Relation of Tryptophan and Lysine to Egg Production, Hatchability and Composition of the Protein of Hens’ Eggs

POULTRY SCIENCE November, 1950, Vol. XXIX, No. 6 •

G. R. INGRAM, W. W. CRAVENS, C. A. ELVEHJEM AND J. G. HALPIN Departments of Poultry Husbandry and Biochemistry, University of Wisconsin, Madison (Received for publication April 1, 1950)

T

HIRTY-FIVE years ago Osborne and Mendel (1914) demonstrated the effect of tryptophan and lysine deficiencies on the growing rat and various workers have since shown that a deficiency of any one of the essential amino acids has a similar effect. Extensive studies have established, within rather narrow limits, the requirements of the rat (Rose, 1937, and Frazier el al., 1.949), the chick (Almquist, 1947) and the human (Rose, 1949) for the essential amino acids. However, there has been very little work reported on the requirements of the laying hen. Cravens and Halpin (1946) and Cravens (1948) using nine percent casein and

Published with the approval of the Director of the Wisconsin Agricultural Experiment Station. Supported in part by a grant from Hiram Walker & Sons, Peoria, Illinois. We are indebted to E. I. duPont deNemours Company, Wilmington, Delaware, for the lysine and methionine and to the Dow Chemical Company, Midland, Michigan, for the glycine and tryptophan. 793

seven percent gelatin as sources of protein demonstrated the effect of a leucine deficiency and a multiple amino acid deficiency on the laying hen. Grau and Taylor (1948) reported that zein supplemented with arginine, histidine, methionine, cystine, threonine and valine plus lysine and tryptophan supported fortyone percent egg production compared to seventy-one percent production for a practical ration. When either lysine or tryptophan was omitted, egg production ceased in six days. The relation- between the amino acid composition of the diet fed the hen and the make-up of the egg has been studied by several workers. Csonka, Denton and Ringel (1947) showed a modification of the quantity of cystine and methionine in the egg by dietary means. Gerber and Carr (1930) presented data which indicated that the egg protein of pigeons might be modified by the diet fed the birds. In contrast to these reports, Mc-

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Relation of Tryptophan and Lysine to Egg Production, Hatchability and Composition of the Protein of Hens' Eggs

794

INGRAM, CRAVENS, ELVEHJEM, AND HALPIN

EXPERIMENTAL

Single Comb White Leghorn pullets were used in all experiments. Four birds were fed each ration from B-101 through B-116. Due to expense of the rations, only three hens per lot were used on the remaining rations. The hens were kept in individual laying cages with wire floors and inseminated weekly with pooled semen from New Hampshire males. The eggs were gathered daily and pedigree marked. Settings were made weekly and the hatchability recorded. Feed was kept before the birds at all times. The practical ration used was ration B-l of Robblee et al. (1948). After each experiment the birds were fed the practical ration until they regained any lost weight and had resumed a high rate of production before being used for further experiments. The control ration used throughout these experiments is ration B-106 in Table 1. Since egg production of hens fed this ration was always good, averaging seventy-two percent, the daily production for this group will be omitted from the tables giving the results of each experiment.

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Farlane, Fulmer and Jukes (1930) reported no change in egg proteins as a result of modifications of the diet fed laying hens. The studies reported herein were undertaken to further study the effect of specific amino acid deficiencies on the hen and on the hatchability of eggs produced and to determine the quantitative requirements of the laying hen for certain amino acids. Data on the amino acid composition of eggs as related to diet fed the hen are also presented. Lysine and tryptophan were studied. Zein, corn gluten meal and acid hydrolyzed casein were used as proteins low in these amino acids.

795

EFFECT OF TRYPTOPHAN AND LYSINE ON H E N ' S EGGS

Detailed body weights are not presented since changes were often made in the rations while the experiments were in progress. However, any marked changes in weights will be discussed with each experiment. Some of the eggs laid at the time the hens were going out of production and just as they resumed production after a

determinations. Methionine and cystine were determined by the method of Steele, Sauberlich, Reynolds and Baumann (1949). The following organisms were used: Lysine Tryptophan Cystine Methionine

Leuconostoc mesenteroides p-60 Lactobacillus arabinosus 17-5 Leuconostoc citrovorum 8081 Leuconostoc citrovorum 8081

Days on experiment Lot No.

