Keratin as a Source of Protein for the Growing Chick

EGG TRANSMISSION OF MYCOPLASMA

1257

M. L. Seely and R. E. Corstvet, 1965b. Isolaity of Mycoplasma meleagridis for turkey emtion of "N" Mycoplasma from different sites of bryos. Am. J. Vet. Res. 27: 326-330. the turkey. Poultry Sci. 44: 732-736. Yamamoto, R., C. H. Bigland and H. B. Ortmayer, 1965a Characteristics of Mycoplasma Yoder, H. W. Jr., 1963. Characterization of avian Mycoplasma. Ph.D. Thesis, Iowa State Univermeleagridis sp.n. isolated from turkeys. J. Bact. sity. 90: 47-49. Yamamoto, R., H. B. Ortmayer, C. H. Bigland,

1. AMINO ACID IMBALANCE AS THE CAUSE FOR INFERIOR PERFORMANCE OF FEATHER MEAL AND THE IMPLICATION OF DISULFIDE BONDING IN RAW FEATHERS AS THE REASON FOR POOR DIGESTIBILITY E. T. MORAN, JR., J. D. SUMMERS AND S. J. SLINGER

Departments of Poultry Science and Nutrition, University of Guelph, Guelph, Ontario, Canada (Received for publication March 15, 1966)

T ? EATHER meal has been shown to be *- of value as a protein source when used to replace limited quantities of various protein feedstuffs in a practical ration (Wilder et al., 1955; Lillie et al., 1956; Naber and Morgan, 1956; Sullivan and Stephenson, 1957; Wisman et al, 1958; McKerns and Rittersporn, 1958; Naber et al., 1961; Poppe, 1965). Because of the good replacement value of feather meal for the chick at high dietary protein levels (20-23%) and poor substituting ability at low dietary protein levels (15%) Sibbald et al. (1962) suggested that feather meal was being used as a source of non-specific nitrogen. Routh (1942) found feather meal to be deficient in tryptophan, methionine, lysine and histidine for the rat; however, even with amino acid supplementation only moderate growth was obtained. Feather meal as the primary source of protein for the chick in practical rations was never as good as comparable corn-soybean meal diets, regardless of amino acid supplementation (Naber et al., 1961). In purified diets feather meal as the sole source of pro-

tein with supplemental amino acids supported only moderate growth and then proved to be inferior to isolated soybean protein with methionine (Summers et al., 1965). The former investigators concluded that not only is feather meal protein imbalanced with respect to the chick's requirement for amino acids but also it is poorly absorbed. Approximately 85 to 90% of the protein from feather meal comes from keratin (Harrap and Woods, 1964). The keratins are classified in the sclero-protein group because of their insolubility in aqueous solvents (Fruton and Simmonds, 1960). Xray diffraction data led Schor and Krimm (1961) to postulate a (3-helix as the structural unit of feather keratin, i.e. an extended chain which coils slowly to form a helix of relatively large pitch. Such helices tend to aggregate by hydrogen bonding to form cylindrical units which in turn associate into cable-like structures. The high cystine content of keratin (8.8% of the protein, Block and Boiling, 1951) is of structural importance by virtue of the disulfide

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Keratin as a Source of Protein for the Growing Chick

1258

E. T. MOEAN, JR., J. D. SUMMERS AND S. J. SLINGER

Because of the apparent dependence of digestibility on the structure of feather keratin, studies were initiated to test the ability of various reducing agents to further "avail" the presumably undigested portions

of feather meal protein. Amino acid supplementation of various treated and untreated samples of feather meal was also carried out in an attempt to improve the quality of this keratin protein. GENERAL EXPERIMENTAL PROCEDURE Single Comb White Leghorn male chicks were raised to one week of age on a standard starting ration. They were then weighed individually and placed in respective groups at 5 g. intervals on either side of the total mean. Random distribution of each group over the experimental areas thus reduced between pen variation. Electrically heated raised-wire floor battery brooders housed the birds and permitted ad libitum feeding and water. After a two week experimental period (3 weeks of age) each pen of chicks was weighed and total feed consumption determined. The data were analyzed following the procedure for a completely randomized design (Federer, 1955). Duncan's multiple range test (Duncan, 1955) allowed treatment comparisons at either the 1 or 5% level of significance (see respective Tables). The latin letters after the observed chick weight indicate statistical significance. Those treatments without a common letter are significantly different from each other while those with a common letter are not. SPECIFIC EXPERIMENTAL PROCEDURE AND RESULTS

