TOTAL SULFUR-CONTAINING AMINO ACIDS ISOLEUCINE AND TRYPTOPHAN REQUIREMENTS OF THE FINISHING PIG FOR MAXIMUM NITROGEN RETENTION 1 H. W. Brown 2, B. G. Harmon and A. H. Jensen Illinois Agricultural Experiment Station, Urbana 61801 Summary Four finishing pigs were employed in each of three 4 x 4 latin square designs to study the total sulfur-bearing amino acids, isoleucine and tryptophan requirements. In the total sulfurbearing amino acids and isoleucine studies graded levels of the test amino acid were added to a 14.1% protein basal diet (protein from corn and gelatin) containing 3,700 kcal metabilizable energy per kilogram. In the tryptophan study the diet contained 13.6% protein and 3,727 kcal metabolizable energy per kilo- gram. In all studies essential amino acids, excepting the given test amino acids, were provided in concentrations of 1.2 times the published requirements. In each 4 x 4 latin square design four barrows, four dietary treatments and four I 0-day periods were employed. There were no significant differences in nitrogen retention among the four periods. Amino acid requirements were determined by the method of least squares. Addition of methionine to a basal diet containing.17% total sulfur-bearing amino acids did not significantly increase nitrogen retention. Expressed as a percent of the diet, estimated requirements for isoleucine and tryptophan were, respectively,.31 +-.01% and.059 -+.001%. Introduction Research directed toward determining the dietary requirements for finishing swine for total sulfur-bearing amino acids, tryptophan and isoleucine has been limited. The currently accepted requirements (Becker, Jensen and Harmon, 1966; N.R.C., 1968) for these amino ~This research was supported in part by grants from the Moorman Mfg. Co., Quincy, Illinois and CPC International, Inc., Argo, Illinois. 2The data are from a thesis submitted by the senior author in partial fuffillment of the requirements for the degree of Doctor of Philosophy. acids were calculated (Becker et al., 1966), rather than experimentally determined, using the lysine requirement as a reference for calculating these requirements. It was assumed that the amino acids are required in the same ratios in both growing and finishing pigs. Most of the total sulfur-bearing amino acids studies of finishing swine have been concerned with adding methionine to diets considered adequate or only slightly inadequate in these amino acids. Results reported by Bell et al. (1950), Meade (1956a, b), Welch, Cordts and Vander Noot (1966) and Oestemer et al. (1970) indicate that the methionine plus cystine requirement of growing-finishing swine is lower than the value estimated by Becker et al. (1966). In view of the limitation associated with current calculated requirements for these amino acids, the objectives of the present trials were to determine the total sulfur amino acids, isoleucine, and tryptophan requirements of the finishing pig for maximum nitrogen retention. Experimental Procedure In trial 1, two Yorkshire and two cr barrows with average initial and final weights of approximately 54 and 95 kg, respectively, were used to study the total sulfur-bearing amino acids requirement for maximum nitrogen retention. In trial 2, one Yorkshire and three crossbred barrows having average initial and final weights of approximately 60 and 96 kg, respectively, were used to study the isoleucine requirement. In trial 3, four hybrid barrows having average initial and final weights of approximately 62 and 92 kg, respectively, were used to study the tryptophan requirement. In each trial the 4 x 4 latin square design used four barrows, four levels of the amino acid and four 5-day collection periods. The barrows were randomly assigned to metabolism crates, which were similar to those described by Baker et al. (1967). Room temperature ranged be- 59 JOURNAL OF ANIMAL SCIENCE, vol. 38 no. 1, 19"/4
