7 / L £ - THIS THESIS SUBMITTED BY Titilayo ADSG-BOLA WAS ACCEPTED IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY OF THIS UNIVERSITY THE DATE OF AWARD IS 25th October, 1974 l ' . ..C t u RE UNIVERSITY OF IBADAN LIBRARY 11 0.i3*4- and 0,22g/day/Wi cgQ .734 by the factorial method, The levels of nitrogenous metabolites in the rumen varied with levels of dietary crude protein. Ruminal ammonia was highly correlated (r = 0.99) with blood urea. The amino acids present in lowest concentration in bacterial and protozoal protein are methionine and histidine while there are high levels of lysine and leucine. Isotopic studies with N ammonium chloride and urea shows that 4 - 7 $ of £~ 15n_7 ammonium chloride administered into the rumen was recovered in the faeces, and 3.1$ was recovered in milk. Also 30.5£of f 15n _7 urea administered into the blood was recovered in the urine and the isotope was not recovered in the faeces. Ruminal ammonia contributed 26 - 33$ of the bacterial N and 15 - 19$ of protozoal N ten hours after feeding. Urea was synthesized in the body at the rate of 9.4 to 10.1g/day, and A.7 to 7.3g/day were degraded in the digestive tract of the sheep. The chromic oxide - impregnated paper method showed that 7 2 . of digestible dry matter and 72.£$ of digestiis^ organic matter of the rations were digested in the stomach. The corres­ ponding values for small intestine were 10.1$ and 11 ,ltf0 for dry UNIVERSITY OF IBADAN LIBRARY iii matter and organic matter respectively, while in the ceacum and colon, the values were 1 7. ^ and 16.Cfe for dry matter and organic matter respectively. Substantial amount of N of endogenous origin were secreted in the proximal small intestine but were efficiently absorbed before the distal portion was reached. The results show that the West African dwarf sheep utilize the hay and supplement rations efficiently and are* adapted for survival in areas where the intake of N might be inadequate. UNIVERSITY OF IBADAN LIBRARY iv A C K N O W L E D G E M E N T S The author expresses his sincere appreciation to Professor V.A. Oyenuga, the Head of the Department of Animal Science for the opportunity he has given him to undertake the research work in his department and also for his interest in the progress of the research work of the author. The author wishes Professor Oyenuga the blessing of God. The author also expresses his deep gratitude to his super­ visors, Dr. F.O. Olubajo and Dr. A.U. Mba for their assistance to and guidance of the author throughout the course of the author’s graduate study and research. The author wishes them the blessing of God. The author also acknowledges the financial assistance of the Rockefeller Foundation and the University of Ibadan for the scholarship he held throughout the course of his study. The author acknowledges the help rendered by the Technical staff of the Department of Animal Science, especially Messrs. Paulissen and Okusanya, the animal attendants, Messrs. Ogobo, Ajibogun, Aremu and Adisa, and also Messrs. Onwe, Audu, Adejumo, Chikwe and Ohwarhua of Department of Agric. Economics and Extension who typed the thesis. The author acknowledges the cooperation of Dr. J.U. Akpokodtje and Dr. Ladosu of the Department of Veterinary Medicine and Surgery who fistulated the animals. Special appreciation is extended to the author's wife for her encouragement in the course of the research work. UNIVERSITY OF IBADAN LIBRARY V We certify that this work was carried out by Mr. T. Adegbola in the Department of Animal Science, University of Ibadan. V. A."Uyenuga, B.Sc., Ph.D. (Dunelm), F.R.I.C. F. 0. Olubajo, B.Sc. (Illinois), M.Sc.(Ohio), Ph.D. (Ibadan). UNIVERSITY OF IBADAN LIBRARY TABL2 0? CGKB£?Cg PAGE A B S T R A C T .. .. .. •• i A C K N O W L E D G E M E N T .. • • iv CHAPTER TITLE ONE 1. I n t r o d u c t i o n .. •• 1 1.1 General Introduction .. .. 1 1.1.1 Breeds of sheep in Nigeria .. 5 1.1.2 Importance of sheep to the economy of Nigeria .. .. •• 6 1.1.3 The Management of the West African Dwarf sheep .. .. *« 7 1.2 Literature Review .. •• 8 1.2.1 Microbial fermentation and protein, v digestion in the ruminants .. 8 1.2.1.1 Proteolysis and deamination of amino ac i ds .. ®• .. 8 1.2.1.2 Conversion of food protein in to microbial protein in the rumen •• 10 1.2.1.3 Nutritive values of microbial protein 19 1.2.1.k Factors influencing microbial amino acid availability .. .. 20 1.2.1.5 Relative supply of amino acids and energy .. .. .. 22 UNIVERSITY OF IBADAN LIBRARY rZAPZES vii PAGE CjH 1.2.2 Utilization of non-protein nitrogen by the ruminant .. .. «• •• 27 1.2.2=1 Non-protein nitrogen in ruminantr ations 27 1.2.2.2 Metabolism of ammonia nitrogen by rumen micro-organisms =. .. •« 29 1.2.2.3 Factors affecting the utilization of ammonia in the rumen .. .. 33 1.2.2.4 Absorption of ammonia through the rumgawall 35 1.2.2.5 Influence of ruminal ammonia level on the concentration of ammonia and urea in the blood .. .. .. .. 57 1.2=3 Intestinal digestion of protein-N and utilization in the ruminant .. 42 1.2.3*1 The rate of flow of digesta in the diges­ tive system of ruminant animals .. 45 1.2.4 Isotopic methods of determining the utili­ zation of nitrogen in the ruminant .. 53 1.3 Objectives .. .. .. .. ^2 TWO 2. Ruminal and blood metabolites of the West African dwarf wether sheep maintained on basal hay and concentrate supplements: 63 2.1 I n t r o d u c t i o n .. .. ^3 2.2 Materials and methods .. .. o3 2.2.1 Animals and their management .. 63 2.2.2 Diets •« .. .. .. UNIVERSITY OF IBADAN LIBRARY viii EAPTER TITLES PAGE rvo 2.2.3 Plan of experiment • • 6 4 2.2.4 Collection of faeces and. urine • • 67 2.2.5 Sampling of blood and rumen liquor. • • 68 2.2.5 Isolation of rumen bacteria and protozoa 69 2.2.7 Analytical procedure .. • o 69 2.3 Results o o 71 2.3.1 Total ruminal nitrogen • •* 71 2.3.2 Total ruminal protein nitrogen 0 9 73 2.3.3. Non-protein nitrogen .. .. 9 9 74 2-3.4 Ruminal ammonia nitrogen .. 9 9 75 2.3.5 Ruminal residual nitrogen 9 9 73 2.3.6 Ruminal o(-amino nitrogen 9 a 79 2.3.7 Blood urea nitrogen .. .. • • 81 2.3.8 Amino acid composition of ruminal bacteria and protezoa .. • • 83 2.4 Discussion .. .. .. • 9 85 THREE Isotopic studies of nitrogen metabolism in the West African dwarf wether sheep 0 0 97 3.1 Introduction .. .. .. 0 0 97 3.2 Materials and methods 0 0 98 3.2.1 Animals and their management .. 0 0 98 3.2.2 Diets *. *« .. 0 0 98 3.2.3 Plan of experiment .. .. © © 99 3.2.*+ Collection of faeces, urine, blood , rumen and milk samples .. .. 0 0 100 UNIVERSITY OF IBADAN LIBRARY ix CHAPTER TITLES PAGE THREE 3.2.5 Separation of bacteria and protozoa in the rumen liquor • 0 • • 100 3.2.6 Analytical procedures e 0 0 e 101 3.2.7 Theory 101a a • • 3.3. Results .. 102. . © • • • 3.4 Discussion • • • • 112 FOUR The intake and digestibility of dry matter and N metabolism in the West African dwarf wether sheep maintained on hay and concentrate supplements 0 • 119 4.1 I n t r o d u c t i o n 1190 0 • • 4.2 Materials and methods 0 • 1190 0 4.3 Results 120. . O 0 • 0 4.3.1 The dry matter intake 0 0 1200 • 4.3-2 Intake of nitrogen • 0 • • 128 4.3.3. The digestibility of dry matter 0 a 129 4.3.4 The digestibility of nitrogen 0 • 0 0 130 4.3.5 Absorbed nitrogen O O 0 0 134 4.3.6 Retained nitrogen . . a 0 0 0 135 4.3.7 Nitrogen retention (%) • 0 0 0 137 4.3.8 The metabolic faecal nitrogen • 0 0 • 139 4.3.9 Endogenous urinary nitrogen 0 0 1460 0 4.3.1 0 The biological value of the rations 0 0 147 UNIVERSITY OF IBADAN LIBRARY X CHAPTER TITLES PAGE FOUR It. 3. 11 The coefficient of net utilization 148 4.3.12 The protein requirement for maintenance 148 4.4. Discussion .. • a 0 0 153 FIVE Digestion in the alimentary tract of the West African dwarf sheep . ° .. .. • • 177 5.1 I n t r o d u c t i o n 177 5.2 Materials and methods .. 177 5.2.1 Animals and their management 177 5.2.2 Experimental rations .. .. 178 5.2.3 Plan of experiment .. .. .• 178 5.2.4 Collection of faeces 178 5.2.5 Collection of digests from the compartments of the digestive tract .. .. 179 5.2.6 Analytical procedure .. .. 179 5.2.7 Estimation of digestibility in the compart­ ments of the digestive tract .. .. 179 5.3 Results .» .. .. .. 180 5.3.1 Total digestibility of dry matter and nitrogen 180 5.3.2 Chromic oxide recovery .. .. 182 5.3.3 Digestibility of dry matter in the sections of the digestive tract .. .. 182 5«3»4 Digestibility of organic matter in the various sections of the digestive tract .. 186 5.3.5 Nitrogen intake, distribution and absorption 189 5.4 Discussion .. .. .. .. 191 .6. Summary of Conclusion .. .. 199 7. ..................... 204 8. A p p e n d i x .. .. .. 228 UNIVERSITY OF IBADAN LIBRARY xi LIST OF TABLES PAGE 2.1 Components and chemical composition of rations .. 65 2 .2 Plan of experiment .. .. .. . . 6 6 2.3 Ruminal and blood metabolites of the West African dwarf wether sheep .. .. .. . . 7 2 2.4 The regression equations showing the relationship between ruminal and blood metabolites and nitrogen utilization .. .. .. . . 7 6 2.5 Amino acid composition of ruminal bacteria and protozoa «. .. .. . . 8 4 3.1 Chemical composition of the basal hay .. * * 99 3.2 The recovery of 15Ni n the faeces, urine and milk of the dwarf sheep .. .. .. . . 1 0 3 3.3 Ammonia and urea metabolism in the West African dwarf sheep .. .. .. ..110 4.1 Effect of ruminal fistulation of the West African dwarf wether sheep on digestibility of dry matter and nitrogen contents of basal hay and concentrate supplements. .. .. .. .. 121 4.2 Dry matter intake, digestibility and N metabolism for the West African dwarf wether sheep ..122 4-3.1 The regression equations showing relationships between nutrient utilization in the West African dwarf sheep .. .. .. ..123 4.3.; The regression equations showing relationships between nutrient utilization in the V/est African dwarf sheep .. .. .. ..124 UNIVERSITY OF IBADAN LIBRARY 4.4 Effect of supplementing basal hay with concen­ trates on intake of hay .. .. 126 4.5 True digestibility, biological value and net protein utilization estimates for the West African dwarf wether sheep „. . . .. -j 33 4.6 The percentage of the undigested N, microbial N, water soluble N and non-dietary faecal N in the faeces of the West African dwarf wether sheep 144 4.7 Digestible crude protein requirement of the sheep (N-balance method). 150 4.8 Digestible crude protein requirement by factorial method 152 5«1 A comparison of the total collection and ®3xronio oxide methods for the determination of dry matter and N digestibilities with the West African dwarf sheep .. .. .. .o 181 5.2 Chromic oxide recovery for the West African dwarf wether sheep .. .. .. ■J83 5.3*1 The percentage digestibility of dry matter taking place in the sections of the digestive tract of the West African dwarf wether sheep .. ^g^ 5*3.2 The percentage digestible dry matter taking place in the sections of the digestive tract .. 185 5.4.1 The percentage digestibility of organic matter taking place in the sections of the digestive tract of the West African dwarf wether sheep using the chromic oxide ratio 0 o o • 187 UNIVERSITY OF IBADAN LIBRARY xiii PAGE z.