Experiment 1 was designed to test the chick growth promoting ability of a commercial feather meal sample treated with various levels and types of disulfide reducing agents. Semi-purified diets (Table 1) were calculated to contain 15% protein. Practical corn-feather meal rations (see footnote 7, Table 3) were calculated to contain 20% protein (feather meal, corn meal and alfalfa meal supplied 12.75, 6.91 and 0.34% of

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bonds which apparently stabilize the aggregation of cylindrical units into cables. Conversion of cystine to cysteic acid led to the elimination of the x-ray reflection thought due to cylinder association (Schor and Krimm, 1961). The long known observation that reagents which oxidize (bromine, peracetic acid, etc.) or reduce (sodium sulfide, thioglycolate, etc.) disulfide bonds and also solubilize keratin, would tend to support the implication of cystine in cable formation. The digestion of untreated feathers by cats and owls was observed to be negligible by Mangold and Dubski (1930). Wool keratin was not susceptible to attack by any digestive enzyme (Greenwood and Speakman, 1964). Ball-milling or fine grinding increases susceptibility to trypsin though not extensively (Routh, 1938). Clothes moth larvae readily utilize keratins as a source of protein. The mid-gut of the larvae was characterized by a low redox potential and alkaline pH (Linderstrom-Lang and Duspiva, 1936). The finding of a very active cystine reductase in the moth larvae would explain the formerly observed low redox potential and implicate cysteine as the reducing agent which concerts keratin to soluble kerateine. (Powning and Irzykiewicz, 1959). Kerateine, also formed by sodium sulfide or thioglycolate, is readily degraded by trypsin, pepsin, papain, or chymotrypsin (Geiger et al,., 1941). Allowing reoxidation of the kerateine as amorphous keratin can be formed which is still susceptible to enzyme digestion (Goddard and Michaelis, 1934). Noteworthy is the observation by Draper (1944) that sodium sulfide treatment of feathers resulted in improved utilization by rats and chicks.

1259

KERATIN AS PROTEIN SOURCE TABLE 1.—Composition of semi-purified ration (15% protein) Ingredient

variable 37.50 4.35 2.00 0.40 0.20 0.20 to 100.00

1 Supplies the following in grams per kilogram of complete ration: Ca3(P04)2, 8.5; KH 2 P0 4 , 12.0; NaCl, 4.0; CaCOa, 19.0. 2 Supplies the following in grams per kilogram of complete ration: CoCl 2 -6H 2 0, 0.02; MnS04, 2.5; FeS04> 0.1; MnSCu-ttO, 0.2; KI, 0.006; CuS0 4 , 0.012; ZnCOs, 0.2; NajMoCM• 2H 2 0, 0.01. 3 Supplies the following per kilogram of complete ration: thiamine• HC1, 20 mg.; riboflavin, 12 mg.; Ca pantothenate, 20 mg.; pyridoxine -HC1, 6 mg.; biotin, 0.3 mg.; menadione, 4 mg.; vitamin Bi2, 0.02 mg.; ascorbic acid, 150 mg.; niacin, 60 mg.; folic acid, 3 mg.; vitamin A, 5,000 I.U.; vitamin D s , 595 I.C.U.; vitamin E, 7.5 I.U.

the protein, respectively). The amino acid composition of the feather meal diet shown in Table 2 is that supplied by a 15% protein semi-purified feather meal ration.1 The necessary amino acids and amount supplemented were determined by the deficit between that supplied by the 15% protein feather meal ration and that termed optimal by Dean and Scott (1965). Methionine, lysine, histidine, tryptophan, and glycine were then calculated first through fifth limiting amino acids respectively. Tyrosine was not supplemented because of the near equivalent surplus of phenylalanine. The limiting amino acids of the 20% protein corn-feather meal rations were similarly determined; and, lysine, methionine and tryptophan were calculated to be first, second and third limiting (Table 3). Supplementation was at the expense of either glucose or corn. Sodium thioglycolate and sodium sulfide were the two reducing agents used to treat "Amino acid analysis was done through the courtesy of Canada Packers Ltd., Toronto, Ontario.