60 tween 2i and 27 C. Each pig was fed daffy 1.13 kg of diet at 8:00 am and 1.13 kg at 4:00 pm. Water was given before and after each feeding and at mid-day. Composition of the basal diets is shown in table 1. Supplemental DL-methionine (trial 1), L-isoleucine (trial 2) and L-tryptophan (trial 3) were substituted isonitrogenously for L- glutamic acid to provide, respectively, the levels of total sulfur-bearing amino acids, isoleucine and tryptophan shown in tables 2, 3 and 4. Difference in total weight of the diet created by substituting the test amino acid for L-glutamic acid, which differed slightly in weight per unit of nitrogen, was equalized by altering the amount of starch in the basal diet. The amino acid mix was formulated and included in the basal diet to bring the dietary levels of the essential amino acids, except the test amino acid and lysine, to 1.2 times the requirements suggested for finishing swine by Becker et al. (1966). Lysine was included in the amino acid_ mix to provide 1.2 times the requirement ot.62% of the diet as suggested by Brown, Harmon and Jensen (1973) for obtaining maximum feed efficiency in the finishing pig. The diets fed in trials 1 and 2 assayed 14.1% crude protein and 3,700 kcal/kg of metabolizable energy. Those fed in trial 3 assayed 13.6% crude protein and 3,727 kcal/kg cf metabolizable energy. Diets in trials 1 and 2 were pelleted. During the collection period the urine was filtered through glass wool into a plastic bucket containing 20 ml of concentrated HC1. Urine was collected daffy, diluted to 4,000 ml, from which a 100 ml aliquot was taken. The accumulated daffy aliquots were stored under refrigeration until analyzed. Ferric oxide at 1% of the diet was used as a feed marker. Feces were collected daffy, placed in plastic bags and frozen. At the end of the trial the feces were placed in a forced-air drying oven for one week, then removed, allowed to set for 48 hr. for equilibration with air moisture, weighed and ground in a blender. Feed, fecal and urine samples were analyzed for nitrogen by the Kjeldahl procedure as outlined by A.O.A.C. (1965). Regression analyses were used to determine the kind of response to the test amino acids. The method of least squares was used to calculate the requirements for the amino acids showing a broken line response. Results and Discussion Data showing the response to graded levels BROWN, HARMON AND JENSEN of methionine are shown in table 2. Supplementing the basal diet, which assayed.17% total sulfur-bearing amino acids, with DLmethionine did not significantly increase nitrogen retention. Therefore, the total sulfurbearing amino acids requirement estimated from this trial was.17% or less of a 14% protein diet containing 3,700 kcal/kg of metabolizable energy. Curtin et al. (1952), Backer et al. (1955) and Baker et al. (1969) reported cystine replacement values of 40%, 53% and 56%, respectively, of the methionine requirement of swine. Since the total sulfur-bearing amino acids in the basal diet of this trial were 80% methionine and 20% cystine, all of the cystine TABLE 1. COMPOSITION OF BASAL DIETS Trials Ingredient 1 2 3 Ground yellow corn 20.00 20.00 80.00 Gelatina 10.80 10.80 6.04 Cornstarch 42.61 42.64 9.30 Sucrose 10.00 10.00... Wood cellulose 3.00 3.00... Corn?il 4.00 4.00... WaterO 3.00 3.00... Dicalcium phosphate... 2.00 Trace-mineralized salt c... 0.50 Mineral mixd 4.00 4.00... Vitamin mix e _ 1.00 1.00 1.00 Antibiotic premix f 0.10 0.10 0.10 Amino acid mix ~flg 1.00 1.00... Amino acid mi~ #2h... 1.00 L-glutamic acid 1 0.27 0.27 0.06 DL-methionine... 0.19... L-isoleucine 0.22... 100.00 100.00 100.00 aucopco Gelatin, United Chemical & Organic Products, Calumet City, Illinois. bwas added to improve pelleting of diets. CContained as a % of the mix: salt, 97.0; zinc, 0,800; manganese, 0.400; iron, 0.330; copper, 0.048; iodine, 0.011 ; and cobalt, 0.022. dcontained per kilogram (in g): CaHPO4, 650.05; NaC1 (iodized), 159.96; K2CO~ 139.95; MgCO3 32.74; FeSO4"H20, 3.00; CoC12-6H~O, 1.00; CuSO4, 1.00; NaF, 0.20; ZnCOa 2.00 and KI, 0.10. econtained per kilogram: vitamin A, 500,000 IU; vitamin Ds 284,000 IU; vitamin E, 12,000 IU; vitamin K, 20 mg; riboflavin, 660 mg; d-pantothenic acid, 2.64 g; nicotinic acid, 4.4 g; vitamin Bl 2, 4.4 mg; choline chloride, 220 g; folic acid, 200 rag; biotin, 20 mg; ascorbic acid, 30 g; pyridoxine, 250 rn~; thiamin, 260 rag; and powdered cornstarch to 1 kilogram. feach kilogram contained 33 g of streptomycin.. {as streptomycin sulfate) and 11 g of Procaine penicillin. gcontained per kilogram (in g): L-histidine, 29.94; L-lysine HC1, 219.54; L-tryptophan, 69.85; L-threonine, 79.84 and cornstarch, 600.83. hcontained per kilogram (in g): DL-methionine, 49.90; L-isoleucme, 59.88; L-lysine HC1, 299.40; and starch, 590.82. idl-methionine, L-isoleucine and Irtryptophan were substituted isonitrogenously for L-glutamic acid in trials 1, 2 and 3, respectively, to form three additional diets in each trial.