-,2 The percentage digestible organic matter taking place in the sections of the digestive tract 188 5.5 Nitrogen intake, distribution and absorption at different sites of the alimentary canal 190 UNIVERSITY OF IBADAN LIBRARY XXV LIST OF FIGURES FIGURE PAGE 2»1 Level of ruminal ammonia and blood urea in the sheep .. .. .. oo 82 3.1 Enrichment of ruminal ammonia, blood urea, bacteria and protozoa, after intra-ruminal infusion of f 15« J ammonia .. .» 105 3.2 Enrichment of rtiainal ammonia, blood urea,!?. bacteria and pretozoa after intra-ruminal infusion of £ HkJ ammonia .. .. .» 106 3«3 Enrichment of blood urea and ruminal ammonia after A C intravenous injection of £ urea.. 107 3.^ Enrichment of blood urea and ruminal ammonia after intravenous injection of /_ 15NJ urea 108 ^.1 The relationship between absorbed N and re^ttined N 136 b-,2 The regression of fnitrogen balance on nitrogen intake .. .. .. .. 138 *+.3 The regression of faecal nitrogen on nitrogen intake .. .. .. .. 140 ^.4 Regression of faecal N on crude protein contents of rations 142 UNIVERSITY OF IBADAN LIBRARY CHAPTER ONE . INTRODUCTION 1 General Introduction. It is generally agreed that the most pressing problem in the developing countries like Nigeria is the shortage of proteins, partieul the animal proteins in the human diets. The developing countries of the world have a population of 2,100 million persons of an estimated world population of 3»000 million people. The FAO (i960) reported that the developing countries and the developed countries had an average daily intake per head of about 58g and 90g respectively of total protein in I960, and that the daily animal protein intake per head in developing countries was about 9g compared with about 45g in the developed countries. Thus the average person in the developed countries has about five times as much animal protein as the average person in developing countries. The ratio of plant protein to animal protein (p/a ) in developing and developed countries are 5«3 and 0.35 respectively. This shows that the developing countries consume approximately 5»3 times as much plant protein as animal protein. Wright (l96l) raised two main objections to rhe use of plant proteins to help to solve the world food problem: (i) the foods containing them are less concentrated sources of protein on dry weight or calorie basis than meat and milk. (ii) the quality of animal protein is stated to be superior to that of plants. Cereals have been found to be deficient in essential UNIVERSITY OF IBADAN LIBRARY 2 amino acids like lysine and tryptophan, and this results in low biological value for these proteins. Mitchell (1927) showed that the biological value of whole egg i3 94 and that of whole corn is 60. This shows that proteins from animal products have higher biological value than proteins from plant sources. In the developing countries there is a-protein-deficiency disease called kwashiokor in infants who have been weaned on atarchy 'diet~low in protein or of high calorie/protein ratio. The FAO targets for world food production in 1975 visualize a total annual consumption in human diets of about 125 million tonnes of protein of which about 38 million tonnes will be animal protein giving a P/A ratio of about 2.3* The huge increases over the I960 figure are to be obtained by an increase of about 13 million tonnes of animal protein. The major problem is the production of enough animal protein to meet the needs of those in developing countries. The FAO has found that the proportion of livestock to humans is higher in developing countries than in the developed countries. The problem therefore is not that of increasing the number but the productivity of the livestock. Some of the factors militating against livestock production in the tropics are lack of properly managed pastures, disease pests, unfavourable climatic conditions, lack of technical know - how, poor economic condition of the people and their government, and cultural factors. One of the most important of these UNIVERSITY OF IBADAN LIBRARY 3 factors in Nigeria is lack of properly managed pastures. By far the ■ost important natural grassland of the country is the savannah, which can be divided according to the luxuriance of the growth of the grasses, the rainfall, the length of the dry season and humidity, into the Derived guinea* the Guinea, the Sudan and the Sahel savannah. The savannah as a whole is characterized by a continuous sea of grasses with scattered trees. Oyenuga (1957) noted that while most of Nigeria (8jfi) is covered by natural grassland no serious attempts have been made to bring these grasses into cultivation. Nigerian pastures are left in their wild natural condition without the application of the well-tested findings on the cultivation of pastures that are in use in other parts of the world. In consequence, the natural grass species rapidly decline in nutritional value, becoming fibrous and coarse since they are neither grazed nor cut for fodder at their optimum stages of growth. Moreover, these tough, dry grasses are subject, during the dry season, to periodical burnings which result in an appreciable destruction of organic matter. Burning also destroys the surface organic matter of the soil, damages the trees scattered all over the savannah land, annihilates low bush, removes the shade which protects the land against the sun to conserve moisture and thus intensified soil ersion. If pastures are properly managed, then the protein levels in them could be increased. Most of the cattle are extensively managed and are still in the hands of the pastoral herdsmen whose system of management is largely UNIVERSITY OF IBADAN LIBRARY 4 traditional. Although most of the 3heep and goats in the Northern states of Nigeria belong to settled farmers, these animals have not become integrated into the farm economy. The direction of movement tends southwards as the dry season approaches and northwards during the rainy season. During this period, the animals put on weight, most of which is lost during the dry season when the grasses are dry and the crude protein content is very low. The FA0 (1962) studies on livestock production in Rhodesia, showed that considerable economic losses occur because of the seasonal variation in the availability of pastures for cattle. They also showed that rapid gain in summer is lost during winter. If the loss per animal is multiplied by the number of animals which suffer such losses, the overall economic losses would be appreciated. Oyenuga (1957) lias shown that the tropical grass species are low in crude protein and high in crude fibre when compared with temperate grasses cut at similar stages of growth. Protein is an essential body - building component of the animal. Maynard and Loosli (1969) showed that protein contributes 15 - 21/c of the gross composition of the body of domestic animals. Protein is required for maintenance, growth and production. The maintenance requirement must replace the endogenous urinary and the metabolic faecal losses. While the urinary losses are considered to be reasonably constant per unit metabolic size . 0.734,), the faecal losses are variable according to the nature of the ration and dry matter consumed. UNIVERSITY OF IBADAN LIBRARY 5 It is directly proportional to the dry matter consumed by the animal. To prevent undue weight I033 in domestic animals during adverse conditions and to enhance maximal growth during the rainy season, protein supplements ai® often supplied to livestock. Such concentrates as soybean meal, groundnut cake, palm kernel cake and other by-products of industry are often fed to ruminants. It is known that the rumen microorganisms convert these into high quality microbial protein which is digested in the intestines. The amino acids derived from the microbial protein are absorbed. The endowment of the ruminant animals with the capability of converting such plant proteins and even non-protein nitrogenous substances into proteins of high biological value is a great asset for the development of the livestock industries. Conversion of grass and other forages which are normally not required by other farm animals and certainly not acceptable to man will go a long way to increased livestock production and increased output of meat and milk to augment the much needed animal proteins in developing countries like Nigeria. 1.1.1 Breeds of sheep in Nigeria. The breeds of sheep in Nigeria are classified into two main groups: (a) the West African long-legged sheep of which the West African Uda is an example. (b) the Jest African dwarf sheep. The West African long-legged shee^ i There are many types within this group, such as the Arab, the UNIVERSITY OF IBADAN LIBRARY 6 Tuareg, and the Uda or the Fulani. Thejjare all characterized by their long legs, and are kept in the savanna north of latitude 10 oh by the nomads. There were some 3.2 million head# of Uda or Fulani in Northern Nigeria in I960 (Oyenuga, 1967). The Uda stands between 65 cm and 90 cm in height with body weights varying from 30 kg to 50 leg.T hey vary in colour between brown and white and white spotted. The Nest African dwarf sheep: The West African dwarf 3heep is widely scattered around human settlements in the forest and derived Guinea savanna zones of West Africa south of latitude 14°N. 'The Federal office of statistics estimated the population of the breed in southern Nigeria to be 1.82 million in i960. The West African dwarf sheep is small, varying between 40 cm and 60 cm in height and beween 20 kg and 30 kg in weight. The Colour m y be white, white spotted with black, black or brown. The rams are heavily maned at the neck and chest and have close, spiral hdms. The ewe is hornless. .1.2 Importance of sheep to the economy of Nigeria. The Federal Office of Statistics estimated the livestok population of Nigeria in 1960 to be 10 million cattle, 15 million goats and 5 million sheep. It wa3 also estimated that 1.6 million cattle, 6 million goats and 3.2 million sheep are slaughtered annually UNIVERSITY OF IBADAN LIBRARY 7 in Nigeria. The great importance of sheep to the economy of Nigeria would be appreciated if the cost of the sheep slaughtered is given a modest estimate of MJ2 million. Shaw and Colvile (1950) reported that 1.5 million raw untanned hides of sheep were exported from Nigeria in 1947. Thus, the sheep is not only a supplier of the much - needed animal protein in Nigeria but the hides also earn foreign exchange to strengthen the economy of Nigeria. .