1 101 102 103

4 3 4

2 1 1

3 2 2 2

4 1 2 2

5

6

7

1 2

8 * * *

9

10

11

12

13

14

15

16

1

3 2 2

1 1 1

* Placed on practical ration.

pause, were saved and assayed for certain amino acids. The eggs were held in a refrigerator until ready for assay. This holding period was usually about two weeks. After boiling for ten minutes in order to precipitate the protein, the yolk and albumen were separated and the nitrogen determined by the Kjeldahl method. Two samples of each yolk and albumen were weighed out. One sample was subjected to acid hydrolysis with 3N HCl for ten hours (5N NaOH for fifteen hours for tryptophan determination) and the amino acids determined microbiologically. The second sample was mixed with water and homogenzied in a Waring Blender for five minutes and then filtered through a coarse filter paper. The residue was discarded and microbiological amino acid determinations made on the filtrate. The values obtained from the samples subjected to the latter procedure are called throughout this paper the "free" amino acids. The method of Henderson and Snell (1947) was used for lysine and tryptophan

RESULTS AND DISCUSSION

Experiment #/.—The rations fed in the first experiment were B-101, B-102 and B-103 shown,in Table 1. All the rations contained zein which is known to be deficient in lysine and tryptophan. Ration B-101 was supplemented with tryptophan, B-102 with lysine and B-103 with both amino acids. The effect of these rations on egg production is shown in Table 2. The production ceased on the fourth or fifth day after the hens were placed on the rations. Since it was evident from examinations of the hens that they were definitely out of production, all groups were placed back on the practical ration on the eighth day. Hens in all groups started production again on the fourteenth day and were back to normal four or five days later. During the first week the hens on ration B-101 lost an average of thirty-four grams, those on B-102 lost one hundred and forty-six grams, and those on B-103 lost thirty-one grams. It is evident from the results of this

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TABLE 2.—Daily production for experiment §1 (Number of Eggs)

796

INGRAM, CRAVENS, ELVEHJEM, AND HALPIN

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Experiment §2.—Rations B-107 and B-108 were used in this experiment. B-107 contained zein alone, while ration B-108 was supplemented with five percent casein to raise the level of histidine and threonine. Tryptophan and lysine were included to raise the level of these two amino acids to approximately that found in the control ration. The egg production data in Table 3 show that the hens fed ration B-107 stopped laying on the fifth day, but one egg was laid on the thirteenth day. The hens fed ration B-108 continued to lay for a much longer period, but they ceased production on the fifteenth day. On the eighteenth day five percent gelatin was added to the ration to raise the level of arginine and glycine, since all the other amino acids were present in quantities known to support egg production. The hens started laying five days later and continued to lay intermittently throughout the remainder of the experiment, although production never reached the level of the control hens. The hens fed ration B-107 lost approximately two hundred grams each during the three week experiment. To prevent loss of the bens they were placed back on

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experiment that zein.. when supplemented with both lysine and tryptophan at the levels used in this study is still deficient in some nutrient, presumably an amino acid, and does not support egg production in hens. In fact the hens on the supplemented ration stopped production as quickly as did the hens on the rations supplemented with only one of the two amino acids. From a comparison of the amino acid content of zein with that of our control ration as given by Block and Boiling (1947), it appeared that histidine and threonine might be the deficient amino acids and thus the next experiment was designed to test this possibility.

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EFFECT OF TRYPTOPHAN AND LYSINE ON H E N ' S EGGS

Experiment §3.—Since the hens fed B-108 in Experiment 2 ceased laying before gelatin was added, it was thought that if they were placed on this ration while in a high rate of production the ration would support production. Therefore, one group of hens was fed B-108a. Ration B-109 was designed to be deficient in tryptophan using gelatin in combinations with zein to increase the level of arginine, glycine and threonine, while lysine and histidine were added in crystalline form. In ration B-110, zein was supplemented with all the amino acids thought to be low except lysine. The production for all three groups is shown in Table 3. The production of the hens fed ration B-108a dropped severely after four or five days on the ration and continued at a low rate for about two weeks, then improved somewhat, but never did reach the level of the controls. The average production of the lot was forty-four percent. On ration B-109, the hens ceased production on the fourth day. Tryptophan was added on the tenth day and the hens laid a few eggs at intervals during the re-

