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The Availability of Niacin in Corn, Soybean Meal and Wheat Middlings for the Hen1
The Availability of Niacin in Corn, Soybean Meal and Wheat Middlings for the Hen 1 A. G. MANOUKAS,2 R. C. RINGROSE AND A. E. TEERI Department of Animal Sciences and Biochemistry, University of New Hampshire, Durham, New Hampshire 03824 (Received for publication March 30, 1968)
ODICEK (1940a) suggested that a by Ringrose et al. (1965), with various large amount of niacin present in cereal levels of crystalline niacin, was fed to and other grains is in a bound form. Later White Leghorn hens for at least ten weeks it was reported by Kodicek (1940b), Krehl and several variables were studied. It was and Strong (1944), Krehl et al. (1944), found that hatchability of eggs was a recHeuser and Scott (1953), Goldsmith et al. ognizable linear function of niacin intake (1956), Kodicek et al. (1959) and Chau- over a wide range of intake; thus it was dhuri and Kodicek (1960) that the bound chosen as the independent variable to be form of niacin is unavailable or partly estimated in the assay. The regression available to the dog, microorganisms, chick, equation of hatchability (Y) on the niacin duckling, man, pig and rat, respectively. In intake (X) was Y = 3.74 + 90.9 X. The relation to the hen, the common quantities available niacin intake, in mg. per hen per of feedstuffs used in poultry rations should day, was given by the equation, X = meet the need of the hen for niacin based 0.011Y —0.041 (1). In this equation, Y upon minimum requirement studies as re- represents the mean hatchability, in perported by Ringrose et al. (1965). In poul- cent, for a period of ten weeks during try feeds, crystalline niacin is added to which eggs were incubated weekly. Due to meet the requirements for niacin because of the data used to derive equation (1), lack of information concerning the avail- hatchability values smaller than 13 and ability of feedstuffs niacin for the hen. The larger than 86 percent can not be used for present study was undertaken to determine determining the available niacin intake. Assays for available niacin in yellow corn, the availability of the niacin present in yellow corn, dehulled soybean meal and wheat dehulled soybean meal (50 percent protein) and wheat middlings were made. middlings for the hen. White Leghorn hens were used in all experEXPERIMENTAL PROCEDURE iments. A description of the niacin deficient The development of a quantitative bio- diet, experimental design and housing of assay procedure for available niacin was the hens has been reported by Ringrose et described by Manoukas (1967). In that al. (1965). The niacin deficient basal diet study, a niacin deficient basal diet reported was adapted to make the assay diets. An effort was made to maintain the tryptophan 1 Published with the approval of the Director content of all diets as close as possible to of the New Hampshire Agricultural Experiment the minimum requirement and to adjust Station as Scientific Contribution No. 434. Acthe rest of the dietary nutrients to approxicepted as part of a doctoral dissertation by the mately the same levels found in the niacin University of New Hampshire. deficient basal diet. 2 Present address: Department of Poultry Science, Cornell University, Ithaca, New York.
The niacin content of the basal diet and 1836
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K
AVAILABILITY or
meal. The composition of this diet is presented in Table 1 under the designation SN0. The soybean meal in this diet supplied 7.95 mg. of niacin per kg. of feed. The calculated tryptophan content of this diet was 0.157 percent. Crystalline niacin was added to the SN0 diet to provide assay diets containing 3.0, 5.0, 7.0 and 9.0 mg. of crystalline niacin per kg. of feed. These diets, designated SN3, SN5, SN7 and SN9, respectively, were fed to the hens for a period of seven weeks. By the seventh week, at which time four hatches of eggs were completed, it was apparent that most of the soybean niacin was available to the hen. Because of this observation the groups of hens fed diets SN5 and SN7 during the first seven weeks were fed diets SN0 and SNi.s, respectively, for the following five weeks of the experimental period. This change was made to determine the magnitude of the niacin availability in soybean meal without or with very little crystalline niacin supplementation. The groups of hens which were fed diets SN3 and SN9 were kept on the same diets throughout the experiment. In Experiment 3, the basal diet was formulated to contain 7 percent wheat mid-
TABLE 1.—Composition of assay diets Expt. 2
Expt. 3
Zl6.25
SN 0
TN»
%
% —
% — —
Experiment 1 Diet Zs.75
Corn meal Soybean meal Wheat middlings Corn starch Crude casein Gelatin D,l.-Methionine Limestone Di-Ca-phosphate Iodized salt Mineral mix* Vitamin mix* Corn oil Cellulose * Ringrose et al. (1965)
%
Zn.25
%
Zl3.75
%
35.00
45.00
55.00
65.00
— —
— —
— —
— —
30.69 9.54 7.14 0.22 7.65 3.50 0.50 1.60 0.56 1.60 2.00
21.99 9.02 6.76 0.22 7.65 3.50 0.50 1.60 0.56 1.20 2.00
13.29 8.51 6.38 0.21 7.65 3.50 0.50 1.60 0.56 0.80 2.00
4.59 8.00 6.00 0.20 7.65 3.50 0.50 1.60 0.56 0.40 2.00
25.00
—
48.49
—
7.00 0.20 7.65 3.50 0.50 1.60 0.56 4.00 1.50
7.00 54.99 10.00 7.50 0.20 7.65 3.50 0.50 1.60 0.56 5.00 1.50
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studied feedstuffs was determined by the microbiological assay of Barton-Wright (1944). The tryptophan content of the basal diet and corn was determined by the method of Green and Black (1944). Sixty hens were allotted to four treatments, fifteen hens per treatment in each of three separate experiments. In order to obtain hatchability data, the hens were artificially inseminated twice weekly. The eggs were held in a 15°C. egg room and incubated weekly. Individual egg production was recorded daily. Treatment-group feed consumption was recorded at the end of each two-week period. Individual body weight was recorded on the first and last day of each experiment. In Experiment 1, the basal diet was adapted to contain 35, 45, 55 and 65 percent corn. The composition of these diets is presented in Table 1 under the designations Zs.75, Zn.25) Zi3,75 and Zie.25, respectively. Subscripts indicate the niacin content derived from corn, in mg. per kg., for each diet. The tryptophan content of these diets was 0.155 percent. The experiment lasted ten weeks. In Experiment 2, the basal diet was adapted to contain 25 percent soybean
1837
NIACIN
1838
A. G. MANOUKAS, R. C. RINGROSE AND A. E. TEERI
TABLE 2.—Daily feed consumption, egg pro?"-ction, daily feedstuff niacin intake (FNI), daily crystalline niacin intake (CNI), hatchability and feedstuff niacin availability for the hen. (Means of 15 hens per treatment) Diet
Feed consumption g./hen
Zll.25 Zl3.76 Zl6.25
88 94 94 103
SN„*
96
SN,.6*
94
SN3.0
96
SN,.o
92
TN a
85
TN,. 5
84
TN3.„
97
TN4.5
100
^8.75
Egg, production
(1) FNI (2) CNI
mg./hen Yellow corn, Expt. 1 60.2 (1) 0.770 64.5 (1) 1.058 68.6 (1) 1.293 (1) 1.674 79.2 Soybean meal, Expt. 2 57.1 (1) 0.785 (2) 57.9 (1) 0.747 (2) 0.141 54.1 (1) 0.763 (2) 0.288 56.5 (1) 0.731 (2) 0.828 Wheat middlings, Expt. 3 56.0 (1) 0.940 (2) 60.5 (1) 0.929 (2) 0.128 (1) 1.073 71.9 (2) 0.291 76.1 (1) 1.106 (2) 0.450
%
* For the last five weeks of the Experiment, see text. ** This figure was obtained by direst comparison.
