of the high protein control fed birds. When the 25. tranquilizer was orally administered in combination with Lipamone. this effect was noted only in the males. A statistical analysis of the body weights at 13 weeks of age for both males and females receiving all eight dietary treatmens is found in Table 7. These data indicate a highly significant difference in weight gain due to sex while there was no statistical difference between the treatments; thus, the addition of Lipamone or Quietrol had little or no bene— fit on weight gain. Table 8. A statistical analysis of body weights at 13 weeks of age between the males receiving the eight dietary treatments (Table 9) showed the males receiving high protein (16 percent) inisal plus Lipamone and Quietrol in combination and Lipamone BIJDDB were slightly heavier than the other treatments. This wo111d infer that Lipamone plus a protein intake of 16 percent is zas efficient at this age as any other combination inves- tigxited. An analysis of the body weights of the females .huiicates a significant difference between the two levels of Inmrtein administered and apparently the additives had little or 110 influence on body weight between 9 and 13 weeks of age. The effect of treatment on mean body weights of males Eindfemales is given in Table 10. This table is devoid of statisstical analysis in an attempt to show the influence of the dimetary treatments on mean body weights for a simple Compalfiison.of the results at both ages. A. comparison of-the weight gain and feed efficiency 0f the males during the growing period. 5 to 9 weeks of age. Table 4 — Analysis of variance for 9—week body weights as replication and affected by sex, treatment. interaction. 26. F value Source of Variance d.f. m.s. F p‘<;0 05 Total variance 1,016 Sub—class 31 539.267 27.80** .47 Sex 1 14.676 838 756.58** .85 Treatment 7 269.938 13.92** .02 Interaction 7 21.552 1.11 .02 Replication 1 560 0.0285 .85 EFITH‘ 985 19.399 1 I? value with two asterisks indicates significance at P <;(LCH.level using the Analysis of Variance. 2 .moanhm> mo m_mz~m:< one wcwm: ~m>o~ Ho.Qv.m an mosmowquwwm m epwowccw mamwampmw 039 new: os~w> hm .mpmme m oaafle~sz was mwnmm maafipasz amocsn one 22 Ho>o~ Ho.0uvm pm pzmammmwc mdpcmomw_:wwm mam scam oswm map an cohoomamczz p0: manoZH **m.em mmO. meo~ maQH meo_ mama omma omma emma see: me we as me me we so we mesa: we sepazz a a . a a z 2 z 2 New mesan> a can can; on; an; on; one; on; an; paeapnese gAmawhw m.sm m.n:ama no Moshe .cpmv house wcmcamommc :« vegans Awashw :Hv mama: quooama ©~V :«maosn 30H wcw>weoes .cmqoon mmpmoflanmh .mm~wamm can mean: **m.em oaaa mafia mmaa «mam mama emma mmma emma new: we we we we we no em we new“: we tenasz a a a a z z z z , New me=~e> a on: on: one: am: one can an: 04mm sceaenone HAmEMMw v.vm m names mo Moshe .cpmv Macho wawcaoommc :u coacmh Awashw :fiv mums: ApcmOth omv :mopoam aw”: wcfi>wmooa .coaooa moanew~nwh .mwamsmm was mean: ll ‘ I’ll! , \‘II ‘II' mmm no ammo: a 9m mcmxomao mo wnwwmz kuon :0 ~m>m~ Camacha ho mHOQMHo 0:5 II n sagas 28. .eosmflsw> mo mwmxfimc< map wcwm: ~c>o~ ~o.oth pm moanonwcwwm casewccw mxmwamemw 03p new: m=~m> b N .mpmme m mHQfleasz cam omens mdawpfisz amoczn $59 %n ~m>o~ Ho.ouvm as acmhmmmwc kfipamofiww:m_m was mcw~ mean one an concomumcas we: mane: H **mm.~ mmo.~ mvo.~ mwo.~ who.~ o-.~ m-.— mm~.~ ¢m~.H saw: mm mm we mm so so me we was“: mo honszz mesaa> a ens one; ans an; an: on: ‘ one: an: eneapneee ~Amamuw Hm.¢~ w.n:mma no scape .cpmv Macho waflcsmomou am vegans Awashw :wV use: topmowcqfi acoaumoap .co~ooa mopMOMHQmH new: we~mamm **mb.m mbm.~ 0mm.~ 0mm.~ vmm.~ mwm.~ nmm._ mmm.~ mmm.~ . 2mm: mm ww we am am no we vw mchmn MO #03852 m03~m> & on; can: on; AAA om: mam am: can: unoEPane Awashw m©.m~ m.u:mos we gonna .oemv house wcwunoomoc :H vex:8h Amawsw may meme: cemeMUQw uamapmehp .mmaoom meQOMHQoM sew: mean: II 1“ . O mwfl H mMmmS m an mmamsmh vnm mmawfi %0 p£ww®3 zcon no pfimfipmohv Ho muomhho 03% I w UHQQE Table 7 — Analysis of variance for l3—week body weights as affected by sex. treatment. replication and interaction. Source of variance d.f. m.s. F1 g Béége Total variance 501 Sub—class 31 1,020,090 2.93** 1.49 Treatment 7 920,373 2.65* 2.03 Sex 1 23.762.271 68.43** 3.86 Replication 1 13,516 0.03 3.86 T x S 7 90.685 T x R 7 49,167 11 x S 1 347.248 17.x R x S 7 11,170 IError 470 53.239 F value with one asterisk indicates significance at P4{0.05 level, two asterisks indicate significance at P<10.01 level using the Analysis of Variance. 