mainder of the experiment. After the production of each egg, there was a delay of several days before another egg was laid. The hens fed B-110 ceased production in seven days and lysine was added on the tenth day. Production started seven days later and continued at a somewhat higher rate than was obtained with tryptophan supplemented B-109. The hens on ration B-108a maintained their weight throughout the experiment, while the hens fed the other two rations lost weight. On ration B-109, deficient in tryptophan, the hens lost one hundred grams each before tryptophan was added to the ration. The hens then maintained their weight for two weeks and toward the end of the experiment they again lost weight. The average loss was two hundred and forty grams each during the fortyfour days of the experiment. The birds fed ration B-110 which was deficient in lysine lost fifty grams each before the addition of lysine. These hens then maintained their weight for three weeks before again losing weight. The average loss for this group during the experiment was one hundred and fifty grams. Since the hens fed B-110 came back into production upon the addition of lysine, it was thought that if the level of the amino acids added to the zein were increased, the ration would be adequate to support maximum egg production. However, a group of hens fed ration B-llOa stopped production on the eighth day and went into a severe molt. It is possible that an imbalance of the amino acids was responsible for the poor results of this group. Experiment §4.—From the results of the previous experiments it was evident that zein, even when supplemented with the deficient amino acids, was an inade-

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the practical ration after twenty-five days. It required about two weeks for these hens to regain their weight and to come into production again. All hens, except one, fed ration B-108 maintained their weight, even before the addition of gelatin, and by the end of the experiment the three had gained an average of one hundred and forty-seven grams per hen. The one hen gradually lost weight and died on the forty-fifth day. The fact that three of the hens were able to maintain their weight would indicate that the requirement of the hen for amino acids are much more critical for egg production than for the maintenance of body weight. This supports the observations of Cravens (1948).

797

798

INGRAM, CRAVENS, ELVEHJEM, AND HALPIN

would result in complete cessation of production. The poor results obtained with B-116 when compared with B - l l l may have been due either to an amino acid inbalance or to the toxic action of the form of isoleucine used in ration B-116. Ration B-115 which contained soybean oil meal as the source of protein supported egg production at the same level as casein supplemented with gelatin, and since the effect of the protein on egg production is very rapid, it appears that soybean

TABLE i.~Daily production for experiment §4 (Number of Eggs) Days on experiment Lot N o . Ill 112 113 114 115 116

•1

2

3

4

3 4 4 3 3 2

2 3 2 2 4 3

2 1 1 1

3 4 3 3 4 1

-3

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6

7

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4 1

3 3

3 2

8

9

10 11 12 13 14 IS

16 17

18 19 20 21 22 23 24 25 26 27 28

2

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2 2 1

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2 1

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1 2 3

2 2

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3 1

4 1

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1 1

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4

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1 2 1 1 3

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being low in corn gluten meal. Ration B-115 with soybean oil meal as the source of protein was used for comparison with the corn gluten meal. The daily production supported by these rations is shown in Table 4. The percent production was as follows: B-lll B-112 B-113 B-114 B-115 B-116

47

% 36% 34% 14% 68% 32%

From these results it is evident that corn gluten meal was improved by the addition of lysine and tryptophan. It is of interest to note that the addition of either one of these amino acids improved the ration for production and that the extent of improvement was almost the same. We realize that only a partial deficiency was obtained with corn gluten meal alone, because a complete deficiency of either one of these two amino acids

oil meal as the sole source of protein would continue to support egg production indefinitely. The weights of the hens fed these rations showed very little change with the exception of those receiving B-115. The hens on this ration gained one hundred and thirty grams each during the experiment. Experiment §5.-—Acid hydrolyzed casein prepared according to a modification of the method of Henderson et al. (1947) was used as the basic protein for studying the effect of a tryptophan deficiency. The sample of acid hydrolyzed casein was tested with rats and found to give satisfactory results when supplemented with tryptophan. The rations used were B-117 through B-119. Rations B-117 and B-118 contained five percent gelatin in addition to the acid hydrolyzed casein. Ration B-117, which was designed to be deficient in tryptophan, was supplemented

