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The Relationship of Calcium-Phosphorus Ratios to the Utilization of Plant and Inorganic Phosphorus by the Chick1
METABOLIZABLE ENERGY ACKNOWLEDGMENTS
The authors gratefully acknowledge the technical assistance of Mr. W. F. Pepper of the Department of Poultry Husbandry and Mrs. J. Czarnocki, Miss A. Wilson and Mr. D. Arthur of the Department of Nutrition. Our thanks are also due to Professor E. V. Evans of the Department of Nutrition for assaying the diets of Experiment 1 for pantothenic acid. REFERENCES Association of Official Agricultural Chemists, 1955. Methods of Analysis, 8th Edition. Baldini, J. T., 1960. The effect of dietary deficiency on the energy metabolism of the chick. Poultry Sci. 39:1232. Brambila, S., M. C. Nesheim and F. W. Hill, 1960. Studies of the effect of trypsin on the utilization of a raw soybean oil meal by the chick. Poultry Sci. 39: 1237. Czarnocki, J., I. R. Sibbald and E. V. Evans, 1960. Unpublished procedure. Gray, J. A., and E. V. Evans, 1958. Unpublished procedure. King, E. J., 1932. Colorimetric determination of phosphorus. Biochem. J. 26: 292-297. Nair, K. R., 1948. The studentized form of the ex-
951
treme mean square test in the analysis of variance. Biometrika, 35: 16-31. Skeggs, H. R., and L. D. Wright, 1944. The use of Lactobacillus arabinosus in the microbiological determination of pantothenic acid. J. Biol. Chem. 156: 21-26. Sibbald, I. R., J. D. Summers and S. J. Slinger, 1959. Factors affecting the metabolizable energy content of poultry feeds. Poultry Sci. 38: 1247. Sibbald, I. R., J. D. Summers and S. J. Slinger, 1960a. Factors affecting the metabolizable energy content of poultry feeds. Poultry Sci. 39: 544556. Sibbald, I. R., S. J. Slinger and G. C. Ashton, 1960b. A synergistic relationship between tallow and undegummed soybean oil. Poultry Sci. 39: 1295. Sibbald, I. R., S. J. Slinger and G. C. Ashton, 1961a. Factors affecting the metabolizable energy content of poultry feeds. 2. Variability in the M.E. values attributed to samples of tallow and undegummed soybean oil. Poultry Sci. 40: 303308. Sibbald, I. R., S. J. Slinger and G. C. Ashton, 1961b. Factors affecting the metabolizable energy content of poultry feeds. 3. The influence of Kaolin and Alphacel when used as ration diluents. Poultry Sci. 40: 454-458. Snedecor, G. W., 1956. Statistical Methods, Fifth Edition, Iowa State College Press, Ames, Iowa.
The Relationship of Calcium-Phosphorus Ratios to the Utilization of Plant and Inorganic Phosphorus by the Chick 1 J. M. VANDEPOPULIERE,2 C. B. AMMERMAN3 AND R. H. HARMS4 Florida Agricultural Experiment Stations, Gainesville (Received for publication September 19, 1960)
C
ONSIDERABLE interest has developed in the past 20 years in determining the availability of phosphorus from plant and inorganic sources. In order to 1 Florida Agricultural Experiment Stations Journal Series No. 1120. 2 Present address Ralston Purina Co., St. Louis, Missouri. 3 Department of Animal Science. 4 Department of Poultry Science.
properly evaluate results from such experiments it is necessary to understand some of the factors which may influence utilization of phosphorus such as calcium-phosphorus (Ca:P) ratio, levels of ascorbic acid and fat. Bethke et al. (1928) and Hart et al. (1930) reported that the optimum or near optimum Ca:P ratio for growing chicks was between 2:1 and 4:1. Wilgus (1931) stated
952
J. M. VANDEPOPULIERE, C. B. AMMERMAN AND R. H. HARMS TABLE 1.—Composition of basal diet* Ingredients
White corn meal (degerminated) Soybean oil meal (50% protein) Dehydrated alfalfa meal (17% protein) Micro-ingredients1 Iodized salt Ground limestone
Percent 64.10 31.18 3.00 0.90 0.40 0.42 100.00
* Basal diet contains 0.28% phosphorus and 0.28% calcium. 1 Supplies per pound of feed: 2268 I. U. vitamin A, 10 meg. vitamin Bi 2 , 340 I. C. U. vitamin D 3 , 2 mg. riboflavin, 9 mg. calcium pantothenate, 18 mg. niacin, 261 mg. choline chloride, 140 mg. methionine, 10 mg. Terramycin, 2 mg. Oleandomycin, 57 mg. Santoquin, 9 mg. iron, 0.9 mg. copper, 90 meg. cobalt, 5 mg. iodine, 45 meg. zinc, 80 mg. manganese sulphate, and 35 mg. manganous oxide.
