INVESTIGATION OF THE IMPORTANCE OF PROTEIN SOURCE AND PROTEIN LEVEL IN DAIRY CALF STARTERS Thesis for the Degree of M. .S. MICHIGAN STATE UNIVERSITY DELBERT KENT NELSON 1964 THESIS LIBRAR Y Michigan State Universnty (J ABSTRACT INVESTIGATION OF THE IMPORTANCE OF PROTEIN SOURCE AND PROTEIN LEVEL IN DAIRY CALF STARTERS by Delbert Kent Nelson Seventy-two dairy calves were used to investigate the importance of protein source in calf starters at three different levels of crude protein. All calves were removed from their dams and placed on experiment between two and four days of age. A stepwise weaning program was employed whereby each calf was weaned by three weeks of age and none received more than 125 pounds of‘milk. The experiment was conducted for 8# days. All starter rations were pelleted and varied thy with regard to level and source of'protein. Four 13% crude protein rations were fed in Experiment I. The various nitrogen sources of the respective rations were urea. soybean oil meal. fish meal and a combination of soybean oil meal and fish meal. Two levels of crude protein were fed in Experiment II. At 17.5% crude protein the main nitrogen sources were urea. soybean oil meal. and fish meal. and at 22% crude protein the main nitrogen sources were soybean oil meal and fish meal. The evaluating criteria for Experiments I and II were dain gain. wither height increase. heart girth increase. milk consumption. 2 Delbert Kent Nelson starter intake. and feed conversion. Analysis of variance of each criterion at each crude protein level showed no significant differ- ence among the protein source groups. Protein level effects were exposed by combining the data of the two experiments and perfbrming analysis of variance on a factorial arrangement. This analysis indicated that daily gain, wither height, and feed conversion were improved by protein level, while heart girth, milk consumption, and starter intake were not. Observations of blood composition of the calves in Experiment II show that at 17.5% crude protein urea caused significantly higher plasma urea nitrogen than did soybean oil meal or fish meal. Plasma protein levels were significantly greater for the fish meal group than for the soybean oil meal group. A digestion trial was conducted in conjunction with Experiment II. Analysis of the data from three collection periods indicated that dry matter digestibility was significantly greater for the soy. bean oil meal than for the fish meal. Nitrogen digestibility sig- nificantly improved with crude protein level. while nonsignificant trends of greater dry matter digestibility. energy digestibility. and nitrogen retention were observed as crude protein level increased. INVESTIGATION OF THE IMPORTANCE OF PROTEIN SOURCE AND PROTEIN LEVEIIIN DAIRY CALF STARTERS By Delbert Kent Nelson A THESIS Submitted to Michigan State University in.partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Impartment of Dairy Science 1964 ACKNOWLEDGMENTS The author wishes to thank Dr. L. D. Brown for his council and guidance during the duration of this experiment. Gratitude is also expressed to Ralph Ried for his loyal care of experimental animals. The author also deeply appreciates the help and.moral support given by his wife during the preparation of this thesis. ii TABLE OF CONTENTS Page INTRODUCTION 1 REVIEW OF LITERATURE 2 Protein Levels for Growing Swine 3 Protein Quality and weanling Swine 4 Amino Acid Supplementation of WBanling Swine Rations 6 Importance of Protein Level in Ruminant Rations 13 Protein Quality in Ruminant Rations 17 The Effect of Protein Nutrition on Blood Urea 22 Effect of Diet on Plasma or Serum Protein 24 EXPERIMENTAL PROCEDURE 26 Assignment, Management, and Feeding 26 Preparation and Composition of Rations 27 Procurement of PerfOrmance Data 28 Analytical Procedures 31 Statistical Treatment of the Data 33 RESULTS AND DISCUSSION 37 Experiment I 3? Experiment II 40 Growth. feed consumption, and feed conversion results #0 Digestion trial results 49 Blood plasma studies 61 iii Page sum 69 1319:10um 71 APPENDIX 81 iv Table 1. Table 2. Table 3. Table 4. Table 5. Table 6. Table 7. Table 8. Table 9. Table 10. Table 11. Table 12. LIST OF TABLES Effect of source of protein on mean growth, feed consumption. and feed conversion at the 13% crude protein level Effect of source of protein on mean growth. feed consumption and feed conversion at the 17.5% crude protein level Effect of source of protein on.mean growth. feed consumption and feed conversion at the 22% crude protein.level Mean daily weight gains at 13% and 17.5% crude protein Mean daily weight gains at 13%. 17.5%. and 22% crude protein Mean increase in wither height (in.) at 13% and 17.5% crude protein Average increase in wither height (in.) at 13%. 17.5%. and 22% crude protein Average values for increase in heart girth. milk consumption. and starter intake for each crude protein.1evel Mean feed conversion (lb. of starter per lb. of gain) at 13% and 17.5% crude protein Mean feed conversion (1b. starter per lb. gain) at 13%. 17.5% and 22% crude protein Mean coefficients of apparent dry matter digestibility at 17.5% crude protein Mean coefficients of apparent digestibility of dry matter at 17.5% and 22% crude protein Page 1+1 #2 43 47 50 51 Table Table Table Table Table Table Table Table Table Table Table Table Table 130 1A. 15. 16. 17. 18. 19. 20. 21. 22. 23. 2h. 25. Mean coefficients of apparent digestibility of energy at 17.5% crude protein Nean coefficients of apparent digestibility of energy at 17.5% and 22% crude protein Mean nitrogen digestibility coefficients at 17.5% crude protein Mean nitrogen digestibility coefficients at 17.5% and 22% crude protein Mean nitrogen retention as percent of nitrogen intake at 17.5% crude protein Mean nitrogen retention as percent of nitrogen intake at 17.5% and 22% crude protein Mean grams of nitrogen retained per day per hundred pounds of body weight at 17.5% crude protein Mean grams of nitrogen retained per day per hundred pounds of body weight at 17.5% and 22% crude protein Average grams of dry matter consumed per day during collection Kean plasma urea nitrogen (mg %) at 17.5% and 22% crude protein Mean plasma.urea nitrogen (mg %) at 17.5% crude protein Mean plasma protein (grams %) of calves fed 17.5% crude protein rations Mean.plasma protein (grams %) at 17.5% and 22% crude protein Page 52 53 54 55 56 58 59 61 62 65 66 Figure 1. Figure 2. IIST OF FIGURES Arrangement of the combined and factorialized starter groups for analysis of growth data Arrangement of the blood composition data for factorial analysis vii Page 35 36 Table 26. Table 27. Table 28. Table 29. Table 30. IIST 0F APPENDIX TABLES Composition of calf starters Proximate analysis of calf starters Growth perfOrmance criteria of calves in Experiment I and Experiment II Apparent coefficients of digestion and nitrogen retention values Plasma protein and plasma urea nitrogen content of the blood for calves in Experiment II 9.1 Page 82 81+ 85 90 9A INTRODUCTION Raising dairy calves for herd replacements is one of the dairy- man's most costly and time consuming operations, and feeding is the most expensive phase of this operation. In order to reduce the cost of feeding baby calves, dairymen have replaced milk in the diet with a less expensive milk substitute or removed milk from the calves' diet at an early age. Early weaning. however. places a hardship on the calf since the calf is dependent upon the milk for the quantity and quality of’protein which it contains. In order to alleviate this hardship, calf starters containing high quality protein and having the optimum crude protein level may be needed in the diet of’these young calves. The primary purpose of these trials was to determine whether or not protein source was an important factor in calf starters fed to early weaned calves. various protein levels were also examined in conjunction with protein source. since protein level and protein quality are not completely independent of each other. REVIEW OF LITERATURE Exact protein requirements of the dairy calf are complicated by the dynamic character of the calf's digestive development. At birth a calf is not actually a ruminant, for the rumen has developed little. Instead, its digestive process resembles that of the simple stomached animals and its protein requirements are similar to those of the dog or pig (Savage and McCay. 1942; Morrison. 1956). From birth the calf undergoes a progressive transition from a simple stomached animal to a ruminant. and reaches the latter status at about one month of age (Lengemann gtflgl.. 1959; Bryant and Small, 1960; Godfrey, 1961). This age is dependent. of course. upon the particular ration fed during the development period (Wardrop and Coombe. 1961; Church gt'gl.. 1962). Because of’the lack of'infbrmation about the protein level and protein quality requirement during the "simple stomached" and transitional stages, it is necessary to review experimental data concerning the protein requirements of the pig:which may be applied to the very young calf. Protein level values for growing swine are not completely applicable to the young calf. However. the work in this area should be briefly examined in order to lay the fOundation fer protein quality discussion. Protein nggl§_fgg Growing Swing Becker 32.3l, (1963) has outlined the protein requirements for the pig at different stages of growth. Expressed as percent of the ration, these requirements are: suckling (5 to 30 pounds). 20%: weaned (10 to 30 pounds). 22%; grower (30 to 100 pounds). 16%: and finisher (100 to 200 pounds). 12%. The recommendation of 22% for weaned pigs (10 to 30 pounds) is high compared.to the results of Jensen gtflgl. (1957) who found 17% crude protein to be the optimum for weaned pigs between the age of 1A to 56 days or 8 to 40 pounds. The work of Rutledge gt El. (1961) also indicates that a lower crude protein level met the requirements of’pigs‘between the age of 3 weeks and 8 weeks or 12 and 45 pounds. He found that a 16% ration supported growth as well as 20%. 