Hatchability
Niacin availability
%
%
23.2 31.3 37.6 58.6
28 29 29 36
93.6
100**
96.8
—
91.9
—
93.7
—
32.7
34
51.0
42
67.3
38
83.7
39
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dlings. The composition of this diet is were made at the 0.05 level of probability. presented in Table 1 under the designa- The linear equation (1) was used to estition TN 0 . The middlings in this diet sup- mate the available niacin present in corn, plied 11.06 mg. of niacin per kg. of feed. soybean meal and wheat middlings. Diet TN 0 constituted one treatment. NiaEXPERIMENTAL RESULTS cin was then added to diet TN 0 to provide AND DISCUSSION three additional diets containing 1.5, 3.0 and 4.5 mg. of crystalline niacin per kg. of feed. In Experiment 1, diets Zs.7f„ Z u . 2 5, Z13.75 These diets were designated TNi. 5 , TN:t.0 and Z16.25 were used which contained 35, and TN4.5, respectively. The calculated 45, 55 and 65 percent corn, respectively. tryptophan content of these diets was A summary of mean daily feed consump0.148 percent. tion, egg production, daily corn niacin inFor brevity and convenience in reporting take, hatchability and estimated niacin and discussing the data, the symbols used availability in corn is presented in Table 2. to designate the diets are also used to rep- Groups Z8.75, Zu.25 and Z13.7r, lost 128, 106 resent the respective groups of hens to and 68 grams of body weight per hen, rewhich the diets were fed. spectively, during the experiment. Group The analysis of variance (completely Z16.25 gained 9 grams of body weight per randomized design) as outlined by Snedecor hen during the experiment. (1956) and the multiple range test of DunStatistical analysis of the data showed can (1955) were used to evaluate the ex- that hatchability of eggs obtained from perimental data. All tests of significance group Z8.75 differed from that obtained
AVAILABILITY OF NIACIN
under their experimental conditions. Consideration of all results obtained with corn diets shows that the niacin in corn is approximately 30 percent available to the hen. The low availability of corn niacin for the hen is in agreement with experimental results reported by Kodicek (1940b), Krehl and Strong (1944), Goldsmith et al. (1956), Kodicek et al. (1959) and Chaudhuri and Kodicek (1960). Ghosh et al. (1963) reported that 90 percent of corn (Zea mays) niacin is unavailable to the microorganism Leuconostoc mesenteroides. Luce (1965) reported that corn niacin is largely unavailable to swine. In Experiment 2, dehulled soybean meal diets SNS and SN9 were tested during a twelve-week experimental period and diets SN5 and SN7 during the first seven weeks of the experimental period. Diets SN0 and SN1.5 substituted for diets SN5 and SN7, respectively, for the following five weeks of the experiment. A summary of mean daily feed consumption, egg production, daily niacin intake, hatchability and estimated niacin availability in soybean meal is presented in Table 2. Groups SN3, SNS, (SNo), SN r , (SNLS) and SN9 gained 176, 188, 183 and 145 grams of body weight, respectively, during the twelve-week experimental period. Statistical analysis of the data showed that no one group differed from another in any of the reported functions. Hatchability and feed consumption were not affected by the dietary niacin intake and all groups gained body weight. Egg production was equivalent but lower than expected for all groups. The low egg production may be due partly to the low egg production of the experimental stock at the beginning of the experiment and partly to the relatively high temperature of the summer months. On the basis of the data obtained, it is apparent that all of the niacin found in
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from groups Z13.7r, and Z16.25. Also hatchability from groups Z n . 2 5 and Z13.75 differed from that of group Z16.,5. Egg production and feed consumption of groups Z8.75, Zn.25 and Zi3.75 differed from that of group Z16.25. No other differences were found among the groups. Hatchability data from all groups clearly demonstrates that corn niacin is partly available to the hen. Application of the mean hatchability percentage obtained for groups Zs.75, Zn.25, Z13.75 and Z16.25 to equation (1), gave availability of niacin equal to 28, 29, 29 and 36 percent, respectively. The greater availability of corn niacin in diet Z16.25 may be attributed to variation and/or to greater tryptophan intake; the additional tryptophan being converted to niacin. The minimum requirement of the hen for tryptophan is approximately 135 mg. per day as reported by Johnson and Fisher (1958). The tryptophan intake of groups Z8.75 and Z16.25 was 136 mg. and 161 mg. per hen per day, respectively, a difference of 25 mg. Application of the hatchability figure of 58.6 percent for diet Zi6.25 to equation (1) gave an available niacin intake of 0.604 mg. per hen per day. Applying the estimate of 28 percent available niacin, obtained for diet Z8.75, to the daily niacin intake of 1.674 mg. per hen fed diet Z16.25, one derives an available niacin intake of 0.470 mg. If the greater hatchability and niacin availability percentage for corn in diet Zi6.25 relative to diet Zs.75 was due to more niacin, then 0.134 mg. (0.604-0.470) of niacin per hen per day must have come from the 25 mg. of additional tryptophan available per hen in group Z ie . 