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Methionine, Choline, Betaine and Fish Products in a Simplified Ration1,2
Methionine, Choline, Betaine and Fish Products in a Simplified Ration12 D. H. MISHLER, C. W. CARRICK AND S. M. HAUGE Departments of Poultry Husbandry and Agricultural Chemistry, Purdue University Agricultural Experiment Station, Lafayette, Indiana (Received for publication June 15, 1948)
moting properties in a purified diet and is not effective for perosis prevention, whereas choline supplementation resulted in excellent growth and complete perosis prevention. McKittrick (1947) has divided the choline and methionine requirements for the stimulation of growth into two parts, namely, the essential and the replaceable parts—the essential function of one of these supplements will not be replaced by the other. Glycine betaine would not substitute for choline or methionine in the essential part of the requirements but would substitute for either one in the replaceable increment. McGinnis, Norris and Heuser (1942) found that the occurrence of perosis in chicks was greatly reduced by the supplementation of their basal diet, which was not a purified diet, with betaine-hydrochloride. In fact, the incidence and severity of perosis, amount of gain, and mortality were approximately the same with either choline or betaine supplementation. McGinnis (1946) working with turkeys found that additions of gelatin increased the incidence of perosis, depressed growth and reduced the perosis-preventing function of betaine. It is the purpose of this report to present evidence concerning the growth promoting effects of fish products, betaine, choline, and methionine when used as
REVIOUS investigations at this Station have indicated a supplementary value for choline and methionine in a simplified corn and soybean oil meal ration. Gerry et al.- (1948) found these factors to be interchangeable to a certain extent, but that methionine was of somewhat more value than choline. However, Clandinin et al. (1946) using a somewhat more complex ration concluded that choline and methionine cannot be considered interchangeable. Using four different soybean oil meals they found that only one was improved by choline, whereas all four were improved by methionine additions. However, their experiment was terminated when the chicks were four weeks of age. Gerry et al. found that 4 weeks was not sufficient time for bringing out these relative values. Almquist and Grau (1945) state that even if high levels of choline are employed the addition of cystine cannot compensate for a methionine deficiency. These authors have indicated (1944) that cystine may be formed in the chick from choline or methionine. Moyer and du Vigneaud (1942) and Jukes and Welch (1942) have reported that betaine has only alight growth pro1 Journal Paper No. 359 of the Purdue University Agricultural Experiment Station. 2 This investigation was supported in part by a grant from The Borden Company.
24
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P
METHIONINE, CHOLINE, BETAINE AND F I S H PRODUCTS
supplements in a simplified ration, consisting largely of corn and soybean oil meal. EXPERIMENTAL
ground yellow corn, 25 soybean oil meal, 5 dehydrated alfalfa leaf meal, 5 distillers' dried solubles, 4 liver meal, 3 dried whey, 3 meat and bone scraps, 1 ground limestone and 0.5 steamed bone meal and 1 of a salt mixture consisting of 90 percent iodized salt and 10 percent manganese sulphate. D-activated animal sterol was included to provide 1 A.O.A.C. unit per gram of ration. The percentage composition of the basal experimental rations was 35 soybean oil meal in Trail I and 38 in Trials I I and III, 1.5 each of steamed bone meal and ground limestone, 0.5 of feeding oil (3,000-A, 400-D) 0.5 of a salt mixture (90 percent iodized salt and 10 percent manganese sulphate) and ground yellow corn to bring each ration to 100. Synthetic riboflavin was included in all of the experimental rations at a level of 300 micrograms per hundred grams. Supplements of 3 milligrams of nicotinic acid and 1 milligram of calcium pantothenate per hundred grams were included in all of the experimental rations in Trials I and III. Additional ingredients are listed in the tables. Each ration contained at least 21 percent protein. RESULTS Trial I. This trial, which was conducted during July and August 1947 was a paired experiment. It was designed to measure the comparative supplementary value of condensed fish solubles and a non-protein extract from the same amount of fish solubles, as well as the supplementary value of various levels of choline chloride and betaine. The variables in the rations and the results obtained are given in Table 1. The four-to-eight and six-to-eight week gains were treated statistically by analysis of variance. Trial II. This trial was conducted during September and October 1947, to test the comparative value of semi-solid
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Approximately 50 Barred Plymouth Rock chicks of both sexes were used in each lot within each trial. In Trial I, the chicks were fed the simplified diet without either fish or methyl group supplements for a period of four weeks, after which they were sorted into blocks on the basis of the two-to-four week gains and distributed into the experimental lots, where they were fed the experimental diets for the remaining four weeks. In Trials II and III the chicks were distributed into the various lots according to initial weight and sex. The all-mash rations and tap water were supplied ad-libitum and the chicks were reared in battery brooders with wire floors. The synthetic vitamins were dissolved in hot ethyl alcohol and water, mixed with ground yellow corn and dried. The DL-methionine was finely ground and mixed carefully with ground yellow corn. The condensed sardine fish solubles analyzed 33.3 percent protein and approximately fifty percent solids. Although some loss of moisture occurred in the stored samples, the additions of this product to the rations were adjusted to equivalent amounts on a dry matter basis. The semisolid fish product is a product resulting from the hydrolysis of undecomposed fish and/or fish trimmings from which oil may or may not have been removed. The sample used in Trial I I analyzed 34.2 percent protein and 50.92 percent solids. The amounts of fish solubles and semi-solid fish product will be expressed on the dry matter basis in both tables and discussion. The percentage composition of the positive control ration in each trial was 53