2b% or 28% crude protein rations. Aunan.§t|§l. (1961). using a different approach. weaned pigs at 3 weeks and fed.them a 20% crudejprotein ration up to 8 weeks of age (approximately A0 pounds). Hereafter they fed a 17% and a 15% crude protein ration.unt11 the pigs reached 120 to 130 pounds at which time they decreased the protein level to 15% and 11%. respectively. Gains and efficiencies between the groups were not significantly different. Using the same technique they compared 16%. 1A% and 12% crude protein rations. The 12% ration was the only one that was inadequate up to 125 pounds of body weight. Thereafter. it yielded normal gains. This Observation agrees with the recommendation for finishing swine by Becker gt a}. (1963). Abernathy gt a}. (1958), canparing the growth of pigs from 40 to 110 pounds. found faster gains resulted from feeding an 18% crude protein ration rather then a 14% ration. However. Kennington gt a}; (1958) found no difference between a 14% and a 20% crude protein ration fed to 30 pound pigs. The results reviewed thus far indicate that the optimum crude protein level must be near 14% for the growing pig. The variation in performance of animals among experiments may be partially explained on the basis of protein quality. Unless the protein in the ration is of highest quality. or in other words contains the essential amino acids in the exact proportions, minimum requirements for protein cannot be established (Lucas and Lodge. 1961; Becker gt; 3}}. 1963). Protein Quality and Weanling m Becker gt 3.1. (1954) clearly demonstrated the importance of protein quality by comparing a corn-soybean oil meal ration with a corn-menhaden fish meal ration. Both rations were fed to weaner pigs between 40 and 100 pounds. At 18% crude protein the rations were equal with regard to gain and feed conversion efficiency. But the minimum requirement of crude protein for satisfactory performance was 14% for the corn-soybean oil meal ration and 16% for the corn- fish meal ration. These results indicate that soybean meal is of higher quality than menhaden fish meal. Using white fish rather than menhaden. Smith and Lucas (1957) found that a 15% ration supplemented with fish meal produced 13% more gain and was 14% more efficient than a 17% ration supplemented with extracted decorticated groundnut meal. Evans (1962a) reported that a 14.08% crude protein ration containing white fish meal was superior in gain and efficiency to a 12.35% crude protein ration containing extracted soybean oil meal. Additionally. extracted decorticated groundnut meal was found to be inferior to extracted soybean meal when both were added to approximately 12.5% crude protein rations. At 18% crude protein. white fish meal rations have proved to be superior in regard to nitrogen retention when compared to groundnut meal rations with supplemental lysine (Jones gt _al_.. 1960; 1962). ‘Kifer and Young (1961) fed corn-soybean oil meal rations contain- ing 14%. 15% or 16% crude protein. At each level a portion of the soybean oil meal was replaced by menhaden fish meal. However. there were no significant differences between levels or sources. The author concludes that the 14% crude protein ration. with or without fish meal, was in excess of the pig's requirement. Using very young weaners from 10 through 25 pounds. Blair (1961) fed rations supplenented with white fish meal or soybean oil meal at the crude protein levels of 28%. 23% and 18%. White fish meal supplementation at 23% crude protein significantly increased gflns over the 18% fish meal and the 18% or 23% soybean oil meal rations. The 28% rations showed no superior performance over the 23% rations and in the case of soybean oil meal proved to be detrimental. A definite level-quality interaction is indicated here. The results thus far are inconsistent and inconclusive. therefore a further look into protein quality or more specifically amino acid supplementation is necessary. M 5313 Supplementation g; Weanling _S_wi_§g Rations Becker gt_ gl__. (1963) states that corn plus fish meal or meat and bone scrap is less effective than corn plus soybean oil meal. However. additions of tryptophan to the former rations will yield gains superior to those of the corn-soybean oil meal rations. The results of work by Terrill 53-: 11, (1954) are in complete agreement with this. At an 18% crude protein level. corn supplemented with meat and bone scrap was inferior to corn supplemented with soybean oil meal. but superior in the case where 0.1% tryptophan was added to the meat and bone scrap. Similarly. Miner gt 3;. (1955) reported no improvement in gain by adding fish solubles or tryptophan alone to a corn-cottonseed ration. but when these supplements were added in combination gains were significanthr improved. Henson gt gl_. (1954) also found tryptophan to be limiting in corn. meat—byoproduct rations for growing swine. and postulated the require- ment of tryptophan to be greater than 0.1% of the diet. In ayeement with this Shelton gt gl_. (1951b) had found that 0.2% Dip-tryptophan gave a higher growth response than a ration containing 0.1% DI.- tryptophan. The levels of 0.132% (Meade. 1956) and 0.137% tryptophan (Meade and Teter. 1956) were both found to be adequate in 15.9% and 14.2% crude protein rations. respectively. Becker gt E}: (1954) determined that 0.13% tryptophan was adequate. Later Becker gt 51; (1955a) did an extensive study on the tryptophan requirement of the young pig and found it to be 0.115% in a diet containing 15.3% crude protein. and postulated this value to be very near the minimum requirement. This value is comparable to the recommendation for the rat. which is 0.11% tryptophan (Rama Rao gt gl_.. 1959). Amino acid supplementation of a corn-tankage ration by Pfander and Tribble (1957) resulted in satisfactory gains n‘om 0.11% tryptophan in the diet. However. superior performance resulted from adding a canbination of amino acids to the diet. No response was obtained from separate additions of tryptophan. methionine or lysine. Similar- ly. Clawson and Matrone (1963) found no response from additions of tryptophan. lysine or isoleucine separately to an 8% crude protein corn-soybean oil meal ration. but the combination of all three significantly improved performance. In contrast. Pfander and Tribble (1955). feeding a corn-soybean oil meal ration at 18%. 16% and 14% crude protein. found that L-lysine. DL-methionine or DL-tryptophan supplementation in m combination failed to yield gains equal to that of L-lysine additions alone. This indicates that lysine may be deficient in rations of this type. Agreeing with this. Chance gt g_1_. (1960) found lysine to be the most limiting amino acid in a 12% protein corn-soybean oil meal ration. Yet Becker gt: _a_l_. (1963) claims that soybean oil meal is an cutstand. ing source of supplementary amino acids for growing swine when fed with corn at a 16% crude protein level. In agreement with both Chance g3 gl_. (1960) and Becker gt gl_. (1963). Acker gt gl_. (1959) found that 23 pound pigs responded to lysine supplementation when fed a 12% crude protein ration. but not when fed a 14% crude protein ration. This indicates that the 12% ration was lysine deficient. but the 14% ration was not. Catron g_t_ gl_. (1953) reported a similar response with pigs of the same age. Further information which indicates that lysine additions are important at low levels of crude protein. but not at high levels in corn-soybean oil meal rations. is provided by Pond g1; gl_. (1953) and Neilsen gt 2}: (1959). Hagruder g3 g1_. (1961) observed similar results when lysine was added to corn-cottonseed-dried whey rations. In this particular case a lysine supplemented 12.5% crude protein ration produced gains equal to an unsupplemented 14% crude protein ration and did it more efficiently. lysine supplementation had previously been shown to improve corn-cottonseed rations (Miner g3 31-." 1955). Using wheat and barley. plus soybean oil meal. Bowland (1962) promoted gains by lysine supplementation of a 13% crude protein ration. These gains were equal to those produced by a 16% ration which had no additional lysine. Similarly. Jones gt g. (1962) found that a 12% crude protein barley-groundxmt meal ration. supplemented with 0.2% L-lysine monohydrochloride. produced gains equal to an 18% crude protein ration without added lysine. Schnarre and Tribble (1962). unlike the two previous groups. noticed a depression in gain of pigs from 30 to 125 pounds in body weight when 0.1% lysine was added to 20%. 16% and 12% crude protein corn-soybean oil meal rations. They blamed this depression on the already existing amino acid imbalance. Henson gt gl_. (1954) noticed no beneficial result from additions of lysine to a 1h. 5% crude protein corn-meat-by-product ration. They concluded that the basal rations. which alreachr contained as little as 0. 63% lysine. were not deficient in this amino acid. The 0. 63% level is higher than that recommended by Hutchinson gt gl. (1957) who found 0.52% lysine to be adequate for weaning pigs fed a 11.69% crude protein ration. Germann gt gt. (1958). feeding a 12.9% crude protein and a 13.14% crude protein ration to 6 week old pigs. found the minimum requirement of lysine to be near 0.6% of the diet or 11.7% of the protein portion. This is in close agreement with Becker gt gt. (1951+) who noticed that 0.63% lysine in the diet or 1+. 5% in the protein supported growth of pigs between ’40 and 100 pounds. Satisfactory nitrogen retention was realized when Meade and Teter (1956) fed a 11+. 2% crude protein corn-soybean oil meal ration which contained 0.62% lysine. At a slightly higher crude protein level of 15.9%. 0.69% lysine appeared to be adequate (Meade. 1956). This is comparable with the recommendation of 0.70% to 0.90% lysine in a 16% crude protein ration for weaning pigs. by Mitchell gt gt. (1962). 