TABLE 2.—Amino acid composition and deficiencies of semi-purified feather meal diets (15% protein) Amino acid Ammo acid Threonine Cystine Valine Methionine Isoleucine Leucine Tyrosine Phenylalanine Lysine Histidine Arginine Tryptophan Glycine 1

F Requirement' *a*f mert^ % dief.

0.65 0.35 0.82 0.45* 0.80 1.20 0.63 0.68 1.10 0.30 1.10 0.225 1.60

0.68 0.50 1.41 0.08 0.81 1.40 0.45 0.85 0.35 0.12 1.14 0.10 1.41

% of

Requirement

,

105 143 172 18 (l) 6 101 117 71 125 31(2) 40(3) 104 44(4) 88(5)

Based on the values of Dean and Scott, 1965. As supplied from a 15% protein ration. The percentage of the requirement of each amino acid as supplied by a 15% feather meal protein diet. 4 The recommended value of 0.55% was lowered by the safety factor to 0.45 because of the apparent plethora of cystine. ' Figures in parentheses refer to the calculated order of limitation. 2

3

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Protein source Corn starch Macro mineral mix 1 Corn oil Micro mineral mix2 Vitamin mix 3 Choline chloride Glucose monohydrate

% of diet

the feather meal. Treatment entailed the addition of 3 liters of distilled water to 1 kg. of feather meal in a 6 liter glass flask. The pH of the mixture was then increased to 11 with a concentrated sodium hydroxide solution. Although the slurry became considerably more viscous constant agitation was still permitted with an electric stirrer. Assuming all the protein of feather meal was keratin, the molecular weight of keratin was 10,400 (Herrap and Woods, 1964), and 8.8% of the protein was cystine (Block and Boiling, 1951), %, \ and one times the molar cystine equivalent of the reducing agents were used to treat the meal; i.e. for every 10,400 g. of feather keratin there were 4 moles of cystine (MW-240) which resulted in the treatment of the keratin with 1, 2 and 4 moles of sodium thiogycolate (MW 114) or 1 mole of sodium sulfide (Na2S.9H20, MW 240). Upon addition of the reducing agent to the slurry a further increase in viscosity was apparent. The mixture was allowed to stand 3

1260

E. T. MOEAN, JR., J. D. SUMMERS AND S. J. SLINGER TABLE 3.—Effect

of disulfide reducing agents and amino acid supplementation on the quality of feather meal protein for the chick (Experiment 1) Treatment 1

Level & source of protein 13% soybean 15% feather

6

None Treated W/O reagent 1 M Thioglycolate 2 M Thioglycolate

1 M Sodium sulfide 20% Corn-feather7

None 4 M Thioglycolate 1 M Sodium sulfide

3 week wt., g.3

G/F*

+

141 e» 75 ab 161 g 73 ab 155 fg 76 ab 158 g 76 ab 118 d 70 a 80 b 75 ab 148 ef 198 h 98 c 191 h

0.34

—

+ + — + + + — + + + +

—

0.43

—

0.40

—

0.41

—

0.29

— — —

0.40 0.49 0.14 0.47

1

See text for explanation. Amino acid supplementation: isolated soybean protein, 0.2% DL-methionine, 0.9% glycine; feather meal, 0.4% DL-methionine, 0.8% L-lysine • HCl, 0.18% L-histidine • HCl • H 2 0, 0.12% L-tryptophan, 0.2% glycine; corn-feather meal, 0.1% DL-methionine, 0.6% L-lysine• HCl, 0.05% L-tryptophan. 3 Average starting weight at 1 week of age was 80 g. The 3 week wt. represents an average of 3 replicate pens of 10 chicks per pen. 4 Gain/feed consumed. 5 The isolated soybean diets contained only 13% protein due to an error in calculation. 6 Latin letters refer to Duncan's multiple range test at the 5% level of significance (see text). ' The percentage composition of the 20% corn-feather meal ration was: Feather meal, 15.00; alfalfa meal (17%), 2.00; ground corn, 78.55; dicalcium phosphate, 2.30; limestone, 1.25; iodized salt, 0.3; vitamin mix (see footnote 3, Table 1), 0.2; micro mineral mix (see footnote 2, Table 1), 0.40. 2