25 . It takes, therefore, 187 mg. of tryptophan to supply 1 mg. of niacin. This conversion ratio is not in agreement with results reported by Briggs (1945), West (1952) and Petterson (1956) who state that it takes 55 to 60 mg. of tryptophan to replace 1 mg. of niacin for chicks,
1839
1840
A. G. MANOUKAS, R. C. RINGEOSE AND A. E. TEERI
consumption, egg production, niacin intake, hatchability and estimated niacin availability in wheat middlings is presented in Table 2. Groups TN 0 , T N ^ , TN3.0 and N4.5 gained 16, 47, 45 and 62 grams of body weight, respectively, during the experiment. Statistical analysis of the data showed that diets TN 0 and TN li5 significantly depressed feed consumption, hatchability and egg production as compared to diets TN3.0 and TN4..5. Hatchability results of all groups differed significantly from each other. No other differences were found among the groups. Hatchability data from all groups clearly shows that the niacin in wheat middlings is partly available to the hen. Application of the mean hatchability percentage obtained from groups TN 0 , TN1.5, TN3.0, and TN4.5 to equation (1) gave availability of niacin equal to 34, 42, 38 and 39 percent, respectively. With a conversion ratio of 187 mg. of tryptophan to 1 mg. of niacin, derived from Experiment 1, it can be calculated that the availability of niacin in wheat middlings for groups TN3.0 and TN4.S is 34 and 33 percent, respectively, because of greater tryptophan intake comNo data could be found on the availabil- pared to groups TN 0 and TNi.5, The ity of soybean niacin for other species. niacin availability derived for group TN1.5 Ghosh et al. (1963) reported that peas cannot be explained on the basis of trypto(Pisum sativum) and other pulses do not phan conversion to niacin. contain a bound form of nicotinic acid. But Taking into consideration the conversion groundnut {AracMs hypogea) and other oil of tryptophan to niacin, it was concluded seeds contain about 40 percent of the total that the availability of niacin in wheat midniacin content in a bound form which is dlings was 36 percent for the hen. No inunavailable to Leuconostoc mesenteroides. formation could be found on the availabilIt should be noted that the soybean meal ity of wheat middlings niacin for other speused in this experiment was a heat treated cies with the exception of ducklings. commercial product for direct use in poul- Heuser and Scott (1963) suggested that try feeds. Heat treatment may affect niacin the niacin present in wheat middlings apavailability. peared to be relatively unavailable to the Diets TN„, TNj.s, TN3.0 and N4.5 con- duckling. Luce (1965) indicated that the taining wheat middlings were tested in in niacin present in wheat was only partially Experiment 3. A summary of mean daily feed available to swine.
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soybean meal was available to the hen. The average soybean niacin intake of group SNo was 0.78S mg. per hen per day and the average hatchability recorded was 93.6 percent for the five weeks. Combined data reported by Manoukas (1963) and Ringrose et al. (1965) shows that 0.830 mg. of niacin intake per hen per day from a purified diet supported hatchability equal to 83.4 percent during a period of five weeks. This information indicates that 0.785 mg. of niacin could not support 93.6 percent hatchability for a period of five weeks. Thus it appears that the required niacin was obtained from the conversion of tryptophan to niacin. Group SN 0 consumed 155 mg. of tryptophan per hen per day of which 20 mg. may be converted to niacin. With a conversion ratio of 187 mg. of tryptophan to 1 mg. of niacin, derived from Experiment 1, it can be calculated that 20 mg. of tryptophan can supply 0.107 mg. of niacin. This quantity of niacin increases the available niacin intake to 0.892 mg. per hen per day which could maintain the hatchability recorded for this group. This evidence demonstrates that all niacin determined by microbiological analysis in the soybean meal was available to the hen.
