Factors Influencing the Response of Methionine, Choline and Inorganic Sulfate Supplementation to Practical Poultry Diets1, 2

Factors Influencing the Response of Methionine, Choline and Inorganic Sulfate Supplementation to Practical Poultry Diets 1

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sulfate resulted in a 66.1 % growth response over the basal diet (371.5 g vs. 617 g).These results indicated that the bird could satisfy part of its total sulfur requirement with inorganic sulfate. Ross and Harms (30) were able to show a response to inorganic sulfate in a cornsoybean meal based diet. Working with purified diets other investigators have also shown a similar response to inorganic sulfate (1,16). A very important point which was brought out in this research was that as the methionine level in the diet increased the response to inorganic sulfate decreased. Further work by other investigators (3, 32, 33, 35, 36) showed that inorganic sulfate spares about 15% of the physiological need for cystine rather than methionine when added to a cystine deficient diet containing less than 200 ppm sulfate. Therefore, from 7 to 9% of the sulfur amino acid requirement can be met as inorganic sulfate when cystine is deficient in the diet. Other factors can be shown to influence the results in studies involving methionine and inorganic sulfate. Plavnik and Bornstein (26) reported that age of the bird influenced the magnitude of the response of chicks to inorganic sulfate. Using a soybean meal -grain sorghum based diet in three experiments they found that between the third and the fifth week, the growth response to sulfate was higher than between the first and fourth week. Growth responses as great as 10% were obtained when 0.12% inorganic sulfate was supplemented to the basal diet. Of course, in practical diets, the growth response to supplemental inorganiC sulfate would not be expected to be as great as when purified diets are used because these diets usually contain some sulfate . It must also be realized that the bioavailability of the inorganic source of sulfate has to be considered. Tillman and Pesti (37) stated that the response to supplemental choline was dependent on the dietary sulfate content of the diet. These authors reported that the greatest response to inorganic sulfate by broiler chicks occurred when the basal diet contained high levels of choline. These results somewhat supported the hypothesis of Miles et a/ (18, 19) that the responses to sulfate and choline were interdependent. Tillman and Pesti (37) also pointed out that a lot of the variability in experimental results in studies involving methionine, choline and inorganiC sulfate may be caused by the composition of the basal diet.

by R. D. Miles, N. Ruiz and R. H. Harms Poultry Science Department Institute of Food and Agricultural Sciences University of Florida Gainesville, Florida 32611 Introduction If nutrient interrelationships did not exist the nutritionist's job would be much easier. However, interrelationships do exist among the dietary nutrients and most practical nutritionists are aware of the majority of them. There exists in animal nutrition an optimal level for each nutrient in relation to the levels of some others in order to obtain the most efficient desired animal response. When two or more nutrients are interrelated it becomes critical for nutritionists to consider the practical implications of these interrelationships. One three-way interrelationship which has been recently defined is that among methionine, choline and inorganic sulfate. The objective of this manuscript is to review and summarize the relationships of these three nutrients. Methionine and Inorganic Sulfate Interrelationship Gordon and Sizer (8) were the first investigators to report a growth response from the addition of sodium sulfate to a diet deficient in sulfur amino acids. These authors published the results of an experiment in which a casein-gelatin based purified diet, deficient in cystine)0.08% cystine; 0.51 % methionine) and sulfatefree, was fed to growing chicks. The duration of the study was five weeks. The addition of 0.50% sodium sulfate to the basal diet resulted in a 31.4 % growth response over the basal diet (371.5 g gain vs. 488.1 g) at the end of the fifth week. Nevertheless, inorganic sulfate could not replace dietary cystine or methionine for protein synthesis. Thus, the addition of 0.22% methionine gave a 39.1 % growth response over the basal diet (371.5 g vs. 521 g). The simultaneous supplementation with 0.22% methionine and 0.50% sodium , Florida Agricultural Experiment Stations Journal Series No. 7552 2Reviewed by: Robert L. Harrold and William A. Dudley Cash

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34 Methionine and Choline Interrelationship

MILES, RUIZ, AND HARMS sulfate occurred in a practical corn-soybean meal diet formulated to be low in sulfate. However, in practical situations the sulfate form of trace minerals is usually used. This would mean that the sulfate content of the diet is