that a 2.5:1 ratio was borderline and 3.3:1 was disastrous. Carver et al. (1946) found that a 1:1 or 2:1 ratio produced the most .favorable growth in Leghorn cockerels. Briggs et al. (1944) reported that ascorbic acid fed to chicks receiving various purified diets promoted small but consistent increases in growth rate. They suggested that chicks fed a highly purified diet were not able to synthesize sufficient ascorbic acid to permit maximum growth. March and Biely (1953) reported a growth stimulus from the addition of vitamin C to a diet of natural ingredients. The work of Thornton and Moreng (1958) indicated that the addition of ascorbic acid to the diet of hens improved calcium utilization. Almquist (1954) suggested that the phosphorus requirement of chicks from one day to four weeks of age was 0.45% from a readily available source. Singsen and Mitchell (1944) reported that 0.45% phosphorus from plant source was adequate. However, McGinnis et al. (1944) reported that 0.58% phosphorus from plant source was required for maximum calcification. It is generally accepted that the total level of dietary phosphorus required will depend on the amount which is derived from plant
source. Lowe et al. (1939) showed that the addition of phytin to a low phosphorus basal diet, thereby increasing the phosphorus content of the basal diet by 50%, resulted in no significant improvement in the calcification of the bones of the chick. This study was designed to determine the influence of different Ca:P ratios on the utilization of phosphorus from plant source, feed grade dicalcium phosphate and soft phosphate. Ascorbic acid and added fat were incorporated as variables to determine their effect on calcium and phosphorus utilization. EXPERIMENTAL PROCEDURE
Day-old Vantress X White Plymouth Rock chicks were used in these experiments; seven males and seven females were used per replication in Experiment 1, and 5 males and 5 females per replication in Experiment 2. Each treatment was replicated 2 times in Experiment 1, and 3 times in Experiment 2. In Experiment 1, phosphorus levels 0.28, 0.44, and 0.58% were compared in Ca:P ratios of 1:1, 2:1, 4:1, and 8:1. Calcium and phosphorus levels were obtained by the addition of feed grade dicalcium phosphate and ground limestone. Each of the above experimental diets was fed with and without the addition of 0.05% vitamin C. In Experiment 2, phosphorus levels of 0.44, 0.58, and 0.72%, produced by the addition of feed grade dicalcium phosphate, were compared in Ca:P ratios of 1.0:1, 1.5:1, and 2.0:1. These comparisons were made with two different diets which contained 960 and 1,060 Calories of productive energy per pound of feed. Soft phosphate was studied at phosphorus levels of 0.58, 0.65, and 0.72% with Ca:P ratios of 1.50: 1, 1.75:1, and 2.00:1. The soft phosphate diets contained 960 Calories of energy per pound of feed. The composition of the basal diet is
PHOSPHORUS UTILIZATION
953
shown in Table 1. This basal diet contained 0.28% calcium and 0.28% phosphorus. In PHOSPHORUS 0.2B_, order to obtain the desired level of protein 0.44 and energy in the various experimental 0.5 8 diets, adjustments were made in amounts of i < corn and soybean oil meal used, and animal fat was added when necessary. Energy and (- 300. protein levels of experimental diets were i o calculated using values of Titus (1955). The chicks were housed in metal bat>• 2 0 0 . Q teries with raised wire floors, and water and 0 (0 feed were supplied ad libitum. The temperature was thermostatically controlled. •/o MORTALITY Birds were individually weighed and feed 35-* 50-** utilization calculated at the end of the 4 75+O90-+** week experiment period. Percent bone ash II 2:1 4:1 8:1 of the right tibia was determined as outCALCIUM:PUO&PUORUS RATIO lined by A.O.A.C. (1955). Statistical analyses of Experiment 1 were made by the least FIG. 1. Body weight of chicks receiving diets squares method according to Anderson and containing various calcium-phosphorus ratios (ExBancroft (1952). Data in Experiment 2 periment 1). were subjected to analyses of variance according to Snedecor (1957) and significant ity on all other treatments was less than differences among treatment means were 5%. . determined according to Duncan's Multiple Growth rate increased in a linear fashion Range Test (1955). with all phosphorus levels as the Ca:P ratio narrowed to 1:1 (Figure 1). With diets RESULTS AND DISCUSSION containing 0.58% phosphorus there was no Experiment 1: During the first week significant difference between the 1:1 and chicks fed the basal diet, containing 0.28% 2:1 ratio; however, all other treatments plant phosphorus with Ca:P ratios of 4:1 were significantly different from the 1:1 and 8:1, developed severe rickets and were ratio at the .01 level of probability. unable to reach the feed and water troughs. The mean adjusted bone ash for ExperiAll chicks on these two treatments were fed ment 1 was 43.7%. It can be seen (Figures and watered for the remainder of the ex- 1 and 2) that the bone ash data paralleled periment in containers which they could the growth data very closely, with the exreach while sitting on the wire mesh floor. ception of the chicks receiving the basal Even with this special handling 75 and diet. The slope of the bone ash line of the 90% mortality was observed with the basal diet was not as steep as that of the chicks receiving diets with 4:1 and 8:1 other diets. Since the slope of all growth ratios, respectively. When the ratio was curves were comparable, it is possible that narrowed to 2:1, the mortality decreased to the phosphorus was available for increased 35%, and only 7% died when the ratio was growth but not adequate to give a marked 1:1. When the phosphorus level was in- increased in bone ash. All treatments excreased to 0.44% with an 8:1 Ca:P ratio cept the 0.58% phosphorus in a Ca:P mortality was approximately 50%. Mortal- ratio of 2:1 differed significantly from the
954
J. M. VANDEPOPULIERE, C. B. AMMERMAN AND R. H. HARMS
50
"It, PHOSPHORUS
n n 0 44 0.58..
•"
\
\
40.
\
\
\
\ \
,0
I
\
\
N
4
N
\
U
N
*»
z
0 a
30.
• • . . .
~~
x " ^
20. |:| 2:| 4:1 C A L C I U M : P H O S P H O R U S RATIO
8:1
FIG. 2. Tibia ash of chicks receiving diets containing various calcium-phosphorus ratios (Experiment 1).
