Protein and Sulfur Amino Acid Requirement of the Laying Hen as Influenced by Dietary Formulation

Protein and Sulfur Amino Acid Requirement of the Laying Hen as Influenced by Dietary Formulation R. H. HARMS AND B. L. DAMRON Florida Agricultural Experiment Station, Gainesville, Florida 32601 (Received for publication June 10, 1968)

T

Florida Agr. Exp. Sta. Journal Series No. 2999.

indicated that these levels of sulfur amino acids were marginal for the laying hen and that a consistent increase in egg production, egg weight and body weight gain was obtained by feeding slightly higher levels. The purpose of this study was to determine the influence of various total protein and sulfur amino acid levels upon the methionine requirement of the laying hen. EXPERIMENTAL PROCEDURE Hy-Line 934-H pullets, purchased from a commercial hatchery, were used in these experiments. The chicks were hatched on July 10, 1965, and August 7, 1966, respectively, for experiment one and two. They were debeaked, dubbed and vaccinated intra-ocularly for infectious bronchitis and Newcastle disease at one day of age. Fowl pox vaccination was given at four weeks of age and infectious bronchitis and Newcastle vaccinations were repeated at eight and 22 weeks of age. For the first eight weeks of life the pullets were fed a starter diet containing 22 percent protein and 2,079 kilocalories of productive energy per kilogram. At eight weeks of age the protein level was reduced to 16 percent and this diet was given until the birds were moved into laying cages at 22 weeks of age. At this time they were changed to a commercial egg production type laying mash containing 17 percent protein (Waldroup and Harms, 1964). At 28 weeks of age the birds were randomly distributed into laying cages in groups of five individually caged pullets. Fifteen experimental diets, containing

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HE National Research Council (1966) suggests that laying hens require 15 percent protein in a diet containing 2,850 kilocalories of metabolizable energy per kilogram. They have also suggested that the diet should contain 0.28 percent methionine and 0.25 percent cystine. In the past the protein requirement of the laying hen has been reported to be as high as 17 percent and as low as 11 percent. A review of the literature concerning protein requirement of the laying hen was made by Harms et al. (1962) in a paper in which it was reported that the energy content of the diet and season of the year would influence this requirement. In a later study (Harms et al., 1966), it was found that each decrease of 22 kilocalories of productive energy per kilogram of feed resulted in a one percent increase in the amount of feed consumed per hen per day. This change in feed intake resulted in a one percent change in the methionine or sulfur amino acid required, expressed on a per unit of feed basis. In this study it was also reported that a daily intake of approximately 0.28 grams of methionine was necessary to support maximum egg production. A slightly higher level was required for maximum egg weight and body weight gain. This requirement was met by a level of 0.271 percent methionine and 0.475 percent total sulfur amino acids in a diet containing 2,112 kilocalories of productive energy per kilogram. However, a subsequent study by Harms et al. (1967)

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AMINO ACID REQUIREMENT OF HENS

TABLE 1.—Protein, methionine and sulfur amino acid content of experimental diets Sulfur %of Protein Methionine amino methionine (%) Acid requirement (%)

Formulation

(%)

Corn-soybean meal

70 85 100 115 130

11.93 14.69 17.46 20.24 23.05

0.188 0.228 0.268 0.308 0.348

0.374 0.453 0.533 0.612 0.691

Corn-soybean mealfish meal

70 85 100 115 130

10.40 13.16 15.93 18.69 21.47

0.188 0.228 0.268 0.308 0.348

0.344 0.424 0.503 0.583 0.662

70 85 100 115 130

10.61 11.84 13.07 14.30 16.00

0.188 0.228 0.268 0.308 0.348

0.355 0.412 0.470 0.528 0.592

Corn-soybean mealmethionine*

* Methionine supplied as methionine hydroxy analogue.

mulation. All diets were maintained isocaloric at 2,887 kilocalories of metabolizable energy per kilogram. The diets containing 0.268 percent methionine were selected as the diets expected to meet the laying hens requirement; therefore, these

TABLE 2.—Composition of diets CSM4

CSF 3

CS2 Ingredients 70

130

70

130

(%) 74.91 55.89 28.23 3.35 2.50 2.50 1.82 1.00 6.46 6.89

130

74.57 7.34 2.50 1.00 6.85

66.32 19.29 2.50 1.00 6.89

—

2.01 0.35 0.50 4.86

1.89 0.35 0.50 1.13

—

0.024

0.126

—

—

21.47 0.348 0.662

10.61 0.188 0.355

16.00 0.348 0.592

Yellow Corn Soybean Meal (50% protein) Alfalfa Meal (20% protein) Stabilized Animal Fat Ground Limestone Defiuorinated Phosphate (18% P & 32% Ca) Iodized Salt Micro-ingredients1 Builders Sand Methionine-hydroxy Analogue (90%) Menhaden Fish Meal (60% protein)