hours after which the pH was lowered to 6 with concentrated hydrochloric acid. This treatment of the feather meal slurry decreased the viscosity markedly with the result that it tended to flocculate out. Cheese cloth permitted an easy separation of meal and solution. The filtered meal was then washed with a liter of distilled water, refiltered, and dried in a forced-draft oven at 75°. The dried product was incorporated into the respective rations. The data from Experiment 1 are illustrated on Table 3. All semi-purified feather meal rations without amino acid supplementation gave weights which were not significantly different from each other. The untreated feather meal, the meal treated but without reducing agent, and that meal reacted with \ the molar quantity of cystine with thioglycolate, all promoted significantly better chick growth when supplemented with amino acids than was ob-

served with the 13% protein isolated soybean ration. Higher treatment levels of thioglycolate and the only level of sodium sulfide would seem to have been toxic based on the respectively poorer chick growth. The 20% protein corn-feather meal practical rations in which the feather meal was untreated or sodium sulfide treated promoted weight gain considerably superior to that observed in any of the semi-purified diets. A significant depression in growth was obtained with the meal reacted with the highest level of thioglycolate. The observation that the sodium sulfide treated meal depressed growth in the semi-purified but not in the practical ration may be explained in part by the fact that the 20% protein ration contained less absolute quantities (3%) of the treated feather meal. This 3 % differential is significant if one considers that the depression is marginal. The difference in other dietary in-

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4 M Thioglycolate

Amino acid2

1261

KERATIN AS PROTEIN SOURCE

gredients between semi-purified and practical rations, e.g., 78% corn, could also be a factor in the expression of sodium sulfide toxicity.

The chick growth data illustrated in Table 5 agree with the calculated order of amino acid limitation when feather meal is the sole source of protein. Methionine, ly-

for Experiment 21 Corn-soybean

ingredient Soybean meal (50% protein) Alfalfa meal (17% protein) Corn meal

Corn-soybean-

~W0^ protein

20% protein

^ - ^ 20% protein

17.82

28.00

17.82

2.00

2.00

2.00

65.35

64.35

65.35 5.88

Feather meal (85% protein) Glucose monohydrate

—

—

7.93

—

3.91

Cellulose

2.25

—

0.39

Constant*

4.65

4.65

4.65

1

All diets were calculated to be isocaloric and contain 2,933 Kcal. of metabolizable energy per kg. of diet. 2 All diets contained the following percentage of ingredients: dicalcium phosphate, 2.30; limestone, 1.25; choline chloride (25%), 0.20; iodized salt, 0.30, vitamin mix (see footnote 3, Table 1), 0.20; micro mineral mix (see footnote 2, Table 1), 0.40.

sine, histidine, and tryptophan would appear to be the first through fourth limiting amino acids, respectively; however, the chick did not apparently require additional glycine. These data verify the results of Experiment 1; feather meal with appropriate amino acid supplementation can support chick growth and feed efficiencies at least equivalent to that of soybean protein with methionine. The results when feather meal was used in practical rations further support the contention of its high protein value with appropriate amino acid balance. There were no significant differences between chicks fed the 20% protein corn-soybean meal diet with or without methionine and glycine supplementation. Removing 5% of the protein (all from soybean meal) resulted in a significant depression in chick growth. Returning the 5% protein in the form of feather meal increased chick growth to that observed with the 20% corn-soybean meal diets; and, amino acid supplementation proved of no value. However, removal of all the soybean protein and its replacement with feather meal and corn to 20% of protein led to a growth depression significantly greater than observed with dropping the protein level to 15%. Addition of methio-