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quate protein for egg production, and thus it was decided to use corn gluten meal as the protein in rations to study tryptophan and lysine deficiencies. Rations B - l l l through B-116 were fed to hens in this experiment. In ration B - l l l the corn gluten meal was supplemented with both tryptophan and lysine, in B-112 with lysine and in B-113 with tryptophan. Ration B-114 contained corn gluten meal alone, while B-116 contained a supplement of all the amino acids calculated as

799

EFFECT OF TRYPTOPHAN AND LYSINE ON H E N ' S EGGS 1

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The weight of the hens varied considerably when fed these rations. On B-117 there was an average loss of five hundred grams per hen. Even the addition of tryptophan did not stop this loss of weight. The hens were too run down and as noted above, failed to eat. The hens on B-117a gained an average of one hundred and thirty grams. This demonstrates again the critical requirement of the hen for protein to support egg production. The hens on B-118 lost an average of two hundred and fifty grams. There was a slight loss of weight on ration B-119, but this was due to one hen. The others just about maintained their weight. The results of this experiment are hard to explain on the basis of amino acid composition of the rations, since the amino acid level of the acid hydrolyzed casein

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with this amino acid after production of the hens ceased. When the hens on B-117a did not lay as well as was expected, additional methionine was added to this ration. Rations B-118 and B-119 contained five percent casein and five percent gelatin in addition to the acid hydrolyzed casein. Methionine was added to both rations and tryptophan was added to B-119. Production for these groups is shown in Table 5. The hens on ration B-117 went out of production in five days and the addition of tryptophan did not bring them back into production. A loss of appetite was noted for the hens fed this ration and very little feed was eaten even after the addition of tryptophan. B-117a also failed to maintain normal production. The addition of methionine improved production some, but it remained very low. The hens fed B-118 also stopped production on the fifth day. However, a single egg was obtained toward the end of the experiment. The hens fed B-l 19 continued to lay, but at a much reduced rate.

800

INGRAM, CRAVENS, ELVEHJEM, AND HALPIN

that the lowered hatchability noted in experiment #2 was caused by the same deficiency. Since according to the work of Sunde et al. (1950) the original level of folic acid was more than sufficient to support maximum egg production, it is assumed that the variation in egg production in both experiments was caused by some deficiency of the protein. There was no indication in any of the other experiments that hatchability was affected. The eggs which were laid as the hens were going out of production, or as they were coming back into production after a pause, hatched as well as the control eggs. Since we were unable to design a deficient ration which when supplemented would support maximum egg production, we did not feel justified in attempting a quantitative study of the requirements of the laying hen for lysine and tryptophan. We were successful in obtaining definite amino acid deficiencies in certain rations. From the results of the experiments in which zein was used as the basic protein, it is evident that the ability of zein to support egg production in the hen cannot be calculated from its amino acid composition. In rations which had all the essential amino acids of zein raised to the level of the control ration, production still did not equal that of the controls. These results are probably explained on the basis of incomplete release of the amino acids of the zein by the enzymes of the digestive tract. Our results are similar to those obtained by Geiger (1949) with rats fed zein. He obtained very poor growth when the essential amino acids of zein were raised to the level of casein. Digestion of the zein with pancreatin for two days before feeding did not help. When acid hydrolyzed, and properly supplemented with the deficient amino acids, the zein gave results comparable to casein.

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was determined microbiologically and supplemented to the minimum level known to support egg production. The hens on rations B-118 and B-119 were killed and an examination made of some of the internal organs. The only abnormalities noted were in the ovaries. One of the hens from B-118 showed small, round, transparent cysts in the ovaries. All of this group exhibited the loose, flabby yolks reported by Cravens (1948). These yolks appeared to be partially reabsorbed. The ovaries of the hens on ration B-119 did not have the flabby yolks found in the previous group. One hen showed the typical ovaries of a hen out of production with no developing ovum. The other two hens had ovaries with fully developed yolks and from appearance, one would have thought that they were in full production. Hatchability.—The hatchability of the eggs from the hens fed the experimental rations remained as high as those from the hens receiving the control ration in all experiments. In experiment #2 the hatchability of the eggs from the hens receiving ration B-108 remained high for the first three weeks and then suddenly dropped to about fifty percent and remained at approximately this level for the remainder of the experiment. There was a simultaneous drop in the hatchability of the control eggs. A vitamin deficiency was suspected although by the time the drop in hatchability was detected, the birds were off the experiment. In experiment #3 the eggs from ration B-108a and the controls again showed a marked drop in hatchability similar to that noted in experiment #2. An examination of the dead embryos indicated a folic acid deficiency (Sunde et al., 1950), therefore the level of folic acid was raised to 3 mgs. per kg. of ration. This raised the hatchability to normal. It is assumed