10 Jones gt gl_. (1962). feeding an 18% crude protein ration to weaners. obtained maximum gains and efficiency when lysine levels were near 1.0%. The lysine requirement. when expressed as percent of the ration. appears to increase as percent of crude protein increases (Pfander and Tribble. 1957). Further evidence of this is given by Chance gt gl_. (1958). They found that the requirement of lysine increased from 0.7% at 10% and 15% crude protein to 0.9% at 20% crude protein. McWard g 31; (1959). when feeding send-purified diets to 30 pigs. concluded that the lysine requirement for a 12.78% and a 21.71% crude protein ration was 0.71% and 0.95% respectively. When expressed as a percent of the crude protein. the respective values were 5.55% and 1+. 38%. This agrees with Beoker gt gt. (1963) who points out that the requirement of lysine in the protein decreases as the protein portion of the ration increases. Some work has been done with lysine supplementation in combination with methionine. Brooks and Thomas (1959) noticed improved gains when lysine or lysine plus methionine was added to corn-peanut oil meal rations. Where 0.15% L-lysine had no affect on corn-soybean oil meal rations Pfander and Tribble (1953) improved gains and feed conversion efficiencies by adding 0.014% methionine with the lysine. Dyer gt gt. (1952) noticed the same effect when lysine and methionine were added to a corn-cottonseed ration while additions of L-lysine alone produced only satisfactory gains. other experiments have shown the benefit from adding lysine and methionine in combination 11 to low protein diets (Evans. 1961. 1962a). In a detailed study. Evans (1960) provided evidence that lysine and methionine each have their optimum values for different stages of growth. Berry gt gl_. (1962). unlike the previous researchers. noticed no benefit when adding lysine and methionine to a low protein soybean oil meal supplemented diet. However. methionine additions alone proved to be very beneficial. Their only explanation was that methionine is the most limiting amino acid in soybean protein. Becker gt gl_. (1963) agrees with this. but considers it to be only slightly lacking in corn-soybean oil meal rations. Benefits from methionine supplementation of a 16% crude protein corn-soybean oil meal ration have been observed by Long gt gl_. (1962). As percent methionine increased. feed intake. gains. dry matter digestion and nitrogen digestion all showed significant improvement. 0n the other hand. there is much work indicating little or no benefit from additions of methionine to corn-soybean oil meal rations at low crude protein levels (Catron gt gl_. . 1953; Sewel and Keen. 1958; Acker gt g1_.. 1959) or at high protein levels (Maner gt 2}.” 1961). It has also failed to improve groundnut meal protein (Jones gt gl_.. 1962; Jones gt gl_.. 1960). USing purified diets. Becker gt gt. (1955b) calculated the methionine-cystine requirement of a 12.6% crude protein ration to be 0.112% of the diet or 3. 33% of the protein. Kroening gt gl_. (1961) 12 also obtained.maximum performance with these values at 12% protein. Feeding a 16% crude protein ration. Pfander and Tribble (1955) found that 3.5% methionine.cystine in the protein or 0.56% in the diet met the requirements of the weanling pig. An unusually low level of 0.27% methionine in the diet satisfied requirements in a 15.9% crude protein corn-soybean oil meal ration (Meade. 1956). Becker gt_g;, (1954) also found very low levels would satisfy the weaning pigs needs. These values were 0.23% of’the diet or 1.65% of the protein. The only possible explanation for these last two low requirement values is that neither group considered the cystine content of the diet. At the high level of 21% crude protein Shelton‘gt gt, (1951a) calculated the methionine-cystine requirement to be 0.6% of the diet. This high level is acceptable because the methionine requirement. when expressed as percent of the diet. increases with the crude protein percent (Lucas and Lodge. 1961). One of the less popular but important amino acids is isoleucine. Meade and Teter (1956) found it to be limiting in a 12.1% crude protein corn-soybean oil meal ration. However. the same ration at 1#.2% crude protein.provided 0.63% isoleucine which satisfied require- ments for ample nitrogen retention. This value closely agrees with the value of’0.60% given by Evans (1962b) which is a minimum level for maximum growth. 13 Studying two levels of protein. 13. 35% and 26.7%. Becker gt gJ_._. (1957) found isoleucine requirements to be 0.46% and 0.65% of the respective diets or 3.11% and 2.14% of the protein. respectively. This and other trials point out that the isoleusine requirement. when expressed as percent of the diet. increases with crude protein levels. But when expressed as percent of the protein. decreases as crude protein levels increase (McWard gt 2‘2.” 1959: Becker gt gl_.. 1963; Lucas and Lodge. 1961). Iygortance g1: Protein tgzg; ta Ruminant Rations Compared to the extensive and detailed studies of swine protein nutrition. the protein data for ruminants are very scarce. This is especially true for the immature ruminant (Morrison. 1956). Brown gt gl_. (1958) conducted a study with Jersey and Holstein calves during the period from two days of age to 86 days of age. Milk was fed at the rate of 8% of body weight for the first three weeks; thereafter it was fed at the rate of 6%. 5%. 3% and 2% for the fourth. fifth. sixth and seventh week. respectively. Starter intake was limited to four pounds for the Jerseys and five pounds for the Holsteins. Alfalfa-brome hay was fed ad libitum. All starters contained the same ingredients. with the protein concentrate varied in each to give the desired crude protein levels of 24. 3%. 20.2%. 16.6% and 12.2%. There were no significant differences in average daily gain. increase in height or heart girth and starter intake. But with regard to efficiency. the 16.2% crude protein 14 rations were superior to 24.3% or 20.2% crude protein rations. The digestibility data showed no significant differences in average daily nitrogen retention. However. nitrogen digestion was lowest for the 12.2% crude protein ration. In a similar second trial they compared a wider range of crude protein levels. These were 23.7%. 20.0%. 16.2%. 13.0% and 8.5%. Results proved the 16.2% crude protein ration to be superior with regard to gain and efficiency. and the 8.5% ration was inferior with regard to the same criteria. The 8. 5% crude protein ration also resulted in the reduced nitrogen retention and had the lowest crude protein digestion coefficient. General observations of both digestion trials were that dry matter and crude protein digestion coefficients decreased with age. while nitrogen retentions increased. Using the same hay and milk feeding practices of Brown gt gt. (1958). Everett gt g_l_. (1958) compared protein levels of 6.3%. 8.6%. 10.1%. 12.2% and 14.2%. Up to six weeks of age the protein percent of the rations had no effect. due to the amount of milk common to all diets. Hereafter. however. an increase in consumption. gain. and efficiency was noticed for each increase in protein content of the diet. Digestion data showed that crude protein digestibility increased with the protein content of the diet and that calves on the 1#.2% crude protein ration retained significantly more nitrogen than those on the 6. 3% crude protein ration. 15 .More recently. Brown and Lassiter (1962) using Holstein and Guernsey calves compared 14%. 16% and 18% crude protein rations. In this trial milk was fed fer #0 days to total 228 pounds for the Holsteins and 169 pounds for the Guernseys. Alfalfa hay was added to the grain ration and the entire mixture was pelleted. The results of growth studies indicated no difference in body weight gains. how. ever. the 1w% crude protein ration was more efficient than the other rations. In the previously cited studies milk was fed during a.major portion of the trials which probably influenced the amount of protein required in the grain ration. Whitelaw gt gl_. (1961a) removed some of the influence of milk by weaning their calves at three weeks of age. However. they did not begin their trial until the calves reached 11 weeks of age. The experiment lasted only 10 days. five of which the calves were on digestion trials. Using ten.parts of corn-oats-ground barleyegroundnut meal and one part dried hay. they fermulated rations to contain 1#.9%. 16.9%. 19.14% and 21.4% crude protein on a dry matter basis. These rations were fed at the rate of 8% of‘metabolic weight. Dry matter and nitrogen digestibility increased with increasing levels of dietary protein. In the nitrogen balance studies the 19.9%1and the 21.6% crude protein rations yielded greater nitrogen retention than the 14.9% or 16.9% crude protein rations. When nitrogen retention was expressed as percent of dietary protein. there were no significant 16 differences even though the values tended to be lowest at the higher levels of protein intake. In a second trial 11.6%. 18.8%. 20.4% and 22.6% crude protein levels were compared. Again nitrogen retention was maximum for the two high groups. and digestibility of dry matter and nitrogen increased with protein level. Data from both trials indicate the 20.4% or 19.4% crude protein on a dry matter basis is the minimum for maximum nitrogen retention. When 19.4% crude protein is converted to an air dry basis it becomes 16.3% crude protein. which agrees very closely with the minimum values given by the previous workers. Results of protein research with sheep are comparable to the results given thus far for dairy calves. Hinds gt gt. (1961) found that a 11.6% crude protein ration produced better gains than did a 15.62% or 19.62% crude protein ration. However. this trial was conducted after lambs were weaned at nine weeks of age. Jones and Hogue ( 1960) compared two protein levels for older lambs weighing about 70 pounds. The 11.2% digestible protein ration. on a dry matter basis. resulted in greater feed consumption and greater gains than the 8.4% digestible protein rations. Using ewe lambs from 70 to 100 pounds Griffith gt gt. (1959) found a 12.7% crude protein ration to be superior to a 10.9% crude protein ration and equally as effective as a 14.7% or 16.0% crude protein ration. 