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Experiment 2 was conducted in order to: (1) verify the results with amino acid supplementation obtained in Experiment 1; (2) to determine whether the calculated order of amino acid limitation in feather meal is correct; and (3) to test the effectiveness of feather meal in practical rations as a soybean meal protein replacement with and without amino acid supplementation. The semi-purified feather meal ration was the same as that used in Experiment 1 (Table 1) with the exception that the rations were maintained isonitrogenous by appropriate manipulation of diammonium citrate at the expense of amino acids and/ or glucose. The practical corn-soybean and corn-soybean-feather meal rations were maintained isocaloric (2933 Kcal. M.E./kg.) by appropriate glucose or cellulose manipulation (Table 4). A constant amount of protein was supplied by corn and alfalfa meal (6.09%); thus, protein level (15 or 20%) was manipulated by soybean (50%) or feather meal (85%). The 20% protein corn-feather meal diet (see footnote 7, Table 3) was little different from the soybean meal containing rations (7.25% protein from corn and alfalfa, 12.75% protein from feather meal; 3001 Kcal. M.E./kg.). Necessary amino acid supplementation was calculated in the same manner for the practical rations as demonstrated for the semi-purified ration (Table 2) with the exception that "book values" of amino acids were used for ingredients other than feather meal (e.g. corn, alfalfa and soybean meal).

TABLE 4.—Composition of practical rations

1262

E. T. MORAN, JR., J. D. SUMMERS AND S. J. SLINGER TABLE 5.—Ejfecl of amino acid supplementation on the growth of chicks fed feather meal containing semi-purified and practical rations (Experiment 2) Supplemented amino acids1

*t W7f*fAr"

•J VV CClV

ijcvci bourse ui protein 15% soybean 15% feather

Methionine

Lysine

Histidine

Tryptophan

0.9

0.1 0.8 0.8 0.8 0.8

0.18 0.18 0.18

0.12 0.12

0.2

20% corn-soybean 0.8

0.2 15% corn-soybean 20% corn-soybean-feather

0.15 0.15 0.15

0.2

0.20

0.6

0.4 0.4

20% corn-feather 0.05

wt., g. 160 e4 67 a 73 a 83 b 95 c 166 e 168 e 207 g 214 g 187 f 215 g 208 g 215 g 216 g 112 d 207 g

G/F 3 0.43 0.03 0.13 0.21 0.44 0.45 0.48 0.50 0.42 0.49 0.48 0.51 0.51 0.32 0.50

1 The forms of the supplementary amino acids were: DL-methionine, L-lysine• HC1, L-histidine • HC1 • H 2 0, L-tryptophan. 2 Average starting weight at one week of age was 71 g. The 3 week wt. represents an average of 3 replicate pens of 10 chicks per pen. 3 Gain/Feed consumed. 4 Latin letters refer to Duncan's multiple range test at the 1% level of significance (see text).

nine, lysine and tryptophan completely overcame the depression and resulted in growth equivalent to that observed with the 20% corn-soybean and corn-soybeanfeather meal diets. Experiment 3 was designed to determine: (1) whether commercial feather meals very in quality; (2) if raw feathers are a source of available protein; and (3) whether autoclaving or reducing agent treatment affect the quality of the resultant meal. The same purified diets were employed as described on Table 1. The rations were not maintained isonitrogenous and amino acid supplementation was at the expense of glucose. Whole raw feathers were obtained from a feather meal plant, frozen, and lyophilized. The raw, untreated feather meal constituted only ground freeze-dried feathers. Autoclaving entailed the use of a standard laboratory steam-operated sterilizer. Raw ground lyophilized feathers were placed on aluminum foil and autoclaved for 30

minutes and 18 hours at 121°C. The raw meal autoclaved for 30 minutes was not physically different from the starting product, i.e. they both required considerable volume per unit weight. Autoclaving for 18 hours resulted in an obviously different product. The odor of sulfur was quite predominant upon reduction of autoclave pressure and the meal was darker in color, gelatinous while still hot and wet, and formed a desirable free flowing meal upon drying. All meals were lyophilized again prior to their incorporation into the ration to insure a low moisture feed. Sodium sulfide was the only reducing agent used to react with the raw feather meal. The reaction procedure was different than described in Experiment 1 because of the problem of very low ground feather density. One kg. of the raw meal was mixed with approximately 3 liters of 0.1 N sodium hydroxide. After a slurry was formed, the pH was adjusted to 11, the meal was allowed to soak for 12 hrs., and then