‘.5 The Management of the West African dwarf sheep. The West African dwarf sheep owe their existence to their ability to survive periods of drought and semi - starvation. The sheep in Nigeria have not been subjected to any good management but are left to roam around feeding on poor grasses supplemented with waste products available from kitchens or traditional food - processing industries, such as the bran from the milled maize, Guinea corn or millet or the peel from cassava■* *»and yams or other root crops. Considerable energy is expended in seeking food, and this reduces from the limited food intake the amount available for productive purposes. They bed down at night in market sheds or other secluded places, by the trunk of shade trees, in the dry season, outside. It is evident from this lack of management programme that the economy of their living is not such as results in rapid growth and early maturity. If Nigeria is to supply her rapidly increasing population with adequate animal protein, the present state of lack of management of sheep UNIVERSITY OF IBADAN LIBRARY a should not continue. There is a need for intensive management programme. This will bring about rapid growth and early maturity, and decreased mortality of lambs due to diseases and poor nutrition. Efforts aimed at intensive management of sheep have been few and confined to government agricultural stations and the universities. Reports of the performances of the West African dwarf sheep have been few but rate of live weight gain of 1 . 1 kg per week had been obtained with lambs (Oyenuga, 1967). This shows that with good management, the quality and quantity of sheep and sheep products could be increased in Nigeria. 1.2 LITERATURE REVIEW 1.2.1 MICROBIAL FERMENTATION AND PROTEIN DIGESTION IN THE RUMINANTS. The digestion, absorption, metabolism and utilization of nitrogenous compounds in the ruminant is very much dependent upon the rumen microbial population. Nitrogenous compounds are degraded to varying extents in the rumen and some of the products of microbial degradation are metabolized by the microorganisms, some are absorbed.' through the rumen wall, while othersmove down the alimentary tract. 1.2.1 ,1 Proteolysis and deamination of amino acids. McDonald (194S) demonstrated that the rapid degradation of casein and the much slower degradation of zein was accompanied by the formation of ammonia and fatty acids in the rumen. C 14 - labelled casein used in in vitro bacterial studies resulted in the release of r - labelled UNIVERSITY OF BADAN LIBRARY fatty acids as well as ammonia. The proteolytic action of' the - > rumen micro-organisms resulted in the liberation of amino acids from protein when toluene was added to the reaction mixture to inhibit the deaminase which otherwise would have caused the liberation of ammonia. However, amino acids are not all rapidly deaminated in the rumen. Of the amino acids tested individually, aspartic acid was observed by Lewis (1955) to be more rapidly deaminated by washed bacterial cells than other amino acids. El - Shazly (1952) observed that when alanine and proline were incubated together with washed rumen bacterial cells, more ammonia production occurred than when they were incubated separately. This suggests the possibility of a "stickland type" of reaction which involves the oxidative deamination of one member of an amino acid pair and the reductive deaminatinn of the second member. Decarboxylases have been known to effect the decarboxylation of lysine and ornithine. The 0̂ - amino groups of these two amino acids were removed by rumen micro-organisms to yield the corresponding amine derivative of the fatty acids. Tryptophan was found to yield indole. Looper, Stallcup and Reed (1959) shOi?ed that in vitro deamination of amino acids occurs rapidly with asp( - ketobutyric acid + NE + h2 ‘.2.1.2 Conversion of Feed Protein to Microbial Protein in the Rumen. The reports of McDonald (1954) and McDonald and Hall (1957) indicate that the extent of conversion of dietary protein to microbial protein can vary considerably among the protein supplements. These investigators used zein to determine the extent of conversion of zein to microbial protoin. The use of zein facilitates the separation of feed protein and microbial protein because zein is soluble in aqueous ethanol, and contains no lysine whereas microbial protein is insoluble in aqueous ethanol but contain lysine. ‘The proportion of zein in the abomasal sample is the proportion not degraded in the rumen. UNIVERSITY OF IBADAN LIBRARY + O 11 The investigators found that about 40fo of zein is converted into microbial protein. Ely, Little, YJoolfolk and Mitchell (1967) also estimated the extent of conversion of dietary zein to microbial protein and they obtained the values of 26.3 and 30.57° with high cellulose and high starch rations respectively. The amino acids present in the abomasal fluid result from a mixture of dietarj* and microbial protein present in the fluid. The observed conversion of zein to microbial protein is probably less than would occur in animals fe^d ordinary ration. This is because of the low solubility of zein in rumen liquor, its formation of a glutinous, fibrous mass when warmed to body temperature, thus reducing the surface area for enzymic attack, and also its deficiency in lysine and tryptophan. All these render zein unsuitable for conversion to microbial protein, feller, Gray and Pilgrim (195£>) have determined the extent of conversion of plant nitrogen into microbial nitrogen in the rumen of the sheep. They were able to separate plant nitrogen from microbial nitrogen by using E diaminopimelic acid (DAP), as a marker. The marker has been found to be present in bacteria (Work and Dewey, 1953) but not in protozoa or fodder. Lignin was also used as a marker to determine the extent cf contamination of bacteria by fine plant particles. This is possible because if the amount of lignin is known, and the ratio of lignin to plant nitrogen is also known, the plant nitrogen present in the bacterial sample can be estimated. The following schematic representation shows UNIVERSITY OF IBADAN LIBRARY the distribution of nitrogen in the rumen content of a sheep killed 7 hrs after feeding. NITROGEN IN RUMEN. — ! i jfASEED FIBRE HASHING SOLUBLE NITROGEN • 4 3$ 51 % 6/s Lignin indicates that 12% is plant ~ N jAP indicates that 58fo of N was bacterial r _ ] 1 I Bacterial -N Plant-N Plant-N Microbial N 25$ 13/o 2% 49/a DISTRIBUTION: Plant N - 20% Microbial - N - 74$ Soluble - - 6$ There was a steady increase in microbial N at the expense of plant N as the time after feeding increased. It was assumed that the concentration of LAP - N in bacteria did not vary widely from the value of 0.62fi> used. Microbial N formed 82$ of the total N 16 hrs after feeding and this was the ms.s*5i» for the day, the minimum being UNIVERSITY OF IBADAN LIBRARY ) 13 61$. It indicates that the extent of conversion of plant N to microbial N is between 61 and 82$. Use can also be made ofo(~ E diaminopimelic acid to estimate the quantity of bacterial N entering the duodenum per day from the rumen. In this case, the duodenal samples would be collected over 24 - hr period and the Ns BAP ratio would be determined for the isolated bacterial specimen. The daily flow of bacteria from the rumen to the duodenum is estimated from the following equation: Bacterial N (g/day) = R x DAP (g/day) where R is the N: DAP ratio of isolated bacteria. Certain assumptions have been made in the use of DAP as a marker for bacterial N. They are: (1) DAP is present in bacteria but absent from other nitrogenous components of the rumen. (2) The N: DAP ratio determined for the sample of bacteria is truly representative of the total bacterial population. (3) The isolated bacterial preparation is not contaminated - with feed residues. Since the microbial proteins constitute the major portion of the nitrogen - containing compounds that reach the lower gastro - intestinal tract (Weller et_ al, 1958), factors which affect the microbial population also affect the availability of microbial protein to the ruminant. Blackburn and Hobson ^1960) and Warner (1965) showed chat UNIVERSITY OF IBADAN LIBRARY 14 the changes in the rations fed to ruminants can exert a modifying effect on the rumen microbiota. Both semi - purified diets and antibiotics hare been shown to influence concentrations of protozoa in the rumen (Bryant and Snail, i960) and to modifjr the bacterial population or metabolic activity (Purser, Klopfenstein and Cline,1965). Changes in ration did not however, modify the amino acid compositions of rumen bacteria or rumen protozoa (Weller, 1957; Meyer, Bartley, Deyoe and Colenbrander, 1967). However, the amino acid composition of protozoa has been found to vary (Poley, 1965; Heller and Harmeyer, 1964) while such values for bacteria have not been shown to vary (Purser and Buechler, 1966). Bergen, Purser and Cline (1968) investigated the effects of different rations on the amino acid composition, pepsin digestibility and protein quality of the rumen bacteria (b ), rumen protozoa (?) and the recoverable rumen miorobial cell mass (P + B). They found that the total protozoal counts were not significantly affected by ration changes. Within a microbial preparation, the amino acid compositions were not affected significantly by rations. The lysine content was higher in protozoa than in the bacteria. It was also suggested that for conventional type of rations, the microfauna represented a higher percentage of the total microbial protein whereas for a semi - purified diet, the microflora represented most of the total microbial protein. However, this suggestion cannot be accepted unequivocally without data UNIVERSITY OF IBADAN LIBRARY 15 on total bacterial and protozoal distributions and recovery from rumen contents. It m s found that the pepsin digestibilities of protozoa and protozoa plus bacteria preparations from sheep fed semi - purified rations were significantly lower than the pepsin digestibilities of equivalent preparations from sheep fed conventional type of rations. There were no notic^ble differences in protein quality of rumen bacterial preparations on different rations. This may be due to a failure to establish different bacterial population or to an incomplete recevery of bacteria of widely divergent protein quality. The cellulolytic bacteria which are generally more difficult to remove from rumen contents may be of relatively low protein quality (Bergen et al.j 1967). Meyer et al. (1967) shewed that feed processing does not alter the quality of the protein of bacteria or protozoa but affect the quantity of protein synthesized in the rumen. Hume, Moir and Somers (1970) investigated the quantity of microbial protein produced by the rumen micro-organisms by measuring the daily flow of protein through the omasum from the rumen. Thcyfed virtually protein - free diets at levels of 2 , 4, 9, l6g nitrogen/day in the form of urea in the ration. Casein hydrolysate was infused continuously into the abomasum through a length of surgical quality vinyl tubing. The rate of flow of digesta was estimated by reference to polyethylene (Pgr*) glycol ' ' when steady state conditions have been closely approached in the rumen. The marker wa3 injected into the rumen, and rumen fluid UNIVERSITY OF IBADAN LIBRARY 16 samples were taken for analysis before injection (To) and 1 , 2, 4, 8, 1 2 , 16, 20, 24 hr after injection. Prom this, the rate of flow of digesta was estimated. When a protein-free diet is given under equilibrium conditions, the