AMINO ACID REQUIREMENTS IN FINISHING PIGS 61 should contribute to the total sulfur-bearing amino acids requirement. Using the method of Williams etal. (1954), and assuming a dietary lysine requirement of.62% of the diet, the total sulfur-bearing amino acids requirement calculated from the amino acid analysis and ratios of the pig carcass is.20% of the diet. This agrees closely with the requirement of.17% or less of the diet as observed in this trial. Studies from the Minnesota station (Meade, 1956a, b; Meade, Dukelow and Grant, 1966a, b) indicate that the total sulfur-bearing amino acids requirement of the finishing pig is below or equal to the requirement of.30% of the diet reported by Becker et al. (1966). Welch et al. (1966) reported that methionine supplementation of a 12% crude protein cornsoybean meal diet containing..27% methionine plus cystine and fortified with crystalline lysine and tryptophan did not increase nitrogen retention of the finishing pig. To the authors' knowledge no finishing pig studies have been conducted at low enough total sulfur- containing amino acid levels to set a minimum requirement. Data showing the response to graded levels of isoleucine are shown in table 3. The isoleucine requirement calculated from this trial was.31 +.01% of a 14% crude protein diet containing 3,700 kcal/kg of metabolizable energy. This estimated requirement does not differ greatly from the requirement of.35% reported for the finishing pig by Becker et al. (1963). Their requirement was determined with a 13.4% protein diet containing 3,330 dcal/kg of metabolizable energy. Data showing the response of the finishing pig to graded levels of tryptophan are shown in table 4. The estimated tryptophan requirement of the finishing pig receiving a 13.6% protein diet containing 3,727 kcal/kg of metabilizable energy is approximately.059 -+.001% of the diet and.43% of the protein. These values are slightly less than the estimated requirements of.07% of the diet and.58% of the protein of Becker et al. (1966). Assuming a dietary lysine sulfur amino acids %a TABLE 2. NITROGEN RETENTION BY FINISHING BARRO~VS FED GRADED LEVELS OF DL-METHIONINE (TRIAL i) Period b Barrow c Intake d Urine Feces Retention.17 A 2 51.65 32.68 2.96 16.01 B 1 51.65 36.98 2.68 11.99 C 3 51.65 34.05 2.93 14.66 D 4 51.65 30.83 2.61 18.21 51.65 33.64 2.80 15.22.26 A 3 51.65 31.13 3.25 17.27 B 2 51.65 30.43 3.19 18.03 C 4 51.65 34.25 2.99 14.40 D 1 51.65 35.87 3.02 12.76 51.65 32.92 3.11 15.62.35 A 1 51.65 36.17 3.14 12.33 B 4 51.65 32.84 2.82 15.99 C 2 51.65 34.56 3.50 13.59 D 3 51.65 32.54 3.02 16.08 51.65 34.03 3.12 14.50.44 A 4 51.65 31.94 2.70 17.01 B 3 51.65 33.25 3.00 15.40 C 1 51.65 34.15 2.75 14.74 D 2 51.65 32.28 3.05 16.31 51.65 32.91 2.88 15.87 apercent of diet... bperiods were consecutive and consisted of a 5-day pretest portion followed by a 5-day collection portion. ~Foux barrows were used in a 4x4 latin square design. Feed intake was 1.13 kg twice daily.