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0.4 0.4 0.4 0.4 0.4

Glycine

1263

KERATIN AS PROTEIN SOURCE

TABLE 6.—Effect of amino acid supplementation, autoclaving, and sodium sulfide treatment of raw feathers on the quality of the meal fed to chicks {Experiment 3) Treatment Isolated soybean protein Raw feather meal Feathers, 30 min., 121° Feathers, 18 hours, 121° Feathers, sodium sulfide Commercial feather meal A H C

Amino1 acids

3 week2 wt., g.

' + + + + + + + +

164 C 58 a 53 a 62 a 58 a 60 a 125 b 51 a 55 a 154 c 157 c 156 c

1 Supplementing amino acids see footnote 2, Table 3. 2 Average starting weight at 1 week of age was 70 g. The 3 week wt. represents an average of 3 replicate pens of 10 chicks per3 pen. Gain/Feed consumed. 4 See footnote 6, Table 3.

studies indicate that commercial feather meal, appropriately supplemented, is an excellent source of protein and a desirable free-flowing product. The only difference between the raw and processed meals is the heating process. Extensive autoclaving of raw feathers (18 hours, 121°) in Experiment 2 led to an obvious elimination of sulfur from the meal. Commercial processing of feathers had been shown to result in extensive cystine degradation (8.8 to 3.6% of protein) while there was little effect on other amino acids (Gregory et al., 1956). The most probable reason for cystine destruction is disulfide bond cleavage by heat. Wool keratin has been observed to become considerably more soluble when heated (205°) almost solely because of crosslink cleavage with probable formation of kerateine and loss of sulfur (Menefee and Yee, 1965; Menfee, 1965). The role of cystine in keratin structure and the concomitant increase in nutritional value of feathers with disulfide destruction would strongly implicate this sulfur amino acid in digestibility.

DISCUSSION The observation that raw feathers are Protein disulfide bonds can be cleaved at poorly digested and utilized as a source of an alkaline pH by treating the protein with protein is well substantiated (Naber et al., excess reagent disulfide in the presence of 1961; Mangold and Dubski, 1930). Present catalytic thiol (Smithies, 1965). The high

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reacted mole for mole with sodium sulfide (refer to the procedure of Experiment 1). After 2 hours the pH was lowered to 6, the meal filtered off with cheese cloth, washed with distilled water, refiltered. and then freeze-dried. The lyophilized meal was incorporated directly into the ration. The chick growth data from Experiment 3 was illustrated on Table 6. Raw feathers with or without amino acid supplementation failed to support any growth. Autoclaving the raw feathers for 30 minutes also failed to support any growth, regardless of amino acid supplementation; however, autoclaving for 18 hours and amino acid supplementation resulted in a meal which supported reasonably good growth. Without amino acid supplementation the 18 hour autoclaved meal was no better than any other unsupplemented meal. Sodium sulfide treatment of the feathers was without effect in improving protein quality since the respective meals also did not produce a weight response. All there commercial samples of feather meal proved to be not significantly different from one another and were at least equal to soybean protein in their ability to promote growth when appropriately supplemented. A large proportion of the ground feathers in the raw, 30 minute autoclave, and sodium sulfide treatments would seem to have been poorly digested. From gross observation it appeared that most, if not all, of the meal in the aforementioned treatments passed through the chick without any apparent physical or chemical change; however, feces of chicks offered the 18 hour autoclaved meal indicated almost complete digestion and absorption.