amount of protein flowing from the rumen to the omasum daily may be equated with the daily production of microbial protein in the rumen. The small amount of protein entering the rumen in the saliva (McDonald, 1948) and by desquamation of the rumen epithelium (Phillipson, 1964) will introduce little error into this assumption. The amount of protein flowing from the rumen daily is then the product of the rate of flow of digesta and the concentration of protein in the digesta. Hume (1970) found that there was a linear respors e in the rate of flow of protein out of the rumen to increasing levels of IT intake ranging from 2 to 9g nitrogen/day but there was no further increase between 9 and I6g ll/day intake. In the protein - free diet, the protein flowing out of the rumen has low concentration of four and five - carbon branched chain and straight chain volatile fatty acids (VFA), parti­ cularly, isobutvric. It was suggested by Allison (1965) that branched - chain carbon skeletons cannot be synthesised by most rumen cellulo­ lytic bacteria. Hence protein synthesis in the rumen may have also been limited by the supply of these nutrients. The ration used by Hume (1970) contained 418g dry organic matter (O.M.) ingested daily. Assuming a protein yield of 15 - I6g/l00g O.M. digested in the rumen, UNIVERSITY OF IBADAN LIBRARY 17 the potential production of protein could be as much as 67g. However, the investigators found that only 48g protein were synthesized. Protein production in the rumen could be limited by factors other than energy and nitrogen. Sulphur and branched - chain volatile fatty acids have been suggested. The amount of protein producetion a protein - free diet i3 capable of satisfying the animal's requirement for protein if the intake of energy is adequate. Hume (1970) found that the supplementation of a virtually protein - free diet with a mixture of higher VFA resulted in the increased production of microbial protein from 71g to gflg/day. The flow of total N out of the rumen was also increased. There were no differences in N balance values. A negative correlation was found between acetic acid proportions and protein production (r = - 0.62). The addition of VFA did not result in the increase in efficiency of protein production from the energy available. The increased production of microbial protein when protein - free diets are supplemented with VFA has been explained as being due to the fact that the cellulolytic bacteria require branched chain fatty acids which could only arise from amino acids (El-Shazly, 1952). It is suggested that it was the higher VFA that was limiting on protein - free diets. Hume (1970) showed that dietary protein also affects microbial protein synthesis in the rumen. He found that protein production in the rumen of sheep fed on a virtually protein - free diet supplemented with UNIVERSITY OF IBADAN LIBRARY 18 urea and higher VT’A and yielding 6OOg organic matter/day amounted to 90g/day. When gelatin was substituted for the higher VFA and 5Q& of the urea - N, microbial protein production remained at similar level (91g/d), with casein, production increased to I01g/day, and with zein to lC>4g/d. The protein production on VFA/urea and on gelatin ration may have been limited by the rate of synthesis of one or more amino acids by the rumen bacteria. This has been suggested to be the case with methionine by Loosli and Harris (1945)• Zein, being resistant to microbial proteolysis could contribute a great percentage of the protein passing the abomasum. It is however, deficient in lysine and tryptophan. Casein has amino acid composition not very different from that of microbial protein ( B.V 79). However, its high degree of degradation in rumen, and consequent loss through rumen wall makes a portion of it unavailable fVT microbial protein synthesis. The investigation by Parser (1970) on the micro-organisms as a source of portein for ruminants has been divided into three general areas. (a) a consideration of the amino acid composition and nutritive value of the rumen micro-organisms. (b) a discussion of the factors influencing the availability of the amino acids to the host animal's metabolic system. (c) a discussion of the importance of supply of energy and protein to the host animal UNIVERSITY OF IBADAN LIBRARY 19 .1.3 Nutritive values of microbial proteins. McNaught, Owens, Henry and Eon (1954) obtained the values of 74, 81 and 60 for bacterial true digestibility ('ID), biological value (b v ) and net protein utilization (NHj) repectively, and 90, 80 and 73 for \ protozoal TD, BV and NPU respectively. Loosli, Williams, Thomas, Ferris and Maynard (1949)> Black, Weiber, Smith and Stewart (1957) and Donnes (1961) showed that the amino acids isoleucine, leucine, lysine, methionine, phenylalanine, tyrosine, threonine, valine and histidine were metabolica.lly essential to the ruminant and could be synthesized from urgflby the rumen microbial population. The reported amino acid compositions of rumen bacteria are strikingly similar. Similar agreement between the amino acid composition of microbial preparations from animals receiving' different rations has also been reported (We Her, 1957; Purser and Buechler, 1966). L. similar type# of relationship has been observed vrith the amino acid composition of rumen protozoa by Weller (195T) and Purser and Buechler (1966). Slight differences are apparent in the amino acid composition of protozoa and bacteria. Lysine, leucine, phenylalanine and tyrosine are slightly higher in protozoa than in bacteria. These differences have been used by a number of investigators to explain the difference between the NHJ values of protozoa and bacteria. However, this is an incorrect interpretation as the BV's of the fauna and flora do not differ; their digestibilities only differ, giving rise to higher NPU for protozoa UNIVERSITY OF IBADAN LIBRARY 20 1.2.1.4 Factors influencing Microbial Amino Acid Availability Limiting amino acids of rumen mircobial proteins have been determined using the plasma amino acid score (PAAS) technique of McLaughlan (1964) which is reasonably reliable when these proteins were fed to ra't^s (Bergen et al., 1968a). The application of such results to ruminants nay not be strictly applicable. Extremely low plasma levels for histidine in the rats fed protozoal protein were recorded. Valine was also low in ra t#s on this dietary treatment. Histidine was in fact indicated as the limiting amino acid in animals fed protozoal protein and cystine gave the lowest value for the animals fed bacterial protein, with arginine, histidine, leucine and lysine also being low. These results implicate histidine, cystine, leucine, arginin/e, and lysine as potentially limiting amino acids in rumen microbial proteins. Protein utlization and hence the quality may be influenced by a number of factors. (a) In ruminants, energy (VFl) and amino acids are absorbed from 4 different sites in the alimentary tract, that is, from the rumen and from the intestine whereas in a monogastric animal they are absorbed from the scene site (intestine). (b) rate of release of specific amino acids from microbial protein. (c) amino acid composition effect upon absorption, and (d) selective absorption of essential amino acids as compared with non-essential amino acids. UNIVERSITY OF IBADAN LIBRARY 21 Miller and Payne (1964) have discounted the practical importance of the timing of nutrients in large animals, but since amino acids and energy are absorbed from different sites in the ruminant, this aspect of nutrient timing and a possible effect upon nutrient utilisation is worthy of investigation. Differences in rates of release of three amino acids from protozoal, bacterial, and egg protein have been studied by digesting each in pepsin for 180 min and then in pancreatin^ (Purser 1970). It was shown that pepsin is particularly ineffective in releasing arginine from protozoal and bacterial proteins but arginine was freely released from egg protein. Glutamic acid was released fairly steadily over the entire digestion period. Alanine was released veiy easily from bacterial protein, about twice the rate of- release from protozoal and egg protein. The composition of the amino acids presented to the intestine can markedly influence both the rate of absorption and the composition of the amino acids absorbed. The amino acid content of both the duodenum and ileum was expressed as a percentage distribution and the ileal value expressed as a percent of that of the duodemun. The duodenal values are thus shown as 100. It was found that the essential amino acids were lower in the ileal content and the non-essential amino acid higher. Consequently in the passage of digesta from duodenum to ileum, a greater relative quan­ tity of essential amino acids (EAA) than non-essential amino acids (NEAA) were absorbed. The EAA comprise about 50% of the total duodenal amino acids and only 44% of the ileal amino acids. UNIVERSITY OF IBADAN LIBRARY 22 There is some evidence for the existence of an interraction between amino acid utilization and specific metabolic energy as shown by Porter, Purser and Cline/ (1968). In their investigation, a specific energy source was infused into the carotid artery of sheep and the plasma amino acid changes then expressed as ratios of original values (Plasma amino acid indices),Glucose was found to be more effective than propionic acid in decreasing the amino acid concentration of blood. Butyric acid was less effective than propionic acid, and acetic acid has no effect on the plasma amino acids. This means that of all the sources of energy mentioned above, glucose is best utilized in the metabolism of amino acids. 