25
26
D. H. MISHLEE, C. W. CAERICK AND S. M. HAUGE
fish product and condensed fish solubles in the simplified corn and soybean oil meal ration containing supplements of riboflavin, nicotinic acid, calcium pantothenate, choline chloride and DL-methionine. Since it is possible that some deterioration might occur in fish solubles during long periods of storage, three samples of
and was a further study of the supplementary inter-relationship of choline, betaine and methionine in a corn-soybean oil meal ration containing synthetic riboflavin, nicotinic acid and calcium pantothenate. Previous results in this laboratory had shown that choline and methionine exerted some supplementary interaction, although methionine was of more
TABLE 1.—A comparison of choline, betaine and fish solubles (both sexes) Variable Ingredients (percent)
Lot 1
Av. gain Males 4-8 wks. (gms.)2 Females 4-8 wks. (gms.)2 Both Sexes 4-8 wks. (gms.)2 Males 6-8 wks. (gms.)2 Females 6-8 wks. (gms.)2 Both Sexes 6-8 wks. (gms.)2 Number of chicks at 8 weeks 1 2
Lot 3
Lot 4
Lot 5
Lot 6
0.1
0.2
Pos. Control
1.0
Lot 7
Lot 8
Lot 9
0.1
0.2
0.3
291 280 286 142 145 144 47
346 307 327 171 155 163 47
382 365 373 172 172 172 47
.07 288 238 263 147 112 129 47
379 340 360 210 179 194 47
321 284 302 177 148 162 47
466 355 411 230 162 196 47
Non protein extract from 1.0 percent fish solubles Approximate least significant differences: 5% level 4-8 wk. Males Females Both Sexes 6-8 wk. Males Females Both Sexes
fish solubles were compared. One sample had been stored for a year in a refrigerator at 36°F. (Lot 13); a second sample from the same barrel was stored for a year in a garage (Lot 14); and a third sample was from a new supply (Lot 15), which was the same as that used in other lots in this trial. Since the work of Mishler et al. (1948) had indicated that male and female chickens responded differently to additions of fish solubles, both sexes of chickens were used. The variables in the rations and the results are given in Table 2. Trial III. This trial was conducted from December 1947 to February 1948,
47 31 26 34 23 18
grams grams grams grams grams grams
346 282 314 167 130 149 47
309 283 296 144 144 144 47
1% level 62 41 34 44 31 24
grams grams grams grams grams grams
value than choline as a supplement to a corn-soybean oil meal ration with fish solubles. Since the results of Trial I indicated the effectiveness of higher levels of betaine a further test of this product in the simplified ration was undertaken. The variables in the rations and the results are given in Table 3. DISCUSSION
The corn and soybean oil meal basal ration supplemented with synthetic riboflavin, nicotinic acid, and calcium pantothenate, and used as a depletion ration for all chicks the first four weeks, resulted in
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Choline chloride Betaine Fish solubles Fish solubles extract1
Lot 2
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METHIONINE, CHOLINE, BETAINE AND FISH PRODUCTS TABLE 2.-—A comparison of various levels offish products (both sexes) Variable Ingredients (percent) Fish solubles1 Semi-solid fish pdt. Nicotinic acid Ca pantothenate Choline chloride DL-methionine
Lot 11
3.0
3.0
+ +
Lot 12
Lot 14
Lot 15
Lot 16
Lot 17
Lot 18
Lot 19
0.5
0.5
0.5
0.5
1.0
1.5
4.0
+ + + + + + + + 0.15 0.15 0.15 0.15
+ +
0.15 0.15
0.15 0.15
569 465 517 869 679 774 49
574 492 533 851 740 796 49
417 418 417 646 559 602 49
Lot 13
525 495 510 824 747 786 49
561 479 520 880 737 809 49
0.15
0.15
0.15
0.15
554 544 549 880 838 859 52
605 551 578 927 811 869 50
644 536 590 985 801 893 51
633 542 588 977 799 888 53
589 490 540 913 747 830 52
Lot 20 Pos. Control
620 514 567 949 774 862 51
1 Samples used in Lots 13 and 14 were stored one year in a refrigerator and garage respectively. All other samples were from a new supply. 2 Approximate least significant differences:
5 % level 58 grams 49 grams 38 grams 86 grams 70 grams 55 grams
6 weeks Males Females Both sexes 8 weeks Males Females Both sexes
1% level 77 grams 64 grams 50 grams 114 grams 92 grams 73 grams
TABLE 3.—A comparison of various levels of choline, betaine and methionine (both sexes) Variable Ingredients (percent)
Lot 21
Choline chloride Betaine DL-methionine Average weight Males 6 wks. (gms.)1 Females 6 wks. (gms.)1 Both Sexes 6 wks. (gms.) 1 Males 8 wks. (gms.)1 Females 8 wks. (gms.)1 Both Sexes 8 wks. (gms.)1 No. of chicks at 8 weeks 1
Lot Lot 22 23
Lot Lot 24 25
0.1
0.3
0.2
0.2
357 310 334 533 447 490 48
442 358 400 649 527 588 46
437 393 415 671 582 627 47
427 392 410 680 581 631 50
430 431 431 658 640 649 49
Lot 26
Lot 27
Lot 28
0.3
0.4
0.3 0.15
43.9 443 441 647 655 651 50
485 443 464 738 656 697 50
443 494 469 696 734 715 50
Lot 32
Lot Lot 29 30
Lot 31
0.1 0.2
0.15 Pos. 0.15 Con0.15 trol
0.3 457 430 444 687 633 660 48
395 385 390 626 584 605 48
472 443 458 744 673 709 49
536 522 538 873 752 813 46
Approximate least significant differences: 6 weeks Males Females Both sexes 8 weeks Males Females Both sexes
significantly retarded growth of the chicks (Lot 1) throughout the eight-week period. A supplement of one percent condensed fish solubles (Lot 2) produced gains which
5% level 60 grams 43 grams 36 grams 82 grams 60 grams 50 grams
1% level 79 grams 57 grams 48 grams 108 grams 79 grams 66 grams
were highly significant over those of the negative control (Lot 1). A. nonprotein extract of fish solubles (Lot 3) also resulted in highly significant gains over the