72.55 10.27 2.50 1.00 6.86

48.85 36.03 2.50 3.61 6.42

1.98 0.35 0.50 3.99

1.74 0.35 0.50

—

1.50 0.35 0.50 6.00

— —

— —

3.00

3.00

% Protein % Methionine % Sulfur Amino Acids

11.93 0.188 0.374

23.05 0.348 0.691

10.40 0.188 0.344

—

70

1.25 0.35 0.50

1 Supplied per kilogram of diet: 6,600 I.TJ. vitamin A, 2,200 I.C.U. vitamin D 3 , 500 mg. choline chloride, 40 mg. niacin, 4.4 mg. riboflavin, 13 mg. pantothenic acid, 22 meg. vitamin B12, 125 mg. ethoxyquin, 20 mg. iron, 2 mg. copper, 200 meg. cobalt, 1.1 mg. iodine, 100 meg. zinc, 71 mg. manganese and 2.2 mg. sodium bisulfite. 2 Indicates corn and soybean meal varied to supply desired protein level. 3 Indicates corn, soybean meal and fish meal varied to supply desired protein level. 4 Indicates corn, soybean meal and methionine hydroxy analogue varied to supply desired protein level.

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various levels of protein, methionine and sulfur amino acids from three different formulations, were fed (Table 1). Sixteen and eight replicates of five individually caged hens were fed each diet in experiments one and two, respectively. One series of diets was formulated with the major portion of the protein coming from corn and soybean meal (CS). Another series was formulated with the protein furnished from corn, soybean meal and 2.5 percent menhaden fish meal (CSF). In the third series, the protein was supplied from corn, soybean meal and a synthetic source of methionine (CSM). The composition of diets containing the lowest and highest levels of protein and sulfur amino acids is shown in Table 2. All diets were formulated using linear programming techniques. Nutrient analyses of ingredients according to Maddy et al. (1963) were used as a basis of for-

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R. H. HARMS AND B. L. DAMRON

TABLE 3.—Rale of egg production, egg weights, feed efficiency, and body weight of hens fed various of protein and/or sulfur amino acid

% .

methionine requirements

CS1

CSF2

CSM3 Average*

Production (%) 52.0 58.1 67.3 68.3 69.6 73.9 72.3 72.5 72.6 74.1

53. 4" 65.5 b 70.6 b " 72.2" 72.9°

70 85 100 115 130

Egg weights (gms.) 55.2 56.4 55.8 56.9 57.0 56.1 57.6 58.1 57.5 58.1 58.3 58.5 58.6 58.8 57.8

55.8* 56.7 ab 57.7" 58.3 b 58.4 b

70 85 100 115 130

Feed efficiency (kg./doz.) 1.75 1.97 2.09 1.58 1.66 1.77 1.56 1.56 1.67 1.51 1.54 1.61 1.50 1.59 1.55

1.94» 1.67b 1.60b« 1.55" 1.54"

70 85 100 115 130

Body wt. (gms.) l,47l" 1,422 1,504 1,522 1,606 1,527 1,595 1,635 1,657 1,628

1,303 1,412 1,519 1,582 1,587

1,399" l,479 b 1,551" l,604 d l,624 d

1 Indicates corn and soybean meal varied to supply desired protein level. 2 Indicates corn, soybean meal and fish meal varied to supply desired protein level. 3 Indicates corn, soybean meal and methionine hydroxy analogue varied to supply desired protein level. * Means with different superscripts are significantly different according to Duncan's multiple range test.

diets were designated as 100 percent of the estimated requirement and will be referred to in that manner hereafter. Daily egg production records were kept on each pullet throughout the 280-day experimental period. Rate of egg production was calculated on a hen-day basis. One egg was weighed from each hen at the end of each 28-day period, and these weights were averaged for each group. The feed consumed by each group was measured at 28-day intervals. Using these data, feed efficiency and feed consumed per hen per day were calculated. Each

RESULTS AND DISCUSSION

Egg Production Period. A significant linear response was obtained in egg production from increasing the level of methionine and sulfur amino acids in the diet (Table 3). Using Duncan's multiple range test a maximum rate of egg production was obtained with a level of 0.268 percent methionine and 0.533 percent total sulfur amino acids when corn and soybean meal served as the source of protein. However, using the method of orthogonal polynomials it was found that 0.317 percent methionine was required for maximum production. A higher level of methionine was required for maximum production with the diets containing fish meal or synthetic methionine. This difference was attributed to the difference in the total sulfur amino acid content in the three different diet series. In the CS series a level of 0.533 percent total sulfur amino acids were present as compared to a level of 0.503 and 0.470 percent with the CSF and CSM diets, respectively. These data agree very well with the requirements as suggested by NRC (1966) and Harms el al. (1967) in that the methionine requirement for maximum egg production is not in excess of 0.28 percent in a diet containing approximately 2850 kilocalories of metabolizable energy per kilogram, provided that at least 0.23 percent cystine is present. Although the diet formulation X amino