1.2.1.5 Relative Supply of Amino Acids and Energy. It has been found necessary to know the interraction between amino acid utilization and availability of energy source. The following assumptions have been made. (1) Microbial cell material synthesized in the rumen contain 65.4f3 crude protein/ (Hungate, 1966). (2 ) Microbial protein passed from the rumen to the alimentary tract has a digestibility of 80$/ (McNaught et al,1954). (3) Abomasal secretions amount to 1 to 2g N/day and must be deducted from quantities calculated from duodenal material (Kogan and Pnilipson, 1969). (4) Adjustment has to be made for some protein that escapes rumen microbial degradation and digested in the intestine. In most caes, UNIVERSITY OF IBADAN LIBRARY 23 maximal conversion of available dietary protein to microbial protein was nearly achieved. (5) It is assumed that 75$ of the energy absorbed by the animal i3 absorbed from the rumen and 25$ from the rest of the tract (Hogan and Weston, 1967a). (6) That 1g of digestible dry matter contains 4*3 kcal. Hungate (1966) stressed the fact that the rumen system is aerobic which places a limitation upon the maximum possible conversion of dietary nitrogenous material to microbial cellular material. Aerobic fermentation takes place with a maximum cell yield of 15$ ^Hungate, 1966) whi*h is equivalent to 9.84$ (65.4f° of 15) yield of protein in material leaving the rumen. Hungate (1966) has calculated the ratio of digestible protein to energy to be 18: 1 when protein is expressed in grams and energy in megacalories. He calculated it as follows using the assumptions previously mentioned that is assuming 11fy cell yield per lOOg dry matter fermented in the rumen. (1) Protein yield per 100 g fermented = 15 x .65f = 9.84 g. (2) Digestible protein per 100 g fermented = 9.84 x 80 = 7.87 g. (3) Digestible protein per 438 (lOOO x 4.3) digestible kilocalories = 7.87 g. (4) Therefore, digestible protein per 1000 digestible kilocalories = 1403000 x 7.87 = 18.3 g. UNIVERSITY OF IBADAN LIBRARY 24 Therefore, digestible protein per digestible megacalorie = 18.3g. tome of the factors influencing the conversion of dietary N to microbial N in the rumen according to Hogan and Weston (1967a) are: (a) the time spent by the feed particles in the rumen (the longer the time, the greater the conversion). M The resistance of dietary N source to deaminative degradation, the more resistant, the less the conversion. (c) Availability of N for microbial protein synthesis. (d) Energy availability for rumen fermentaion. (e) Presence of growth factors for instance, minerals such as cobalt, and also vitamins such as B12. (f) The population composition of rumen micro-organisms. Ruminant animals are poorly suited to the use of sources of nitrogen when present in large amounts because of the degradation of these nitrogenous sources and subsequent loss of N throjagh the rumen epithelia in the form of ammonia. However, they efficently utilize sources of N when these are present only in adequate amounts. When ruminants are fed once daily, there are a number of phases of digestion in the fore - stomach which correspond to the fermentation of various constituents of the ration at different rates (Wallcer, 1965). Conseqeatly, during the day, there are periods when energy becomes available to the microbial population at differing rates. Using rumen fluid, Walker (1965) has shown that when an excess of readily fermentable carbohydarate is UNIVERSITY OF IBADAN LIBRARY 25 available as is the case shortly after feeding, only a small proportion of the energy made available by fermentation to VFA is used for growth, the greater part being used for intracellular polysaccharide synthesis, A similar low rate of protein synthesis in relation to energy supply has been demonstrated in whole rumen contents collected shortly after feeding, while later in the day, the protein/energy ratio increased (walker and Nader, 1968), Walker (1965) introduced the concept of an interrelationship between rumen microbial cell synthesis and adenosine triphosphate (ATP) made available during the degradation of feed materials. The initial and considerable decline in the protein/ATP ratio in the early part of the feeding period indicated wastage of available energy or diversion to purposes other than microbial growth. Part of the available sugar is utilised by rumen micro-organisms for the synthesis of intracellular polysaccharide. There has been found a dramatic increase in the proportion of polysaccharide per unit of DMA in the rumen liquor organism during the first phase of digestion and this corresponds to the part of the decline in protein/A'TP ratio. Unfortunately, because of the lack of methods for distiguishing between microbial and plant polysaccharde, present in whole rumen content, it is not possible to quantitate polysaccharide synthesis and relate the energy used for this purpose to that used for protein synthesis. The protein/energy ratio increases between 8 and 12 hrs after UNIVERSITY OF IBADAN LIBRARY 26 feeding. At the same time the polysaccharide content of the cells returns slowly to the pre-feeding level, which would suggest that the environment of the microflora is becoming energy-limiting. Under such conditions, it would be expected that reserve polysaccharide could be used as a readily available energy source. Forest (1969) has shown for a great many organisms in pure culture that under energy-limiting conditions, the growth of an organism results in the production of 10 - 11g dry weight of cell material per mole of ATP available. ,Since in general, bacteria contain about 60$ protein, 6 - 6.6g protein per mole of ATP would be expected. If 81$ of the crude protein of bacteria is true protein, then the maximum truC- protein to ATP ratio is 5.3 g (0.81 x 6.6) protein per mole of ATP. UNIVERSITY OF IBADAN LIBRARY 27 2 UTILIZATION OP NON - PROTEIN NITROGEN BY TEE RUMINANT. 2.1 Non - Protein Nitrogen in ilmn.mn.t Rationa The concept that micro-organisms play a '.useful role in protein metabolism was put forward by Zuntz (1991) who expressed the view that rumen bacteria use by preference amides, amino acids and ammonium salts instead of protein, and that the protein supplied by a given ration was augmented as a result of the formation of protein in the bodies of bacteria and protozoa which were later digested. These early observations showed that the protein requirement of animals especially Eerbivora could be met in part by such non-protein nitrogenous (NPN) compounds as asparagine, urea and ammonium salts. Loosli _et_ al_ (1949) obtained specific evidence that microbial action in the rumen can synthesize from urea all of the ten amino acids which are essential .for rat growth. In so far as the microbial protein arises from NPN compounds such as urea, a distinct gain in amino acids available to the body results. The microbial protein is of high biological value (BY) as measured by rat growth. McNaught et al. (1954-) got the values of 81 and 80/C for the biological values of bacteria and protozoa respectively. This means that through the rumen microbial activities, rations of poor quality are enhanced in quality. Amino acids deficient in the ration are supplied by microbial synthesis. This explains why the protein quality of tho rations as fed is much less important in the case of the ruminant than in non-ruminant animals. Hoever, the microbial action results UNIVERSITY OF IBADAN LIBRARY 28 in some losses also. Some of the ammonia produced, in the rumen by protein degradation or from KPN compounds such as urea is absorbed into the blood stream and converted to urea in the liver. Most of the urea is lost in the urine and the rest is recycled to the rumen via the saliva and the walls of the rumen. Virtanen (1967) had shown that milk production could be maintained in cows given purified, protein-free feed using urea and ammonium salts as the sole sources of nitrogen provided energy and minerals are adequate. Deif, El - Shazly and Abou Akkada (1968) fed urea, casein and gluten in the diet of the sheep at levels which supplied 1.33g, 3*33g, 5»33g» 7.33g, 11,33g and 14.33g nitrogen per day to each animal. A nitrogen - balance experiment was carried out for each nitrogen level with each of the three sources of the nitrogen supplements. They found that the faecal nitrogen was lowest when area or casein was given whereas it was highest ■with gluten at levels of 11.33 and 14-.33g/day. This is to be expected because urea and casein are rapidly degraded in the rumen with the formation of ammonia, some of which is used for the synthesis of microbial protein while the rest is absorbed through the rumen wall into the blood stream and converted to urea in the liver. Thus, a large portion of the urea and casein nitrogen is lost in the rumen, hence, the low faecal nitrogen on these two diets. Gluten is not degraded in the rumen to a great extent and this accounts for greater faecal nitrogen on this diet than on diets of urea and casein. The fact that casein and urea are highly degraded in the rumen, UNIVERSITY OF IBADAN LIBRARY 29 and gluten is not, also explains why the urinary nitrogen was higher on casein and urea than on gluten. It also explains why absorbed nitrogen is greater on casein and urea than on gluten, k linear relationship existed between nitrogen intake and nitrogen retention up to nigrogen xntakes of 5.33, 11.33 and 7.33g/day for urea, casein and gluten respectively .i. linear relationship was also found to occur between absorbed nitrogen and nitrogen retention up to levels of 1.33, 5.33 and 7.33g/day for gluten, area and casein respectively. Leibholz and Naylor (1971) using early weaned calves found that the replacement of 20.1 and 39.2/6 of the meat meal protein nitrogen by urea was associated with a significantly greater weight gain of calves between 5 and 11 weeks of age. The inclusion of urea in the ration to 5 5 . °f the total nitrogen depressed both weight gain and the intake of the concentrate mixture. The source of carbohydrate was sorghum and at levels of 62.3 to 77.8^ of the ration. Also the faecal nitrogen was lowered, and the urinary nitrogen greater than in other urea rations. The concentration of branched chain amino acids in plasma was low on urea rations, so also was the concentration of free essential amino acids. Limiting factors in the experiment might have been carbohydrate to provide energy and carbon skeleton. .2.2.2 Metabolism of Ammonia Nitrogen by Rumen micro-organisms. The manner in which the liberated ammonia from protein and non-protein is utilized in the synthesis of amino acids ia poorly understood but available evidence suggests that ammonia is a starting UNIVERSITY OF IBADAN LIBRARY 30 material for the synthesis of amino acids which are subsequently incorporated into microbial protein (Loosli et al.1949). On the basis of available information on the synthesis of amino acids by animal tissues and bacteria, it seems probable that in the prresence of ammonia and a keto acid such as ,X~ ketoglutaric acid, rumen micro-organisms synthesize glutamic acid through reductive aminaiion. The occurence of many keto - acids including pyruvic acid and ketoglutaric acid in the rumen liquor may be offered in support of this view. Synthesis of other amino acids would be expected to occur through transamination reactions involving the appropriate keto acids and glutamate Evidence of transaminase activity has boen presented by Otogald., Black, Goss and Kleiber (1955)• Investigations by Allison and Bryant (1963) have shown that cellulolytic rumen bacteria, Ruminococcus flavefaciens required either isovalerate or isobutyrate for growth but that neither 2 - ketoisovalerate, 2 - ketoisocaproate nor leucine supported the growth of these organisms. The organism failed to incorporate labelled leucine into protein but labelled isovalerate or isobutyrate is required because of inability of the organism to synthesize isopropyl group, Suphur - containing amino acids (SAA) are structural units of rumen micro-organisms as well as of ruminant tissue protein. It has been shown that these amino acids can be synthesized by the rumen micro-organism • t utilizing inorganic sulphur to synthesize cysteine, cystine and methionine. UNIVERSITY OF IBADAN LIBRARY 31 Lambs fed rations containing urea and inorganic sulphur as th® sole source of sulphur were found to produce normal wool growth. Orally administered labelled sulphur has been