62 BROWN, HARMON AND JENSEN TABLE 3. NITROGEN RETENTION BY FINISHING BARROWS FED GRADED LEVELS OF L-ISOLEUCINE (TRIAL 2) L-isoleucine %a Period Barrow Intake Urine Feces Retention.22.30.38.46 A 7 51.29 32.95 2.38 15.97 B 5 51.29 32.24 2.96 16.09 C 8 51.29 38.08 2.46 10.75 D 6 51.29 34.25 2.49 14.55 51.29 34.38 2.57 14.34 A 8 51.29 36.37 2.60 12.33 B 7 51.29 29.42 2.70 19.17 C 6 51.29 32.95 2.58 15.77 D 5 51.29 29.72 2.70 18.87 51.29 32.12 2.65 16.54 A 6 51.29 32.44 2.64 16.21 B 8 51.29 35.67 2.66 13.07 C 5 51.29 29.32 2.76 19.21 D 7 51.29 31.33 2.80 17.16 51.29 32.19 2.69 16.41 A 5 51.29 29.92 2.88 18.49 B 6 51.29 30.63 2.64 18.02 C 7 51.29 30.93 2.65 17.71 D 8.51.29 33.95 2.74 14.60 51.29 31.36 2.73 17.21 asee footnotes of table 2. TABLE 4. NITROGEN RETENTION BY FINISHING BARROWS FED GRADED LEVELS OF L-TRYPTOPHAN (TRIAL 3) L-tryptothan %a Period Barrow Intake Urine Feces Retention.032.047.062 A 2 34.15 10.04 5.16 B C 3 1 27.50 26.60 8.29 7.55 13.56 16.20 D 4 29.52 7.59 12.24 29.44 8.37 11.54 A 4 27.81 5.40!6.14 B 1 24.89 7.33 17.14 C 3 27.10 7.51 14.74 D 2 28.61 7.73 13.01 27.10 6.99 15.26 A 3 25.59 7.71 16.05 B 4 25.39 6.74 17.22 C 2 24.99 7.87 16.49 D 1 21.16 6.96 21.23.077 asee footnotes of table 2. 24.28 7.32 17.75 A B 1 2 22.67 22.87 7.37 8.42 19.32 18.07 C 4 23.98 6.82 18.55 D 3 22.97 7.61 18.77 23.12 7.56 18.68
AMINO ACID REQUIREMENTS IN FINISHING PIGS 63 requirement of.62% of the diet, the tryptophan requirement calculated from the amino acid analysis and ratios of the pig carcass (Williams et al., 1954) is.056% of the diet, which agrees rather closely with our estimated dietary requirement of.059%. Pond, Kroening and Gallo (1964) observed that feeding corn containing.06% tryptophan and 9% protein supplemented with vitamins, minerals, antibiotics and lysine to finishing swine produced poor gains and efficiencies. The addition of.04% L-tryptophan to the diet produced gains, feed conversions, loin-eye areas and percentage of lean cuts not significantly different from those obtained from feeding a 12% protein corn-soybean meal diet. Gallo and Pond (1968) reported that a corn diet supplemented with.28% L-lysine HC1 and.02% L-tryptophan produced gains and efficiencies nearly equal to those obtained from feeding a 16% protein corn-soybean meal diet. Additional tryptophan gave no further benefit. Boomgaardt and Baker (1971) reported that the tryptophan requirement expressed as a percent of the dietary protein either increases or remains constant with increasing dietary protein level. Thus, dietary protein level may affect conclusions relative to the tryptophan requirement expressed as a percent of the diet. Our results show that the finishing pig required.059 +.001% dietary tryptophan for maximum nitrogen retention when the diet contained 13.6% crude protein and 3,727 kcal metabolizable energy per kilogram. Our estimate of the dietary isoleucine requirement for maximum nitrogen retention by finishing swine fed a 14% crude protein diet containing 3,700 kcal/kg of metabolizable energy is.31 -+.01%. Nitrogen retention was not increased by addition of methionine to the diet containing.17% total sulfur-bearing amino acids. It is concluded that finishing pigs fed a diet containing 14% crude protein and 3,700 kcal of metabolizable energy require.17% or less dietary total sulfur-bearing amino acids for maximum nitrogen retention. Literature Cited A.O.A.C. 1965. Official Methods of Analysis (10th Ed.). Association of Official Agricultural Chemists. Washington, D. C. Baker, D. H., W. W. Clausing, B. G. Harmon, A. H. Jensen and D. E. Becker. 