17 Recent research regarding protein level for mature ruminants is minimal. Lassiter gt g. (1957) found that 10.3% or 11.9% crude protein met the requirements for the milk production. body weight and nitrogen retention of lactating dairy cows. Hale gt g_l_. (1959) found similar>minimum values when an 11.5% crude protein ration yielded 15% more gain than an 8.6% crude protein ration fed to beef cattle. Although there is some variation among experiments. the trend toward lower protein requirements as the ruminant developes is clearly displayed. Protein gtality tt.Ruminant Rations Where crude protein requirements are highest for young calves. protein quality requirements are also expected to be the greatest (Morrison. 1956). Halter (1956) compared two 16% calf rations; one with soybean oil meal as the protein source and one with linseed oil meal. soybean oil meal. alfalfa meal. dried skimmilk. and dried distillers soluable as the protein source. No difference in gains of the calves fed the two rations was observed. The results may have been influenced.by feeding 350 pounds of milk. Large amounts of milk were also fed by Lambert gt 9;. (1955) and Hibbs gt p_l_. ( 1953) and neither group noticed any difference between a simple or complex starter. Hurley 25.§i9 (1958). however. limited milk feeding through 35 days of age and achieved no extra weight or height increase from 18 feeding a complex starter. Weaning the calves at 24 days of age. Pardue and Jacobson (1961) found no significant difference in daily gain. height or circumference when comparing 16.5% rations which contained either soybean oil meal or dried skim milk as the protein source. Pardue gt gl_. (1962) in a similar trial replaced some of the soybean oil meal with dried skim milk. but no improvement of growth was noted. However. the dried skim milk ration did increase dry matter digestibility. but had no affect on digestible crude protein or digestible energ. It was also noticed that nitrogen retention for all groups was higher at 12 than at 8 weeks. Preston gt gt. (1960) fed a milk replacer to calves until 20 days of age. and canpared a corn—oats—groundnut meal ration to a corn-oats-groundnut meal-fish meal ration. In the simple ration the groundnut meal provided 50% of the total nitrogen. whereas. in the second ration 19% of the groundnut meal nitrogen was replaced by fish meal nitrogen. The youndnut plus fish meal ration was superior in efficiency and wither height increase. Benefits from fish meal supplementation were also noted by Whitelaw gt gl_. (1961b) when fed to six 80-day old calves in a 3 x 3 latin square design. The protein concentrates added to the corn-oats- grass meal ration were: groundnut meal. heated groundrmt meal. and peruvian fish meal. All rations were near 16.5% crude protein. No difference in dry matter digestibility occurred. but the live weight 19 gain and nitrogen retention were highest for the fish meal and poorest for the groundnut meal. Variation in plant protein sources appears to have little effect upon the value of calf starters. Loosli EEMEL' (1952). however. illustrated that differences exist in efficiency of utilization of plant protein concentrates when fed to dairy cows. Brundage and Sweetman (1963) compared a commercial ration against a simple grain mix with plant and animal protein sources added. The former yielded more milk while the latter resulted in greater body weight gains. Loosli (1956) found no difference in fat corrected.milk production between rations containing either urea or corn distiller dried grains as a nitrogen source. No significant differences in milk production or weight gains were noticed when Loosli gt'gt. (1963) compared simple rations to progressively complex rations. or to rations in which urea.made up a portion of‘the nitrogen. Some work has been done with urea additions to calf starters. Reid (1953) claims that calves as young as two.months of age can utilize some urea. Loosli and McCay (1943) conducted some of the initial research with urea in calf rations. Whole milk was fed from birth to 2 months and then gradually decreased.until 4'months when it was removed. The low ration diet was 4.4% crude protein. The high diet was the same 20 ration with enough urea added to make a 16.2% crude protein equivalent ration. All calves gained at the same rate until 2 months of age. Hereafter. the low'protein group gained at a decreasing rate and practically ceased to grow approximately two weeks after milk feeding ceased. 0n the other hand. at feur months of age the calves on the urea ration were 75 to 90% of normal weight. Brown gt_gt, (1956) conducted a similar trial. but reduced.milk feeding after 21 days and weaned all calves at 47 days of age. Two conventional rations were formulated; one at 6.7% crude protein and one at 15.2% crude protein. A second 15.2% crude protein ration was formulated such that 54.2% of the ration nitrogen was supplied by urea. There were no differences between the two 15.2% crude protein rations with regard to gain. height. circumference. feed efficieney. nitrogen retention. and coefficients for digestibility of dry matter or crude protein. The 6.7% crude protein ration was inferior in all the criteria.mentioned. In a more recent trial Brown gt git. (1960) compared corn-oat- starch-urea.rations which had crude protein equivalent levels of 6.5%. 9.4%. 12.1% and 15.3%. Milk was fed at the rate of 8% of body weight for the first three weeks and reduced two percentage points each week thereafter ceasing at 6 weeks of age. All calves perfOrmed equally up to 3 weeks of age; thereafter gains. height. circumference and feed intake all increased with protein percent up to 12.1%. No significant difference existed 21 between the 12.1% group and the 15.3% group. Generally. crude protein and dry matter digestion and nitrogen retention increased with crude protein percent. Using growing Guernsey heifers which weighed more than 300 pounds. Campbell g§|§l3 (1963) found no difference in gain when urea replaced soybean oil meal in a corn-soybean oil meal-alfalfa meal ration. The use of urea in mature ruminant feeding is a common practice. In some cases. methionine has been added to improve the quality of urea containing rations. Noble £t_al, (1955) added methionine to 11% crude protein sheep ration which contained urea or soybean oil meal as the nitrogen source. The addition of‘methionine improved gains in both cases. however. these were significant in only the soybean oil meal group. Gallup 22,3}, (1952) reported small nonsignificant improvement in nitrogen utilization.when methionine was added to awurea containing ration fed to feeder lambs. Feeding ewe lambs from 70 to 100 pounds in body weight Griffith 33,3l. (1959) found that a 10.9% crude protein barleyooats-sqybean oil meal ration was inadequate compared to 12.7%. 14.7% and 16.0% crude protein ration. However. it was superior to all when supple- mented with EL-methionine. Contrary to this Hinds gt_§l. (1961) observed no improvement from adding methionine to sheep rations. However. slight improvement in gain and efficiency was noticed when 22 lysine was added in combination with methionine. Gossett gt 39:0 (1962) observed no improvement in gain when adding lysine and methionine to protein supplements for beef steers. but improvements in efficiency were noticed. They also promced this same situation when 5 or 10 gm. of lysine per day were added to a 64% crude protein concentrate. Feeding 10 an. of L-lysine l'wdrochloride improved efficiency when Hale _e_t_ 13;. (1959) added it to an 8.6% crude protein beef ration. However. when it was added to an 11.5% ration it improved both gains and efficiency. Sherman gt 53;. (1959) also added 900 gm. of L-lysine lurdro. chloride per ton to a corn-alfalfa meal urea ration for fattening lambs. The lysine supplemented group gained .43 lb. per day. while controls gained only .32 lb. per day. Lysine addition. in some cases. have been shown to be of no benefit when added to sheep rations (Harbors gt g” 1961; Meacham gt 3.1.. 1961) or beef cattle rations (Kolari _e_t_ 51;. 1961). .T_h_e_ £13332 9;. Protein Nutrition 23 M 21:93 There are several factors that effect the value of protein for ruminants other than quality. They are solubility. degree of denaturation. particle size. nitrogenous compounds. protein level. and digestibility. All except digestibility affect rumen ammonia production. we ammonia is absorbed into the portal system and 23 converted to urea in the liver. This urea is partly returned to the rumen via the saliva (Lewis. 1957). Houpt (1959) has also shown that urea is returned to the rumen via diffusion. Lewis (1957) points out that wastage of nitrogen by these pathways is reflected by the urea level in the peripheral blood. Lewis (1957) and Perkins (1960) have shown a high relationship between protein level and blood urea concentrations. Everett gt a}... (1958) observed that calves on a 14.2% and 12.2% crude protein ration had higher blood urea levels than did calves on a 6.3% crude protein ration. Brown at 31; (1956. 1960) noticed that additions of urea to rations in order to increase the nitrogen resulted in high blood urea concentrations. Packett and Groves (1963) produced only moderate changes in blood urea when urea was added to the ration. but when ten to twenty grams of urea were placed in the rumen. a definite elevation in blood urea occurred. At a constant protein level of about 16. 5%. Whitelaw £3 31; (1961b) noticed lower blood urea levels as a result of using fish meal rather than groundrmt meal as a protein supplement. The authors postulated that the fish meal was superior in amino acid balance or was less soluble than the groundnut meal. Nitrogen retention values were inversally related to blood urea levels. but the relationship was not significant. 