801

EFFECT OF TRYPTOPHAN AND LYSINE ON H E N ' S EGGS

been responsible for some of the results obtained in these studies. Throughout all of these experiments, it was noted that some of the hens on the deficient rations would appear from physical examination to still be in production for some time after egg production had ceased and at times would lay an occasional egg after a prolonged pause. If the deficiency was acute, the birds would be thrown into a severe molt.

TABLE 6.—Amino acid content of eggs {percent of protein) Lysine

Cystine Lot No.

Days*

Yolk Total Free

106 Practical

6 17

Yolk

Albumen

Total F r e e f t Total Free Total Free

1.51 1.46

.039 .042

2.29 2.01

— —

Ave. of 4 eggs . 1.42

6.6 6.2

.28 .26

5.7 5.8

Methionine

Tryptophan

Albumen

.085 .084

Yolk

Albumen

Yolk

Albumen

Total Free

Total Free

Total Free

Total Free

1.42 1.24

.071 .043

1.57 1.32

0.02 0.02

2.62 2.87

.088 .056

3.87 3.67

.055 .056

.031

2.42

—

6.4

.39

5.8

.087

1.48

.120

1.46

0.81

2.43

.104

3.85

.043

109

16

1.54

.038

2.87

—

6.6

.26

5.6

.087

1.22

.053

1.39

0.02

2.58

.055

3.87

.055

111

21

1.54

.007

2.38

—

6.4

.31

5.7

.084

1.38

.10

1.52

0.02

2.71

.086

3.91

.047

112

1 4 7 14

1.51 1.48 1.51 1.60

.053 .054 .050 .041

'2.42 2.48 2.37 2.42

— — — —

6.7 6.6 6.8 6.3

.22 .25 .26 .24

5.8 5.8 6.0 5.9

.081 .092 .079 .084

1.32 1.28 1.42 1.40

.055 .060 .075 .071

1.52 1.53 1.40 1.46

0.02 0.02 0.03 0.02

2.62 2.65 2.52 2.58

.088 .106 .084 .071

4.00 3.82 3.91 3.76

.060 .065 .055 .055

113

1 6 18 20 21

1.58 1.49 1.48 1.53 1.55

.051 .042 .031 .045

2.21 2.32 2.48 2.51 2.51

— — — — —

6.6 6.5 6.5 6.4 6.4

.24 .21 .23 .26 .29

5.6 5.6 5.9 5.7 5.8

.083 .091 .182 .105 .125

1.37 1.39 1.42 1.45 1.48

.065 .059 .10 .073

1.49 1.46 1.47 1.52 1.43

0.01 0.01 0.03 0.02 0.02

2.62 2.59 2.68 2.58 2.55

.081 .084 .068 .077 .060

3.62 3.83 3.87 3.67 3.76

.050 .060 .055 .060 .060

19 20 22

1.58 1.49 1.51

.041 .052 .045

2.46 2.44 2.48

— — —

6.5 6.3 6.3

.29 .26 .23

5.7 5.7 5.6

.082 .142 .101

1.43 1.46 1.45

.070 .066 .060

1.53 1.55 1.49

0.02 0.02 0.02

2.62 2.64 2.48

.047 .042 .047

3.76 3.67 3.70

.048 .070 .055

114

* Day egg was laid after experiment started, t t No cystine (free) found.

feces and whole zein was isolated from the stools. The possibility also exists that an amino acid imbalance was obtained in certain cases, especially in ration B-llOa, since this ration gave lower production than B-110. Henderson et al. (1947) and Hankes et al. (1949) have demonstrated a growth inhibition in rats caused by the addition of certain amino acids to a tryptophan deficient ration. An effect similar to this may explain our results on rations B-118 and B-119. The possible toxic effect of the D amino acids may also have