0.58% phosphorus and 1:1 Ca:P ratio. Adjusted bone ash of males was 0.8% less than that of females; this difference was significant at the .01 level of probability. Addition of ascorbic acid to the diet did not affect growth or bone ash. Feed effi-
ciency was not calculated due to excessive mortality. An examination of Figures 1 and 2 indicates that the growth rate was as good a criteria as bone ash to use in determining the phosphorus adequacy of diets. This was especially true when the phosphorus requirement of the chick was being approached. Experiment 2: The range of phosphorus levels and Ca:P ratios used in this experiment was narrowed to meet the requirement of the fast growing broiler type chick. The most rapid gain was obtained with chicks receiving diets containing 0.72% phosphorus (Table 2). This growth rate was significantly greater than that obtained when diets contained 0.58% phosphorus. A further reduction in growth was observed with chicks receiving diets containing 0.44% phosphorus. Only the 1.5:1 ratio produced a significant improvement (P < 0.05) over the 2:1 ratio. The high energy diet (1,060 Cal./lb.) produced a highly significant increase in weight gain over the low energy diet (960 Cal./lb.). There were no significant energy X phosphorus level or energy X ratio interactions, indicating that the phosphorus levels and Ca:P ratios studied were not influenced
TABLE 2.—Four week chick weights when fed diets containing various levels of supplemental dicalcium phosphate in three different calcium-phosphorus ratios (Experiment 2) Ca:P ratio Energy level Calories/lb.
960 1060 Average 960 and 1060 1
Phosphorus Levels %»
0.44 0.58 0.72 0.44 0.58 0.72 0.44 0.58 0.72
1 5-l
1.0:1
2 0"1
M
F
AV.
M
F
AV.
M
F
AV.
432 446 475 409 504 538 420 475 506
Grams 395 375 419 386 418 448 390 396 434
414 410 447 398 461 493 405 436 470
413 478 509 455 512 475 434 495 492
Grams 390 415 431 395 439 441 392 427 436
402 446 470 425 476 458 413 461 464
402 473 472 418 490 470 410 482 471
Grams 364 425 404 377 422 447 370 424 426
383 449 438 398 456 458 390 453 448
This includes 0.28% phosphorus from plant sources.
PHOSPHORUS
955
UTILIZATION
TABLE 3—Feed per unit gain ofchicks when fed various levels of supplemental phosphate from dicalcium phosphate and soft phosphate in several calcium-phosphorus ratios (Experiment 2) Phosphorus source
Phiisphorus 1 /o
Cal. prod. energy/lb. feed
Ca:P ratio 1.0:1
1.5:1
1.69 1.61 1.60
1.60 1.57 1.54
1.63
1.57
1.59 1.53 1.51
1.49 1.47 1.45
1.54
1.47
1.75:1
2.00:1
Av.
1.58 1.59 1.57
1.62 1.59 1.57
1.58
1.59
1.51 1.46 1.47
1.53 1.49 1.48
—
1.48
1.50
E
1.60 1.56 1.55
1.61 1.75 1.57
1.68 1.71 1.55
1.63 1.67 1.56
—
1.57
1.64
1.65
1.62
Grams Dicalcium phosphate
0.44 0.58 0.72
960
Av. Dicalcium phosphate
0.44 0.58 0.72
1060
Av. Soft phosphate
0.58 0.65 0.72
960
Av.
—
This includes 0.28% phosphorus from plant sources.
by the increased energy content of the diet. Feed efficiency of chicks was improved when the level of phosphorus from dicalcium phosphate was increased from 0.44 to 0.58%, and as the Ca:P ratio was widened from 1.0:1 to 1.5:1 (Table 3). Highly significant differences in feed efficiency were observed between the groups receiving the two dietary levels of energy. The magnitude of the change in feed efficiency with increasing calcium levels was similar between the two energy levels. This is not in agreement with the report by Pepper et al. (1955) who found an improvement in feed efficiency when the calcium level was increased from 1.0 to 1.2% in diets containing 5-10% added fat, and no improvement in the absence of the added fat. As in Experiment 1, the males grew more rapidly than the females, and a significant phosphorus level X sex interaction was observed. Examination of the data showed that the male gained more than the female with each added increment of phosphorus, which indicates that the requirement for phosphorus is greater for male than for female.
A direct comparison of growth responses was made (Figure 3) between dicalcium and soft phosphate, at 2 phosphorus levels and 2 Ca:P ratios. Highly significant differences were observed with sex, Ca:P ratio, phosphorus level and phosphorus source. The 1.5:1 Ca:P ratio was superior to the 2:1 ratio, and 0.72% phosphorus produced more rapid gains than did 0.58% phosphorus. Body weight of birds receiving dicalcium phosphate was significantly heavier than birds receiving soft phosphate. There was a greater differential in growth between the phosphorus sources for the male than for the female. Hence, it appears that the soft phosphate, when fed at levels used in this experiment (Table 4), comes nearer to meeting the phosphorus requirement of the female than of the male. The ratio X phosphorus source interaction indicates that the wide ratio was more detrimental to the soft phosphate which had less available phosphorus than it was to the dicalcium phosphate. Since the phosphorus from soft phosphate is less available than that from dicalcium phosphate, the levels fed were well below the requirement level
956
J. M. VANDEPOPULIERE, C. B. AMMERMAN AND R. H. HARMS
5 0 0 ,-
DICALCIUM PHOSPHATE SOFT P H O S P H A T E
a ""••a
Si
2 < a i?400
°/o PUOSPMORUS 0.44 • 0.58 O 0.65 A 0.72 •
h 3 UJ
^ ^ ^ ^ ^ ^ ^ ^
BODY
5
^
"
\
^
^ ^ ^ \ ^
300
1.5:1 C A L C I U M : PHOSPHORUS
2.0:1 RATIO
FIG. 3. Comparison of chick weights when fed diets containing dicalcium and soft phosphate (Experiment 2).