24 In addition to blood urea. plasma or serum protein levels are expected to reflect the protein welfare of the non-ruminant. Longnecker and Hause (1959) demonstrated that plasma amino acid changes in the adult dog. after feeding. were directly dependent upon the amino acid composition of the diet. On the other hand. Perkins (1960) found no difference in blood total amino acid nitrogen between groups of dairy cattle fed high or low protein rations. With regard to plasma protein. Brown gt _a_1_. (1956) found no difference due to protein levels or nitrogen sources fed to dairy calves. Bedrak gt §_J_._. (1957). however. noticed some trends in relationship between extremely low protein levels and serum protein values for hereford heifers. Brown gt 31; (1958) while comparing ration that varied from 12.2% crude protein to 21+. 3% crude protein observed that the 12.2% crude protein group had the highest serum protein. even though it had the lowest digestion coefficients for crude protein. Observing the chick. Leveille gt 31; (1960) noticed that serum protein levels were significantly depressed on low protein diets and that albumin levels followed the same trend. Similarly Wright _et gt. (1962) were able to lower or increase the plasma protein in ewes by dropping or raising the protein in the ration. These workers also noticed that this change was primarily due to albumin level changes. 25 Cahilly gt gt. (1963) were able to depress albumin levels by feeding a lysine deficient ration to swine. This is in agreement with other trials which increased albumin levels with additions of lysine (Cahilly gt 13.3. 1960; Brooks gt git” 1961). EXPERIMENTAL PROCEDURE Assiggtent. Mangggment. 229 Feeding For this thesis two experiments were conducted. In Experiment I. thirty-two male and female Holstein calves from the Michigan State University and the Southern Michigan Prison dairy herds were divided into four groups of eight calves each. The calves were assigned to their respective groups by filling all groups simultaneously as calves became available over a ten week period. Forty male and female Holstein calves from the University and Prison dairy herds and from surrounding dairy fanms were used in Experiment II. The calves were divided into five groups of eight calves each. Three bull or steer calves from each of the five groups in Experiment II that were born on or near January 6. 1963 or on or near January 20. 1963 were designated to be used in digestion trials. This was done to facilitate the simultaneous procurement of digestion data from several calves of the same age with a limited number of digestion crates. The remaining five calves per group in Experiment II were assigned to growth studies in such a manner that the average birth wmdght of the groups would be as nearly equal as possible. The last three of these five calves to be assigned were designated to be used for blood studies. 26 27 All calves in both experiments were separated from their dams between two and four days of age and placed on experiment where they remained for eighty-four days. The calves were retained in individual free stalls. bedded with dry wood shavings. Bucket milk feeding was begun twenty-four hours after the calf had been removed from the cow. Whole milk was fed at a rate of 9% of body weigzt the first week. 6% of body weight the second week. and 3% of body weight the third week. At the end of the third week each calf was weaned. All calves received less than 125 pounds of milk. All starters were fed in the pelleted form from approximately 3 days of age. The calf feeder encouraged each calf to eat the pellets by placing a small amount in the calf's mouth after feeding milk. This was continued until pellet consumption became voluntary. Hereafter. the pellets were fed 93. tit. Water was available at all times. Preparation _a_n_d_ Compgsition gt Rations No hay was fed to the calves except that which was incorporated into the starter pellet. The following percentages of ingredients were common to all rations in both experiments: 20% ground alfalfa... brome hw. 45% ground shelled corn. 7. 225% liquid cane molasses. 1.0% trace mineral salt. 1.0% dicalcium phosphate. 0.70% aurofac-ZA. and 0.075% vitamin A and D supplement. The remaining 25% of each 28 ration was made up of dried beet pulp and the main nitrogen source of the particular ration. These last two ingredients were blended so as to give the desired nitrogen.level in each ration. In Experiment I all rations were mixed to approximately 13% crude protein and varied only with regard to the main nitrogen source. The ration numbers and the respective nitrogen sources were: i36. urea; #37. soybean oil meal; #38. fish.meal; and #39. fish meal and soybean oil meal. Soybean oil meal and fish meal supplied equal amounts of nitrogen in ration #39. With regard to Experiment II. the rations varied in crude protein percent as well as nitrogen source. At approximately 17.5% crude protein. the ration numbers and the respective nitrogen sources were: ##1. urea; #42. soybean oil meal; and ##3. fish meal. At approximately 22%. the ration numbers and respective nitrogen sources were: #44. soybean oil meal; and {#5. fish meal. The complete composition for all rations is given in Table 26 of'the appendix. The proximate analysis of the rations are given in Table 27 of the appendix. Procurement gt Perfbrmance‘ggtg In Experiment I growth was observed by measuring body weight. heart girth circumference. and wither height. These criteria were measured as each calf was placed on experiment and at weekly intervals thereafter. Daily milk and pellet consumption were also recorded. The criteria for performance in.Experiment II was the same as that observed in Experiment I. In addition to these. blood studies 29 were conducted on three of the five animals in each group. The initial blood sample was taken from each calf at one week of experi- mental age. and the remaining samples were taken at two week intervals thereafter. up through eleven weeks of age. Forty to fifty milliliters of blood were drawn free the jugular vein three to four hours after the morning feeding. The collection vials were heparinized prior to bleeding to prevent coagulation. The blood was centrifuged within oneohalf hour after collection at two- thousand rpm for twenty-five mimtes. The plasma was then aspirated from the centrifuged sample and frozen. Another phase of Experiment II was to determine the digestible energy. digestible dry matter. digestible nitrogen. and the determi- nation of retainable nitrogen of the five rations when fed to growing male Holstein calves. Three male calves from each of the five groups were fed and raised in the same manner as all calves in Experiment II. up to five weeks of experimental age. it this time they were placed in digestion crates. where they remained for eight dws. The first three days were used for an adjustment period. and fecal and urine collections were taken on the remaining five days. Pellets were fed once daily. Orts were removed and weighed prior to each feeding. Composite samples of fresh pellets and arts were saved daily for later analysis. 30 Fecal collections were taken daily. weighed. and stored in large polyethylene containers. Thymol crystals were added to these storage vessels along with each fecal collection to prevent mold growth. At the end of the collection period. the accumulated collections were thoroughly mixed. and one composite sample was taken from each vessel. The wet samples were immediately analyzed for nitrogen. Energy was determined after the sample had been dried. Urine was collected in polyettwlene pails daily. Ten milliliters of concentrated hydrochloric acid and twenty milliliters of toluene were added to the empty pails before each daily collection commenced. Two percent of each daily urine excretion was saved and added to a composite sample bottle. These accumulated samples were retained in a refrigerator until the end of the five day collection period and 'were then immediately anakyzed fer nitrogen content. After the collection period the calves were returned to the calf barn and cared fbr in.the same manner as the other calves in the experiment. These same calves were returned to the digestion crates on the eighth week of experiment and again on the eleventh week of experiment. All practices were exactly the same in these two periods as in the first period. except for the adjustment period which was shortened from three to two dws. 31 Analytical Procedures All rations underwent proximate analysis according to procedures outlined in L0.A.C. (1955). Dry matter of all rations. except for those pellets fed in the digestion trial. was determined by drying two gram samples for five hours at 100 degrees centigrade. Blood plasma samples were anakyzed for protein and urea nitrogen content. The frozen.plasma samples were thawed overnight in a refrigerator and analyzed for protein by the following colorimetric technique developed by Lowry (1951). One milliliter of each plasma sample was diluted.400 times. and one.milliliter of the diluted plasma 'was pipetted into a test tube. Five milliliters of alkaline copper solution were added to the diluted sample. and the contents of the test tube were immediately mixed and allowed to stand for ten minutes. The alkaline copper solution consisted of 1.0 milliliter of 2% sodium tartarate. 1.0 milliliter of‘1%lcopper sulfate pentahydrate. and 100 milliliters of 2% sodium.carbonate in 0.1 Normal sodium.hydroxide. Fdlin Ciocalteu Phenol reagent (0.5 milliliter) was added to the test tube. and the contents were again thoroughly mixed. After standing for thirty minutes. the absorbance was read in a Beckman B-2 spectra- photometer at a wave length of 760 mu. Versatol. a serum standard produced by General Diagnostics. was diluted to different concentrations and used as standards in protein determinations. 