found to appear in the cystine of wool. Block, Stekol and Loosli (1957) reported that S 35 fed as Sodium sulphate to a lactating goat was detected in cystine and methionine of mi11c protein. These investigators also showed that was incorporated into rumen micro-organisms of the sheep. Emery, Smith and Huffman (1957) found that S"^inorganic sulphate was synthesized more rapidly into cystine than into methionine. Lewis (1954) reported that reduction of sulphate to sulphide was brought about by rumen micro-organism, and the sulphide was believed to be ’used in sulphur amino acid synthesis. Although the ability of rumen microbial population to synthesize amino acids from ammonia nitrogen has been shown (Loosli e_t al. 1949), evidence exists to indicate that some supplementary organic nitrogen is required for maximum nitrogen utilization. The nitrogen requirement of most bacteria can be met by ammonia but some bacteria also require amino acids and evidence has been presented to show that growth stimulation of some species may be brought about by peptide (Bryant' and Robinson, 196l). Supplementation of high urea ration with organic nitrogen may result in the development of a broader spectrum of rumen bacteria by providing nutrients required by some of the most fastidious species. The possibility also exists that a general improvement in rumen microbial metabolism might occur by virtue of the supplementary organic nitrogen UNIVERSITY OF IBADAN LIBRARY 32 supplying a irate-limiting nutrient. Ammonia is an essential nutrient for the growth of Bacteroid.es succinogenes, Ruminococcus flavefacicn3, JAminococcus alhus. Bacteriodes amylophylus, Nethanobacterium .rumiiiantium and Bubacterium ruminantium (Bryant and Robinson, 1963; Hungate, 1966). Addition of nitrogenous sources yielding ammonia stimulated in vitro digestion of cellulose and starch. Both cellulo^tic and amylolytic activities in vitro of mixed rumen micro-organisms were increased when urea replaced soybean meal as the sole crude protein supplement, showing that ammonia is important in the nutrition of both cellulolytic and amylolytic rumen bacteria. Synthesis of amino acids from ammonia by rumen micro-orgamsn3 requires the presence of ammonia, carbon skeleton and energy. Utilization of carbon from carbohydrate, (Hoover, Kesler and Flipse, 1963), carbon dioxide (Huhtanen, Carleton and Roberts 1954; Otogaki, at al 1965), Isovaleric acid, acetate and other volatile fatty acids (Hoover et al. 1963) indicates that carbon from a wide variety of sources could be used for synthesis of amino acids. However, synthesis of leucine from isovalerate (Allison, Bucklin and Robinson, 1966), isoleucine from 2 - methylbutyrate (Hungate, 1966), valine from isobutyrate (Allison and Bryant, 1963), phenylalanine from phenylacetate (Allison, 1965) and tryptophan from Indole - 3 - acetate (Allison and Robinson, 196?), indicates a requirement for certain specific carbon skeleton in the synthesis of certain amino acids. UNIVERSITY OF IBADAN LIBRARY 33 Energy for amino acid synthesis is provided by carbohydrates and other organic compounds in the form of ATP. Hungate (1966) estimated that 1.1g microbial nitrogen is utilized for synthetic purposes for ©ach 100g of carbohydrate fermented. •2.3 Factors Affecting the Utilization of Ammonia in the Rumen. Recent studies have been concentrated on factors which will promote the maximum bacterial synthesis of protein in the rumen to provide for the more effective use of rations of poor quality protein and parti­ cularly non - protein sources of nitrogen such as urea. A readily available source of energy i3 necessary for the efficient utilization of the end-products of protein fermentation. Pure starch or starch feeds such as cereals, cassava and potatoes are usually most satisfactory. Molasses or sugars are less satisfactory because they pass out of the rumen too rapidly. On the other hand, cellulose is made available too slowly. Rations low in protein and high in readily available carbohydrate are most favourable to protein synthesis in the rumen. In ruminants, t is generally considered that soluble carbohydrates exert a positive influence on protein metabolism. Addition of readily available carbohydrate to protein - rich rations fed to ruminants, wa3 followed by a depression in the concentration of ammonia in the rumen (Chahers and Synge, 1954)* The yield of protein produced by incubating ammonium salts with rumen liquor can be markedly increased by the addition of readily available carbohydrate. Nitrogen retention also increases consistently by supplement of readily available carbohydrate. Lower UNIVERSITY OF IBADAN LIBRARY • -J 34 concentrations of blood urea are also observed in ruminant receiving a supplement of readily available carbohydrate. The observed inhibition of ammonia accumulation in the rumen has been explained by the fact that unionised ammonia molecules pass through rumen epithelia much quicker than the ionised form (Lewis, Hill and Annison 1957). At high pH, the ammonia molecules are mostly present in the unionised form. The presence of glucose or its derivative, lactic acid, lowers the pH and the ammonia molecules are mostly present in the ionized form and their passage through the rumen epithelia i3 much delayed, giving time for the rumen micro-organism to incorporate ammonia for microbial protein. The pH of the rumen liquor also affe^cts utilization of ammonia. The pH affects the production of ammonia and also the absorption of ammonia. Reis and Reid (1959) found that high pH favours ammonia production in the rumen. The optimum pH for ammonia production in the rumen varied between 6.0 and 7.0. The observed effect of pH is on deamination as well as on proteolysis. The enzymes concerned with the deamination of amino acids are affected by the pH of rumen liquor. Warner (1955) Las shown that pH affects the rate of proteolysis and that optimum pH range is 6.5 to 7. The pH also affects the rate of growth of bacteria in the rumen. Annison (1956) showed that the formation of ammonia in the rumen varies with the type of protein - rich supplement. He compared casein, UNIVERSITY OF IBADAN LIBRARY 35 groundnut meal, herring meal and flaked maize gluten. He found that the groundnut meal can be deaminated to an extent equal to or greater than casein.* This is because groundnut meal is also very soluble in rumen liquor. Maize gluten yielded very low level of ammonia. Herring meal is intermediate between groundnut meal and maize gluten. Protein supplements such as casein and groundnut meal are easily degraded giving high levels of ruminal ammonia. Urea is easily hydrolysed by the urease of the rumen giving high levels of ammonia. Therefore, the more soluble the protein supplement in the rumen liquor the more ammonia is produced. The method of processing of the protein supplement also affects the rate of degradation of the protein supplements in the rumen. Formaldehyde - treated casein has been shown to be less soluble than untreated casein and therefore gives lower levels of ammonia in the rumen that* untrsated casein. 1.2,2.4 Absorption of Ammonia Through the Rumen Wall. The absorption of ammonia across the rumen irall was reported by McDonald (1948). It is influenced by both the concentration gradient (Lewis, Hill and Annison, 1957) and pH of the rumen liquor. Ammonia is a weak base with a pKa of 8.80 to 9«15. An increase in pH causes the ammonium ion (NH4+ ) to be converted to ammonia (NH^), and this is rapidly absorbed. Absorbed ammonia is carried via the portal circulation to the liver where it is converted to urea, Hogan (1961) has estimated that UNIVERSITY OF IBADAN LIBRARY 36 if efferent blood contains 1.5 mg NHj - N per 100 ml and the rate of flow is 200 ml/minute, then ammonia absorption is 4.3 g/day. The rate of saliva urea secretion has also been estimated at 0.5g/day (Hogan, 1961). The deamination of protein and the hydrolysis of (NPN) substance such as urea in the rumen forms large amounts of ammonia which if allowed to accumulate would be highly toxic to the animal. Ruminant blood contains about 1.5 mg - N/100 ml blood in the ruminal vein but only traces about 0.1 m - mole/litre in the peripheral circulation (Chalmers, 1954). It has been shown that a concentration of 0.4 to 0.5 m - mole per litp© is toxic to the sheep. It is therefore, important that the animal detoxifies this ammonia in the liver before releasing it into the systemic circulation. This is done by its conversion into urea. Krebs and Henseleit (1932), working with liver slices, established the general chemical mechanism by which ammonia is converted to urea. They discovered that the rate of urea production in"liver slices incubated with ammonium salts, bicarbonate and lactate, was increased by addition of ornithione or citrulline, and that arginine was an intermediate product of the reaction. It was also observed that the quantity of ammonia disappearing was equivalent to the urea formed. On the basi3 of these observations, Krebs and Henseleit (1932) proposed a cyclic mechanism for urea synthesis, involving ornithine, citrulline, arginine, ammonia and carbon dioxide. It was found that 2 molecules of ammonia and UNIVERSITY OF IBADAN LIBRARY 37 1 molecule of carbon dioxide are converted to a molecule of urea for each turn of the cycle and the ornithine is regenerated. Therefore, synthesis of urea involves the primary fixation of carbon dioxide and ammonia. It must be known that of the two nitrogen atoms present in a molecule of urea, an atom comes from ammonia, and the other from aspartic acid and could be shown as follows: KHj + hco3" COOH C\OOH \CH RH0 C H 2 ______ > | — NHp if •‘ 2 H 2 ° * 'C OOH COOH I FUMARIC ACID ASPARTIC ACID UREA 1.2.2.5 Influence of Ruminal Ammonia Level on the Concentration of Ammonia and Urea# in the Blood Comparatively, little attention has been given to the quantitative treatment of the relationship between the ammonia concentration in the rumen and in the portal blood or to the loss of dietary nitrogen in the form of ammonia. Lewis (1955) showed that there was a correlation between rumen ammonia and portal blood ammonia concentration over a wide range of rumen ammonia concentration and that spill - over UNIVERSITY OF IBADAN LIBR ~— AR0 Y ii 0 of ammonia into the systemic circulation occurred at relatively low portal ammonia concentration. At higher rumen ammonia levels, it was possible to correlate rumen, portal and peripheral blood ammonia and to relate the levels to the onset and developmentof toxic symptoms. Lewis (1955) determined rumen ammonia, portal blood ammonia, peripheral blood ammonia and blood urea of sheep fed rations giving rise to varying levels of ammonia during fermentation in the rumen, ie, high, medium and low levels of ruminal ammonia. He found that changes in rumen ammonia concentration were paralleled by changes in portal blood ammonia concentration although the ammonia concentration in the rumen and portal blood differed widely. There was'no'significant change in the ammonia content of the arterial