30. mwmzfimc< one mamm: ~o>o~ #0. ~e>e~ Ho.o.vm em eceeeeaae .oosmwsw> Ho eqvm an oocwo_mw:w~m easemuzw mammawgmw 03¢ :pH3 03Hm> L m .mpmoe h manwpfisz can owned mamwefizz :moszn one x: z~esmo~mwcw~m ohm mafia mesa one A: newcomaocas we: memo: a mesa amen mama Heom avom seem can: mm an mm mm Hm mm meeap we nonezz a a z z z z xem on; an; as; one; egg on; eceaenene no sense .cpmv nacho wcwccmommc am vegans Amawhw GMV mane: A9200hmm mfiv :~owo&g 3o~ wcw>fiooea .cofioom mepwow~amh .moamaom was mean: **©m.m~ coma mmGH om Om m m me=_e> a on; one; mamaw . .uswma aA me ow m **mh.mm mafia boma mm 0m 5 m me=~n> a am: on: HAmaasw ©m.mm \ «cams omma oama mama mmmm mmmm mwmm use: an mm mm am mm mm message senasz a a z z z 2 wow one: an: 0mm 0mm am: can: geeseneee mo posse .cwmv sense w:_c:eommc cw cexcws Amamhm :wv memo: Austhom odv :Hmpoaa aw“: w=~>~oomu .cefioon mmpmo-nmh .mm~wsmm was mw~mz I) '1' l (If! I 1" )III‘I“ .mww Ho mx®m3 m~ pm unwwmz kuon :0 ~®>0~ Samsonm Ho mpomwhm was I m QHQSE 31. we n_ns~nc< ecu w:_n: ~e>e_ _o. an _e>ea Ho.OJVn an asenema_e .mocawhm> Ouvm pm eocmo_w_:w_m manages“ mxmwhmemm 039 new: o=~m> & m .mwmoa & oaawpfisz cam swam: mfinfiwHSZ smocsn one z~pcmOmmqnwwm ohm o:_~ oswm one %Q cmaoomaoccz we: mane: ~ *xmv.m ©O©~ mmw~ mmofi ~©©~ Hmvfi somfi wmmfi ova saw: 0m 0m mm ~m mm on fin mm was“: mo ampasz Mosaw> m on; can; 0&4 and 0mm can out: an: pcoapwose Amsaaw mm.mm m nucmos Ho aoaao .cpmv Macao wcwccmomoc cw cmxdwu Amamhw cwv mane: ~ copmoficzq pcmspmmap .cedoon mmpwo_~noa new: meHwaem **mo.o~ mwm~ deem seem snow m~mm mmmm mmmm mmmm awe: mm mm an mm mm Hm mm mm moses no smnasz me=~n> a as; ones on; on; om: can an: 04mm eneaeeeee HAmeam ~©.>¢ m.u=wma «0 sense .cumv house wcflczmomoc a“ vegans Amamaw :wv mane: oopmowcqw pamaumohp .cmmoon magnum~nmh new: me~a2 .ewm me name: mH pm mwfimSmm can mmfiwa Ho p=m_w3 xcon no pCoaemmhp Ho mpoomme 0:5 I m mfinaB meeH seem meoa omma ans mama awom maoa ommm one; ~e©~ mmm~ whoa amma an; coma swam mmoa mama on; ame_ mmmm o-~ sand on: emma mmmm mmaa emma one: cams mmmm amaa mmma 4mm soma mamm mafia mama om: hmamawv “manhwv Amaahmv Awashwv meansmm mafia: mmawaem mean: escapwese own mo mxem3 mm 9< 0mm Ho mxem3 m p< 1.111....“ m— can a pm mmmmamm can mo~ma mo uawmm3 kcoa Emma map :o HuwapmeWwHWOHMWWW% o 1 ~ eases 33. Table 11 — The effect of treatment on weight gain and feed efficiency of males and females during the grow— ing period (5 to 9 weeks of age) Average gain Percent Treatment in grams* Feed/Gain improvement Males (in descending order of efficiency) _ HPLQ 755 3.10 18.6 HPL 758 3.21 15.7 HPQ 722 3.29 13.6 HPC 716 3.30 13.4 LPQ 668 3.54 7.1 IJ“) 672 3.60 5.5 LPLQ 661 3.61 5.2 IQPC 645 3.81 O IPemales (in descending order of efficiency) HPQ 579 3.44 17.5 IIPL 587 3.47 16.8 11F“) 575 3.59 13.9 HPLQ 570 3.63 12.9 LJ’L 524 . 3.91 6.2 LPLQ 496 4.09 1.9 LPQ 501 4.12 1.2 LIDCi 486 4.17 0 * Ebttremely small birds (runts) were eliminated in the calcu— lEltion.of average gain but included in the calculation of feed efficiency. 34. Table 12 — The effect of treatment on weight gain and feed efficiency of males and females during the finish— ing period (9 to 13 weeks of age) Average gain Percent Treatment in grams* Feed/Gain improvement Males (in descending order of efficiency) HPL 945 3.76 19.7 HPQ 913 3.86 17.5 HPC 905 3.90 16.7 HPLQ 957 3.94 15.8 LPLQ 777 4.38 6.4 LPQ 788 4.50 3.8 LPL 723 4.64 0.9 LPC 783 4.68 0 Females (in descending order of efficiency) IIPL 703 4.22 19.3 IIPQ 678 4.30 17.8 HPC 693 4.53 13.4 HPLQ 709 4.61 11.9 LPL 619 4.87 6.9 LPQ 614 5.09 2.7 LPC 570 5.15 1.5 LPLQ 578 5.23 O *— Iixtremely small birds (runts) were eliminated in the (lalculation of average gain but included in the calcula— 1Dion of feed efficiency. 35. is shown in Table 11. The high protein (20 percent) produced a 6 to 11 percent improvement in feed conversion compared to males receIVing the low protein (16 percent) basal. The addition of Lipamone in combination with Quietrol may be advantageous when added to a high protein ration. The addi— tion of Quietrol to a low protein (16 percent) ration suggests a protein sparing effect as evidenced by a 7 percent improve— inent in feed conversion over that exhibited by the low protein 'basal control. A similar comparison of the weight gain and feed effi— (3iency of the females during the growing period. 5 to 9 weeks (3f age, is shown in Table 11. Again the higher protein in— ‘take produced poultry meat more efficiently and the addition ()f Quietrol administered singly proved the most efficient. 1918 addition of Lipamone appears to be slightly less efficient and the combination of Lipamone and Quietrol less efficient tllan either administered alone as an additive to a high P170tein ration for females. When a 16 percent protein ration is; fed with Lipamone. feed efficiency is improved 6 percent (vaer the low protein basal control. Quietrol alone and in combination with Lipamone did not improve feed efficiency muOh over that demonstrated by the low protein basal control. A comparison of the weight gain and feed conversion of tller males during the finishing period. 9 to 13 weeks of age. is sihown in Table 12. The high protein basal proved to be 9 'tC) 13 percent more efficient than any treatment with the low protein basal. The oral administration of Lipamone and/ 36. or Quietrol as an additive to a high protein basal improved feed efficiency to a greater extent than the high protein basal control and the latter was slightly more efficient than the combination of Lipamone and Quietrol. Again the addition of Quietrol to a lower protein basal ration seemed to exert a protein sparing effect. This is evidenced even in combination with Lipamone which in this particular study was the most efficient when added to a low protein basal and fed to males. A similar comparison of the feed efficiency of females (during the finishing period. 