32 The plasma urea was determined by a colorimetric method developed by Brown (1959). One milliliter of each unknown plasma was mixed with 7.0 milliliters of water. Zinc sulfate heptahydrate (1.0 milliliter of a 10% solution) was added to the samples and thoroughly mixed. One milliliter of approximately 0.5 Normal sodium rwdroxide was added and again the contents were immediately mixed. After standing for at least 15 minutes the mixtures were centrifuged at 2000 rpm for 25 minutes. A 2.0 milliliter aliquot of each filtrate was transferred to clean test tubes and 2 milliliters of p-dimfihlaminobenzaldelwdasulmric acid color reagent were added and thoroughly mixed. After standing for 10 minutes the absorbance of the solutions was read at 2440 mu in the Beckman spectrophotaneter. Standard urea solutions were prepared by varying the concentration of reagent grade urea in distilled water. With regan‘d to the digestion trial. dry matter for the pellets and arts was determined by drying approximately 200 gram samples at 80 degrees centigrade for 72 hours. Fecal samples were dried for 96 hours at 80 degrees centigrade. The nitrogen content of the urine. wet feces. and the dried pellets was determined by the improved Kjeldahl method as prescribed by A.0.A.C. (1955). The energy values of the dried feces and the dried pellets were determined in a standard banb calorimeter. 33 Statistical Treatment 3; _t.h_e. Qata- The methods employed in statistical analysis of the data are those described by Snedecor (1956). In Ebcperiment I. beginning bochr weigit. daily weight gain. increase in wither height. increase in heat girth. starter consump- tion. milk consumption and feed conversion were analyzed by single classification analysis of variance. Feed conversion or feed consumed per pound of gain and daily weight gain were also analyzed by covariance to adjust for differences in initial body weight. The data of Experiment II were divided so that the protein quality could be analyzed at each protein level pg; 52. The growth data were subjected to the same analyses employed in Ebcperiment I. Additionally. the growth data from both experiments were combined and analyzed so that saw interactions or differences in protein level could be exposed. This attempt to examine protein levels may be somewhat misleading since performance differences between experiments might be due to enviromental factors other than crude protein level 29; 32. Because of the unbalanced nature of the combined experiments it was necessary that the combined data be analyzed as two separate 2 x 3 factorial experiments. In one case the soybean oil meal. and fish meal as nitrogen sources. were one factor. while 13%. 17.535 and 22% crude protein were the other factor. In the other case. 13% and 17.5% crude protein were one factor. and urea. soybean oil meal. 31. and fish meal as nitrogen sources were the other factor. These factorial arrangements are diagrammed in Figure 1. Increase in wither height. increase in chest circumference. starter intake. and.milk consumption underwent analysis of variance. while feed conversion was analyzed by covariance to correct for variation due to differences in initial weight. Dairy gain was adjusted by covariance for initial weight and starter intake. The blood composition data of Experimant II were also analyzed as two separate experiments. First of all. the data fran all calves on the 17.5% crude protein rations were anaLyzed as a 3 x 6 factorial design. The three nitrogen sources were one factor and the six different ages at bleeding were the other factor. Secondly. the data were analyzed as a 2 x.2 x 6 factorial to expose differences in composition due to nitrogen source. crude protein levels. or ages of bleeding. These arrangements are diagrammed in Figure 2. The digestion data followed the same type of analysis. except that the three collecting ages replaced.the six bleeding ages. Wherever analysis of variance exposed a significant,main effect or interaction. the means of’these significantky different values were compared by The New Multiple Range Test developed by Duncan (1955). 35 me* me* mm* Home new“ msfi mme Hues seam 33* N:* mm* Hues awe «3* mh$ Hues ado concave cmonhon m.o N condom ’3‘ one some dwanoaomm Nmnqw.m. .mw H.m .m .m.o WNW eonsom \fi Hmanopoeuiw.mfl .np N.m .m a: e mi 91 we. dams ta it «1 E. a3 db madame» I mm* new: ad N .md w“; condom emcwnsoo damn momenm Ho mathmem new museum hookup» vouaamanoaomm new nocaneoo on» mo pcoeomcmnn< .w madman 36 Figure 2. Arrangement of the blood composition data for factorial analysis U._S.fF x age factorial at 17.5% C.P. Protein source Age (wks) 1 3 5 7 9 11 urea #41 ##1 #41 {#1 #41 fh1 soybean oil meal ##2 ##2 #42 ##2 ##2 #42 fish meal #0 #43 #43 £43 #43 #43 S. F x 17.5. 22 xgage factorial Protein Age i C.P. source gwksz 12:2. ‘22 1 #42 {an 3 #42 #14 5 #42 M Soybean oil meal 7 #42 #4“ 9 ##Z #44 11 {#2 get 1 #3 #5 3 #43 #45 5 #43 #45 Fish meal 7 #“3 {“5 9 #43 {#5 1 1 #3 1M5 RESULTS AN D DISCUSSION Experiment'I As mentioned in the introduction. the main purpose of this study was to determine the importance of’protein source in calf starters. Urea. soybean oil meal. and fish meal were chosen not only because they are practical nitrogen sources. but also because they cover a wide range of’protein quality. The term protein.quality generally refers to the degree to which a feed stuff'provides the proper level and balance of amino acids which are necessary in the diet fer optimum maintenance and growth. Unfortunately. no essential amino acid require- ments have ever been determined for dairy calves. Using swine research as a guide fer amino acid requirements. the literature-shows that tryptophan. methionine-cyctine. lysine. and isoleucine are the amino acids most commonly deficient in the rations which mainly consist of corn. Since urea adds no amino acids to the ration it does not improve the protein.qualiby of the basic corn ration. Soybean oil meal contains each of these critical amino acids. and thus improves the protein.quality of the ration. Fish meal contains a greater quantity of these amino acids than does soybean oil meal and therefOre should further improve protein quality (Pfander and Tribble. 1957). The combination of fish meal and soybean meal is expected to provide the highest protein quality because of the wide spectrum of 38 amino acids contained in the mixture. The crude protein percent of the rations was to be set at the level where protein quality would have it' s greatest effect. Accord- ing to the research done by Brown at El’ (1958). Everett 31; El: (1958), Brown and Lassiter (1962). and Whitelaw gt 3;. (1961a). the optimma crude protein level for calf starters is near the range of 14% to 16%. In addition to this. the results of several swine studies indicated that lysine (Catron gt _;_a_l_.. 1953: Pond gt §_1_.. 1953: Acker gt 31;. 1959; Nielsen gt _a_l_.. 1959: Chance gt 3.1-." 1960; Magruder st 53,. 1961; Becker gt ill" 1963) and isoleucine (Meade and Teter. 1956) are generally limiting in low protein diets. but not in high protein diets. Additionally. Kifer and Young (1961) noticed no protein quality effect when protein was fed in excess of the pigs requirement. Therefore. if protein quality differences are to be exposed. it will probably be done at crude protein levels just below the calf's minimum protein requirement. With this background in mind the crude protein level of 13% was chosen for Experiment I. The results of Experiment I expressed in Table 1 indicate no significant differences among groups with regard to growth rate or feed consumption. even though starter intake was highest for the urea group. Feed conversion improved from urea to soybean oil meal to fish meal. however these differences were not significant at the 5% level of probability. 39 Table 1. Effect of source of protein on mean growth. feed consumption. and feed conversion at the 13% crude protein level Starter Groups urea SBOM fish meal SBOM & fish meal 36 37 38 39 Growtha Initial weight (lb.) 86.5 87.4 91.8 84.9 Daily gain (1b.) 1.08 1.02 1.05 1.09 Increase in height at withers (in.) 3.21 3.56 3.53 MOB Increase in heart girth (in.) 7. 63 7.16 7.06 7.84 Feed consumptiona Milk (1b.) 109.9 110.9 117.6 106.8 Starter (1b.) 258 2141+ 240 255 Feed conversiona (lb. feed/lb. gain) 2. 92 2.85 2. 71+ 2. 82 aNo significant differences among groups. The failure of Experiment I to produce any significant performance differences may have been caused by an overall protein deficiency in all calves. which was brought about through a combination of early weaning and low crude protein level in the ration. It was suspected that the protein deficiency retarded normal growth to the extent of masking am effects resulting from differences in protein quality. Experiment I; To avoid possible protein deficiencies in Experiment II. ap- proudmately 17.5% and 22% crude protein levels were used. As pointed out in the experimental procedure. the results of Experiment II were analyzed separately according to protein level. m. _f_e_e_d_ consumption. and £9.93. conversion results Table 2 contains the growth. feed consumption. and feed conversion data from Experiment II at the 17. 5% crude protein level. The actual daily gain values were higher for the urea group than for the soybean oil meal or the fish meal groups. However these values as well as those corrected for initial body weight were not significantly different. There were no significant differences among groups for any of the remaining criteria. even though heart girth increase appeared to improve with protein quality and starter consumption responded inversely. The data from the 22% crude protein level are shown in Table 3. Fish meal was superior to soybean oil meal. with regard to daily gain. heart girth increase and feed conversion. but none of these or the other criteria were significantly different. The failure of protein source to affect gains in these experiments reflects the results obtained by Halter (1956). Lambert 33 21.: (1955). Hibbs gt 5;. (1953)o Pardue and Jacobson (1961). Pardue gt a]: (1962). and Preston 93; fl. (1960). all who found protein source not to be an important factor for improving bochr weight gains. 