Klose and Almquist (13) reported that methionine was essential for chick growth and that homocysteine can replace methionine in the diet when adequate choline is present. However, this important finding by Klose and sulfate would probably not result in a significant growth Almiquist should not be misunderstood as saying that response . From the above referenced four experiments choline can substitute for (replace) methionine in the diet. it was concluded that inorganic sulfate and choline would A net synthesis of methionine cannot occur in the animal furnish a margin of safety in the diet when the sulfur without an exogenous source of homocysteine. In pracamino acid content of the diet was slightly below the tical nutrition an exogenous homocysteine source usually animal's requirement. When the sulfur amino acid level in does not exist because practical diets contain very little, the diet was in excess of the animal's requirement the if any, homocysteine. In fact, the majority of the choline and sulfate response would not be as great. homocysteine in animal cells arises from the metabolism The influence of inorganic sulfate on the interrelationof methionine present in the diet. ship between methionine and choline has been reported An important point must be understood regarding in rats (6) and humans (40). Recently, Lovett et al. (14) choline and methionine supplementation to the diet. reported the influence of sulfate and choline in weanling Numerous articles have been published that imply that swine. Vemury et al. (40) reported that in adult humans choline can "replace" methionine in the diet. Such a the combinations of choline plus methionine or combinastatement is very misleading and can result in costly contions of methionine , choline and inorganic sulfate showed sequences in the field. that choline tended to be retained by the body and thus No compound can repiace the intact moiecuie of perhaps spared methionine as indicated by the positive methionine with regards to its specific amino acid funceffect on nitrogen balance compared to methionine suption during protein synthesis. However, a certain amount plementation alone. They concluded that although addiof choline can "spare" the need for supplemental tions of choline and inorganic sulfate do not have the methionine to the extent that its labile methyl group is beability to replace the methionine need in the body, these ing used since the two principal methyl donors in the supplements exerted a sparing action on methionine animal are choline and methionine. This is to say that when methionine was being used for purposes other than once the requirements for methionine and choline, as inprotein synthesis. tact molecules, are met the metabolic requirement for methyl groups can be met by either methionine or The Choline Requirement of Laying Hens choline. The term "replace" should be used only when referrThe response of laying hens to supplemental choline in ing to the intact molecule, and "spare" should be used the diet has been a controversial issue for decades. when referring to a molecule's functional groups. When Numerous reports are available showing that laying hens reading publications in which the term "replace" is used respond to choline supplementation . Evidence can also indiscriminately, careful consideration should be given to be found showing no response in layers consuming diets - - -""::t:':" he = r'::'e:"'s:":' ::u:':lti':': :':' n:':' g'=':c"'--o'-n-='s=e:"":q:':'u'='e'-n::":c::": e':': s:':'o='f::":a..=P:.::.:P-=.ly'-':i-= n-" g-=-t-=h.:.: e'-r=-e=-c""'o"' m:....:m "-e ' -n :.::-- - -supp1ementmtwith- choline . It becomes- appar ell t fr orn dations to practical situations. reading the literature that certain factors must be considered if a response is expected from supplementing choline to laying hen diets. First, a response will usually Methionine, Choline and Sulfate Interrelationship occur if the diet is deficient in total sulfur amino acids or if Miles et al. (18, 19) were able to show that in poultry a diet marginal in protein is fed. Second, the total feed ina three-way interrelationship existed among these take of hens has to be considered because hens need a nutrients. These authors concluded that the level of certain intake of sulfur amino acids daily. dietary choline should be considered in studies concernThe laying hen is capable of synthesizing a coned with investigating the relationship between methionine siderable amount of choline (7, 15, 24, 29). The NRC and sulfate. Likewise, inorganic sulfate should be con(22) questions whether a dietary choline requirement exsidered in studies involving the sulfur amino acids and ists for the laying hen. The dietary requirement for choline. choline in the laying hen has been studied mainly in relaTwo experiments were conducted with broiler-type tion to its role in egg production and the prevention of fatchicks (18) and two experiments were conducted with ty livers. turkey poults to study the three-way interrelationship Althou gh Nesheim et al. (24) found it difficult to produce (19). The results from these studies by Miles et al. (18, a deficiency of choline in layings hens, there is general agreement among the different reports in the literature 19) indicated that a greater growth response to choline that supplementary choline decreases the liver fat conand sulfate occurred in poults when compared with the tent in laying hens. However, the lipid content of the liver broiler response. This was true when the diet was defiis variable, and high levels of liver lipid are not necessarily cient in methionine . It must be emphasized that the detrimental to laying performance (11, 17). Nesheim et growth response obtained from the supplementation of