1969. Replacement value of cystine for methionine for the young pig. J. Anim. Sci. 29:581. Baker, D. H., W. W. Hiott, H. W. Davis and C. E. Jordan. 1967. A swine metabolism unit. Lab. Pract. 16:1385. Becker, D. E., A. H. Jensen and B. G. Harmon. 1966. Balancing swine rations. IU. Agr. Exp. Sta. Circ. 866. Becker, D. E., A. H. Jensen, S. W. Terriil and H. W. Norton. 1955. The methionine-cystine need of the young pig. J. Anim. Sci. 14:1086. Becker, D. E., I. D. Smith, S. W. Terrill, A. H. Jensen and H. W. Norton. 1963. Isoleucine need of swine at two stages of development. J. Anim. Sci. 22:1093. Bell, J. M., H. H. Williams, J. K. Loosli and L. A. Maynard. 1950. The effect of methionine supplementation of a soybean oil meal purified ration for growing pigs. J. Nutr. 40:551. Boomgaardt, J. and D. H. Baker. 1971. The proteintryptophan interrelationship for growing swine..1. Anim. Sci. 33:1146 (Abstr.). Brown, H. W., B. G. Harmon and A. H. Jensen. 1973. The lysine requirement of the finishing pig for maximum rate of gain and feed efficiency. J. Anita. Sci. 37:708. Curtin, L. V., J. K. Loosli, Jean Abraham, H. H. Williams and L. A. Maynard. 1952. The methionine requirement for the growth of sine. J. Nutr. 48:499. Gallo, J. T. and W. G. Pond. 1968 Anim. acid supplementation to all-corn diets for pigs. J. Anim. Sci. 27:73. Meade, R. J. 1956a. The influence of methionine supplementation of 12, 14 and 16 percent protein corn-soybean oil meal diets upon nitrogen balance of growing swine. J. Nutr. 60:599. Meade, R. J. 1956b. The influence of tryptophan, methionine and lysine supplementation of a corn and soybean oil meal diet on nitrogen balance of growing swine. J. Anim. Sci. 15:288. Meade, R. J., W. R. Dukelow and R. S. Grant. 1966a. Influence of percent oats in the diet, lysine and methionine supplementation, and of pelleting on rate and efficiency of gain of growing pigs and on carcass characteristics. J. Anim. Sci. 25:58. Meade, R. J., W. R. Dukelow and R. S. Grant. 1966b. Lysine and methionine addition to corn-soybean meal diets for growing swine: Effects on rate and efficiency of gain and carcass characteristics. J. Anita. Sci. 25:78. N.R.C. 1968. Nutrient Requirements of Domestic Animals, No. 2, Nutrient Requirements of Swine. National Research Council, Washin~on, D. C. Oestemer, G. A., R. J. Meade, W. L. Stockland and L. E. Hanson. 1970. Methionine supplementation of opaque-2 corn for growing swine. J. Anim. Sci. 31:1133. Pond, W. G., G. H. Kroening and J. T. Gallo. 1964. Amino acids for swine: dietary requirements for growth and carcass leanness. Proc. Cornell Nutr. Conf. p. 50. Welch, J., R. H. Cordts and G. W. Vander Noot. 1966. Effect of lysine, methione and tryptophan supplementation upon nitrogen retention of barrows. J. Anim. Sei. 25:806. Williams, 16, H. H., L. V. Curtin, J. Abraham, J. K. Loosli and L. A. Mayn~ird. 1954. Estimation of growth requirements for amino acids by assay of the carcass. J. Biol. Chem. 208:277.