Amino acid composition of the eggs.— The results of the amino acid assays of eggs from some of the rations used in these experiments are given in Table 6. If there were any change in the amino acid makeup of the eggs, one might expect to obtain a gradual reduction of the amount of a certain amino acid in the egg over a period of several days when the hens are going out of production from a deficiency of the same amino acid. Such is not the case. In fact the data presented would indicate that none of the diets used in these experiments had any detectable

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This indicates the existence of bonds resistant to the proteolytic enzymes with the result that some of the amino acids of zein are not available to the organism. Somewhat similar results were also obtained with human infants by Albanese et al. (1949). They found that a diet, with tryptophan and lysine supplemented zein as the protein, failed to give satisfactory nitrogen retention. Some fifty percent of the nitrogen of the diet was found in the

802

INGRAM, CRAVENS, ELVEHJEM, AND HALPIN

SUMMARY

A study has been made on the effect of lysine and tryptophan deficient rations on the laying hens. The basic proteins used in these studies were zein, corn gluten meal and acid hydrolyzed casein. It has been shown that zein is not suitable for a study of the tryptophan and lysine requirements of the laying hen. Even when supplemented with the deficient amino acids, zein does not adequately support egg production. The addition of tryptophan and lysine to a ration containing corn gluten meal as the protein improved egg production but normal production did not result. A ration containing soybean oil meal as the sole source of protein supported egg production for a four week period at a normal rate. This indicates that soybean oil meal protein is adequate for the laying hen. Acid hydrolyzed casein when supplemented with all the essential amino acids at levels contained in a purified control diet failed to support egg production. And further, a combination of five percent casein, five percent gelatin and ten percent acid hydrolyzed casein supplemented with tryptophan and methionine failed to support egg production. An amino acid deficiency causes a hen

to cease production in four to five days indicating that the ability of the hen to draw protein from her body for egg production is very limited. On the rations used in these studies there was no change in the tryptophan, lysine, cystine or methionine composition of the eggs detectable by our method of assay. The hatchability of the eggs was not affected by the amino acid make-up of the rations employed. REFERENCES Albanese, A. A., S. E. Snyderman, M. Lein, E. M. Smetak and B. Vestal, 1949. The biological value of corn and wheat proteins in the male infant, with a note on the utilization of D-tryptophan. J. Nutrition 38:215-224. Almquist, H. J., 1947. Evaluation of amino acid requirements by observations on the chick. J. Nutrition 34: 543-563. Block, R. J. and D. Boiling, 1947. The amino acid composition of proteins and foods. Charles C. Thomas, Springfield, Illinois. Cravens, W. W., and J. G. Halpin, 1946. Studies on the protein nutrition of breeding hens. Poultry Sci. 25:400. Cravens, W. W., 1948. The effect of leucine on egg production and hatchability. Poultry Sci. 27: 562-570. Csonka, F. A., C. A. Denton and S. J. Ringel, 1947. The methionine and cystine content of hens' eggs. J, Biol. Chem. 169: 259-265. Evans, J. E., J. A. Davidson and H. A. Butts, 1950. The amino acid content of fresh and stored shell eggs. III. Methionine, cystine and lysine content of eggs from hens fed diets differing in percentage of these amino acids. Poultry Sci. 29:104-108. Frazier, L. E., R. L. Woolridge, C. H. Steffee and E. P. Benditt, 1949. Minimum daily requirements of indispensible amino acids for maintenance of adult rats. Federation Proc. 8: 355-356. Geiger, E., and E. B. Hagerty, 1949. Growth of rats on hydrolyzed zein. Arch. Biochem. 21: 239—241. Gerber, L., and R. H. Carr, 1930. A chemical and immunological study of egg protein obtained under restricted diets. J. Nutrition 3: 245-256. Grau, C. R., and L. W. Taylor, 1948. Experiments on the amino acid requirements of laying hens. Poultry Sci. 27:665. Hankes, L. V., L. M. Henderson and C. A. Elvehjem, 1949. Effect of cystine and threonine on the

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effect on the protein or the "free" amino acid content of the egg. The fact that the eggs from the deficient rations hatched as well as the eggs from the control ration shows that there is no change in the protein composition of the eggs severe enough to affect embryonic development. These results support the work of McFarlane, Fulmer and Jukes (1930) in contrast to that of Csonka, Denton and Ringel (1947) and Gerber and Carr (1930). This is also in agreement with recent work by Evans, Davidson and Butts (1950).