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50.0 61.0 68.2 71.9 72.0

70 85 100 115 130

bird was weighed at the end of the tenth 28-day test period. Since there were no interactions of treatment X year for any measurements obtained, the data have been pooled for this presentation. Statements of probability are based on analysis of variance according to Snedecor (1956), with significant differences between treatment means determined by a modification of Duncan's multiple range procedure as suggested by Kramer (1956).

AMINO A C I D R E Q U I R E M E N T OF H E N S

T h e interaction of month X amino acid level was statistically significant. This was attributed to the spread in rate of lay between the lowest and highest level of amino acids increasing from 14 percent in the first month to 25.4 percent during the last month. The fact t h a t the spread in egg production increased as the laying period progressed indicates t h a t the amino acid requirement of the laying hen increases with length of lay. R a t e of egg production was closely associated with level of protein. However, as previously discussed, this relationship was assumed to be due to the increase in level of methionine a n d / o r total sulfur amino acids as the protein levels were increased. An analysis was made on total egg mass, which was calculated by multiplying monthly egg weights by egg production. N o significant differences were noted with this measurement t h a t were not observed for rate of production; therefore, these values are not reported. Egg Weights. A significant linear response was obtained in egg weights from increasing the level of methionine in the diet (Table 3). Although a significant increase in egg weights was not observed

when the amino acid level was increased from 100 to 130 percent of the requirement, there was a numerical increase. Using the method of orthogonal polynomials, a level of 0.319 percent methionine was required for maximum egg weight. This was relatively constant, indicating t h a t the amino acid requirement for maximum egg weights did not change greatly throughout the production year. These d a t a also indicate t h a t the sulfur amino acid requirement for maximum egg weight is slightly higher than for maximum rate of egg production. Feed Efficiency. As the level of sulfur amino acids was increased, the feed required to produce a dozen eggs was reduced. T h e major portion of this difference was attributed to the different rate of egg production. The level of amino acids supporting maximum egg production also supported maximum feed utilization. Body Weights. A significant linear response was obtained in adult body weight from increasing the level of sulfur amino acids in the diet (Table 3). This difference in body weight was as expected, since it has been previously reported ( H a r m s et ah, 1962, 1967) t h a t an increase in protein level resulted in larger body weight gains of laying hens. I t would appear t h a t the requirement for body weight gain parallels very closely the requirement for maximum egg production and maximum egg weight. Feed and Amino Acid Intake. Feed intake for the five amino acid levels is shown for each of the ten monthly experimental periods in Table 4. Feed intake was slightly reduced on the two lower levels of amino acids and this was considered to be due to the lowered rate of egg production. These d a t a would indicate t h a t the

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acid level was not statistically significant, there were definite trends. At the 70 percent level, the CSF and CSM diets did not support the rate of production equal to t h a t obtained with the CS diet, although each contained equivalent levels of methionine. This may be explained by the level of total sulfur amino acids, since the CS diet contained a level of 0.374 percent sulfur amino acids as compared to 0.344 and 0.355 percent for the CSF and CSM diets, respectively. This would indicate t h a t the lowered rate of egg production for the CSF and CSM diets was due to an inadequate level of total sulfur amino acids.

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R. H. HARMS AND B. L. DAMEON TABLE 4.— Feed consumed

per hen per day {grams) Month

% estimated methionine requirement

1

2

3

4

5

6

7

8

9

10

All

70 85 100 115 130

93 93 92 90 92

94 96 96 93 94

89 94 96 93 97

88 93 87 92 92

84 88 90 90 93

83 88 89 88 93

80 86 89 88 90

82 84 88 91 91

87 90 94 93 93

89 95 97 96 97

87 91 92 92 93

Based on these feed intake data it would appear that when corn and soybean meal are used to supply the protein for laying hens the requirement for methionine is 0.25 to 0.28 grams per day and the total sulfur amino acid requirement is 0.53 grams. This level of methionine intake with the CSF and CSM diet did not support a maximum rate of lay, since the total sulfur amino acid intake did not reach the 0.53 grams per day. Therefore, it would appear that the commercial egg production type hen requires 0.25 to 0.28 grams of methionine per day, provided it is getting an additional 0.25 grams of

cystine or a total of approximately 0.50 sulfur amino acids. The fact that the hens were consuming increased amounts of feed during the ninth and tenth month of the experiment and during the same time a wider spread occurred in rate of production from the low and high levels of amino acid indicates that the hen's requirement for amino acids does increase in the later stages of production. SUMMARY