AVAILABILITY OF NIACIN
SUMMARY
A quantitative bioassay was used for determining the niacin availability in feedstuffs for the hen. Hatchability of eggs was the independent variable used in the assay. A niacin deficient diet containing 0.134 percent tryptophan was adapted to make the assay diets. White Leghorn hens were used in all experiments. It was concluded that the niacin availability in yellow corn, dehulled soybean meal and wheat middlings was 30, 100 and 36 percent, respectively, for the hen. Tryptophan can be converted to niacin by the hen and it was calculated that it requires 187 mg. of tryptophan to supply 1 mg. of niacin. REFERENCES Barton-Wright, E. C., 1944. The microbiological assay of nicotinic acid in cereals and other products. Biochem. J. 38: 314-319. Briggs, G. M., Jr., 1945. Influence of gelatin and tryptophan on nicotinic acid requirement of chicks. J. Biol. Chem. 161: 749-750. Chaudhuri, D. K., and E. Kodicek, 1960. The availability of bound nicotinic acid to the rat. 4. The effect of treating wheat, rice and barley brans and a purified preparation of bound nico-
tinic acid with sodium hydroxide. British J. Nutrit. 14: 35-47. Duncan, D. B., 1955. The new multiple range and multiple F tests. Biometrics, 11: 1-42. Ghosh, H. P., P. K. Sarkar and B. C. Guha, 1963. Distribution of the bound form of nicotinic acid in natural materials. J. Nutrition, 79: 451-453. Goldsmith, C. A., A. G. Gibbens, W. G. Unglaub and O. N. Miller, 1956. Studies on niacin requirements in man. III. Comparative effects of diets containing lime treated and untreated corn in the production of the experimental pellagra. Am. J. Clin. Nutrition, 4: 151-160. Greene, R. D., and A. Black, 1944. The microbiological assay for tryptophan in proteins and foods. J. Biol. Chem. 155: 1-11. Heuser, G. F., and M. L. Scott, 1953. Studies in duck nutrition. 5. Bowed legs in ducks, a nutritional disorder. Poultry Sci. 32: 137-143. Johnson, D., and H. Fisher, 1958. The amino acid requirements of laying hens. 3. Minimal requirement levels of essential amino-acids: Techniques and development of diet. Brit. J. Nutrition, 12 : 276-285. Kodicek, E., 1940a. Estimation of nicotinic acid in animal tissues, blood and certain foodstuffs. I. Method. Biochem. J. 34: 712-723. Kodicek, E., 1940b. Estimation of nicotinic acid in animal tissues, blood and certain foodstuffs. 2. Applications. Biochem. J. 34: 724-735. Kodicek, E., R. Braude, S. A. Kon and K. G. Mitchell, 1959. The availability to pigs of nicotinic acid in "tortilla" baked from maize treated with lime water. British J. Nutrit. 13: 363-384. Krehl, W. A., C. A. Elvehjem and F. M. Strong, 1944. The biological activity of a precursor of nicotinic acid in cereal products. J. Biol. Chem. 156: 13-19. Krehl, W. A., and F. M. Strong, 1944. Studies on the distribution, properties and isolation of a naturally occurring precursor of nicotinic acid. J. Biol. Chem. 156: 1-12. Luce, W. G., 1965. The availability of niacin in certain cereal grains for swine. Ph.D. Thesis. University of Nebraska, Lincoln, Nebraska. Manoukas, T. G., 1963. The niacin requirement of the hen. A Thesis for the degree of Master of Science. University of New Hampshire, Durham. Manoukas A. G., 1967. The availability of niacin in certain feedstuffs for the hen. Ph.D. Thesis. University of New Hampshire, Durham. Petterson, E. B., J. R. Hunt, P. Vohra, L. G.
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On the basis of the information obtained from these experiments and the fact that animal by-product feedstuffs are devoid of a bound form of niacin, the available niacin supplied by most practical poultry feeds is greater than the minimum requirement of the hen for niacin. Therefore, supplementation of practical feeds for laying and breeding hens with synthetic niacin is not necessary, unless factors other than the niacin availability in feedstuffs modifies drastically the practical requirement of the hen for niacin. No convincing evidence was found to support the latter possibility. It is suggested, however, that experiments should be conducted with commercial practical feeds to test this possibility.
1841
1842
A. G. MANOUKAS, R. C. RINGROSF, AND A. E. TEF.RI
Blaylock and J. McGinnis, 1056. The niacin and typtophan requirements of chicks. Poultry Sci. 35: 499-504. Ringrose, R. C , A. G. Manoukas, R. Hinkson and A. E. Teeri, 1965. The niacin requirement of the hen. Poultry Sci. 44: 1053-1065.
Snedecor, G. W., 1956. Statistical Methods. The Iowa State College Press, Ames, Iowa. West, J. W., C. W. Garrick, S. M. Hauge and F. T. Hertz, 1952. The tryptophan requirement of young chickens as influenced by niacin. Poultry Sci. 3 1 : 479-487.
T. S. NELSON, 1 T. R. SHIEH, R. J. WODZINSKI AND J. H. WARE2 Research and Development Division, International Minerals and Chemical Corporation, Libertyville, Illinois 60048 (Received for publication March 30, 1968)
P
HYTATE phosphorus in plants is the mixed calcium-magnesium-potassium salt of phytic acid (Anderson, 1915; Averill and King, 1926). Most feed ingredients of plant origin contain from one-half to three-fourths of their total phosphorus as phytate (Common, 1940; Mollgaard, 1946). The chicks' ability to utilize phytate phosphorus remains a controversial subject despite extensive research to determine its availability. This problem was discussed in reviews by Kastelic and Forbes (1961), Taylor (1965), and Nelson (1967). It was stressed in the two earlier reviews that there was still no general agreement on the extent to which chicks utilize phytate phosphorus. Nelson (1967) concluded that the chick has a limited ability to utilize phytate phosphorus. Soybean meal (SBM), the major protein supplement in poultry feeds, is also a source of phosphorus. However, two-thirds of SBM phosphorus is bound as phytate. This report describes the ability of chicks to utilize the phytate phosphorus in un1
Present address: Department of Animal Sciences, University of Arkansas, Fayetteville, Arkansas 72701. " Present address: Billings, Montana.
treated SBM and in SBM treated with a mold phytase. EXPERIMENTAL
A phytate phosphorus assay diet (1, Table 1) was used in the first five experiments and a semi-practical diet (2, Table 1) was fed in experiment 6. Both diets conTABLE 1.—Composition of basal diets Diet 1
Diet 2
%
%
Yellow corn or 59.1675 Yellow corn meal Soybean meal, 50% protein 33.50 3.00 Corn oil Dried blood fibrin Cond. fish solubles 0.05 DL-methionine Iodized salt 0.25 0.02 MnSGvH 2 0 0.0125 ZnC0 3 0.50 Vitamin mixture 1 2 3.50 Supplements 100.00
57.4175 25.00 5.00 6.00 2.00 0.05 0.25 0.02 0.0125 0.50 3.75 100.00
1 Provided the following/kg. of diet: vitamin A, 5,000 I.U.; vitamin D3, 450 I.C.U.; (in mg./kg.), choline CI., 875; d-a-tocopheryl acetate, 37.5; niacin, 30; d-Ca. pantothenate, 10; riboflavin, 3.5; pyridoxine-HCl, 3.5; thiamine• HC1, 2.0; menadione sodium bisulfite, 1.0; folic acid, 0.6; biotin, 0.1; vitamin B12, 0.015; ethoxyquin, 187.5; chlortetracycline • HC1, 25; and Zn bacitracin, 25. 2 Calcium, phosphorus and sand.