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Average weight Males 6 wks. (gms.)2 Females 6 wks. (gms.)2 Both Sexes 6 wks. (gms.)2 Male 8 wks. (gms.)2 Females 8 wks. (gms.)2 Both Sexes 8 wks. (gms.)2 No. of chicks at 8 weeks
Lot 10
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D. H. MISHLER, C. W. CARRICK AND S. M. HAUGE
and Bacon (1946) showed that levels of fish solubles as high as 12 percent of the ration resulted in proportional increases in growth. Berry et al. (1945) reported no better growth with 4 percent condensed fish solubles than with 2 percent, but the rate of growth was less than ours. When the four synthetic vitamins and DLmethionine in our simplified ration were supplemented with various levels of fish solubles no better growth resulted with a 4.0 percent level (Lot 19) than with a 1.5 percent level (Lot 18) in the case of male chickens. However, with the female chicken optimum growth was reached at the 0.5 per cent level. These results are in agreement with previous observations by Mishler, Carrick and Hauge (1948), which indicated that males responded more favorably than females to increased levels of fish solubles. A taste test was conducted to determine whether a fishy flavor had been imparted to the flesh of chickens fed 4.0 percent of condensed fish solubles (Lot 19) or 3.0 percent of semi-solid fish product (Lot 11). Thirty chickens (ten from each of these lots and ten from the control lot (Lot 20) which received no fish product) were killed, dressed, halved and distributed to fifteen persons. No consumer had any previous knowledge of the rations fed to the chickens which he consumed. Only four persons reported fishy flavors and these were not confirmed by persons consuming the other half of the same chickens. Moreover, two of the reports involved chickens which had been fed the positive control ration containing no fish products. Therefore it is unlikely that any fishy flavor is transmitted from the ration to the meat. These results are in agreement with those of Berry et al. (1945). The soybean oil meal used in Trial III was apparently low in available choline as evidenced by the suboptimal growth in Lot 21. An addition of 0.1 percent choline
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negative control. The significantly slower gains which were made with the extract as compared with the fish solubles may have been due to some of an unknown factor having been removed with the protein. It is clear that the value of fish solubles in this type of ration is not from protein, since the ration is already adequately supplied with all of the essential amino acids, except methionine. A low level of fish solubles would supply very little methionine and the extract practically none. In our laboratory expeller soybean oil meals have varied somewhat in the growth response obtained from additions of choline and methionine. A level of 0.1 percent choline chloride resulted in increased gains which were highly significant over those of the negative control thus indicating the basal ration to be deficient in available choline. A supplementary level of 0.2 percent betaine (Lot 8) proved as effective a supplement as either 0.1 or 0.2 percent of choline, but at the 0.3 percent level (Lot 9) betaine resulted in gains which were highly significant over those produced by lower levels of either choline or betaine. A level of 3.0 percent of semi-solid fish product did not have any supplementary value in a simplified diet containing nicotinic acid, calcium pantothenate, choline chloride and DL-methionine(Lots 10-12). When a low level of 0.5 percent fish solubles (Lot 16) replaced the semi-solid fish products (Lot 11) growth at eight weeks was improved with high significance. When samples of condensed fish solubles were stored for a period of one year either in a refrigerator (Lot 13) or in an unheated garage (Lot 14), the tests indicated that little deterioration had occurred. If any difference was manifested as a result of storage it was shown in the slower growth of the male chickens fed the refrigerated product. Using a more complex ration, Lassen
METHIONINE, CHOLINE, BETAINE AND FISH PRODUCTS
SUMMARY
1. A study was made of the supplementary value of methionine, choline, betaine and fish products in a simplified corn and soybean oil meal ration supplemented only with riboflavin, calcium pantothenate, and nicotinic acid in addition to a simple mineral mixture and vita-
min A and D feeding oil. 2. Choline, betaine and methionine were all effective in supplementing the simplified ration. Betaine at 0.3 and 0.4 percent levels appeared to have higher supplementary value than either choline or methionine. 3. A three percent level of semi-solid fish product plus riboflavin in a corn and soybean oil meal ration was improved with high significance when calcium pantothenate, nicotinic acid, choline and methionine were added, indicating the product to be deficient in one or more of these factors. A three percent level of semi-solid fish product did not improve the simplified ration containing the four synthetic B-complex vitamins and methionine. 4. A level of 4 percent condensed fish solubles (dry basis) resulted in no better growth than 1.5 percent with male chickens. The maximum growth of the female chickens was at the 0.5 percent level (dry basis). None of the fish products imparted a fishy flavor to the flesh of the cooked chickens. ACKNOWLEDGMENT
We are indebted to Mr. R. H. King of the Department of Agricultural Chemistry for the preparation of the fish solubles extract, the Feedingstuff Laboratories of the State Chemist for the protein analysis of the condensed fish solubles and semisolid fish product, to The Borden Company for the betaine hydrochloride and to U. S. Industrial chemicals for the synthetic DL-methionine used in these experiments. REFERENCES
Almquist, H. J., and C. R. Grau, 1944. Interrelation of methionine, choline, betaine and arsenocholine in the chick. J. Nutrition 27: 263-269. Almquist, H. J., and C. R. Grau, 1945. Further studies on cystine, methionine and choline in chick diets. J. Nutrition 29: 219-222.