blood, neither were there any significant differences between the concentrations of urea in the portal and peripheral blood. However, blood urea levels were found to increase with incease in ruminal ammonia levels. The investigator could find no regular pattern between the varying concentrations of rumen ammonia and corresponding concentrations of ammonia in the peripheral, venous and arterial blood samples. Whereas the ruminal ammonia rose rapidly and reached a peak in 2 hrs after feeding and later declined slowly over the next 6 hrs, the portal ammonia concentration showed a distinct lag period of 2 hrs before any increase in ammonia concentration occurred UNIVERSITY OF IBADAN LIBRARY 39 (Lewis 1955). In all these experiments, no significant changes in the ammonia concentration of peripheral blood was reported although relatively high (60 m - moles/litre) rumen ammonia levels were attained on a number of occasions. However, at high ammonia concentrations, some ammonia could pass into the systemic circulation. It has been suggested that a hepatic ammonia threshold exists in the sheep and that if this threshold is exceeded, the liver can no more cope with the high level of ammonia brought to it and therefore the ammonia concentration in the peripheral blood rises sharply. In case where ammonium acetate was used to induce varying levels of ruminal ammonia, no significant changes in arterial ammonia concentration took place until the portal blood contained about 0.8 m - mole NH-j/liter of blood. Above this level, the arterial ammonia concentration increases at almost the same rate as the portal blood. When the arterial ammonia concentration reaches 0.4 to 0,5 m - mole/ lit|r®, respiratory difficulties arise in the animals, and beyond this death occurs, probably due to a disturbance in acid - base equilibrium caused by excessive ions. It is estimated that the amount of ammonia carried to the liver per day is about 14g. Even if a portion of this is returned to the rumen via the saliva and through the rumen epithelia as urea, the nitrogen loss still represents an appreciable proportion of the total nitrogen intake. UNIVERSITY OF IBADAN LIBRARY 40 Lewis et al, (1957) have shown that the level of blood urea in 3heep is relatively constant and is dependent upon the ration. The blood urea tends to reflect the over-all changes in ammonia production in the rumen. They used blood urea determination to assess nitrogen losses following the absorption of ammonia from the rumen and its use has been found to be of practical importance. Blood urea concentration has been found to be uniform over 24 hr period but that greater diurnal variation is found in those instances where the ruminal ammonia is very high, (Lewis et al. 1957). They could find no significant difference between the venous and arterial blood urea concentration but found a close association between ruminal ammonia and blood urea levels, To show that the rise in blood urea level was a direct result of ammonia^ absorption from the rumen and not to changes in total nitrogen intake, Lewis et, al _ (1957) used casein and zein at the same level of nitrogen intake but which give different patterns of ammonia production in the rumen. The same general correlation has been found between ruminal ammonia and blood urea levels whatever the ration is. Chalmers and Synge (1954) suggested a partial reduction of the rate of attack of feed ■protein by rumen micro-organisms by altering the solubility, degree of denaturations and particle size. Houpt (19$) replaced the rumen content of the sheep with saline solution. Arteriovenous urea differences indicate that blood urea moved into the saline solution and was hydrolySed by the traces of bacterial urease present in the UNIVERSITY OF IBADAN LIBRARY 41 m a n , Accumulation of ammonia in the rumen was measured and concurrent absorption of ammonia from the saline was calculated. The total of these two rates equals the urea nitrogen transfer rate into the rumen. He also found that sheep whose rations were supplemented with readily available carbohydrate utilized 53?° of urea injected into the blood streams whereas those whose rations were supplemented with poor quality hay utilized Only-22$* This shows that readily available carbohydrate is essential for utilization of urea. As a result of urea entering the rumen through the saliva and rumen epithelia, the nitrogen intake necessary to maintain life would be considerably lower for ruminant than non-ruminant animals. The feed - back of blood urea across the rumen epithelia depends very much on the levels of dietary N , the lower the level of dietary N, the more blood urea is being recycled; the higher the level of dietary N, the lower the rate of recycling from the blood. The observation that the decrease of blood urea concentration occurs at constant rates has enabled calculation to be made of the estimates of urea transfer into the rumen. The amount of urea which disappeared from the serum minus the amount excreted in the urine is the amount that moves into the rumen. Houpt (1959) using the above method of calculation obtained a value of 8 .3 m - moles as the amount of blood urea recycled to the rumen per hour. The ability of ruminant animals to utilize blood urea would enable their survival time to be prolonged especially those which live in habitats where for most part of the year, the vegetation is mature, dry, tough and contains very little nitrogen. UNIVERSITY OF IBADAN LIBRARY h2 1.2.3 INTESTINAL DIGESTION OP PROTEIN - N AND UTILIZATION IN THE RUMINANT Interest in the digestive tract of ruminant animals has usually centred on the forestomach, for it is the characteristic digestive organ of the ruminants and about two-thirds of the dige­ stible organic matter is fermented there. The ruminant intestine has been neglected because of the assumption that it resembles that of monogastric animals in its functions. However, the oapacity of the rumen and the metabolic activities of its micro­ organisms affect the flow and composition of digesta passing to the intestines to an extent that makes intestinal digestion in ruminants a distinctive process. Kay (1969) showed that (1) food is retained in the rumen for a long time and only flows to the lower gut slowly; (2) microbial activity in the rumen transforms the diversity of protein in the diet to a more uniform product passing to the abomasum; it also removes most of the digestible carbohydrate from the food so that very little sugar is absorbed from the intestine; (3) flow of digesta from abomasum is enormous, almost continuous and fairly constant in consistency and composition; pancreatic secretion is equally continuous, abomasal secretion of diges­ tive fluid is continuous and the intestinal content remains acid throughout the upper part of the small intestine. UNIVERSITY OF IBADAN LIBRARY 43 (4) large amounts of water and salts are secreted into the gut especially by the salivary glands and these must be efficiently re-absorbed mostly in the small and large intestines. The nitrogenous digesta flowing to the duodenum are largely of rumen microbial origin, though variously supplemented with unfermented food residues and digestive scretions. The faotors affecting the digestion of food in the intestine, therefore, are the extent of protein degradation in the rumen, the nature and quantity of microbial protein synthesized from dietary and endogenous nitrogen and the amount of endogenous c protein flowing to the duodenum. Heclcer 0 9 7 0 compared the deaminative, ureolytic and proteo­ lytic activities and rates of cellulolysis, carbon dioxide and methane production in the rumen with that of the large intestine. He found that the proteolytic activity of the large intestine is greater than that of the rumen contents. Some proteolytic activity was present in caecal cell-free liquor. Deaminase activity was greater in rumen than in caecal oontents. The urease activity of rumen contents was greater than that of oaeoal contents. The rate of carbon dioxide and methane production was, however, higher in oaeoal contents than in rumen contents. The rate of cellulose breakdown in vivo were similar for rumen and caecal contents. Thus it is seen that the large intestine, though little studied, is also capable of enormous digestion, the principal difference UNIVERSITY OF IBADAN LIBRARY 4 4 being that since hydrolytic digestion of protein, starches, sugars and fats occur before the digesta reaches the caecum, the amounts of these substances reaching the caecum are likely to be small or negligible. Hogan and Weston (1967) have shown that the amounts of non­ ammonia crude protein (NACP) passing the abomasum was similar whether the ration contained 7.8 or 1 9 .8̂ crude protein (CP), ftrskov, Fraser and McDonald (1971 ) found that the amount of NACP (Y^g/day) disappearing from the small intestine increased with protein intake (X g/day) according to the equation Y 1 = 2.12 X - 0.0057 X2 - 83. reaching a maximum when there was CP in the dry matter of the feed. Andrews and prskov (1970a) showed that when the protein concentra­ tion of the diet was increased at high constant energy intakes, the growth rate and the retention of nitrogen in the body increased The level of protein was found to have n o . significant effect on the disappearance of NACP from the large intestine. The apparent digestibility of crude protein increases with protein concentration It was not known whether increased absorption came from increase in microbial protein or from dietary nitrogen escaping fermentation Since they found that the protein used, soya be®n has higher digestibility than microbial protein, they concluded that the UNIVERSITY OF IBADAN LIBRARY 45 increments were due to soy-bean protein escaping fermentation in the rumen, 1.2.3.1 The Rate of Flow of Digesta in the Digestive System of Ruminant Animals A mathematical study of the movement of particles and solutes through the digestive tract of the ruminant has been presented by Warner (1966). From his study, he showed that in a ’ steady - state' system, F = 0.693V _ _ _ _ _ (1) T where F = rate of flow from the rumen. V = volume of liquid in the rumen. T = Time for the equivalent of half of the liquid in the rumen to be transferred to omasum. When a water - soluble marker is infused continuously into the rumen, it was shown that F = i / c (2) R = v/f = 1.44 T ( 3 ) P = I X R _ _ ( 0 and X = it , (5) where I = rate of infusion of marker into the rumen. UNIVERSITY OF IBADAN LIBRARY h6 C = concentration of marker in the liquid leaving the rumen. R = the mean retention time of a population of marker molecules in the rumen. P = the quantity of marker present in the rumen (the rumen marker pool). K = is the fraction of the rumen volume transferred to omasum per unit time. In a "steady-state system", one estimate of F, V and T may be obtained by administering a single dose of an appropriate marker into the rumen and studying its rate of disappearance. If the marker is infused continuously at a constant rate, a number of estimates of