9 to 13 weeks of age, indicated tine high protein level produced poultry meat with less feed tllan a lower protein level. Lipamone as an additive to the h_igher protein level was slightly more efficient than the acidition of Quietrol but the combination was less efficient t11an.the high protein basal control. The lower protein level (1:3 percent) presented the same pattern as the higher protein ITitzion but to a lesser extent. The Lipamone additive im— Prfiaved feed efficiency approximately 7 percent over that recorded by the females receiving the combination of Lipamone and Quietrol. §EEEEE§ry of live scores: Only the males receiving Quietrol and Lipamone in com— billEition showed any improvement in mean feather score between the ninth and the thirteenth weeks of age. The males fed the reunaining treatments and the females. regardless of treatment adnlinistered, showed a decrease in mean feather score between 37. the two periods. Fleshing was improved by the administration of the high protein basal control to the males, and the high protein basal in any combination with the additives improved 'the fleshing score of the females. Observed pigmentation scores were higher at 13 weeks of age. The males receiving 'the lower protein level exhibited an increased amount of jpigment deposition. This is consistent with serum xanthophyll (ieterminations discussed in another section. Since it is questionable whether any of these trends are statistically EBignificant. the data are not presented. IDressed carcass grade: The results of the carcass grading for both sexes and £111 eight dietary treatments after conversion to a percentage fiigure is found in Table 13. The dressed grade of chickens Iweceiving a high protein ration apparently can be improved ‘b37 the addition of Quietrol to the ration. The males receiv— 111g the high protein basal plus Quietrol had a dressed grade which was significantly higher than those males fed the high IJIWDtein basal control. No other differences were observed. Abdominal fat we ight: A statistical analysis of the abdominal fat. removed and weighed from 12 birds of each sex and representing each C’f' the dietary treatments indicated a non—significant differ— 91109. Table 14. The addition of an estrogen or a tranquilizer It‘3 either a high or low protein ration did not increase fat deposition in the abdominal area of either sex. Serum xanthophyll: The serum xanthophyll content of the blood collected from 9-week old males receiving any combination of the 16 percent protein basal ration was significantly higher than the other groups, Table 15. The females fed the high protein ‘basal displayed the least amount of xanthophyll in the blood serum. At 13 weeks of age, the groups fed the lower protein Iration continued to show the higher serum xanthophyll content. (Phe sexual difference in serum xanthophyll was not as pro— Ilounced at 13 weeks of age as at the earlier age. (ionadal. oviducal and comb weights: The mean weight of the testes removed from the experi— mental chickens at nine weeks of age is shown in Table 16. 'Iflle testicular weight of the cockerels receiving the combi— :ruation of dienestrol diacetate and Quietrol was equivalent ‘t() the testicular weight of the control birds. The testicu— lirr weight of the cockerels fed either additive singly was Siggnificantly lighter than the controls demonstrating a phy— Sitalogical effect of the oral administration of either compound at 'this age. As indicated in this table. protein level was omitted aiui Eln.analysis of the effect of treatment only is observed. A~55'tatistical analysis of testes weight at 13 weeks of age indiOates a justification of the previous assumption that protainlevel of the ration has little or no effect on the weitgkrt of the gonads. The mean weights of the testes from 12 1Jirds on each of the dietary treatments show little effect 39. on testes weight due to protein level. Table 17. The addi- tion of the estrogen. however. decreased testes weight approximately 80 percent and the tranquilizer administered singly reduced testes weight 84 percent as compared to the testes of the control fed birds. The combination of the two additives increased testes weight slightly over 100 percent in the high protein basal and almost 50 percent in the low protein basal treatments . Figure 1 illustrates the histological changes of the testes following dietary additives. Figure l—A represents a section of a normal testis removed from a male receiving the Control ration. Approximately 15 percent of the seminifer— ous tubules contain spermatids. The other tubules exhibit Various stages of spermatozoa. Figure l—B. at a magnifica— tion of 247 times (identical in all photographs) clearly demonstrates the decrease in the size of the tubules and the lack of sperm activity when the male was fed dienestrol diacetate. The majority of seminiferous tubules are in the Drepuberal state, showing small tubules with a single layer of cells and no apparent multiplication of the basal layer. The males fed Quietrol, Figure l—C, are very similar in