1264

E. T. MORAN, JR., J. D. SUMMERS AND S. J. SLINGER

commercial feather meal as a source of protein, substantial growth responses were observed; but, the responses were termed inadequate because of their inability to approach that obtained by soybean meal. The semi-purified rations of Naber et al. (1961) used sucrose (3675 Kcal. metabolizable energy/kg.) as the carbohydrate source and protein supplement was added at the expense of sucrose to the desired level of protein (20%); consequently, when a 50% protein soybean meal (2530 Kcal. metabolizable energy/kg.) or an 85% protein feather meal (2222 Kcal. metabolizable energy/kg.) are incorporated there results two types of rations which differ by virtue of their protein to energy relationship and cannot be effectively compared. Summers et al. (1965) were getting only moderate growth responses with amino acid supplementation due to the fact that the level of added tryptophan was insufficient. Because feather meal is severely inadequate in several amino acids critical to animal nutrition its use as a replacement protein could accentuate existing and/or induce other amino acid deficiencies. The replacement of 5% soybean protein in a 20% corn-soybean ration with feather meal protein gave equally good results.2 Substituting all of the soybean with feather meal and corn in a 20% protein diet resulted in a severe growth depression which was completely overcome by supplementary methionine, lysine and tryptophan.3 The results would appear to explain why the inclusion of excess feather meal protein without com2

Upon substitution of 5% soybean protein with 5% feather meal protein calculated methionine was reduced 0.38% to 0.32% and lysine 1.17 to 0.94%. 2 Upon substituting 14% soybean protein with feather meal and corn calculated methionine was reduced 0.38 to 0.24, lysine 1.17 to 0.55, and tryptophan 0.28 to 0.18%.

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pH and large quantities of disulfide reducing agent used in Experiment 3 were expected to reduce at least some feather cystine disulfide bonds and avail a part of the protein for digestion; however chick growth trials failed to bear this out. Draper (1944) did get favorable results with sodium sulfide treated feathers. The extent to which feathers should be autoclaved for maximal quality has already been answered. Binkley and Vasak (1950) suggest 142°-1S3°C. (40-60 p.s.i.) for 30 to 60 minutes. One of the commercial meals used in the present studies (Meal B, Experiment 3) in known to have been processed at 142° for 30 minutes. The resultant product was a desirable free flowing meal which supported growth equivalent to that of soybean protein with appropriate amino acid supplementation. Raw feathers autoclaved at 121° for 18 hours were not optimally processed as evidenced by the inability to promote weight gain equal to that the commercial meal. Heating time required to obtain a satisfactory product is known to increase rapidly as the temperature is lowered below 142° (Binkley and Vasak, 1950). Feather protein is unique in that it has four amino acids (methionine, lysine, histidine and tryptophan) which are extremely deficient for the chick while the remaining essential amino acids are adequately supplied and balanced (based on 15% protein) . Consequently, the benefits of supplementation with the first three limiting amino acids are not fully evidenced until the fourth has been adequately supplied (Table 3). Naber et al. (1961) were not, in many cases, obtaining substantial growth responses to amino acid supplements because the histidine level then considered optimal was half the requirement as known today (0.15 vs. 0.30). When appropriate amino acid supplementation was attained in semi-purified rations using the

KERATIN AS PROTEIN SOURCE

pensatory amino acid supplementation would result in growth depressions. SUMMARY

REFERENCES Binkley, C. H., and O. R. Vasak, 1950. Production of a friable meal from feathers. U. S. Dept. Agric, Agric. Res. Service Bull. No. AIC-274. Block, R. J., and D. Boiling, 1951. The Amino acid composition of proteins and foods. Charles C Thomas, Springfield, Illinois.