of the chick indicating that the Ca:P ratio is more critical in diets which contain phosphorus levels that are below the chicks requirement. The significant phosphorus level X phosphorus source interaction suggests that the phosphorus requirement using dicalcium phosphate was near 0.58%, whereas there was a marked improvement when the phosphorus level was increased from 0.58 to 0.72% when soft phosphate was used. SUMMARY
Data from two four-week experiments studying various phosphorus sources and
levels at different Ca:P ratios have been presented. Results from these tests indicate: 1. The importance of Ca:P ratio increases as the dietary phosphorus level decreases from the chick's requirement. This may be due either to absolute quantity or decreased availability of the dietary phosphorus. 2. Plant source phosphorus was readily available to support growth when fed at a Ca:P ratio of 1:1, however, it did not allow for increased bone ash as did dicalcium phosphate when the Ca:P ratios were narrowed from 4:1 to 1:1. This would indi-
TABLE 4.—Four week chick weights when fed diets containing various levels of supplemental soft phosphate in three different calcium-phosphorus ratios (Experiment 2) Ca:P ratio Phosphorus level %
0.58 0.65 0.72 1
J
1.75:1
1.50:1
2.00:1
M
F
AV.
M
F
AV.
M
F
AV.
371 404 442
Grams 345 371 379
358 388 410
366 392 434
Grams 319 348 388
342 370 411
290 381 391
Grams 309 321 351
300 351 371
This includes 0.28% phosphorus from plant sources.
PHOSPHORUS UTILIZATION
cate a differential in the requirement for growth and bone ash. 3. Chicks which had bone ash values of 28% did not exhibit symptoms of rickets when fed a diet containing 0.28% plant phosphorus at a Ca:P ratio of 1:1. 4. It is essential to consider both phosphorus levels and Ca:P ratios when determining phosphorus availability, irrespective of the phosphorus source. 5. The phosphorus requirement of the male chicks appears to be higher than that of the female. A phosphorus level of 0.72% from dicalcium phosphate with a Ca:P ratio of 1:1 to 1.5:1 produced maximum growth in these battery trials. 6. Ascorbic acid did not influence growth rate or bone ash of four-week-old chicks. 7. Increasing the energy level from 960 to 1,060 Calories per pound of feed did not significantly influence the calcium or phosphorus utilization. 8. The criterion, growth, appears to be sufficient to determine the phosphorus adequacy of a diet when the level approaches the requirement. 9. Feed efficiency was improved significantly with increased levels of productive energy in the diet; however, feed efficiency on diets containing comparable levels of soft phosphate and dicalcium phosphate were similar. ACKNOWLED GMENTS
The authors wish to thank Dr. W. L. Reynolds for assistance in the statistical analysis of the data; International Mineral & Chemical Corp., Chicago, Illinois for furnishing the dicalcium phosphate; Soft Phosphate Research Institute, Ocala, Florida for furnishing the soft phosphate, which was from their composite sample of soft phosphate from various Florida sources; and Chas. Pfizer, Terre Haute, Indiana, for furnishing the vitamin A and B vitamins.
957
REFERENCES Almquist, H. J., 1954. The phosphorus requirements of young chicks and poults. A review. Poultry Sci. 3 3 : 936-944. Anderson, R. L., and T. A. Bancroft, 1952. Statistical Theory in Research. McGraw-Hill Book, Inc., New York. Association of Official Agricultural Chemists, 1955. Official Methods of Analysis (8th ed.) Washington, D.C., p. 842. Bethke, R. M., D. C. Kennard, C. H. Kick and G. Zinzalian, 1928. The calcium-phosphorus relationship in the nutrition of the growing chick. Poultry Sci. 8: 257-265. Briggs, G. M., Jr., T. D. Luckey, C. A. Elevhjem and E. B. Hart, 1944. The effect of ascorbic acid on chick growth when added to purified rations. Proc. Soc. Exp. Biol. Med. 55: 130-134. Carver, J. S., R. J. Evans and J. McGinnis, 1946. Calcium, phosphorus, and vitamin D interrelationships in the nutrition of the growing chick. Poultry Sci. 25: 294-297. Duncan, D. B., 1955. Multiple range and multiple F tests. Biometrics, 11: 1-42. Hart, E. B., H. T. Scott, O. L. Kline and J. G. Halpin, 1930. The calcium-phosphorus ratio in the nutrition of growing chickens. Poultry Sci. 9: 296-306. Lowe, J. T., H. Steenbock and C. H. Krieger, 1939. Cereals and rickets. IX The availability of phytin-P to the chick. Poultry Sci. 18: 40-44. McGinnis, J., L. C. Norris and C. F. Heuser, 1944. Poor utilization of phosphorus in cereals and legumes by chicks for bone development. Poultry Sci. 23: 157-159. March, B., and J. Biely, 1953. The effect of ascorbic acid on the growth rate of chicks. Poultry Sci. 32: 768-774. Pepper, W. F., S. J. Slinger and I. Motzok, 1955. Effect of animal fat on the calcium and phosphorus requirement of chicks. Poultry Sci. 34: 1216. Singsen, E. P., and H. H. Mitchell, 1944. Soybean meal chick rations need no inorganic P supplements. Poultry Sci. 23: 152-153. Snedecor, G. W., 1957. Statistical Methods. The Iowa State College Press, Ames, Iowa. Thornton, P. A., and R. E. Moreng, 1958. The effect of ascorbic acid on egg quality factors. Poultry Sci. 37 : 691-698. Titus, H. W., 1955. The Scientific Feeding of Chickens. The Interstate Printers and Publishers, Danville, Illinois. Wilgus, H. S., Jr., 1931. The quantitative requirements of the growing chick for calcium and phosphorus. Poultry Sci. 10: 107-117.