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The Availability of Phytate Phosphorus in Soybean Meal Before and After Treatment With a Mold Phytase
ODICEK (1940a) suggested that a by Ringrose et al. (1965), with various large amount of niacin present in cereal levels of crystalline niacin, was fed to and other grains is in a bound form. Later White Leghorn hens for at least ten weeks it was reported by Kodicek (1940b), Krehl and several variables were studied. It was and Strong (1944), Krehl et al. (1944), found that hatchability of eggs was a recHeuser and Scott (1953), Goldsmith et al. ognizable linear function of niacin intake (1956), Kodicek et al. (1959) and Chau- over a wide range of intake; thus it was dhuri and Kodicek (1960) that the bound chosen as the independent variable to be form of niacin is unavailable or partly estimated in the assay. The regression available to the dog, microorganisms, chick, equation of hatchability (Y) on the niacin duckling, man, pig and rat, respectively. In intake (X) was Y = 3.74 + 90.9 X. The relation to the hen, the common quantities available niacin intake, in mg. per hen per of feedstuffs used in poultry rations should day, was given by the equation, X = meet the need of the hen for niacin based 0.011Y —0.041 (1). In this equation, Y upon minimum requirement studies as re- represents the mean hatchability, in perported by Ringrose et al. (1965). In poul- cent, for a period of ten weeks during try feeds, crystalline niacin is added to which eggs were incubated weekly. Due to meet the requirements for niacin because of the data used to derive equation (1), lack of information concerning the avail- hatchability values smaller than 13 and ability of feedstuffs niacin for the hen. The larger than 86 percent can not be used for present study was undertaken to determine determining the available niacin intake. Assays for available niacin in yellow corn, the availability of the niacin present in yellow corn, dehulled soybean meal and wheat dehulled soybean meal (50 percent protein) and wheat middlings were made. middlings for the hen. White Leghorn hens were used in all experEXPERIMENTAL PROCEDURE iments. A description of the niacin deficient The development of a quantitative bio- diet, experimental design and housing of assay procedure for available niacin was the hens has been reported by Ringrose et described by Manoukas (1967). In that al. (1965). The niacin deficient basal diet study, a niacin deficient basal diet reported was adapted to make the assay diets. An effort was made to maintain the tryptophan 1 Published with the approval of the Director content of all diets as close as possible to of the New Hampshire Agricultural Experiment the minimum requirement and to adjust Station as Scientific Contribution No. 434. Acthe rest of the dietary nutrients to approxicepted as part of a doctoral dissertation by the mately the same levels found in the niacin University of New Hampshire. deficient basal diet. 2 Present address: Department of Poultry Science, Cornell University, Ithaca, New York.
The niacin content of the basal diet and 1836
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K
AVAILABILITY or
meal. The composition of this diet is presented in Table 1 under the designation SN0. The soybean meal in this diet supplied 7.95 mg. of niacin per kg. of feed. The calculated tryptophan content of this diet was 0.157 percent. Crystalline niacin was added to the SN0 diet to provide assay diets containing 3.0, 5.0, 7.0 and 9.0 mg. of crystalline niacin per kg. of feed. These diets, designated SN3, SN5, SN7 and SN9, respectively, were fed to the hens for a period of seven weeks. By the seventh week, at which time four hatches of eggs were completed, it was apparent that most of the soybean niacin was available to the hen. Because of this observation the groups of hens fed diets SN5 and SN7 during the first seven weeks were fed diets SN0 and SNi.s, respectively, for the following five weeks of the experimental period. This change was made to determine the magnitude of the niacin availability in soybean meal without or with very little crystalline niacin supplementation. The groups of hens which were fed diets SN3 and SN9 were kept on the same diets throughout the experiment. In Experiment 3, the basal diet was formulated to contain 7 percent wheat mid-
TABLE 1.—Composition of assay diets Expt. 2
Expt. 3
Zl6.25
SN 0
TN»
%
% —
% — —
Experiment 1 Diet Zs.75
Corn meal Soybean meal Wheat middlings Corn starch Crude casein Gelatin D,l.-Methionine Limestone Di-Ca-phosphate Iodized salt Mineral mix* Vitamin mix* Corn oil Cellulose * Ringrose et al. (1965)
%
Zn.25
%
Zl3.75
%
35.00
45.00
55.00
65.00
— —
— —
— —
— —
30.69 9.54 7.14 0.22 7.65 3.50 0.50 1.60 0.56 1.60 2.00
21.99 9.02 6.76 0.22 7.65 3.50 0.50 1.60 0.56 1.20 2.00
13.29 8.51 6.38 0.21 7.65 3.50 0.50 1.60 0.56 0.80 2.00
4.59 8.00 6.00 0.20 7.65 3.50 0.50 1.60 0.56 0.40 2.00
25.00
—
48.49
—
7.00 0.20 7.65 3.50 0.50 1.60 0.56 4.00 1.50
7.00 54.99 10.00 7.50 0.20 7.65 3.50 0.50 1.60 0.56 5.00 1.50
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studied feedstuffs was determined by the microbiological assay of Barton-Wright (1944). The tryptophan content of the basal diet and corn was determined by the method of Green and Black (1944). Sixty hens were allotted to four treatments, fifteen hens per treatment in each of three separate experiments. In order to obtain hatchability data, the hens were artificially inseminated twice weekly. The eggs were held in a 15°C. egg room and incubated weekly. Individual egg production was recorded daily. Treatment-group feed consumption was recorded at the end of each two-week period. Individual body weight was recorded on the first and last day of each experiment. In Experiment 1, the basal diet was adapted to contain 35, 45, 55 and 65 percent corn. The composition of these diets is presented in Table 1 under the designations Zs.75, Zn.25) Zi3,75 and Zie.25, respectively. Subscripts indicate the niacin content derived from corn, in mg. per kg., for each diet. The tryptophan content of these diets was 0.155 percent. The experiment lasted ten weeks. In Experiment 2, the basal diet was adapted to contain 25 percent soybean
1837
NIACIN
1838
A. G. MANOUKAS, R. C. RINGROSE AND A. E. TEERI
TABLE 2.—Daily feed consumption, egg pro?"-ction, daily feedstuff niacin intake (FNI), daily crystalline niacin intake (CNI), hatchability and feedstuff niacin availability for the hen. (Means of 15 hens per treatment) Diet
Feed consumption g./hen
Zll.25 Zl3.76 Zl6.25
88 94 94 103
SN„*
96
SN,.6*
94
SN3.0
96
SN,.o
92
TN a
85
TN,. 5
84
TN3.„
97
TN4.5
100
^8.75
Egg, production
(1) FNI (2) CNI
mg./hen Yellow corn, Expt. 1 60.2 (1) 0.770 64.5 (1) 1.058 68.6 (1) 1.293 (1) 1.674 79.2 Soybean meal, Expt. 2 57.1 (1) 0.785 (2) 57.9 (1) 0.747 (2) 0.141 54.1 (1) 0.763 (2) 0.288 56.5 (1) 0.731 (2) 0.828 Wheat middlings, Expt. 3 56.0 (1) 0.940 (2) 60.5 (1) 0.929 (2) 0.128 (1) 1.073 71.9 (2) 0.291 76.1 (1) 1.106 (2) 0.450
%
* For the last five weeks of the Experiment, see text. ** This figure was obtained by direst comparison.