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chloride resulted in an improved growth response, which was highly significant. A level of 0.3 percent of choline chloride resulted in still greater growth. At similar levels betaine resulted in somewhat better growth than choline, although the differences were not significant. A supplementary level of 0.4 percent betaine gave a significant difference in growth over a 0.3 percent level in the case of the male chickens. A combination of choline and betaine was improved by the addition of a low level of methionine. However, when 0.1 percent choline and 0.2 percent betaine were added (Lot 29), growth was only slightly better than 0.2 percent betaine alone (Lot 25). When all three products were included at low levels (Lot 31) there was no better growth than when 0.3 per cent betaine and 0.15 percent methionine were included (Lot 28). These results have indicated that betaine is as satisfactory a supplement as choline for rapid growth, and that a combination of betaine and methionine may assume the growth functions of choline in the simplified ration used. The poor results shown with a level of 0.3 percent methionine (Lot 30) may indicate a deficiency of non-replacable choline. The control ration (Lot 32) resulted in a growth response which was highly significant over that produced by any of the experimental lots in Trial III. It is apparent that all of the experimental rations in this trial were deficient in one or more unidentified factors supplied by fish solubles.
29
30
BOOK REVIEWS Poultry Sci. 25:486-491. McGinnis, J., 1946. The influence of gelatin on prevention of perosis in turkeys fed choline and betaine. Poultry Sci. 25: 91-92. McGinnis, J., L. C. Norris, and G. F. Heuser, 1942. Effect of ethanolamine and betaine on perosis in chicks. Proc. Soc. Exp. Biol. Med. 51: 293. McKittrick, D. S., 1947. The interrelations of choline and methionine in growth and the action of betaine in replacing them. Arch, of Biochem. 15: 133-156. Mishler, D. H., C. W. Carrick, and S. M. Hauge, 1948. Condensed fish solubles as a supplement for corn and soybean oil meal chick rations. Poultry Sci. 27: 263-269. Moyer, A. W., and V. du Vigneaud, 1942. The structural specificity of choline and betaine in transmethylation. J. Biol. Chem. 143: 373.
Book Reviews like its predecessors, should have wide 21st edition. The Morrison Publishing distribution among all who are interested Company, Ithaca, N. Y., 1948. 1207 in livestock feeding. pages. Price $7.00.
MORRISON, FRANK B. Feeds and Feeding.
Feeds and Feeding in this 21st edition finally includes poultry. The values of different feeding stuffs for poultry are included in Parts I and II. In Part I I I one chapter is devoted to "General Problems in Poultry Production" and another to "Feeding and Caring for Poultry." These two chapters occupy pages 1028 to 1082 inclusive. The text material for this new edition has been entirely rewritten to include the most recent information concerning livestock feeding and nutrition. New compilations have been made of feed analyses for the appendix tables. In spite of the large number of pages, the book appears to be excellently bound which gives it the appearance of one which should stand considerable wear and use. This edition,
LEONARD. Modern Poultry Husbandry. Crosby Lockwood & Son, Ltd., London. 1948. 528 pa,ges. Price 21 Shillings.
ROBINSON,
This book is written in a clear and readable style. Its purpose is to give a composite picture of the British poultry industry to the poultry farmer or those who contemplate raising poultry. Modern Poultry Keeping is distinctly a British book giving very good descriptions of their poultry industry. Its subject matter content is limited entirely to chickens. A few selected references for each of the chapters are given at the back of the book, as well as a very limited glossary of technical terms and a fairly good index of subject matter.