appearance to the dienestrol diacetate recipient birds in the organization of the tubules and the lack of multiplica— tion and growth of the primary spermatocytes. Figure l—D aptly demonstrates the increase in size of the seminiferous tubules of the tests removed from males administered the Combination of dienestrol diacetate and Quietrol. All of 40. tubules exhibit multilayered epithelium representing various stages of spermatogenesis. From the wall of the tubule to the lumen are found spermatogonia. primary Spermatocytes. secondary spermatocytes, spermatids, the nutritive cells (cells of Sertoli) to which the spermatids are attached, and ‘the spermatozoa. The connective tissue stroma between the ‘tubules contains the interstitial cells (Leydig) and blood vessels. A statistical analysis of ovarian weight at 9 weeks 01? age is given in Table 18. A significant difference (5 peyrcent level) is observed between the ovarian weight of hi rds receiving the tranquilizer compared tithe weight of tlleeovaries removed from birds fed the other treatments. Tllfa additives fed singly tend to depress ovarian growth but When fed in combination indicate a lessening of this sup— p)rwession. These data would suggest that the tranquilizer has more influence on the hypophysis than that exhibited by tile! administration of kienestrol diacetate. The effect of the oral administration of Lipamone or Culiéatrol. fed singly or in combination. on the mean oviducal Weight of the 9—week old pullets is shown in Table 19. Ei— tk"3r' additive, when administered singly. significantly increased Oleilloal weight in the recipient pullets whereas the combina— tiOIl ‘vhen fed to pullets caused the oviduct to be very similar 1“ weight to that exhibited by the control fed birds. In thies ‘table, protein level was omitted and an analysis of the effect of treatment only is observed. A statistical analysis 41. of oviducal weight at 13 weeks of age (Table 20) again in- dicates that protein level of the ration has little or no effect on the weight of the oviduct. Oral administration of estrogen increased the oviduct 'weight of l3—week old females 160 percent or more as did the inclusion of the tranquilizer in the high protein basal com— pared to that observed in females fed a control ration (Lemonstrating the physiological effects of the two additives CHI oviduct weight. The low protein basal plus the estrogen ixlcreased oviduct weight 130 percent or more whereas the tranquilizer increased the weight of the oviduct only 50 per— <3€n1t over that recorded for the low protein basal control. Tileecombination of estrogen and tranquilizer increased ovi- (itmct weight approximately 16 percent. The effect of protein level and treatment on comb ‘veaights of males and females at 9 weeks of age is shown in These data suggest that protein level has little How- VPzible 21. ()I‘ no effect on comb weight of either sex at this age. e'V’er. the addition of Lipamone or Quietrol to the ration Significantly reduced the comb weight of the males below that ‘iXTLibited by the control birds or those fed the combination. Thu? :same trend is apparent in the comb weights of the females at 'tllis age but the differences are not as decisive as in the DISCUSSION Bodygweight gains and feed conversion: The results of body weight gains in this study are in agreement with that observed by other workers, that is, that a higher protein intake resulted in improved rate of gain and improved feed efficiency when compared to that obtained with a lower protein ration. However, the birds fed the lower protein basal control had a dressed grade score as high as any group of birds receiving the other dietary treatments. These data do not agree with that reported by Bird (1948). Who observed lean, blue birds lacking in finish when fed a lower protein ration. Lorenz (1954) reported a greater res— ponse to the various treatments when fed in combination with a lower protein ration. In the experiments reported herein. the dressed grade of male carcasses were slightly improved by the addition of the tranquilizer to a low protein ration indicating a limited reSponse to the treatment. Estrogen administered singly tends to decrease meta— bol ism (Bird. 1946) and activity and the fattening effects may be attributed, in part, to these factors. In combination With a tranquilizer. which is alleged to diminish activity and motivation. further reduction in rate of metabolism can be expected (Wolterink g_t_ a_1_.. 