Hatchability
Niacin availability
%
%
23.2 31.3 37.6 58.6
28 29 29 36
93.6
100**
96.8
—
91.9
—
93.7
—
32.7
34
51.0
42
67.3
38
83.7
39
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dlings. The composition of this diet is were made at the 0.05 level of probability. presented in Table 1 under the designa- The linear equation (1) was used to estition TN 0 . The middlings in this diet sup- mate the available niacin present in corn, plied 11.06 mg. of niacin per kg. of feed. soybean meal and wheat middlings. Diet TN 0 constituted one treatment. NiaEXPERIMENTAL RESULTS cin was then added to diet TN 0 to provide AND DISCUSSION three additional diets containing 1.5, 3.0 and 4.5 mg. of crystalline niacin per kg. of feed. In Experiment 1, diets Zs.7f„ Z u . 2 5, Z13.75 These diets were designated TNi. 5 , TN:t.0 and Z16.25 were used which contained 35, and TN4.5, respectively. The calculated 45, 55 and 65 percent corn, respectively. tryptophan content of these diets was A summary of mean daily feed consump0.148 percent. tion, egg production, daily corn niacin inFor brevity and convenience in reporting take, hatchability and estimated niacin and discussing the data, the symbols used availability in corn is presented in Table 2. to designate the diets are also used to rep- Groups Z8.75, Zu.25 and Z13.7r, lost 128, 106 resent the respective groups of hens to and 68 grams of body weight per hen, rewhich the diets were fed. spectively, during the experiment. Group The analysis of variance (completely Z16.25 gained 9 grams of body weight per randomized design) as outlined by Snedecor hen during the experiment. (1956) and the multiple range test of DunStatistical analysis of the data showed can (1955) were used to evaluate the ex- that hatchability of eggs obtained from perimental data. All tests of significance group Z8.75 differed from that obtained
AVAILABILITY OF NIACIN
under their experimental conditions. Consideration of all results obtained with corn diets shows that the niacin in corn is approximately 30 percent available to the hen. The low availability of corn niacin for the hen is in agreement with experimental results reported by Kodicek (1940b), Krehl and Strong (1944), Goldsmith et al. (1956), Kodicek et al. (1959) and Chaudhuri and Kodicek (1960). Ghosh et al. (1963) reported that 90 percent of corn (Zea mays) niacin is unavailable to the microorganism Leuconostoc mesenteroides. Luce (1965) reported that corn niacin is largely unavailable to swine. In Experiment 2, dehulled soybean meal diets SNS and SN9 were tested during a twelve-week experimental period and diets SN5 and SN7 during the first seven weeks of the experimental period. Diets SN0 and SN1.5 substituted for diets SN5 and SN7, respectively, for the following five weeks of the experiment. A summary of mean daily feed consumption, egg production, daily niacin intake, hatchability and estimated niacin availability in soybean meal is presented in Table 2. Groups SN3, SNS, (SNo), SN r , (SNLS) and SN9 gained 176, 188, 183 and 145 grams of body weight, respectively, during the twelve-week experimental period. Statistical analysis of the data showed that no one group differed from another in any of the reported functions. Hatchability and feed consumption were not affected by the dietary niacin intake and all groups gained body weight. Egg production was equivalent but lower than expected for all groups. The low egg production may be due partly to the low egg production of the experimental stock at the beginning of the experiment and partly to the relatively high temperature of the summer months. On the basis of the data obtained, it is apparent that all of the niacin found in
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from groups Z13.7r, and Z16.25. Also hatchability from groups Z n . 2 5 and Z13.75 differed from that of group Z16.,5. Egg production and feed consumption of groups Z8.75, Zn.25 and Zi3.75 differed from that of group Z16.25. No other differences were found among the groups. Hatchability data from all groups clearly demonstrates that corn niacin is partly available to the hen. Application of the mean hatchability percentage obtained for groups Zs.75, Zn.25, Z13.75 and Z16.25 to equation (1), gave availability of niacin equal to 28, 29, 29 and 36 percent, respectively. The greater availability of corn niacin in diet Z16.25 may be attributed to variation and/or to greater tryptophan intake; the additional tryptophan being converted to niacin. The minimum requirement of the hen for tryptophan is approximately 135 mg. per day as reported by Johnson and Fisher (1958). The tryptophan intake of groups Z8.75 and Z16.25 was 136 mg. and 161 mg. per hen per day, respectively, a difference of 25 mg. Application of the hatchability figure of 58.6 percent for diet Zi6.25 to equation (1) gave an available niacin intake of 0.604 mg. per hen per day. Applying the estimate of 28 percent available niacin, obtained for diet Z8.75, to the daily niacin intake of 1.674 mg. per hen fed diet Z16.25, one derives an available niacin intake of 0.470 mg. If the greater hatchability and niacin availability percentage for corn in diet Zi6.25 relative to diet Zs.75 was due to more niacin, then 0.134 mg. (0.604-0.470) of niacin per hen per day must have come from the 25 mg. of additional tryptophan available per hen in group Z ie . 