A study involving two 280-day laying hen experiments was conducted to determine the methionine and sulfur amino acid requirement of commercial egg production type pullets as influenced by diet formulation. Measurements were obtained on rate of egg production, feed consumed per dozen eggs, body weight gain and egg weights from Hy-Line 934-H pullets. Data from this study indicate that the hen requires 0.25 to 0.28 grams of methionine daily provided she is supplied a total of 0.53 grams of sulfur amino acids. This requirement was met by a level of 0.268 percent methionine and 0.533 percent total sulfur amino acids in a diet containing 2,887 kilocalories of metabolizable energy per kilogram. However, this level of methionine did not support maximum performance when the diet contained lower levels of total sulfur amino acids.

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hen eats to meet its energy requirement and cannot overconsume in order to meet its amino acid requirement. There was a trend for slightly higher feed intakes during the first and second month, and the ninth and tenth month. This was attributed to the cooler weather during these two periods. Although there was a slight difference in feed intake, it did not influence the apparent sulfur amino acid requirement. The feed intake during the first year was approximately 5 percent greater than the second. However, the trend for the entire ten months was the same in both experiments. Although the birds in the first experiment consumed approximately 5 percent more feed, this did not alter their amino acid requirement. It is possible that the assay with laying hens is not sufficiently critical to detect a 5 percent difference.

AMINO ACID REQUIREMENT OF H E N S ACKNOWLEDGEMENT

This work was supported in part by the National Institute of Child Health and Human Development (Grant HD 0029-01A2). The authors are indebted to Dr. W. A. Dudley of Monsanto Company for his assistance in designing these experiments. They are also indebted to Dr. F. B. Martin, Associate Statistician, University of Florida, for assistance in analyzing the data. REFERENCES

1967. Evaluation of the sulfur amino acid requirements of commercial egg production type pullets. Poultry Sci. 46:181-186. Harms, R. H., C. R. Douglas and P. W. Waldroup, 1962. Methionine supplementation of laying hen diets. Poultry Sci. 41: 805-812. Kramer, C. Y., 19S6. Extension of multiple range test to group means with an equal number of replications. Biometrics, 12: 307. Maddy, K. H., R. B. Grainger, W. A. Dudley and F. Puchal, 1963. The application of linear programming to feed formulation. Feedstuffs, 35(15): 28-30, 70-73. National Research Council, 1966. Nutrient requirements of domestic animals. No. 1 Nutrient requirements of poultry, Washington D. C. Snedecor, G. W., 1956. Statistical Methods, 5th edition. Iowa State College Press, Ames, Iowa. Waldroup, P. W., and R. H. Harms, 1964. The effect of dietary protein restriction of laying hens on subsequent performance. Poultry Sci. 43: 792794.

Ovalbumin as a Factor in the Evaluation of Betapropiolactone Inactivated Newcastle Disease Vaccines* JOHN GRUN Department of Animal Sciences, Rutgers, The State University of New Jersey, New Brunswick, New Jersey 08903 (Received for publication June 14. 1968)

M

ACK and Chotisen (1955) showed that beta-propiolactone (BPL) inactivated Newcastle disease virus (NDV) vaccines are feasible. This stimulated work by Sullivan et al. (1958), Winmill and Weddell (1961), Dardiri and Yates (1962), Hofstad et al. (1963), Piercy et al. (1964), Grun and Hudson (1966), and Grun (1966a). Evaluations and comparisons, such as Hofstad's (1967) evaluation of commercial inactivated NDV vaccines, have been an inherent part of this re* Paper of the Journal Series, New Jersey Agricultural Experiment Station, Rutgers, The State University of New Jersey, New Brunswick, New Jersey.

search up to the present time. Winmill and Weddell (1961) and Piercy et al. (1964) mentioned the removal of albumin in their preparations. Grun and Hudson (1966) and Grun (1966a, b) indicated that albumin may be a factor which should be considered because it may have an influence on the course of a Gilbert-Boney (GB) NDV challenge infection. This report describes the activity of a number of BPL inactivated materials in Single Comb White Leghorn (S.C.W.L.) chickens infected with NDV:BPL inactivated amnio-allantoic (A A) fluid; A A fluid+NDV; whole embryo (WE) preparation controls; and various concentra-

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Harms, R. H., B. L. Damron and P. W. Waldroup, 1966. Influence of energy level upon performance and methionine requirement of the laying hen. Proc. VII International Congress of Nutr. 1:160163. Harms, R. H., B. L. Damron and P. W. Waldroup,

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