The authors gratefully acknowledge the technical assistance of Mr. W. F. Pepper of the Department of Poultry Husbandry and Mrs. J. Czarnocki, Miss A. Wilson and Mr. D. Arthur of the Department of Nutrition. Our thanks are also due to Professor E. V. Evans of the Department of Nutrition for assaying the diets of Experiment 1 for pantothenic acid. REFERENCES Association of Official Agricultural Chemists, 1955. Methods of Analysis, 8th Edition. Baldini, J. T., 1960. The effect of dietary deficiency on the energy metabolism of the chick. Poultry Sci. 39:1232. Brambila, S., M. C. Nesheim and F. W. Hill, 1960. Studies of the effect of trypsin on the utilization of a raw soybean oil meal by the chick. Poultry Sci. 39: 1237. Czarnocki, J., I. R. Sibbald and E. V. Evans, 1960. Unpublished procedure. Gray, J. A., and E. V. Evans, 1958. Unpublished procedure. King, E. J., 1932. Colorimetric determination of phosphorus. Biochem. J. 26: 292-297. Nair, K. R., 1948. The studentized form of the ex-
951
treme mean square test in the analysis of variance. Biometrika, 35: 16-31. Skeggs, H. R., and L. D. Wright, 1944. The use of Lactobacillus arabinosus in the microbiological determination of pantothenic acid. J. Biol. Chem. 156: 21-26. Sibbald, I. R., J. D. Summers and S. J. Slinger, 1959. Factors affecting the metabolizable energy content of poultry feeds. Poultry Sci. 38: 1247. Sibbald, I. R., J. D. Summers and S. J. Slinger, 1960a. Factors affecting the metabolizable energy content of poultry feeds. Poultry Sci. 39: 544556. Sibbald, I. R., S. J. Slinger and G. C. Ashton, 1960b. A synergistic relationship between tallow and undegummed soybean oil. Poultry Sci. 39: 1295. Sibbald, I. R., S. J. Slinger and G. C. Ashton, 1961a. Factors affecting the metabolizable energy content of poultry feeds. 2. Variability in the M.E. values attributed to samples of tallow and undegummed soybean oil. Poultry Sci. 40: 303308. Sibbald, I. R., S. J. Slinger and G. C. Ashton, 1961b. Factors affecting the metabolizable energy content of poultry feeds. 3. The influence of Kaolin and Alphacel when used as ration diluents. Poultry Sci. 40: 454-458. Snedecor, G. W., 1956. Statistical Methods, Fifth Edition, Iowa State College Press, Ames, Iowa.
The Relationship of Calcium-Phosphorus Ratios to the Utilization of Plant and Inorganic Phosphorus by the Chick 1 J. M. VANDEPOPULIERE,2 C. B. AMMERMAN3 AND R. H. HARMS4 Florida Agricultural Experiment Stations, Gainesville (Received for publication September 19, 1960)
C
ONSIDERABLE interest has developed in the past 20 years in determining the availability of phosphorus from plant and inorganic sources. In order to 1 Florida Agricultural Experiment Stations Journal Series No. 1120. 2 Present address Ralston Purina Co., St. Louis, Missouri. 3 Department of Animal Science. 4 Department of Poultry Science.
properly evaluate results from such experiments it is necessary to understand some of the factors which may influence utilization of phosphorus such as calcium-phosphorus (Ca:P) ratio, levels of ascorbic acid and fat. Bethke et al. (1928) and Hart et al. (1930) reported that the optimum or near optimum Ca:P ratio for growing chicks was between 2:1 and 4:1. Wilgus (1931) stated
952
J. M. VANDEPOPULIERE, C. B. AMMERMAN AND R. H. HARMS TABLE 1.—Composition of basal diet* Ingredients
White corn meal (degerminated) Soybean oil meal (50% protein) Dehydrated alfalfa meal (17% protein) Micro-ingredients1 Iodized salt Ground limestone
Percent 64.10 31.18 3.00 0.90 0.40 0.42 100.00
* Basal diet contains 0.28% phosphorus and 0.28% calcium. 1 Supplies per pound of feed: 2268 I. U. vitamin A, 10 meg. vitamin Bi 2 , 340 I. C. U. vitamin D 3 , 2 mg. riboflavin, 9 mg. calcium pantothenate, 18 mg. niacin, 261 mg. choline chloride, 140 mg. methionine, 10 mg. Terramycin, 2 mg. Oleandomycin, 57 mg. Santoquin, 9 mg. iron, 0.9 mg. copper, 90 meg. cobalt, 5 mg. iodine, 45 meg. zinc, 80 mg. manganese sulphate, and 35 mg. manganous oxide.