METHIONINE, CHOLINE AND INORGANIC SULFATE SUPPLEMENTATION

at. (23) reported that hens fed choline-deficient diets had liver fat contents lower than values determined in hens dying from fatty liver syndrome. No correlation was found between rate of egg production and level of liver lipid. In other words, it is difficult to implicate choline in fatty liver syndrome. Ruiz et at. (31) conducted studies to investigate the choline requirement of commercial laying hens grouped according to body weight and fed according to feed intake . Hen performance (egg production, feed efficiency, egg specific gravity and egg weight) was not significantly different at any level of choline supplementation. Liver lipid content was significantly higher only in extra-heavy birds fed the basal diet containing no supplemental choline. Results of this experiment support the conclusion that aged layers, regardless of thier body weight, require very little, if any supplemental choline when they are consuming sufficient levels of methionine daily. These findings are in agreement with those of March (17) who reported that a diet containing adequate methionine did not require supplementation with choline and asked what concentration of fat may be considered to be excessive before a liver can be termed "fatty" and the "fattiness" considered to be a pathological condition Nesheim et at. (23) found in several experiments that hens with very high levels of liver lipid were excellent egg producers. Parsons and Leeper (25) also reported that a choline response would occur if a diet contained an inadequate crude protein level. Keshavarz and Austic (12) reported that choline supplementation improved egg production when practical layer diets deficient in methionine were fed to hens. Choline was not as effective in maintaining egg size as methionine. These results agree with those of Tsiagbe et at. (38) and Brooks and Creger (5). Miles et at. (20) suggested the minimum daily choline intake of layers, in the absence of methionine supplementation to the diet, was 11 8 mg per bird for maximum egg production. However, maximum egg size was obtained only when hens consumed diets containing adequate methionine. Practical Considerations

It has already been emphasized in previous paragraphs that the influence of dietary inorganic sulfate was determined in a purified diet deficient in cystine or in a practical diet without sulfate supplementation. These conditions were deliberately established so as to obtain a sulfate response. However, in practical diets, if inorganic sulfate is deficient it will be supplied by oxidation of sulfLlr amino acids within the animal. This would therefore limit the amount of sulfur amino acids available for other body functions. In practical situations it is not likely that inorganic sulfate will be in very short supply. The reason for this is because the water supply may have sulfate present and ingredients such as fish meal, meat and bone meal and whey, which are high in non-amino acid sulfur (2), may be

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present in the diet. In practical diet formulation the important point to consider is to have a sulfate source in the diet. Usually, this can easily be accomplished by using the sulfate form of trace minerals. Several investigators (21, 36) have reported a sulfate response in a corn-soybean meal basal diet containing other natural ingredients which have contributed to the basal sulfate level. Therefore, the maximal response to supplemental inorganic sulfate in these studies could not be assessed. Potter et at (27) reported that supplemental potassium or sodium sulfate failed to influence body weights of young turkeys. These results were contrary to those of several previous studies with turkeys and chickens (9, 34, 36, 39). The basal diet used by Potter et at. (27) contained 12% meat and bone meal and this could explain why no response to inorganic sulfate resulted. Potter and Shelton (28) reported that supplemental sodium sulfate failed to increase final body weights of young turkeys at 7 and 8 weeks of age in two experiments, respectively. The basal diet in this study contained 5% menhaden fish meal. However, in a later study Blair et at (4) reported a significant growth response to 0.1 % supplemental potassium sulfate in methionine deficient diets. The basal diet in this study contained only corn and soybean meal as the natural protein and sulfate source. The trace mineral mix used by Blair et at (4) was also sulfate free. The supplemental sulfate did not result in a growth response when adequate methionine was present in the diet. The importance of conSidering feed intake, or, more importantly, total sulfur amino acid (TSAA) intake in choline studies, is emphasized by the recent publication of Hennig et at. (10). These researchers concluded from several studies with mature laying hens that layers do not respond to, or have a requirement for, dietary choline. However, the only reference to feed intake in this article was in a study involving a casein-gelatin-corn starch basal diet. It is plausible that the high feed intake (102-120 grams/hen/day) in this study coupled with the dietary TSAA level of .55% can explain why no choline response occured. Considering the feed intake and TSAA level of the diet, the TSAA intake would exceed 600 mg/bird/day. This level of sulfur amino acid intake would be more than enough to meet or exceed the hen's requirement and provide for any additional need for methyl groups. Therefore, a choline response would not be expected in this case. These researchers indicated that the choline preparation used was bound to shredded, dried sugar beet slices. Sugar beets are a very rich source of betaine and betaine is the actual methyl donor during ct"l line metabolism. Therefore, the carrier itself may have been furnishing methyl groups to the hens. If this was the case, the response to dietary choline supplementation would be expected to be reduced. The above discussion regarding a choline response in mature laying hens has emphasized the importance in practical situations to consider the daily intake of total feed, protein and sulfur amino acids in laying hens.