41 Table 2. Effect of source of protein on mean growth. feed consumption and feed conversion at the 17. 5% crude protein level Starter Groups urea SBOM fish meal 41 42 43 Growtha Initial weight (lb.) 93.6 90.0 90.4 Daily gain (1b.) 1.17 1.14 1.07 Increase in height at the withers (in.) 4.30 4. 26 “JD Increase in heart girth (in.) 7.20 7.40 7.80 Feed consumptiona Milk (1b.) 123 117 119 Starter (1b.) 272 241 233 Feed conversiona (lb. feed/lb. gain) 2.83 2.52 2. 61 3No significant differences among groups. In order to examine the effect of protein level on calf perform- ance. the factorial arrangements explained in the experimental procedure were employed. Investigation of the average daily weight gains by an urea. soybean oil meal. fish meal X 13%. 17. 5% factorial comparison (here- after designated U. S. F x 13. 17.5 factorial) as shown in Table 4 42 Table 3. Effect of source of‘protein on mean growth. feed consumption and feed conversion at the 22% crude protein.level Starter Groups SBOM fish meal #4 45 Growtha Initial weight (lb.) 90.0 93.2 Daily gain (1b.) 1.21 1.36 Increase in height at the withers (int) b.80 3.80 Increase in heart girth (in.) 3.30 9.30 Feed consumptiona Milk (1b.) 113 121 Starterhub. ) 246 2% Feed conversiona (lb. feed/lb. gain) 2.1411 2.16 aNo significant differences among groups. shows no significant differences among sources. levels. or inter- action when the actual data are used. However. when the data are anakyzed by covariance to adjust fbr starter intake an interaction does exist. where fish meal at the 17.5% crude protein level is superior to urea at 13% and 17.5% crude protein. Body weight gains adjusted fer starter intake. however. is merely an expression of feed conversion and should not be confused with actual weight gains. 43 Table 4. Mean daily weight gains at 13% and 17.5% crude protein % crude protein Nitrogen sourcea 13b 17.51, average Actual( adjusted) d Actual( adjusted)d Actual ( adj )<= (1b.) Urea 1.08(o.98)d 1.17(o.99)d 1.11(1.03) Soybean oil meal 1.02(1.06) 1.14(1.11) 1.o7(1.10) Fish meal ‘ 1.o#(1.05) 1.o7(1.15)d 1.06(1.11) Average 1.05 1.13 aNo significant difference among groups. bApproximate. cAdjusted for starter intake. (11.15 is significantly greater than 0.98 and 0.99 (P<0.05). Analysis of covariance to adjust for initial body weight exposed no significant differences in weight gain. therefore the mean gain values were not adjusted. The soybean oil meal. fish meal at 13%. 17.5%. 22% factorial (hereafter called the S. F x 13. 17.5. 22 factorial) canparison of average daily weight gains is shown in Table 5. The analysis of the actual data and the data adjusted for differences in initial body weight shows that the 22% crude protein level was significantly superior (P< 0.05) to the 13% crude protein level. When the data 44 Table 5. Mean daily weight gains at 13%. 17.5%. and 22% crude protein A;% Crude_protein Nitrogen sourcea 13b 17.5b 22b average (1b.) Soybean oil meal 1.02 1.14 1.21 1.11 Fish meal 1.05 1.07 1.36 1.14 Average(actual) 1.03B 1.11AB 1.29A Average(adjusted)c 1.03** 1.14""I 1.27** Average(adjusted)d 1.04B 1.12AB 1.27A aNo significant differences among groups. bApproximate. cAdjusted for starter intake. dAdjusted for initial weight. MSignificantly different (P< 0.01 ). Avalues with the same large superscript represent a homogeneous group (P<0.05). 1 are adjusted for starter intake the crude protein.levels are all significantly different (P<<0.01); the daily gains improving with each level of crude protein. Here again the intake corrected values simply reflect efficiency. The difference in gain due to protein source was not significant for the actual data. This confirms the results obtained earlier in the analysis of each crude protein level for differences in sources. 45 The results of this 5. F x 13. 17.5. 22 factorial comparison do not agree with results obtained by Brown and Lassiter (1952) and the first trial conducted by Brown 93 fl. (1958). In both cases these workers noticed no effect of protein level on body weight gains. Additionally. in a second trial. Brown 93 31; (1958) found that a 16.2% crude protein ration was superior to 20.0% and 23.7% crude protein rations. One possible explanation that can be given for the difference in results is that milk was fed through the seventh week by Brown _e_t_ §._l_. (1958). and their calves were not as dependent upon the protein in the ration during the fourth. fifth. sixth. and seventh week. The calves in this trial were weaned at three weeks of age and were solely dependent on the pellet rations hereafter. in which case the high protein.rations would be of greater value to the calves. Brown 23:39? (1958) also point out that calves fed the 16.2% crude protein ration consumed more starter than the other groups. and the 16.2% ration had what they considered to be an optimum.protein~energy ratio. Investigation of the increase in wither height with a U. S. F x 13. 17.5 factorial comparison. as shown in Table 6. indicates a.highly significant advantage of the 17.5% crude protein.level over the 13% crude protein.1evel. This protein level effect on wither height in Table 6 was repeated in the S. FIX 13. 17.5. 22 factorial comparison. as can be seen in Table 7. However. it was not significant in this factorial. 46 Table 6. Mean increase in wither height (in.) at 13% and 17. 55% crude protein % Crude protein Nitrogen sourcea 13b 17.5b average Urea 3.21 4.30 3.25 Soybean oil meal 3. 56 4. 26 3. 91 Fish meal 3. 53 4. 30 3.91 Average 3, 43am 4. 29*... 8”No significant difference among groups. bApproximate. MSignif:’1.cantly different (P< 0.01). Table 7. Average increase in wither height (in.) at 13%. 17.5%. and 22% crude protein j CrudeLprotein Nitrogen sourcea 13b 17. 5b 22b average (lb. ) Soybean oil meal 3. 56 4. 26 4. 80 4. 21 Fish meal 3. 53 4. 30 3. 80 3. 88 Average 3. 55 4. 28 4. 30 aNo significant difference among groups. bApproximate. 117 There was no protein source effect in either factorial investigation of wither height. when increase in heart girth. milk consumption and starter in. take data were analyzed by the two factorial comparisons. no significant differences among crude protein levels were exposed. These average values are shown in Table 8. Table 8. Average values fer increase in heart girth. milk consumption. and starter intake fer each crude protein level gfi_Crud§_protein __¥ Criteria 13 17.5 22 Increase in heart girth (in.)a 7.3 7.4 8.8 Milk consumption (1b.)a 112 119 117 Starter intake (1b.)a 247 249 246 aNo significant differences among groups. The last criteria of growth perfOrmance to be considered is feed conversion. which was measured as pounds of starter consumed per'pound of gain. The U. S. F'x 13. 17.5 factorial arrangement exposed no significant differences among actual values or values adjusted for initial body weight. However. the efficiency appeared to improve with protein quality and with protein level as can be seen in Table 9. 48 Table 9. Mean feed conversion (lb. of starter per 1b. of gain) at 13% and 17.5% crude protein % Crude groteinL Nitrogen sourcea 13b 17.5b average (1b.) Urea 2L92 2.83 2.89 Soybean oil meal 2.85 2.52 2.73 Fish meal 2.74 2.61 2.69 Average 2.84 2.66 aNo significant difference among groups. bApproximate. Similarly the S. F x 13. 17.5. 22 factorial analysis showed an insignificant improvement in efficiency as protein quality increased (Table 10). A significant improvement in efficiency occurred with each increase in crude protein level. Both actual values and values corrected for initial body weight followed the same trend. however. the corrected.values were all significantly different at the 1.0 percent level. Here again. the efficiency results like the daily gain results disagree with the observation of Brown 23 3;. (1958) and Brown and Lassiter (1962). but since gain and efficiency are highly related this would be expected. and the reasons given fer the differences in gain would apply to the differences observed in efficiency. 49 Table 10. Mean feed conversion (lb. starter per lb. gain) at 13%. 17. 5% and 22% crude protein $ (rude protein Nitrogen sourcea 13b 17. 5b 22b average (lb. ) ‘ Soybean oil meal 2. 85 2. 52 2. 44 2. 65 Fish meal 2. 74 2. 61 2. 16 2. 54 Average( actual) 2. 80M 2. 56Bel 2. 30b Average( adjusted) c 2. 79” 2. 56'MI 2. 32" aNo significant differences among groups. bApproximate. cAdjusted covariance for initial body weight. “Significantly different (P<0.01). Values with the same large subscript represent a homogeneous group (P<0.05). Mean with the same small subscript represent a homogeneous group (P 40.01). Digestion 21a}. results The statistical approach used to analyze the digestibility data are those outlined in the experimental procedure and diagrammed in Figure 2. The statistical analysis of the coefficients of apparent digest- ibility of dry matter at 17. 5% crude protein using the urea. soybean oil meal. fish meal by age factorial (hereafter denoted by U. S. F x age factorifl) exposed a nonsignificant increase in apparent dry matter digestibility at each successive age (Table 11). 50 Table 11. Mean coefficients of apparent dry matter digestibility at 17.5% crude protein Age (URS. )b Nitrogen sourceb 5 8: 11 average (1%) Urea 72.63 76.72 76.66 75.3# Soybean oil meal 78.47 76.49 81.89 78.95 Fish.meal 72.54 7#.44 77.80 7h.93 Average 74.54 75.88 78.78 aApproximate. bNo significant differences among groups. The soybean oil meal, fish meal by 17.5%. 22% by age factorial (hereafter referred to as the S. F‘x 17.5. 