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Association of American Feed Control Officials, 1948. Berry, E. P., C. W. Carrick, Roy E. Roberts, and S. M. Hauge, 1945. Condensed fish press water and fish liver meal in chick rations. Poultry Sci. 24: 270-276. Clandinin, D. R., W. W. Cravens, J. G. Halpin, and E. B. Hart, 1946. Supplementary value of methionine, cystine and choline in a practical soybean oil meal starter ration. Poultry Sci. 25: 509-516. Gerry, R. W., C. W. Carrick, and S. M. Hauge, 1948. Methionine and choline in a simplified chick ration. Poultry Sci. 27: 161-168. Jukes, T. H., and A. D. Welch, 1942. The effect of certain analogues of choline on perosis. J. Biol. Chem. 146: 19. Lassen, Sven, and E. Kyle Bacon, 1946. The use of condensed fish solubles in poultry nutrition.
moting properties in a purified diet and is not effective for perosis prevention, whereas choline supplementation resulted in excellent growth and complete perosis prevention. McKittrick (1947) has divided the choline and methionine requirements for the stimulation of growth into two parts, namely, the essential and the replaceable parts—the essential function of one of these supplements will not be replaced by the other. Glycine betaine would not substitute for choline or methionine in the essential part of the requirements but would substitute for either one in the replaceable increment. McGinnis, Norris and Heuser (1942) found that the occurrence of perosis in chicks was greatly reduced by the supplementation of their basal diet, which was not a purified diet, with betaine-hydrochloride. In fact, the incidence and severity of perosis, amount of gain, and mortality were approximately the same with either choline or betaine supplementation. McGinnis (1946) working with turkeys found that additions of gelatin increased the incidence of perosis, depressed growth and reduced the perosis-preventing function of betaine. It is the purpose of this report to present evidence concerning the growth promoting effects of fish products, betaine, choline, and methionine when used as
REVIOUS investigations at this Station have indicated a supplementary value for choline and methionine in a simplified corn and soybean oil meal ration. Gerry et al.- (1948) found these factors to be interchangeable to a certain extent, but that methionine was of somewhat more value than choline. However, Clandinin et al. (1946) using a somewhat more complex ration concluded that choline and methionine cannot be considered interchangeable. Using four different soybean oil meals they found that only one was improved by choline, whereas all four were improved by methionine additions. However, their experiment was terminated when the chicks were four weeks of age. Gerry et al. found that 4 weeks was not sufficient time for bringing out these relative values. Almquist and Grau (1945) state that even if high levels of choline are employed the addition of cystine cannot compensate for a methionine deficiency. These authors have indicated (1944) that cystine may be formed in the chick from choline or methionine. Moyer and du Vigneaud (1942) and Jukes and Welch (1942) have reported that betaine has only alight growth pro1 Journal Paper No. 359 of the Purdue University Agricultural Experiment Station. 2 This investigation was supported in part by a grant from The Borden Company.
24
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P
METHIONINE, CHOLINE, BETAINE AND F I S H PRODUCTS
supplements in a simplified ration, consisting largely of corn and soybean oil meal. EXPERIMENTAL
ground yellow corn, 25 soybean oil meal, 5 dehydrated alfalfa leaf meal, 5 distillers' dried solubles, 4 liver meal, 3 dried whey, 3 meat and bone scraps, 1 ground limestone and 0.5 steamed bone meal and 1 of a salt mixture consisting of 90 percent iodized salt and 10 percent manganese sulphate. D-activated animal sterol was included to provide 1 A.O.A.C. unit per gram of ration. The percentage composition of the basal experimental rations was 35 soybean oil meal in Trail I and 38 in Trials I I and III, 1.5 each of steamed bone meal and ground limestone, 0.5 of feeding oil (3,000-A, 400-D) 0.5 of a salt mixture (90 percent iodized salt and 10 percent manganese sulphate) and ground yellow corn to bring each ration to 100. Synthetic riboflavin was included in all of the experimental rations at a level of 300 micrograms per hundred grams. Supplements of 3 milligrams of nicotinic acid and 1 milligram of calcium pantothenate per hundred grams were included in all of the experimental rations in Trials I and III. Additional ingredients are listed in the tables. Each ration contained at least 21 percent protein. RESULTS Trial I. This trial, which was conducted during July and August 1947 was a paired experiment. It was designed to measure the comparative supplementary value of condensed fish solubles and a non-protein extract from the same amount of fish solubles, as well as the supplementary value of various levels of choline chloride and betaine. The variables in the rations and the results obtained are given in Table 1. The four-to-eight and six-to-eight week gains were treated statistically by analysis of variance. Trial II. This trial was conducted during September and October 1947, to test the comparative value of semi-solid
Downloaded from http://ps.oxfordjournals.org/ by guest on May 27, 2015