that a 2.5:1 ratio was borderline and 3.3:1 was disastrous. Carver et al. (1946) found that a 1:1 or 2:1 ratio produced the most .favorable growth in Leghorn cockerels. Briggs et al. (1944) reported that ascorbic acid fed to chicks receiving various purified diets promoted small but consistent increases in growth rate. They suggested that chicks fed a highly purified diet were not able to synthesize sufficient ascorbic acid to permit maximum growth. March and Biely (1953) reported a growth stimulus from the addition of vitamin C to a diet of natural ingredients. The work of Thornton and Moreng (1958) indicated that the addition of ascorbic acid to the diet of hens improved calcium utilization. Almquist (1954) suggested that the phosphorus requirement of chicks from one day to four weeks of age was 0.45% from a readily available source. Singsen and Mitchell (1944) reported that 0.45% phosphorus from plant source was adequate. However, McGinnis et al. (1944) reported that 0.58% phosphorus from plant source was required for maximum calcification. It is generally accepted that the total level of dietary phosphorus required will depend on the amount which is derived from plant
source. Lowe et al. (1939) showed that the addition of phytin to a low phosphorus basal diet, thereby increasing the phosphorus content of the basal diet by 50%, resulted in no significant improvement in the calcification of the bones of the chick. This study was designed to determine the influence of different Ca:P ratios on the utilization of phosphorus from plant source, feed grade dicalcium phosphate and soft phosphate. Ascorbic acid and added fat were incorporated as variables to determine their effect on calcium and phosphorus utilization. EXPERIMENTAL PROCEDURE
Day-old Vantress X White Plymouth Rock chicks were used in these experiments; seven males and seven females were used per replication in Experiment 1, and 5 males and 5 females per replication in Experiment 2. Each treatment was replicated 2 times in Experiment 1, and 3 times in Experiment 2. In Experiment 1, phosphorus levels 0.28, 0.44, and 0.58% were compared in Ca:P ratios of 1:1, 2:1, 4:1, and 8:1. Calcium and phosphorus levels were obtained by the addition of feed grade dicalcium phosphate and ground limestone. Each of the above experimental diets was fed with and without the addition of 0.05% vitamin C. In Experiment 2, phosphorus levels of 0.44, 0.58, and 0.72%, produced by the addition of feed grade dicalcium phosphate, were compared in Ca:P ratios of 1.0:1, 1.5:1, and 2.0:1. These comparisons were made with two different diets which contained 960 and 1,060 Calories of productive energy per pound of feed. Soft phosphate was studied at phosphorus levels of 0.58, 0.65, and 0.72% with Ca:P ratios of 1.50: 1, 1.75:1, and 2.00:1. The soft phosphate diets contained 960 Calories of energy per pound of feed. The composition of the basal diet is
PHOSPHORUS UTILIZATION
953
shown in Table 1. This basal diet contained 0.28% calcium and 0.28% phosphorus. In PHOSPHORUS 0.2B_, order to obtain the desired level of protein 0.44 and energy in the various experimental 0.5 8 diets, adjustments were made in amounts of i < corn and soybean oil meal used, and animal fat was added when necessary. Energy and (- 300. protein levels of experimental diets were i o calculated using values of Titus (1955). The chicks were housed in metal bat>• 2 0 0 . Q teries with raised wire floors, and water and 0 (0 feed were supplied ad libitum. The temperature was thermostatically controlled. •/o MORTALITY Birds were individually weighed and feed 35-* 50-** utilization calculated at the end of the 4 75+O90-+** week experiment period. Percent bone ash II 2:1 4:1 8:1 of the right tibia was determined as outCALCIUM:PUO&PUORUS RATIO lined by A.O.A.C. (1955). Statistical analyses of Experiment 1 were made by the least FIG. 1. Body weight of chicks receiving diets squares method according to Anderson and containing various calcium-phosphorus ratios (ExBancroft (1952). Data in Experiment 2 periment 1). were subjected to analyses of variance according to Snedecor (1957) and significant ity on all other treatments was less than differences among treatment means were 5%. . determined according to Duncan's Multiple Growth rate increased in a linear fashion Range Test (1955). with all phosphorus levels as the Ca:P ratio narrowed to 1:1 (Figure 1). With diets RESULTS AND DISCUSSION containing 0.58% phosphorus there was no Experiment 1: During the first week significant difference between the 1:1 and chicks fed the basal diet, containing 0.28% 2:1 ratio; however, all other treatments plant phosphorus with Ca:P ratios of 4:1 were significantly different from the 1:1 and 8:1, developed severe rickets and were ratio at the .01 level of probability. unable to reach the feed and water troughs. The mean adjusted bone ash for ExperiAll chicks on these two treatments were fed ment 1 was 43.7%. It can be seen (Figures and watered for the remainder of the ex- 1 and 2) that the bone ash data paralleled periment in containers which they could the growth data very closely, with the exreach while sitting on the wire mesh floor. ception of the chicks receiving the basal Even with this special handling 75 and diet. The slope of the bone ash line of the 90% mortality was observed with the basal diet was not as steep as that of the chicks receiving diets with 4:1 and 8:1 other diets. Since the slope of all growth ratios, respectively. When the ratio was curves were comparable, it is possible that narrowed to 2:1, the mortality decreased to the phosphorus was available for increased 35%, and only 7% died when the ratio was growth but not adequate to give a marked 1:1. When the phosphorus level was in- increased in bone ash. All treatments excreased to 0.44% with an 8:1 Ca:P ratio cept the 0.58% phosphorus in a Ca:P mortality was approximately 50%. Mortal- ratio of 2:1 differed significantly from the
954
J. M. VANDEPOPULIERE, C. B. AMMERMAN AND R. H. HARMS
50
"It, PHOSPHORUS
n n 0 44 0.58..
•"
\
\
40.
\
\
\
\ \
,0
I
\
\
N
4
N
\
U
N
*»
z
0 a
30.
• • . . .
~~
x " ^
20. |:| 2:| 4:1 C A L C I U M : P H O S P H O R U S RATIO
8:1
FIG. 2. Tibia ash of chicks receiving diets containing various calcium-phosphorus ratios (Experiment 1).