25 . It takes, therefore, 187 mg. of tryptophan to supply 1 mg. of niacin. This conversion ratio is not in agreement with results reported by Briggs (1945), West (1952) and Petterson (1956) who state that it takes 55 to 60 mg. of tryptophan to replace 1 mg. of niacin for chicks,
1839
1840
A. G. MANOUKAS, R. C. RINGEOSE AND A. E. TEERI
consumption, egg production, niacin intake, hatchability and estimated niacin availability in wheat middlings is presented in Table 2. Groups TN 0 , T N ^ , TN3.0 and N4.5 gained 16, 47, 45 and 62 grams of body weight, respectively, during the experiment. Statistical analysis of the data showed that diets TN 0 and TN li5 significantly depressed feed consumption, hatchability and egg production as compared to diets TN3.0 and TN4..5. Hatchability results of all groups differed significantly from each other. No other differences were found among the groups. Hatchability data from all groups clearly shows that the niacin in wheat middlings is partly available to the hen. Application of the mean hatchability percentage obtained from groups TN 0 , TN1.5, TN3.0, and TN4.5 to equation (1) gave availability of niacin equal to 34, 42, 38 and 39 percent, respectively. With a conversion ratio of 187 mg. of tryptophan to 1 mg. of niacin, derived from Experiment 1, it can be calculated that the availability of niacin in wheat middlings for groups TN3.0 and TN4.S is 34 and 33 percent, respectively, because of greater tryptophan intake comNo data could be found on the availabil- pared to groups TN 0 and TNi.5, The ity of soybean niacin for other species. niacin availability derived for group TN1.5 Ghosh et al. (1963) reported that peas cannot be explained on the basis of trypto(Pisum sativum) and other pulses do not phan conversion to niacin. contain a bound form of nicotinic acid. But Taking into consideration the conversion groundnut {AracMs hypogea) and other oil of tryptophan to niacin, it was concluded seeds contain about 40 percent of the total that the availability of niacin in wheat midniacin content in a bound form which is dlings was 36 percent for the hen. No inunavailable to Leuconostoc mesenteroides. formation could be found on the availabilIt should be noted that the soybean meal ity of wheat middlings niacin for other speused in this experiment was a heat treated cies with the exception of ducklings. commercial product for direct use in poul- Heuser and Scott (1963) suggested that try feeds. Heat treatment may affect niacin the niacin present in wheat middlings apavailability. peared to be relatively unavailable to the Diets TN„, TNj.s, TN3.0 and N4.5 con- duckling. Luce (1965) indicated that the taining wheat middlings were tested in in niacin present in wheat was only partially Experiment 3. A summary of mean daily feed available to swine.
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soybean meal was available to the hen. The average soybean niacin intake of group SNo was 0.78S mg. per hen per day and the average hatchability recorded was 93.6 percent for the five weeks. Combined data reported by Manoukas (1963) and Ringrose et al. (1965) shows that 0.830 mg. of niacin intake per hen per day from a purified diet supported hatchability equal to 83.4 percent during a period of five weeks. This information indicates that 0.785 mg. of niacin could not support 93.6 percent hatchability for a period of five weeks. Thus it appears that the required niacin was obtained from the conversion of tryptophan to niacin. Group SN 0 consumed 155 mg. of tryptophan per hen per day of which 20 mg. may be converted to niacin. With a conversion ratio of 187 mg. of tryptophan to 1 mg. of niacin, derived from Experiment 1, it can be calculated that 20 mg. of tryptophan can supply 0.107 mg. of niacin. This quantity of niacin increases the available niacin intake to 0.892 mg. per hen per day which could maintain the hatchability recorded for this group. This evidence demonstrates that all niacin determined by microbiological analysis in the soybean meal was available to the hen.