Approximately 50 Barred Plymouth Rock chicks of both sexes were used in each lot within each trial. In Trial I, the chicks were fed the simplified diet without either fish or methyl group supplements for a period of four weeks, after which they were sorted into blocks on the basis of the two-to-four week gains and distributed into the experimental lots, where they were fed the experimental diets for the remaining four weeks. In Trials II and III the chicks were distributed into the various lots according to initial weight and sex. The all-mash rations and tap water were supplied ad-libitum and the chicks were reared in battery brooders with wire floors. The synthetic vitamins were dissolved in hot ethyl alcohol and water, mixed with ground yellow corn and dried. The DL-methionine was finely ground and mixed carefully with ground yellow corn. The condensed sardine fish solubles analyzed 33.3 percent protein and approximately fifty percent solids. Although some loss of moisture occurred in the stored samples, the additions of this product to the rations were adjusted to equivalent amounts on a dry matter basis. The semisolid fish product is a product resulting from the hydrolysis of undecomposed fish and/or fish trimmings from which oil may or may not have been removed. The sample used in Trial I I analyzed 34.2 percent protein and 50.92 percent solids. The amounts of fish solubles and semi-solid fish product will be expressed on the dry matter basis in both tables and discussion. The percentage composition of the positive control ration in each trial was 53
25
26
D. H. MISHLEE, C. W. CAERICK AND S. M. HAUGE
fish product and condensed fish solubles in the simplified corn and soybean oil meal ration containing supplements of riboflavin, nicotinic acid, calcium pantothenate, choline chloride and DL-methionine. Since it is possible that some deterioration might occur in fish solubles during long periods of storage, three samples of
and was a further study of the supplementary inter-relationship of choline, betaine and methionine in a corn-soybean oil meal ration containing synthetic riboflavin, nicotinic acid and calcium pantothenate. Previous results in this laboratory had shown that choline and methionine exerted some supplementary interaction, although methionine was of more
TABLE 1.—A comparison of choline, betaine and fish solubles (both sexes) Variable Ingredients (percent)
Lot 1
Av. gain Males 4-8 wks. (gms.)2 Females 4-8 wks. (gms.)2 Both Sexes 4-8 wks. (gms.)2 Males 6-8 wks. (gms.)2 Females 6-8 wks. (gms.)2 Both Sexes 6-8 wks. (gms.)2 Number of chicks at 8 weeks 1 2
Lot 3
Lot 4
Lot 5
Lot 6
0.1
0.2
Pos. Control
1.0
Lot 7
Lot 8
Lot 9
0.1
0.2
0.3
291 280 286 142 145 144 47
346 307 327 171 155 163 47
382 365 373 172 172 172 47
.07 288 238 263 147 112 129 47
379 340 360 210 179 194 47
321 284 302 177 148 162 47
466 355 411 230 162 196 47
Non protein extract from 1.0 percent fish solubles Approximate least significant differences: 5% level 4-8 wk. Males Females Both Sexes 6-8 wk. Males Females Both Sexes
fish solubles were compared. One sample had been stored for a year in a refrigerator at 36°F. (Lot 13); a second sample from the same barrel was stored for a year in a garage (Lot 14); and a third sample was from a new supply (Lot 15), which was the same as that used in other lots in this trial. Since the work of Mishler et al. (1948) had indicated that male and female chickens responded differently to additions of fish solubles, both sexes of chickens were used. The variables in the rations and the results are given in Table 2. Trial III. This trial was conducted from December 1947 to February 1948,
47 31 26 34 23 18
grams grams grams grams grams grams
346 282 314 167 130 149 47
309 283 296 144 144 144 47
1% level 62 41 34 44 31 24
grams grams grams grams grams grams
value than choline as a supplement to a corn-soybean oil meal ration with fish solubles. Since the results of Trial I indicated the effectiveness of higher levels of betaine a further test of this product in the simplified ration was undertaken. The variables in the rations and the results are given in Table 3. DISCUSSION
The corn and soybean oil meal basal ration supplemented with synthetic riboflavin, nicotinic acid, and calcium pantothenate, and used as a depletion ration for all chicks the first four weeks, resulted in
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Choline chloride Betaine Fish solubles Fish solubles extract1
Lot 2
27
METHIONINE, CHOLINE, BETAINE AND FISH PRODUCTS TABLE 2.-—A comparison of various levels offish products (both sexes) Variable Ingredients (percent) Fish solubles1 Semi-solid fish pdt. Nicotinic acid Ca pantothenate Choline chloride DL-methionine
Lot 11
3.0
3.0
+ +
Lot 12
Lot 14
Lot 15
Lot 16
Lot 17
Lot 18
Lot 19
0.5
0.5
0.5
0.5
1.0
1.5
4.0
+ + + + + + + + 0.15 0.15 0.15 0.15
+ +
0.15 0.15
0.15 0.15
569 465 517 869 679 774 49
574 492 533 851 740 796 49
417 418 417 646 559 602 49
Lot 13
525 495 510 824 747 786 49
561 479 520 880 737 809 49
0.15
0.15
0.15
0.15
554 544 549 880 838 859 52
605 551 578 927 811 869 50
644 536 590 985 801 893 51
633 542 588 977 799 888 53
589 490 540 913 747 830 52
Lot 20 Pos. Control
620 514 567 949 774 862 51
1 Samples used in Lots 13 and 14 were stored one year in a refrigerator and garage respectively. All other samples were from a new supply. 2 Approximate least significant differences:
5 % level 58 grams 49 grams 38 grams 86 grams 70 grams 55 grams
6 weeks Males Females Both sexes 8 weeks Males Females Both sexes
1% level 77 grams 64 grams 50 grams 114 grams 92 grams 73 grams
TABLE 3.—A comparison of various levels of choline, betaine and methionine (both sexes) Variable Ingredients (percent)
Lot 21
Choline chloride Betaine DL-methionine Average weight Males 6 wks. (gms.)1 Females 6 wks. (gms.)1 Both Sexes 6 wks. (gms.) 1 Males 8 wks. (gms.)1 Females 8 wks. (gms.)1 Both Sexes 8 wks. (gms.)1 No. of chicks at 8 weeks 1
Lot Lot 22 23
Lot Lot 24 25
0.1
0.3
0.2
0.2
357 310 334 533 447 490 48
442 358 400 649 527 588 46
437 393 415 671 582 627 47
427 392 410 680 581 631 50
430 431 431 658 640 649 49
Lot 26
Lot 27
Lot 28
0.3
0.4
0.3 0.15
43.9 443 441 647 655 651 50
485 443 464 738 656 697 50
443 494 469 696 734 715 50
Lot 32
Lot Lot 29 30
Lot 31
0.1 0.2
0.15 Pos. 0.15 Con0.15 trol
0.3 457 430 444 687 633 660 48
395 385 390 626 584 605 48
472 443 458 744 673 709 49
536 522 538 873 752 813 46
Approximate least significant differences: 6 weeks Males Females Both sexes 8 weeks Males Females Both sexes
significantly retarded growth of the chicks (Lot 1) throughout the eight-week period. A supplement of one percent condensed fish solubles (Lot 2) produced gains which
5% level 60 grams 43 grams 36 grams 82 grams 60 grams 50 grams
1% level 79 grams 57 grams 48 grams 108 grams 79 grams 66 grams
were highly significant over those of the negative control (Lot 1). A. nonprotein extract of fish solubles (Lot 3) also resulted in highly significant gains over the