1958). A reduction in meta—- bolic rate and the fattening effects may be expressed in the Slight improvement of feed efficiency exhibited when Lipamone 43. and Quitrol are fed compared to the feed efficiency of the birds not receiving the additives. Numerous investigators have established that the ad— ministration of an estrogenic compound to chickens decreases the secretion of FSH and a balance is therefore maintained depressing the secretion of androgen accounting for the capon-like appearance of hormonized chickens. A smooth "silky" texture is imparted to the dressed carcass by the increased subcutaneous fat and by the effects of estrogen on skin structure (Lorenz, 1945). The relative effect of estro- gen treatment on gain in body weight increases with increasing age of birds. Estrogenic substances mixed in the feed in moderate dosages have had no effect. on the average. on rate of gain in growing birds (Lorenz, 1945). The results of this immediate study show a similar response was obtained from the administration of dienestrol diacetate and Quietrol. singly and in combination. The mechanisms of these effects are undoubtedly complex. Stimulation of lipogenesis may exert either a positive or a negative effect on total growth, depending on whether fat is deposited in addition to, or substituted for, other tissue. The Stimulation of lipogenesis will have a net negative effect on feed efficiency in either case, because of the higher CaloI‘ic content of adipose tissue. The increased feed con— sumption together with decreased energy expenditure due to the inhibition of aggressiveness, should favor accumulation or Weight and higher feed efficiency. A hypothesis suggested 44. by Bird (1949) is the possible relation of estrogen to the basal metabolism rate; a low rate of "metabolic efficiency" is requisite for the deposition of fat. Another theory is the stimulation of growth-hormone production or an adrogen— like action of estrogens in promoting an increased anabolism of protein, accompanied by loss of fat (Kumaran and Turner. 1949). Abdominal fat: Numerous investigators have reported that estrogen in— <3reases the amount of fat deposited in the tissue. particu— ;1arly abdominal fat. and improves the grade of the carcass ()f the male. Begin and Grainger (1957) observed the amount ()f abdominal and subcutaneous fat found in the carcass was ggreatest in the diethylstilbestrol—treated birds. The above Eiuthors stated that the fat was more evenly distributed in tdle birds fed dienestrol diacetate. Warden (1958) reported tllat DES implants significantly increased the amount of abdominal fat over that exhibited by birds fed dienestrol Ciiaacetate. The results reported herein, substantiate the Ilrwavious findings by the slightly heavier body weight of therbirds receiving the estrogenic compound singly and in CCHDIJination with the tranquilizer and the non—significant difference in abdominal fat weights shown to exist between tre atments and between sexes. This even distribution of fa13 11as economic significance by improving carcass grade and appearance besides a slightly improved rate of gain compared tc’ IJirds not fed either additive for at least a four week 45. period before slaughter. Serum xanthophyll: Blood serum from the male showed a higher concentration of xanthphyil than that found in the serum of the female. independent of the protein content of the feed. Also a lower protein level fed to males resulted in a higher xan- thophyll concentration than males fed a slightly higher protein level and slightly higher than that found in the blood serum of the females. The differences are possibly due to the greater amount of yellow corn in the lower protein ra- tion. This gives further evidence that xanthophyll content of the feed is one factor responsible for the amount of pig— mentation that appears on the finished product. The sexual differences in serum xanthophyll may be partially explained by the greater amount of feed normally consumed by the males. The males consumed approximately 10 percent more feed than the females during the 5 to 9 week period and 20 percent more feed during the 9 to 13 week period. The percent difference in serum xanthophyll between sexes at 9 weeks of age was 28 percent and 8 percent at 13 weeks of age. Obviously the expression of some other mechanism is involved. Preliminary research done in this laboratory (unpub— lished) suggests that capons have approximately the same :xanthophyll content as intact males, virtually eliminating arutrogen as the factor responsible for the sexual differences shown to exist. The oral administration of an estrogenic compound had little or no effect on pigment deposition or 46. serum xanthophyll suggesting the apparent elimination of an estrogenic influence on the sex differences in xanthophyll. The xanthophyll content of the blood serum remains at a higher concentration in the male compared to female at least to one year of age (unpublished), which would allow sufficient time for the expression of hormonal secretion of either sex. The author concludes from these data that the sexual differ- ences in visual pigmentation and serum xanthophyll concentra- tion are specific and a conclusive explanation for these results cannot be expressed until further experimentation is done on this particular occurrence. Gonadal and oviducal weights: Kumaran and Turner (1949a and 1949b) studying the histology of the testes of White Plymouth Rock males at various ages. reported that during the first 5 weeks, the seminiferous tubules are organized. followed by multiplica— tion of the basal layer of cells (spermatogonia). The primary spermatocytes begin to appear about the sixth week. During the next 2 or 3 weeks, growth of the primary sperma— tocytes takes precedence over the further multiplication