0.58% phosphorus and 1:1 Ca:P ratio. Adjusted bone ash of males was 0.8% less than that of females; this difference was significant at the .01 level of probability. Addition of ascorbic acid to the diet did not affect growth or bone ash. Feed effi-
ciency was not calculated due to excessive mortality. An examination of Figures 1 and 2 indicates that the growth rate was as good a criteria as bone ash to use in determining the phosphorus adequacy of diets. This was especially true when the phosphorus requirement of the chick was being approached. Experiment 2: The range of phosphorus levels and Ca:P ratios used in this experiment was narrowed to meet the requirement of the fast growing broiler type chick. The most rapid gain was obtained with chicks receiving diets containing 0.72% phosphorus (Table 2). This growth rate was significantly greater than that obtained when diets contained 0.58% phosphorus. A further reduction in growth was observed with chicks receiving diets containing 0.44% phosphorus. Only the 1.5:1 ratio produced a significant improvement (P < 0.05) over the 2:1 ratio. The high energy diet (1,060 Cal./lb.) produced a highly significant increase in weight gain over the low energy diet (960 Cal./lb.). There were no significant energy X phosphorus level or energy X ratio interactions, indicating that the phosphorus levels and Ca:P ratios studied were not influenced
TABLE 2.—Four week chick weights when fed diets containing various levels of supplemental dicalcium phosphate in three different calcium-phosphorus ratios (Experiment 2) Ca:P ratio Energy level Calories/lb.
960 1060 Average 960 and 1060 1
Phosphorus Levels %»
0.44 0.58 0.72 0.44 0.58 0.72 0.44 0.58 0.72
1 5-l
1.0:1
2 0"1
M
F
AV.
M
F
AV.
M
F
AV.
432 446 475 409 504 538 420 475 506
Grams 395 375 419 386 418 448 390 396 434
414 410 447 398 461 493 405 436 470
413 478 509 455 512 475 434 495 492
Grams 390 415 431 395 439 441 392 427 436
402 446 470 425 476 458 413 461 464
402 473 472 418 490 470 410 482 471
Grams 364 425 404 377 422 447 370 424 426
383 449 438 398 456 458 390 453 448
This includes 0.28% phosphorus from plant sources.
PHOSPHORUS
955
UTILIZATION
TABLE 3—Feed per unit gain ofchicks when fed various levels of supplemental phosphate from dicalcium phosphate and soft phosphate in several calcium-phosphorus ratios (Experiment 2) Phosphorus source
Phiisphorus 1 /o
Cal. prod. energy/lb. feed
Ca:P ratio 1.0:1
1.5:1
1.69 1.61 1.60
1.60 1.57 1.54
1.63
1.57
1.59 1.53 1.51
1.49 1.47 1.45
1.54
1.47
1.75:1
2.00:1
Av.
1.58 1.59 1.57
1.62 1.59 1.57
1.58
1.59
1.51 1.46 1.47
1.53 1.49 1.48
—
1.48
1.50
E
1.60 1.56 1.55
1.61 1.75 1.57
1.68 1.71 1.55
1.63 1.67 1.56
—
1.57
1.64
1.65
1.62
Grams Dicalcium phosphate
0.44 0.58 0.72
960
Av. Dicalcium phosphate
0.44 0.58 0.72
1060
Av. Soft phosphate
0.58 0.65 0.72
960
Av.
—
This includes 0.28% phosphorus from plant sources.
by the increased energy content of the diet. Feed efficiency of chicks was improved when the level of phosphorus from dicalcium phosphate was increased from 0.44 to 0.58%, and as the Ca:P ratio was widened from 1.0:1 to 1.5:1 (Table 3). Highly significant differences in feed efficiency were observed between the groups receiving the two dietary levels of energy. The magnitude of the change in feed efficiency with increasing calcium levels was similar between the two energy levels. This is not in agreement with the report by Pepper et al. (1955) who found an improvement in feed efficiency when the calcium level was increased from 1.0 to 1.2% in diets containing 5-10% added fat, and no improvement in the absence of the added fat. As in Experiment 1, the males grew more rapidly than the females, and a significant phosphorus level X sex interaction was observed. Examination of the data showed that the male gained more than the female with each added increment of phosphorus, which indicates that the requirement for phosphorus is greater for male than for female.
A direct comparison of growth responses was made (Figure 3) between dicalcium and soft phosphate, at 2 phosphorus levels and 2 Ca:P ratios. Highly significant differences were observed with sex, Ca:P ratio, phosphorus level and phosphorus source. The 1.5:1 Ca:P ratio was superior to the 2:1 ratio, and 0.72% phosphorus produced more rapid gains than did 0.58% phosphorus. Body weight of birds receiving dicalcium phosphate was significantly heavier than birds receiving soft phosphate. There was a greater differential in growth between the phosphorus sources for the male than for the female. Hence, it appears that the soft phosphate, when fed at levels used in this experiment (Table 4), comes nearer to meeting the phosphorus requirement of the female than of the male. The ratio X phosphorus source interaction indicates that the wide ratio was more detrimental to the soft phosphate which had less available phosphorus than it was to the dicalcium phosphate. Since the phosphorus from soft phosphate is less available than that from dicalcium phosphate, the levels fed were well below the requirement level
956
J. M. VANDEPOPULIERE, C. B. AMMERMAN AND R. H. HARMS
5 0 0 ,-
DICALCIUM PHOSPHATE SOFT P H O S P H A T E
a ""••a
Si
2 < a i?400
°/o PUOSPMORUS 0.44 • 0.58 O 0.65 A 0.72 •
h 3 UJ
^ ^ ^ ^ ^ ^ ^ ^
BODY
5
^
"
\
^
^ ^ ^ \ ^
300
1.5:1 C A L C I U M : PHOSPHORUS
2.0:1 RATIO
FIG. 3. Comparison of chick weights when fed diets containing dicalcium and soft phosphate (Experiment 2).