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Average weight Males 6 wks. (gms.)2 Females 6 wks. (gms.)2 Both Sexes 6 wks. (gms.)2 Male 8 wks. (gms.)2 Females 8 wks. (gms.)2 Both Sexes 8 wks. (gms.)2 No. of chicks at 8 weeks
Lot 10
28
D. H. MISHLER, C. W. CARRICK AND S. M. HAUGE
and Bacon (1946) showed that levels of fish solubles as high as 12 percent of the ration resulted in proportional increases in growth. Berry et al. (1945) reported no better growth with 4 percent condensed fish solubles than with 2 percent, but the rate of growth was less than ours. When the four synthetic vitamins and DLmethionine in our simplified ration were supplemented with various levels of fish solubles no better growth resulted with a 4.0 percent level (Lot 19) than with a 1.5 percent level (Lot 18) in the case of male chickens. However, with the female chicken optimum growth was reached at the 0.5 per cent level. These results are in agreement with previous observations by Mishler, Carrick and Hauge (1948), which indicated that males responded more favorably than females to increased levels of fish solubles. A taste test was conducted to determine whether a fishy flavor had been imparted to the flesh of chickens fed 4.0 percent of condensed fish solubles (Lot 19) or 3.0 percent of semi-solid fish product (Lot 11). Thirty chickens (ten from each of these lots and ten from the control lot (Lot 20) which received no fish product) were killed, dressed, halved and distributed to fifteen persons. No consumer had any previous knowledge of the rations fed to the chickens which he consumed. Only four persons reported fishy flavors and these were not confirmed by persons consuming the other half of the same chickens. Moreover, two of the reports involved chickens which had been fed the positive control ration containing no fish products. Therefore it is unlikely that any fishy flavor is transmitted from the ration to the meat. These results are in agreement with those of Berry et al. (1945). The soybean oil meal used in Trial III was apparently low in available choline as evidenced by the suboptimal growth in Lot 21. An addition of 0.1 percent choline
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negative control. The significantly slower gains which were made with the extract as compared with the fish solubles may have been due to some of an unknown factor having been removed with the protein. It is clear that the value of fish solubles in this type of ration is not from protein, since the ration is already adequately supplied with all of the essential amino acids, except methionine. A low level of fish solubles would supply very little methionine and the extract practically none. In our laboratory expeller soybean oil meals have varied somewhat in the growth response obtained from additions of choline and methionine. A level of 0.1 percent choline chloride resulted in increased gains which were highly significant over those of the negative control thus indicating the basal ration to be deficient in available choline. A supplementary level of 0.2 percent betaine (Lot 8) proved as effective a supplement as either 0.1 or 0.2 percent of choline, but at the 0.3 percent level (Lot 9) betaine resulted in gains which were highly significant over those produced by lower levels of either choline or betaine. A level of 3.0 percent of semi-solid fish product did not have any supplementary value in a simplified diet containing nicotinic acid, calcium pantothenate, choline chloride and DL-methionine(Lots 10-12). When a low level of 0.5 percent fish solubles (Lot 16) replaced the semi-solid fish products (Lot 11) growth at eight weeks was improved with high significance. When samples of condensed fish solubles were stored for a period of one year either in a refrigerator (Lot 13) or in an unheated garage (Lot 14), the tests indicated that little deterioration had occurred. If any difference was manifested as a result of storage it was shown in the slower growth of the male chickens fed the refrigerated product. Using a more complex ration, Lassen
METHIONINE, CHOLINE, BETAINE AND FISH PRODUCTS
SUMMARY
1. A study was made of the supplementary value of methionine, choline, betaine and fish products in a simplified corn and soybean oil meal ration supplemented only with riboflavin, calcium pantothenate, and nicotinic acid in addition to a simple mineral mixture and vita-
min A and D feeding oil. 2. Choline, betaine and methionine were all effective in supplementing the simplified ration. Betaine at 0.3 and 0.4 percent levels appeared to have higher supplementary value than either choline or methionine. 3. A three percent level of semi-solid fish product plus riboflavin in a corn and soybean oil meal ration was improved with high significance when calcium pantothenate, nicotinic acid, choline and methionine were added, indicating the product to be deficient in one or more of these factors. A three percent level of semi-solid fish product did not improve the simplified ration containing the four synthetic B-complex vitamins and methionine. 4. A level of 4 percent condensed fish solubles (dry basis) resulted in no better growth than 1.5 percent with male chickens. The maximum growth of the female chickens was at the 0.5 percent level (dry basis). None of the fish products imparted a fishy flavor to the flesh of the cooked chickens. ACKNOWLEDGMENT
We are indebted to Mr. R. H. King of the Department of Agricultural Chemistry for the preparation of the fish solubles extract, the Feedingstuff Laboratories of the State Chemist for the protein analysis of the condensed fish solubles and semisolid fish product, to The Borden Company for the betaine hydrochloride and to U. S. Industrial chemicals for the synthetic DL-methionine used in these experiments. REFERENCES
Almquist, H. J., and C. R. Grau, 1944. Interrelation of methionine, choline, betaine and arsenocholine in the chick. J. Nutrition 27: 263-269. Almquist, H. J., and C. R. Grau, 1945. Further studies on cystine, methionine and choline in chick diets. J. Nutrition 29: 219-222.