22 x age factorial) analysis, of the apparent dry matter digestibility coefficients. also exhibits an insignificant increase in value for each age (Table 12). In all cases, the apparent dry matter digestibility coefficient was greater for the soybean oil meal source than for the fish meal source. When the protein levels were combined. this difference was significant at the 5% level. The coefficients of apparent digestibility of energy followed the same pattern as the apparent dry matter digestibility coefficients, with the values increasing with age and with protein percent. Here 51 Table 12. Mean coefficients of apparent digestibility of dry matter at 17.5% and 22% crude protein fi Crude proteina Nitrogen source 17.5b 22b Average Age(wks) Average Age(wks) Average ($3 5 78.47 5 78.92 Soybean oil meal 8 76.49 78.95 8 80.74 79.53 79.26* 11 81.89 11 79.05 5 72.54 5 76.n6 Fish meal 8 74.44 74.93 8 78.15 77.88 76.87* 11 77.80 11 79.02 Average 75.94 78.72 Age Averages AgG‘VkSO )a 5 8 11 75. 50 7'7. 45 79.44 aMo significant difference among groups. bApproximate. *Siglfificantly different (P <0. 05). 52 also. the soybean oil meal source had the highest apparent energy digestibility coefficient. These values are shown in Tables 13 and 14. and asindicated none are statistically significant. Table 13. Mean coefficients of apparent digestibility of energy at 17.53%a crude protein Age(uks)b Nitrogen sourceb 5 8 1 1 average Urea b9. 46 73. 81 73. 9O 72. 39 Soybean oil meal 75.97 73. 90 79. 80 76.58 Fish meal 71.46 72.14 76.96 73.52 Average 72. 29 73. 30 76. 89 aApproocimate. b No significant differences among groups (1340.05). Nitrogen digestibility of all rations significantly increased with each successive age (Table 15 and 16). In contrast Brown gt :4. (1958) found that crude protein digestibility coefficients decreased with age. but this was probably due to the reduction of milk feeding as their trial progressed. The 22% crude protein level had a significantly (P<0.05) higher nitrogen digestibility coefficient than did the 17. 5% crude protein level. This confirms the results 53 Table 14. Mean coefficients of apparent digestibility of energy at 17.5% and 22% crude protein % (rude proteina Nitrogen source 17. 5b 22b Average Age(wks) Average Age(wks) Average (i) 5 75. 97 5 76. 61 Soybean oil meal 8 73.96 76.58 8 78. 65 77.46 77.02 1 1 79. 80 1 1 77. 12 Fish meal 8 72.14 73.52 8 77. 65 77.04 75. 28 1 1 76.96 1 1 78. 1+0 Average 75.05 77. 25 Age Averages “3(731‘50 )a 5 8 11 74.78 75. 60 78.07 8LNo significant differences among groups (P40.05). bApproximate. 54 Table 15. Mean nitrogen digestibility coefficients at 17. 5703 crude protein Agebrks) Nitrogen sourceb 5 8 1 1 average (fi) Urea 62. 60 73. 43 74. 96 70. 33 Soybean oil meal 70. 26 71.96 78. 95 73.72 fish meal 59. 49 65. 44 75. 20 66. 71 Average 64. 11Bb 7o. 27Bab 76. 14Ml aApproximate. b1110 significant difference among groups (P40.05). Values with the same large superscript represent a homogeneous group (P<0.05). Values with the same small superscript represent a homogeneous group (P<0.01). obtained by Brown 33 _a_l_. (1958). Everett 2!; g. (1958). Whitelaw £2 59;. (1961a) and Brown gt g. (1960). all who noticed that nitrogen digestibility increased with the protein content of the diet. Again the digestibility coefficients were the greatest for the soybean oil meal ration in both factorials. however. these values were not significantly greater. In order that the nitrogen welfare of the groups might be studied more thoroughly. retainable nitrogen was expressed as a percent of nitrogen intake. Data in Table 17 indicate that calves 55 Table 16. Mean nitrogen digestibility coefficients at 17.5% and 22% crude protein i_Crudegprotein Nitrogen sourceb 17.5a 22a Average Age(wks) Average Age(wks) Average (16) 5 70.26 5 73.36 Soybean on meal 8 71.96 73.72 8 79.90 77.53 75.63 11 78.95 11 79.33 5 59.49 5 67.51 Fish meal 8 65.44 66.71 8 76.33 74.15 70.43 11 75.20 11 78.61 Average 70.22B 75.811“ ___A_gA_Ag§,Averages AgG‘UkSe) 5 8 11 67.6613 73.40"”3 78.02.“ a’Approxima‘te. b No significant difference among groups. Values with the same large superscript represent a homogeneous group (IMO-05)- 56 Table 17. Mean nitrogen retention as percent of nitrogen intake at 17. 5% crude protein . Age(wkS)b Nitrogen source 5 8 1 1 average (75) Urea 17.77 34.09 31.92 27.93Mb Soybean oil meal 16. 68 18. 22 19. 40 18.10]3b Fish meal 24. us 28. 59 35. 95 29. 66‘La Average 19. 63 26. 97 29. 09 a‘Approximate. b No significant differences among groups (P< 0.05). Values with the same large superscript represent a homogeneous group (P<0.05). Values with the same small superscript represent a homogeneous group (P<0.01). on the soybean oil meal ration retained significantly less nitrogen than those on urea (P<0.05) and less thanthose on fish meal (P< 0.01). These results seem unusual since calves fed the soybean oil meal ration had the highest nitrogen digestibility coefficients. However. the dry matter intake during collection averaged over the three ages. for the urea. soybean oil meal. and fish meal groups were 1.714 m. 1.060 gm and 1.27? gm. respectively. Whether this difference in intake is of the magnitude to affect the digestibility and the retainibility of the nitrogen is questionable. The effect of dry matter intake on 57 nitrogen digestibility has been reported by Kumta and Harper (1962). Table 18 shows the S. F x 17.5. 22 period factorial where level and source effect were rendered insignificant due to a source level interaction. Calves fed the 17.5% crude protein ration containing fish meal and the 22% crude protein ration containing soybean oil meal retained significantly more nitrogen than those fed the 17.5% crude protein ration containing soybean oil meal (P< 0.01). Another method used to express retained nitrogen was grams of nitrogen retained per day per hundred pounds of body weight. These values are given in Tables 19 and 20. and show the same pattern as the percent nitrogen retained values. Here again. at the 17.5% crude protein level. the value for soybean oil meal was significantly (P< 0.01) less than for urea or fish meal. Similarly. this value was less than the 22% crude protein soybean oil meal value (P< 0.01) or the 22% crude protein fish meal value (P<0.05). In addition the 22% crude protein fish meal group retained significantly less (P<0.01) nitrogen than the 22% soybean oil meal group. If the daily dry matter intake values listed in Table 21 are examined. the relationship between nitrogen retention and dry matter intake is easily recognized. Computation of the correlation coef- ficient of dry matter and percent nitrogen retained using individual observations and disregarding quality. quantity. and age effects gives a coefficient of 0.75. 58 Table 18. Mean nitrogen retention as percent of nitrogen intake at 17 . 5% and 22% crude protein ' % Crude proteina NitrOgen . sourcea 17. 5b 22b Average Age (wks) Average Agehtcs) Average (5) 5 16. 68 5 30. 22 Soybean b a oil meal 8 18.22 18-10 8 32-56 30.13 24.12 11 19.40 11 27.60 5 24.45 5 16.33 Fish meal 8 28.59 29.66a 8 29.08 22.47ab 26.07 11 35.95 11 22.00 Average 23. 88 26. 30 Age Averages Age (wks. )a 5 8 11 21.92 27.12 26.24 aNo significant difference among groups (P<0.05). bApproximate. Values with the same small subscript represent a homogeneous group (P<0.05). 59 Table 19. Mean grams of nitrogen retained per day per hundred pounds of body weight at 17.55% crude protein Age(wks)b Nitrogen source 5 8 11 average — (an) Urea 5.81 13.28 11.56 10.21a Soybean oil meal 4.53 4.13 4.38 4.35b Fish meal 7.31 8.71 11.51 9.18“ Average 5.88 8.71 9.15 a Approximate. b No significant differences among groups. values with the same small subscript represent a homogeneous group (P<0.01). Since this high relationship did exist. percent nitrogen retention was analyzed with covariance to correct for differences in dry matter intake. As a result. none of'flhe significant dif- ferences in nitrogen retention in Table 18 remained. Whitelaw 31 El. (1961b) overcame the intake effect by controlling consumption. With this accomplished they noticed an.improvement in nitrogen retention when fish meal was included in the ration. Visual obser- vation of the data indicates no relationship between average nitrogen retention and average daily gain of the groups. although both gain and nitrogen retention generally increased as the crude protein per- cent increased. 60 Table 20. Mean grams of nitrogen retained per day per hundred pounds of body weight at 17.5% and 22% crude protein % Crude proteina Nitrogen sourcea 17. 5b 22b Average Age(wks) Average Age(wks) Average — (adv 5 4. 53 5 11. 66 Soybean Bo Aa Oil meal 8 4.13 4.35 8 13.17 12.04 8.19 11 4.38 11 11.29 5 7. 31 5 4. 69 Fish meal 8 8. 71 9.18Mb 8 1o. 50 7. 66“” 8. 42 11 11.51 11 7.78 Average 6. 76 9. 85 Age Avergges Age(Wk50 )a 5 8 11 7.05 9.13 8. 74 —_ aNo significant differences among groups. bApproximate. Values with the same large superscript represent a homogeneous group (P<0005)0 Values with the same small superscript represent a homogeneous group (P<0.01). 61 Table 21. Average grams of dry matter consumed per day during collection % Crude jrotein Nitrogen source 17 22 (an) Soybean oil meal 1.060 1.365 Fish meal 1.277 1.105 M M studies Whitelaw gt _8_l_. (1961b) observed an insignificant yet inverse relationship between nitrogen retention and blood urea levels. This was not the case in this experiment since calves fed the 22% crude protein ration containing soybean oil meal had the highest plasma urea nitrogen level as well as the highest nitrogen retention values (Table 22). The only explanation that can be given for the high level of plasma urea nitrogen produced by the soybean oil meal ration at 17.5% crude protein is that solu- bility and the high nitrogen level of this starter acted together to produce large amounts of rumen ammonia. which after absorption were converted to blood urea in the liver. Like Lewis (1957). Brown 25 a}... (1960). Perkins (1960). and Everett gt ;a_l_. 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