of the chick indicating that the Ca:P ratio is more critical in diets which contain phosphorus levels that are below the chicks requirement. The significant phosphorus level X phosphorus source interaction suggests that the phosphorus requirement using dicalcium phosphate was near 0.58%, whereas there was a marked improvement when the phosphorus level was increased from 0.58 to 0.72% when soft phosphate was used. SUMMARY
Data from two four-week experiments studying various phosphorus sources and
levels at different Ca:P ratios have been presented. Results from these tests indicate: 1. The importance of Ca:P ratio increases as the dietary phosphorus level decreases from the chick's requirement. This may be due either to absolute quantity or decreased availability of the dietary phosphorus. 2. Plant source phosphorus was readily available to support growth when fed at a Ca:P ratio of 1:1, however, it did not allow for increased bone ash as did dicalcium phosphate when the Ca:P ratios were narrowed from 4:1 to 1:1. This would indi-
TABLE 4.—Four week chick weights when fed diets containing various levels of supplemental soft phosphate in three different calcium-phosphorus ratios (Experiment 2) Ca:P ratio Phosphorus level %
0.58 0.65 0.72 1
J
1.75:1
1.50:1
2.00:1
M
F
AV.
M
F
AV.
M
F
AV.
371 404 442
Grams 345 371 379
358 388 410
366 392 434
Grams 319 348 388
342 370 411
290 381 391
Grams 309 321 351
300 351 371
This includes 0.28% phosphorus from plant sources.
PHOSPHORUS UTILIZATION
cate a differential in the requirement for growth and bone ash. 3. Chicks which had bone ash values of 28% did not exhibit symptoms of rickets when fed a diet containing 0.28% plant phosphorus at a Ca:P ratio of 1:1. 4. It is essential to consider both phosphorus levels and Ca:P ratios when determining phosphorus availability, irrespective of the phosphorus source. 5. The phosphorus requirement of the male chicks appears to be higher than that of the female. A phosphorus level of 0.72% from dicalcium phosphate with a Ca:P ratio of 1:1 to 1.5:1 produced maximum growth in these battery trials. 6. Ascorbic acid did not influence growth rate or bone ash of four-week-old chicks. 7. Increasing the energy level from 960 to 1,060 Calories per pound of feed did not significantly influence the calcium or phosphorus utilization. 8. The criterion, growth, appears to be sufficient to determine the phosphorus adequacy of a diet when the level approaches the requirement. 9. Feed efficiency was improved significantly with increased levels of productive energy in the diet; however, feed efficiency on diets containing comparable levels of soft phosphate and dicalcium phosphate were similar. ACKNOWLED GMENTS
The authors wish to thank Dr. W. L. Reynolds for assistance in the statistical analysis of the data; International Mineral & Chemical Corp., Chicago, Illinois for furnishing the dicalcium phosphate; Soft Phosphate Research Institute, Ocala, Florida for furnishing the soft phosphate, which was from their composite sample of soft phosphate from various Florida sources; and Chas. Pfizer, Terre Haute, Indiana, for furnishing the vitamin A and B vitamins.
957
REFERENCES Almquist, H. J., 1954. The phosphorus requirements of young chicks and poults. A review. Poultry Sci. 3 3 : 936-944. Anderson, R. L., and T. A. Bancroft, 1952. Statistical Theory in Research. McGraw-Hill Book, Inc., New York. Association of Official Agricultural Chemists, 1955. Official Methods of Analysis (8th ed.) Washington, D.C., p. 842. Bethke, R. M., D. C. Kennard, C. H. Kick and G. Zinzalian, 1928. The calcium-phosphorus relationship in the nutrition of the growing chick. Poultry Sci. 8: 257-265. Briggs, G. M., Jr., T. D. Luckey, C. A. Elevhjem and E. B. Hart, 1944. The effect of ascorbic acid on chick growth when added to purified rations. Proc. Soc. Exp. Biol. Med. 55: 130-134. Carver, J. S., R. J. Evans and J. McGinnis, 1946. Calcium, phosphorus, and vitamin D interrelationships in the nutrition of the growing chick. Poultry Sci. 25: 294-297. Duncan, D. B., 1955. Multiple range and multiple F tests. Biometrics, 11: 1-42. Hart, E. B., H. T. Scott, O. L. Kline and J. G. Halpin, 1930. The calcium-phosphorus ratio in the nutrition of growing chickens. Poultry Sci. 9: 296-306. Lowe, J. T., H. Steenbock and C. H. Krieger, 1939. Cereals and rickets. IX The availability of phytin-P to the chick. Poultry Sci. 18: 40-44. McGinnis, J., L. C. Norris and C. F. Heuser, 1944. Poor utilization of phosphorus in cereals and legumes by chicks for bone development. Poultry Sci. 23: 157-159. March, B., and J. Biely, 1953. The effect of ascorbic acid on the growth rate of chicks. Poultry Sci. 32: 768-774. Pepper, W. F., S. J. Slinger and I. Motzok, 1955. Effect of animal fat on the calcium and phosphorus requirement of chicks. Poultry Sci. 34: 1216. Singsen, E. P., and H. H. Mitchell, 1944. Soybean meal chick rations need no inorganic P supplements. Poultry Sci. 23: 152-153. Snedecor, G. W., 1957. Statistical Methods. The Iowa State College Press, Ames, Iowa. Thornton, P. A., and R. E. Moreng, 1958. The effect of ascorbic acid on egg quality factors. Poultry Sci. 37 : 691-698. Titus, H. W., 1955. The Scientific Feeding of Chickens. The Interstate Printers and Publishers, Danville, Illinois. Wilgus, H. S., Jr., 1931. The quantitative requirements of the growing chick for calcium and phosphorus. Poultry Sci. 10: 107-117.