of the Spermatogonial layer. The secondary spermatocytes begin to appear at about 10 weeks of age as a result of the Ixniuction division of the primary spermatocytes. Spermatids Kldmmature spermatozoa) begin to appear in the seminiferous tubules at about12 weeks of age, and by the twentieth week guns usually present in all the tubules. Microscopic examination of representative testes 47. removed from males receiving the control basal ration shows Spermatocytes in various stages of development from primary to immature spermatozoa. however. the number of tubules showing spermatozoa was very limited. The oral administra- tion of an estrogenic compound depressed the proliferation of germ cells. The tubules are organized, and very little if any multiplication is observed: In no case was Sperma— tozoa observed. Exogenous estrogen in unphysiological dosa— ges is a potent inhibitor of gonadotrophins produced by the anterior—hypophysis. The dosages of dienestrol diacetate used in this experiment were highly unphysiological, hence complete inhibition was produced and the observed results expected. The result of such treatment is an impaired genital system which simulates that observed in an animal after hypophysectomy. A representative testes from a male receiving the oral administration of Quietrol shows stages of development very similar to that exhibited by the males fed dienestrol dia- cetate. These results present strong evidence that the tranquilizer used in this study blocks the secretion of the hypophyseal gonadotrophins. Pino and Hudson (1953) reported a similar phenomena when Enheptin (2-amino, 5-nitrothiazole) was fed to White Leghorn pullets and cockerels just prior to sexual maturity. These same authors state that Enheptin completely blocks sexual maturation when fed to immature chickens; and. as long as the drug is continued, the birds fail to overcome its effects. These authors also reported 48. that a histological examination of testis sections taken at autopsy substantiated the gross observations of sexual in— volution. While spermatogenesis and testes conditions were normal in the controls, there was a regression in the drug— treated tubules to one layer of cells, presumably Spermato— gonia, lying next to the tubular wall. Tubule size was reduced and the interstitial tissue appeared fibrous and condensed. In addition. these authors found that not only in hypophysis size reduced significantly in Enheptin-treated chickens, but so is the total hypOphysis gonadotrophic hor— mone potency. In this experiment, microscopic examination of repre- sentative testes removed from males receiving the combina— tion show that almost all of the seminiferous tubules contain spermatids. The tubules and interstitial tissue appear normal indicating that the increased size was not due to just FSH or LH secretion alone but rather a simultaneous inten- sified elaboration of gonadotrophins. Abnormal spermato— genic cells are found. In the spermatogonia this is manifes— ted by an excessive hypertrophy. Among the spermatocytes. giant forms are also common, as well as cells with two or more nuclei, sometimes of unequal size. They arise from fusion or abnormal mitosis. The Spermatids fuse to form multi— nucleated giant cells (Maximow and Bloom, 1957). The degenerating and monster spermatogenic cells are carried, with the mature spermia, into the epididyimis, where their substance is perhaps reabsorbed by the epithelium 49. of the excretory duct. The sex cells of the seminiferous epithelium are sensitive to noxious factors of various kinds. In pathological conditions of general (infectious diseases. alcoholism, dietary deficiencies) or local (injury. inflam— mation) character. and under the influence of mental depres— sion, the degenerative changes. especially the formation of multinucleated giant cells by the coalescing spermatids. may become prominent (Maximow and Bloom. 1957). The occurrence of a many—fold differential when two compounds are fed in combination is not uncommon. For exam— ple. the administration of FSH at a given dose level is known to increase testes size. A comparable dosage of LH. fed singly. will result in a similar accretion of testicular tissue. When the two compounds. FSH and LH. are fed in combination. an increase above that of an expected two-fold increase in testes size can be found. The significance of 'the results found in this study are directly the opposite of 'these conventional observations. When dienestrol diacetate (1r Quietrol was fed singly. reduction in testes weight was «dbsemwed. To be specific, the reduction in testes weight ccnupared to control values was 80 percent when dienestrol dieucetate was fed and 84 percent for Quietrol. When fed in confliination the expected results would be an equivalent re— ductxion or possibly a greater suppression than when either compound was fed singly. The data, however, demonstrate that “the additives. fed in combination. counteract each Otku3r' and in addition. stimulate FSH and LH release. hence resulting in nearly a two—fold increase in testes size when compared to the testes size of birds fed the control ration. When