AVAILABILITY OF NIACIN
SUMMARY
A quantitative bioassay was used for determining the niacin availability in feedstuffs for the hen. Hatchability of eggs was the independent variable used in the assay. A niacin deficient diet containing 0.134 percent tryptophan was adapted to make the assay diets. White Leghorn hens were used in all experiments. It was concluded that the niacin availability in yellow corn, dehulled soybean meal and wheat middlings was 30, 100 and 36 percent, respectively, for the hen. Tryptophan can be converted to niacin by the hen and it was calculated that it requires 187 mg. of tryptophan to supply 1 mg. of niacin. REFERENCES Barton-Wright, E. C., 1944. The microbiological assay of nicotinic acid in cereals and other products. Biochem. J. 38: 314-319. Briggs, G. M., Jr., 1945. Influence of gelatin and tryptophan on nicotinic acid requirement of chicks. J. Biol. Chem. 161: 749-750. Chaudhuri, D. K., and E. Kodicek, 1960. The availability of bound nicotinic acid to the rat. 4. The effect of treating wheat, rice and barley brans and a purified preparation of bound nico-
tinic acid with sodium hydroxide. British J. Nutrit. 14: 35-47. Duncan, D. B., 1955. The new multiple range and multiple F tests. Biometrics, 11: 1-42. Ghosh, H. P., P. K. Sarkar and B. C. Guha, 1963. Distribution of the bound form of nicotinic acid in natural materials. J. Nutrition, 79: 451-453. Goldsmith, C. A., A. G. Gibbens, W. G. Unglaub and O. N. Miller, 1956. Studies on niacin requirements in man. III. Comparative effects of diets containing lime treated and untreated corn in the production of the experimental pellagra. Am. J. Clin. Nutrition, 4: 151-160. Greene, R. D., and A. Black, 1944. The microbiological assay for tryptophan in proteins and foods. J. Biol. Chem. 155: 1-11. Heuser, G. F., and M. L. Scott, 1953. Studies in duck nutrition. 5. Bowed legs in ducks, a nutritional disorder. Poultry Sci. 32: 137-143. Johnson, D., and H. Fisher, 1958. The amino acid requirements of laying hens. 3. Minimal requirement levels of essential amino-acids: Techniques and development of diet. Brit. J. Nutrition, 12 : 276-285. Kodicek, E., 1940a. Estimation of nicotinic acid in animal tissues, blood and certain foodstuffs. I. Method. Biochem. J. 34: 712-723. Kodicek, E., 1940b. Estimation of nicotinic acid in animal tissues, blood and certain foodstuffs. 2. Applications. Biochem. J. 34: 724-735. Kodicek, E., R. Braude, S. A. Kon and K. G. Mitchell, 1959. The availability to pigs of nicotinic acid in "tortilla" baked from maize treated with lime water. British J. Nutrit. 13: 363-384. Krehl, W. A., C. A. Elvehjem and F. M. Strong, 1944. The biological activity of a precursor of nicotinic acid in cereal products. J. Biol. Chem. 156: 13-19. Krehl, W. A., and F. M. Strong, 1944. Studies on the distribution, properties and isolation of a naturally occurring precursor of nicotinic acid. J. Biol. Chem. 156: 1-12. Luce, W. G., 1965. The availability of niacin in certain cereal grains for swine. Ph.D. Thesis. University of Nebraska, Lincoln, Nebraska. Manoukas, T. G., 1963. The niacin requirement of the hen. A Thesis for the degree of Master of Science. University of New Hampshire, Durham. Manoukas A. G., 1967. The availability of niacin in certain feedstuffs for the hen. Ph.D. Thesis. University of New Hampshire, Durham. Petterson, E. B., J. R. Hunt, P. Vohra, L. G.
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On the basis of the information obtained from these experiments and the fact that animal by-product feedstuffs are devoid of a bound form of niacin, the available niacin supplied by most practical poultry feeds is greater than the minimum requirement of the hen for niacin. Therefore, supplementation of practical feeds for laying and breeding hens with synthetic niacin is not necessary, unless factors other than the niacin availability in feedstuffs modifies drastically the practical requirement of the hen for niacin. No convincing evidence was found to support the latter possibility. It is suggested, however, that experiments should be conducted with commercial practical feeds to test this possibility.
1841
1842
A. G. MANOUKAS, R. C. RINGROSF, AND A. E. TEF.RI
Blaylock and J. McGinnis, 1056. The niacin and typtophan requirements of chicks. Poultry Sci. 35: 499-504. Ringrose, R. C , A. G. Manoukas, R. Hinkson and A. E. Teeri, 1965. The niacin requirement of the hen. Poultry Sci. 44: 1053-1065.
Snedecor, G. W., 1956. Statistical Methods. The Iowa State College Press, Ames, Iowa. West, J. W., C. W. Garrick, S. M. Hauge and F. T. Hertz, 1952. The tryptophan requirement of young chickens as influenced by niacin. Poultry Sci. 3 1 : 479-487.
T. S. NELSON, 1 T. R. SHIEH, R. J. WODZINSKI AND J. H. WARE2 Research and Development Division, International Minerals and Chemical Corporation, Libertyville, Illinois 60048 (Received for publication March 30, 1968)
P
HYTATE phosphorus in plants is the mixed calcium-magnesium-potassium salt of phytic acid (Anderson, 1915; Averill and King, 1926). Most feed ingredients of plant origin contain from one-half to three-fourths of their total phosphorus as phytate (Common, 1940; Mollgaard, 1946). The chicks' ability to utilize phytate phosphorus remains a controversial subject despite extensive research to determine its availability. This problem was discussed in reviews by Kastelic and Forbes (1961), Taylor (1965), and Nelson (1967). It was stressed in the two earlier reviews that there was still no general agreement on the extent to which chicks utilize phytate phosphorus. Nelson (1967) concluded that the chick has a limited ability to utilize phytate phosphorus. Soybean meal (SBM), the major protein supplement in poultry feeds, is also a source of phosphorus. However, two-thirds of SBM phosphorus is bound as phytate. This report describes the ability of chicks to utilize the phytate phosphorus in un1
Present address: Department of Animal Sciences, University of Arkansas, Fayetteville, Arkansas 72701. " Present address: Billings, Montana.
treated SBM and in SBM treated with a mold phytase. EXPERIMENTAL
A phytate phosphorus assay diet (1, Table 1) was used in the first five experiments and a semi-practical diet (2, Table 1) was fed in experiment 6. Both diets conTABLE 1.—Composition of basal diets Diet 1
Diet 2
%
%
Yellow corn or 59.1675 Yellow corn meal Soybean meal, 50% protein 33.50 3.00 Corn oil Dried blood fibrin Cond. fish solubles 0.05 DL-methionine Iodized salt 0.25 0.02 MnSGvH 2 0 0.0125 ZnC0 3 0.50 Vitamin mixture 1 2 3.50 Supplements 100.00
57.4175 25.00 5.00 6.00 2.00 0.05 0.25 0.02 0.0125 0.50 3.75 100.00
1 Provided the following/kg. of diet: vitamin A, 5,000 I.U.; vitamin D3, 450 I.C.U.; (in mg./kg.), choline CI., 875; d-a-tocopheryl acetate, 37.5; niacin, 30; d-Ca. pantothenate, 10; riboflavin, 3.5; pyridoxine-HCl, 3.5; thiamine• HC1, 2.0; menadione sodium bisulfite, 1.0; folic acid, 0.6; biotin, 0.1; vitamin B12, 0.015; ethoxyquin, 187.5; chlortetracycline • HC1, 25; and Zn bacitracin, 25. 2 Calcium, phosphorus and sand.
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The Availability of Phytate Phosphorus in Soybean Meal Before and After Treatment With a Mold Phytase