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MILES, RUIZ, AND HARMS

16 . Machlin, L. J . and P. B. Pearson . 1956. Proc. Soc. Exp. BioI. Med. 93:204 . If choline or inorganic sulfate are not adequate in the 17 . March, B. E. 1981. Poultry Sci. 60 :818 . diet methionine can furnish the methyl groups or sulfur to 18. Miles, R. D., N. Ruiz and R. H. Harms . 1983a. the metabolic pool. On the other hand, neither choline Poultry Sci. 62 :495 . nor inorganic sulfate can replace the methionine needed 19. Miles, R. D., N. Ruiz and R. H. Harms. 1983b. for protein synthesis. In practical conditions it is likely that Proc. Soc. Exp. BioI. Med. 173:32 . sulfate will be adequate in the diet due to the sulfate 20. ~Y1iles, R. D., ('.~ . Ruiz and R. H. Harms. 1986 . already present in various feedstuffs , water and the Poultry Sci. 65:1760. mineral premix. Under these cond itions a supplemental 21 . Miller, D. M ., G. N. Biddle, P. E. Bauersfeld, Jr. choline or methionine response would be dependent on and S. L. Cuppett. 1974. Poultry Sci. 53 :226. the methyl group and sulfur amino acid status of the diet. 22 . National Research Council. 1984. Nutrient ReWith layers, a choline response would be expected if the quirements of Poultry. 8th ed. National Academy hens were consuming less than adequate total protein or Press. 2101 Constitution Av . Washington, D. C. sulfur amino acids daily . !n the absence of adequate 20418. sulfur amino acid intake it has been suggested that the 23 . Nesheim, M. C., C. A. Ivy and M . J. Norvel. laying hen requires 118 mg choline intake per day. 1969 . pp.36-41 in Proc. Cornell Nutr. Conf. However, choline alone will not maximize egg size wh ich 24 . Nesheim, M . C., M. J . Norvell, E. Ceballos and R. is achieved when the daily sulfur amino acid needs of the M . Leach, Jr. 1971 . Poultry Sci. 50:820. hen are met. Finally, from an economic point of view it is 25 . Parsons, C . M . and R. W. Lepper. 1984. Poultry important that sufficient sulfate and methyl groups be Sci. 63 : 1604. provided so that the sulfur bearing amino acids be used 26 . Plavnik, Y. and S. Bornstein. 197 7. Br. Poultry primarily for protein synthesis. Sci. 18:33 . Literature Cited 27 . Potter, L. M., J. R. Shelton and D. J. Castaldo. 1983. Poultry Sci. 62:2398. 1. Almquist, H. J . 1964. Feedstuffs 36 (24):60. 28. Potter, L. M . and J. R. Shelton. 1984. Poultry 2 . Almqu ist, H. J . 1974. Feedstuffs 46 (1 1) :22 . Sci. 63:98 7 . 3 . Anderson, J. 0., R. E. Warnick and R. K. Dalai . 29. Ringrose, R. C . and H. A. Davis. 1946 . Poultry 1975. Poultry Sci. 54:1122 . Sci. 25 :646 . 4. Blair, M . E., L. M. Potter, B. A. Bliss and 30 . Ross, E. and R. H. Harms. 1970. Poultry Sci. J . R. Shelton. 1986. Poultry Sci. 65:130 49:1605. 5 . Brooks, L. G. and C. R. Creger. 1983. Poultry 31. Ruiz, N., R. D. Miles, H. R. Wilson and Sci. 62: 1391. R. H. Harms. 1983. Poultry Sci. 62: 1028. 6. Byington, M. H., J. M . Howe, and H. E. Clark. 32. Sasse, C . E. and D. H. Baker. 1974a. Poultry 1972. J. Nutr. 102:219. Sci. 53:652. 7. Crawford, J. S., M . Griffith , R. A. Teekell and 33 . Sasse, C . E. and D. H. Baker. 1974b. J Nutr. -----A~afts:_19-6-9~ oultryScr-4--a:6-Z0r. - - - - - - --1 1 e>
SUMMARY