the combination of dienestrol diacetate and the perphenazine derivative was administered. it is postulated that the secretion of FSH and LH is stimulated, increasing gonad size and maturation compared to that expressed by either compound fed singly. This may not necessarily be a stimulated FSH and LH release but could be. theoretically. an increased sensitivity of the end target organ, the testes. to respond to the same amount of circulating gonadotrophins. then either compound is fed singly apparently the secretion ()f hypOphyseal gonadotrophins (FSH) are suppressed resulting in.the conventional decreased gonad size. The results of (lamb weights that were recorded at 9 weeks of age indicate tfliat the combination increased the weight of the comb compared in) the weight of the comb from birds fed either of the com— runxnds singly or the control birds. This would suggest that 13H? increased secretion of gonadotrophin (LH) stimulated the intuerstital cells resulting in an increase of androgen secre— t1°On. hence, comb growth. Apparently the combination permitted some sedation aPPIWDaching mental depression to allow the formation of giant cells; that were found in the seminiferous tubules. The oral administration of dienestrol diacetate in— CreaSed oviducal weight as expected. This occurrence is Inwaslunabiy due to a reduction of hypophyseal gonadotrophin secretion (FSH) acting directly on the ovary which was slightly smaller than the ovarian weight recorded for the birds not receiving the estrogenic compound. The estrogenic— like constituent of dienestrol diacetate replaces the natural estrogen liberated by the ovary. hence. stimulating growth of the oviduct. Ovarian weight of the tranquilizer fed birds Ivas significantly less than the ovarian weight of the con— ‘trol fed birds. These data are strikingly similar to that :recorded for the birds fed dienestrol diacetate indicating 21 suppression of FSH and LH secretion reducing ovarian size {and hence. decreasing normal estrogen production of the ()vary. The oviducal weights. of the females receiving the ‘tranquilizer were heavier than those removed from the birds Ireceiving the control ration. The stimulated oviducal growth amust be due to the tranquilizer. the only additive to the tnasal control in this instance. indicating that the compound luas an estrogen~like property. When the combination of dienestrol diacetate and Quie— tlfol was fed. ovarian weight was very much like that recorded fcxr the birds fed the control ration. This would suggest that tile inhibition of hypophyseal gonadotrophin secretion ex— PIYessed by either compound fed singly was counteracted per— lniirting normal function of the ovary. Since both compounds ‘VGITB fed simultaneously at the same level. the dosage was airtue111y two times that administered when either compound Was fed singly. Possibly one compound blocked the expres— Sicnfl (3f the other and permitted the normal secretion of 52. estrogen from the ovary. giving a similar oviducal size of these birds as compared to those fed the control ration. These data concerning an estrogen and Quietrol. fed singly and in combination. would offer excellent tools in the study of reproduction in the fowl. Since it is possible to interrupt the reproductive cycle. apparently through en- (iocrine interference. a more thorough investigation of endo— (3rine inter-relationship can be made. SUMMARY AND CONCLUSIONS Dienestrol diacetate (a synthetic estrogenic compound incorporated at 21.14 grams per ton of feed) and Quietrol (an oral tranquilizer incorporated at 4 grams per ton of feed) were fed singly and in combination. together with a control at two protein levels. A higher protein intake permitted improved rate of gain and improved feed efficiency when compared to that obtained with a lower protein ration. A slight increase in body weight gain was observed when dienestrol diacetate and/or Quietrol were administered singly or in combination compared to birds fed a control ration. The addition of dienestrol diacetate and Quietrol improved feed efficiency slightly above that observed when a high or low protein basal control was fed. Either administered singly or in combination. the additives gave a non-significant difference in abdominal fat weight between treatments and between sexes. Blood serum from the male showed a higher concentration of xanthophyll than that found in the serum of the female. Also a lower protein level fed to males resulted in a higher xanthophyll concentration than males fed a slightly higher Protein level and slightly higher than that found in the blood serum of the females. These differences are further eVidence that xanthophyll content of the feed is one factor reSponsible for the amount of pigmentation that appears on 54. the finished product and the concentration of xanthophyll found to exist in the blood serum. Dienestrol diacetate physiologically suppresses hypo; physeal gonadotrophin release resulting in decreased testi- cular weight and ultimately comb size in the male and decreased ovarian weight with increased oviducal weight in the female compared to that exhibited by the