Dean, W. F., and H. M. Scott, 1965. The development of an amino acid reference diet for the early growth of chicks. Poultry Sci. 44: SOSSOS. Draper, C. I., 1944. The nutritive value of corn oil meal and feather protein. Iowa Agric. Expt. Station Res. Bull. 326. Duncan, D. B., 1955. Multiple range and multiple F tests. Biometrics, 1 1 : 1-42. Federer, W. T., 1955. Experimental Design. The MacMillan Co., New York. Fruton, J. S., and S. Simmonds, 1960. General Biochemistry. John Wiley and Sons Inc., New York. Geiger, W. B., W. I. Paterson, L. R. Mizelt and M. Harris, 1941. Nature of the resistance of wool to digestion by enzyme. J. Res. National Bureau Standards, 27 : 459^68. Goddard, D. R., and
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Chicks were fed semi-purified and practical rations from one to three weeks of age with soybean and/or feather meal as the primary protein sources. Commercial feather meal as the sole source of protein (15%) supplemented with 4 amino acids proved equal to isolated soybean protein. The order of amino acid limitation was methionine, lysine, histidine and tryptophan, respectively. Ground, raw feathers failed to support growth regardless of amino acid supplementation, as did feathers autoclaved for 30 minutes at 121°, and those treated with sodium sulfide. Feathers autoclaved for 18 hours at 121° and supplemented with amino acids supported moderate chick growth. Feather keratin availability was discussed in terms of the disulfide bond of cystine. Heating apparently results in dissulfide bond cleavage allowing the protein to be utilized. Substitution of 5% protein in a practical 20% protein corn-soybean ration with feather meal protein resulted in equally good chick growth. Replacement of all the soybean protein with corn and feather meal severely depressed three week chick weight; supplementation with methionine, lysine and tryptophan completely overcame the depression. Feather meal is believed to aggravate existing and/or induce deficiencies of methionine, lysine and tryptophan when extensively used as a replacement protein.

1265

1266

E. T. MORAN, JR., J. D. SUMMERS AND S. J. SLINGER J. 1: 489-515. Sibbald, I. R., S. J. Slinger and W. F. Pepper, 1962. The utilization of hydrolyzed feather meal by growing chicks. Poultry Sci. 4 1 : 844-849. Smithies, 0., 1965. Disulfide-bond cleavage and formation in proteins. Science, 150: 1595-1598. Sullivan, T. W., and E. L. Stephenson, 1957. Effect of processing methods on the utilization of hydrolyzed poultry feathers by growing chicks. Poultry Sci. 36:361-365. Summers, J. D., S. J. Slinger and G. C. Ashton, 1965. Evaluation of meat meal and feather meal for the growing chicken. Can. J. An. Sci. 45: 63-70. Wilder, O. H. M., P. C. Ostby and B. R. Gregory, 1955. The use of chicken feather meal in feeds. Poultry Sci. 34: 518-524. Wisman, E. L., C. E. Holmes and R. W. Engel, 1958. Utilization of poultry by-products in poultry rations. Poultry Sci. 37: 834-838.

Effect of Dietary Protein in Holding Rations for Adult Male Chickens1 G. H. ARSCOTT AND J. E. PARKER Department of Poultry Science, Oregon State University, Corvallis, Oregon 97331 (Received for publication March 15, 1966)

A

RSCOTT AND PARKER (1963) have reported that adult White Leghorn males fed diets containing 16.9, 10.7 and 6.9 percent of protein over a 3 3-week period showed no adverse effects on semen volume, fertilizing capacity of semen or hatchability of fertile eggs. Since these results showed that protein levels considerably below those ordinarily fed to breeder flocks had no adverse effects and since the males in this investigation were kept in individual cages, an experiment was initiated to study the effect on reproductive performance of maintaining males together in floor pens on a low-protein diet. "Technical paper No. 2114, Oregon Agricultural Experiment Station. Supported in part by a grantin-aid from the American Poultry and Hatchery Federation.

PROCEDURE

Thirty adult White Leghorn males hatched April 22, 1964 were placed in two pens of a colony house on March 26, 1965. Prior to this the two groups of 15 males were flock-mated to two pens of about 240 similarly managed White Leghorn layers. Initial hatchability and fertility data involving 300 eggs were obtained during this period. Each group of males was kept as a unit after transfer. Each male in the colony house was provided with .405 sq. m. of floor space and .228 sq. m. of porch space for a total of .633 sq. m. of space per bird. The experiment started April 15, 1965 and continued for 22 weeks. The rations used are shown in Table 1. Corn, milo, barley and oats made up the ground grain component of both rations.

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