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chloride resulted in an improved growth response, which was highly significant. A level of 0.3 percent of choline chloride resulted in still greater growth. At similar levels betaine resulted in somewhat better growth than choline, although the differences were not significant. A supplementary level of 0.4 percent betaine gave a significant difference in growth over a 0.3 percent level in the case of the male chickens. A combination of choline and betaine was improved by the addition of a low level of methionine. However, when 0.1 percent choline and 0.2 percent betaine were added (Lot 29), growth was only slightly better than 0.2 percent betaine alone (Lot 25). When all three products were included at low levels (Lot 31) there was no better growth than when 0.3 per cent betaine and 0.15 percent methionine were included (Lot 28). These results have indicated that betaine is as satisfactory a supplement as choline for rapid growth, and that a combination of betaine and methionine may assume the growth functions of choline in the simplified ration used. The poor results shown with a level of 0.3 percent methionine (Lot 30) may indicate a deficiency of non-replacable choline. The control ration (Lot 32) resulted in a growth response which was highly significant over that produced by any of the experimental lots in Trial III. It is apparent that all of the experimental rations in this trial were deficient in one or more unidentified factors supplied by fish solubles.
29
30
BOOK REVIEWS Poultry Sci. 25:486-491. McGinnis, J., 1946. The influence of gelatin on prevention of perosis in turkeys fed choline and betaine. Poultry Sci. 25: 91-92. McGinnis, J., L. C. Norris, and G. F. Heuser, 1942. Effect of ethanolamine and betaine on perosis in chicks. Proc. Soc. Exp. Biol. Med. 51: 293. McKittrick, D. S., 1947. The interrelations of choline and methionine in growth and the action of betaine in replacing them. Arch, of Biochem. 15: 133-156. Mishler, D. H., C. W. Carrick, and S. M. Hauge, 1948. Condensed fish solubles as a supplement for corn and soybean oil meal chick rations. Poultry Sci. 27: 263-269. Moyer, A. W., and V. du Vigneaud, 1942. The structural specificity of choline and betaine in transmethylation. J. Biol. Chem. 143: 373.
Book Reviews like its predecessors, should have wide 21st edition. The Morrison Publishing distribution among all who are interested Company, Ithaca, N. Y., 1948. 1207 in livestock feeding. pages. Price $7.00.
MORRISON, FRANK B. Feeds and Feeding.
Feeds and Feeding in this 21st edition finally includes poultry. The values of different feeding stuffs for poultry are included in Parts I and II. In Part I I I one chapter is devoted to "General Problems in Poultry Production" and another to "Feeding and Caring for Poultry." These two chapters occupy pages 1028 to 1082 inclusive. The text material for this new edition has been entirely rewritten to include the most recent information concerning livestock feeding and nutrition. New compilations have been made of feed analyses for the appendix tables. In spite of the large number of pages, the book appears to be excellently bound which gives it the appearance of one which should stand considerable wear and use. This edition,
LEONARD. Modern Poultry Husbandry. Crosby Lockwood & Son, Ltd., London. 1948. 528 pa,ges. Price 21 Shillings.
ROBINSON,
This book is written in a clear and readable style. Its purpose is to give a composite picture of the British poultry industry to the poultry farmer or those who contemplate raising poultry. Modern Poultry Keeping is distinctly a British book giving very good descriptions of their poultry industry. Its subject matter content is limited entirely to chickens. A few selected references for each of the chapters are given at the back of the book, as well as a very limited glossary of technical terms and a fairly good index of subject matter.
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Association of American Feed Control Officials, 1948. Berry, E. P., C. W. Carrick, Roy E. Roberts, and S. M. Hauge, 1945. Condensed fish press water and fish liver meal in chick rations. Poultry Sci. 24: 270-276. Clandinin, D. R., W. W. Cravens, J. G. Halpin, and E. B. Hart, 1946. Supplementary value of methionine, cystine and choline in a practical soybean oil meal starter ration. Poultry Sci. 25: 509-516. Gerry, R. W., C. W. Carrick, and S. M. Hauge, 1948. Methionine and choline in a simplified chick ration. Poultry Sci. 27: 161-168. Jukes, T. H., and A. D. Welch, 1942. The effect of certain analogues of choline on perosis. J. Biol. Chem. 146: 19. Lassen, Sven, and E. Kyle Bacon, 1946. The use of condensed fish solubles in poultry nutrition.