birds fed the control ration. No noticeable tranquilization was observed when Quie— trol was administered at the given level. However, this level was physiologically active in reducing testicular weight and comb size in the male and decreasing ovarian weight in the female. These data suggest that hypophyseal secretions are depressed when Quietrol is fed. An increase in oviducal weight with a decrease in ovarian weight fol— lowing Quietrol administration suggests an estrogen—like activity by this compound. When dienestrol diacetate and Quietrol were fed in combination ovarian weight was very similar to that exhibi— ted by birds fed the control ration. indicating that the suppression of hypophyseal secretions, when either compound Was administered singly, was relieved. Oviducal weights indicated that when fed in combination. the two compounds counteracted each other eliminating their estrogenic acti— vity and giving ovarian weights similar to the controls. In contrast testicular weight was increased significantly. When the combination was fed. compared to testicular weight of birds fed the control ration. Microscopic examination of a representative testis removed from a male receiving the combination shows that almost all of the seminiferous tubules contain Spermatids at 13 weeks of age. The tubules and interstitial tissue appear normal indicating an intensified elaboration of the gonadotrophins FSH and LB or an increased sensitivity of the end organ. The number of tubules showing Spermatozoa was very limited in a representative testis removed from a male receiving the basal control ration. 56. L3.» de ‘ .k’.l.o -$. - iT; , a? "W‘ .‘a Vlaf’l’il : ’ O . _ I I W l . I a V \ ' . . 7 i I . 4 _ .‘ l‘: ', k ...L. ‘, .1! - ‘Ip'- . C a Figure 1. Cross sections of testes of cockerels showing the effect of the oral administration of Lipamone or Quietrol, fed singly or in combination, on testicular development at 13 weeks of age. (A) The normal or section of a testis from a cockerel fed the control ration. (B) The effect of feeding Lipamone on testes development. 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Table 16 — Effect of the oral administration of Lipamone or Quietrol. fed singly or in combination. on the testicular weights of 9—week old cockerels. l Testicular weight (in milligrams) in descending order (Std. error of mean = t 30.9 mg.) Treatment Combination Control Quietrol Lipamone Sample size 13 10 13 18 Mean weight 507 429 208 190 lMeans not underscored by the same line are significantly different at P<( 0.01 level by the Duncan Multiple Range and Multiple F Tests. 61. .mpmca a m~ampfizz use ewsam canmpfizz :sossn map an ~e>e~ Ho.o.vvm pm esoaomwmc aupswowmmswwm can mama seam one an cahoomaecsz we: mama:H eon mam mam ewe mmem seam seem meme enamez nee: Na Na Na Na Na Na Na Na eNmm eaasem can sea on: an: om: can ones 34;: pseseeeae Amamaww-~s v.v>w H name no Moshe .cpmv Macho w:_cseomec cm Amamawwaawa :MV pswwmz mopmce d .mamacxooo cfio amazimH no pnwmc3 mepmcp so scapmswnsoo s“ so kstwm 6mm Hoapmwsv so msosm9- use sowpma cap mo ambefi :flcpoam no mpommme was I Pd wanwe 62. Table 18 — Effect of the oral administration of Lipamone or Quietrol, fed singly or in combination. on the ovarian weight of 9—week old pullets. Ovarian weight (in milligrams) in descending order . l (std. error of mean 3 i 15-1 milligrams) Treatment Control Combination Lipamone Quietrol Sample size 9 17 14 13 Mean weight 302 295 277 243 1 Means not underscored by the same line are significantly different at P (0.05 level by the Duncan Multiple Range and Multiple F Tests. 63. Table 19. Effect of the oral administration of Lipamone or Quietrol, fed singly or in combination. on the mean oviducal weight of 9—week old pullets. l Oviducal weight (in milligrams) in descending order (Std. error of mean I g 35.04 mg.) Treatment Lipamone Quietrol Control Combination Sample size 14 13 9 17 Mean weight 429 355 185 171 lMeans not underscored by the same line are significantly different at P.4 0.01 level by the Duncan Multiple Range and Multiple F Tests. 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Barrett, 1945. The effect of feeding diethylstilbestrol to cockerels. Poultry Sci. 24:542. Thayer. R. H.. R. G. Jaap and R. Penquite. 1945. Fattening chickens by feeding estrogens. Poultry Sci. 24:483. Umberger. E. J. and G. H. Gass. 1959. Detection of resi- dual estrogenic activity in the edible tissues of chickens fed dienestrol diacetate. Poultry Sci.38:128. Warden. W. K., R. K. Ringer, L. F. Wolterink and P. J. Schaible. 1958. Estrogenic compounds for chickens. Feedstuffs. 30(36)86—87. Wilson, W. 0.. 1956. Identifying non—laying chicken hens. Poultry Sci. 35:226. Wolterink, L. F.. E. Speckmann. K. G. Hood. and R. K. Ringer. 1958. The effects of an oral estrogen—tranquilizer combination on transit through the digestive tract and on the intestinal absorption of radiophosphorous in broilers fed a low protein ration. Poultry Sci. Proc. 1958. EXPER/MENTAL DESIGN MaLe Pens Femate Pens /’“*““*"*r\ (”rrrrrw/“\ LPQ LPLQ HPLQ HPL. LPL- HPLQ HPC LPQ HPL. HPQ LPc. 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