ASSOCIATION NOTES

acids in nutrition and growth. J. Biol. Chem. 17: 325-349. Robblee, A. R.,.C. A. Nichol, W. W. Cravens, C. A. Elvehjem and J. G. Halpin, 1948. The effect of hen diet and chick diet on the need of chicks for an unidentified growth factor. Poultry Sci. 27: 442-147. Rose, W. C , 1949. Amino acid requirements of man. Federation Proc. 8:¥546-552. "Steele, B. F., H. E. Sauberlich, M. S. Reynolds and C. A. Baumann, 1949. Media for leuconostoc mesenteroides P-60 and leuconostoc citrovorum 8081. J. Biol. Chem. 177:533-544. Sunde, M. L., H. W. Bruins, W. W. Cravens, C. A. Elvehjem and J. G. Halpin, 1950. Pteroylglutamic acid requirement of laying and breeding hens. Poultry Sci. 29: 220-226.

Association Notes ANNUAL MEETING—AUGUST 22-25, 1950 The 39th annual meeting of the Poultry Science Association was held at the University of Wisconsin, Madison, on August 22 to 25, 1950. The total registration of members and guests, including wives and children was 652. Officers elected for the Association year, 1950-51 were: President—W. R. Hinshaw, Biological Department, Camp Derrick, Frederick, Md. First Vice-President—E. M. Funk, Poultry Department, College of Agriculture, University of Missouri, Columbia, Mo. Second Vice-President—H. J. Sloan, Poultry Department, University Farm, University of Minnesota, St. Paul 1, Minn. Secretary-Treasurer—T. B. Avery, Poultry Department, Kansas State* College, Manhattan Kan. Directors—J. R. Cavers, Poultry Department, Ontario Agricultural College, Guelph, Ontario, Can. (1949-51). O. E. Goff, Poultry Department, University of Tennessee, Knoxville, Tenn. (1949-51). J. M. Gwin, Poultry Department, University of Maryland, College Park, Md. (1950-52). W. M. Insko, Jr., Poultry Department, Kentucky

Agricultural Experiment Station, Lexington, Ky. (1950-52). H. M. Scott, Poultry Division, Department of Animal Science, University of Illinois, Urbana, 111. (1949-51). Editorial Board of Poultry Science—H. D. Branion, Department of Animal Nutrition, Ontario Agricultural College, Guelph, Ont., Can. (Editor). F. R. Beaudette, Department of Poultry Husbandry, New Jersey Agricultural Experiment Station, Rutgers University, New Brunswick, N. J. (Pathology and Disease). T. H. Canfield, Poultry Department, University Farm, University of Minnesota, St. Paul 1, Minn. (General). W. W. Cravens, Poultry Department, University of Wisconsin, Madison, Wis. (Nutrition). J. M. Gwin, Poultry Department, University of Maryland, College Park, Md. (Marketing and Extension). P. D. Sturkie, Department of Poultry Husbandry, New Jersey Agricultural Experiment Station, Rutgers University, New Brunswick, N. J. (Physiology and Genetics).

The next annual meeting will be held at the University of Tennessee, Knoxville on August 27-31, 1951.

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growth of rats receiving tryptophan-deficient rations. J. Biol. Chem. 180:1027-1035. Henderson, L. M., T. Deodhar, W. A. Krehl and C. A. Elvehjem, 1947. Factors affecting the growth of rats receiving niacin-tryptophan-deficient diets. J. Biol. Chem. 170: 261-268. Henderson, L. M., and E. E. Snell, 1947. A uniform medium for determination of amino acids with various microorganisms. J. Biol. Chem. 172: 1529. McFarlane, W. D., H. L. Fulmer and T. H. Jukes, 1930. Studies in embryonic mortality in the chick. I. The effect of diet upon the nitrogen, aminonitrogen, tyrosine, tryptophan, cystine, and iron content of the proteins and on the total copper of the hen's egg. Biochem. J. 24: 1611-1631. Osborne, T. B., and L. B. Mendel, 1914. Amino

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