rate of flow from the rumen can be made from the concentration of marker in the liquid leaving the rumen by using equation 2. After the continuous infusion is stopped, an estimate of T may be obtained by studying the rate of dis­ appearance of marker from the rumen. The value of T together with the estimates of the rate of flow during the continuous infusion, may be used to calculate the volume (v) of water in the rumen as indicated by equation (l). It was assumed that the concentration of marker in the liquid leaving the rumen and abomasum were the same as those obtained in samples of liquor taken from those organs. Rate of flow from the abomasum was calculated from equation (2) by substituting the marker concentration UNIVERSITY OF IBADAN LIBRARY in abomasal liquor for the marker concentration rumen liquor. The digestibility coefficients of the dietary nutrient for the entire tract have frequently been obtained by determining the ratio of a given food constituent to some indigestible marker in the food itself, such as lignin and the ratio of the cons­ tituent to the marker in the faeces. From them, the percentage of the nutrient digested is given as 100 - f 100 X ̂ T’ — ■*" --------- x ^ This method does not require quantitative collection of faeces, provided representative samples can be obtained. The markers most frequently used in ruminant digestion studies are lignin, polyethylene glycol, and chromic oxide. Chromic oxide is used either in powdered form mixed with the ration (Drennan, Holmes and Garrett, 1970) given in gelatin capsules (Putnam, Loosli and Warner, 1956) or impregnated on to paper (Cowlishaw and Alder, 1963). Chromic oxide has been shown to be associated with the solid phase of intestinal digesta (Harris and Phillipson, 1962) and can be easily and accurately determined. Polyethylene glycol associates itaelf with the liquid phase of digesta (Hyden, 1956) UNIVERSITY OF IBADAN LIBRARY and the method of its determination have not given consistent results. Lignin has the advantage of being a plant constituent but suffers the disadvantage of being an ill-defined entity, the estimation of which is empirical. Johnson, Dinuson and Bolin (1964) examined the concentration of chromic oxide in all the sections of the gut of sheep after feeding and measured the rate of excretion of a single dose when given in paper form or as powder mixed with a pelleted ration. They found that the powder form moved through the gut significantly faster than the paper form and that this difference was established by different rates of passage from the rumen. Prom their results, it seemed as if the passage from omasum to abomasum of the paper form was similar to that of lignin. Consequently, chromic oxide concentration in the abomasum might be used to give an accurate estimation of digestion anterior to this point if the paper form were used. Johnson et al. (1964-) also found that the powder form yielded an abomasal concentration of chromic oxide only of that found when the paper was used and would lead to a large underestimation of digestion. Balch (1957) used the lignin-ratio technique to determine the extent of digestion in the reticulo-rumen of the cow. The results showed that about of the herbage dry matter was digested in the reticulo-rumai, and that in cows fed entirely on hay, the amount of nitrogen flowing out of the reticulo-rumen was UNIVERSITY OF IBADAN LIBRARY 49 greater than the nitrogen intake. Rogertson (1958) also used the lignin-ratio technique to determine partial digestion in sections of the alimentary tract of sheep using a slaughter method. He shaved that 4Qfo> and 1% of the dry matter of hay, mixed diet and concentrate respectively occurred in the rumen. Bines and Davey (1970) also using the same technique found that 6($ of straw diet dry matter was digested in the rumen. Drennan, Holmes and Barrett (1970), and Holmes, Drennan and Garrett (1970) compared the use of lignin and powdered chromic oxide as markers for estimating the magnitude of digestion in the rumen and intestines using slaughter technique in sheep. They found that the results obtained using lignin as marker was higher and more consistent than those obtained using chromic oxide powder, and suggested that the poor results obtained by using powdered chromic oxide might be due to its very rapid or uneven passage from the rumen. They found that about 7Q$ of the organic matter digested occurred in the rumen. For studies with ruminants, chromic oxide paper appears to be suitable and promising, no doubt owing to the slow and sustained release of the oxide as the paper undergoes microbial digestion (Corbett, Greenhalgh, McDonald, and Florence, 1960). This has been confirmed by Langlands, Corbett, McDonald and Reid (1963) and Lambourne and Reardon (1963) who showed that chronic UNIVERSITY OF IBADAN LIBRARY 50 cxide in the form of impregnated paper gave a more even release of marker into the faeces. Digestibility of nutrients in the sections of the digestive tract is also estimated by the techniques of the re-entrant cannulation. For studies of digestion in the reticulo-rumen, the cannula is placed at the abomasum or duodenum so that all digesta from the reticulo-rumen can be collected. Digestibility of a nutrient is then calculated as the difference between the nutrient in food and the nutrient recovered at duodenal collection point. Similarly digestion in the small intestine is determined by placing cannulae at duodenum and terminal ileum, and the nutrient passing through the duodenal cqnnula nanus the nutrient reaching the terminal ileum is the amount of nutrient apparently digested in the small intestine. Digestion in large intestine is the difference between the total nutrient in terminal ileal point and in the faeces. Digesta may be totally collected at the collection points or samples of digesta may be collected at suitable intervals and pooled to give representative samples of digesta flowing through the portion of digestive tract. Chromic oxide either in powdered form or impregnated on to paper, or lignin is usually given so that digesta could be adjusted to percent recovery of the marker. Tills technique of the re-entrant UNIVERSITY OF IBADAN LIBRARY 51 cannulation has been widely used (Topps, Kay and G-oodal, 1968; Nicholson and Sutton, 1969; McRae and Armstrong 1970; and McRae, J970). It is not only useful for determining digestibility in sections of the digestive tract but also in studying biochemical reactions in the sections of the digestive tract, thus Hecker (1971) used sheep with ruminal and caecal cannulae to compare metabolism in the rumen and the caecum. Using the re-entrant cannulation method for determining digestibility, Hogan and Phillipson (i960) found that of the total dry matter digested in the sheep, 7C$ disappeared in the stomach, 11$ in the small intestine and 19$ in the large intes­ tine whereas the corresponding values as obtained by Topps et al. (1968) are 6l$, 22$ and 11$ for hay, and 65$, 11$ and 11$ for ooncentrate - fed animals in the stomach, small intestine and large intestine respectively. Several investigators (Nicholson and Sutton, 1969; Topps at .al., 1968) have reported that more nitrogen is recovered at abomasum than fed when sheep are given diets low in nitrogen but that substantial loss of nitrogen occurs in the rumen when the diet is rich in nitrogen. Ben - Ghedalia, Tagari and Bondi (1974), by means of cannulae placed in portions of the small intestine, were able to show that there were substantial increases in water, dry matter and total nitrogen in the section immediately distal to the UNIVERSITY OF IBADAN LIBRARY 52 pylorus and that these were caused by the inflow of bile, and pancreatic and duodenal juices. The net increase found beyond the entry of the common bile duct was 2.7g protein N and 2.0g non-protein N per day. The region 7 - 1 5m from the pylorus was found to be the region of most intensive absorption of amino acids, 60.5 of the essential, and l*.j$ of the non-essential amino acids passing through the region being absorbed. They also shovred that only small changes occurred in the region after 15m distance from the pylorus. McRae, Ulyatt, Pearce and Hendtlass (1972), in ten 21*. hr. collections of digesta entering the duodenum and eleven 21*. hr. collections of digesta reaching the ileum of sheep given dried grass showed that there were highly significant correlations between the 21*. hr. flows of chromium marker and the corresponding flows of dry matter, organic matter, nitrogen, gross energy, hemicellulose and cellulose at both sites. This has enabled the investigators to estimate the quantitative intestinal digestion in sheep. The reactions in the digestive tract are very complex and a knowledge of how they take place, the products formed, the utili­ zation of the products formed, and the factors that enhance the production is essential for adequate feeding of ruminant animals and hence meat production. UNIVERSITY OF IBADAN LIBRARY 53 ISOTOPIC METHODS OF DETERMINING THE UTILIZATION OF NITROGEN IN THE RUMINANT The isotopes that have been commonly used in nutrition studies are 15 n either in urea or ammonium salt form, 35 S, usually used as sulphate, and 32P usually in the form of phosphates- These isotopes have been used to study the synthesis or utilization of protein in the ruminant. In addition I ifC is also used to study the utilization of car­ bohydrates or other carbon-bearing materials. It is used either as 1 bC in urea or in glucose. White, Steel, Leng and Luik (1969) have used 1 bC glucose to study the kinetics of glucose metabolism in the sheep. Harrison, Beever and Thomson (1972) and Beever, Harrison and Thomson (1972) have used 35S as sodium sulphate to estimate the proportion of food and microbial protein in the duodenum of the sheep, while Landis (1968) had also used sulphur - 35 as sodium sulphate to study quantitative aspects of sulphur metabolism in the ruminant. Mathison and Milligan (1971) and Nolan and Leng (1972) have used 15N as Ammonium chloride or sulphate to study the ruminant digestion, while Land and Virtanen (1959) have used 15 as Ammonium nitrate to study the synthesis of milk protein from Ammonium salts. Lofgreen and Kleiber (1953) used 32P or N: P 32 ratio to determine the value of the metabolic faecal nitrogen of young calves UNIVERSITY OF IBADAN LIBRARY 5k All the isotopes in common use satisfied the basic require­ ment that: (a) the compound studied for instance urea, ammonia, glucose can be labelled in the required position with a suitable isotope, (b) the label is firmly attached to the molecule or or at least to that part of the molecule which is of interest to the investigator, (c) the amount of isotopic material introduced into the initial compound is such that it allows for considerable dilution before the concentra­ tion of the isotope is too low for accurate determination, (d) when radioactive, the rate of decay is sufficiently low to permit all the radioactivity determinations to be made with reasonable accuracy, while on the other hand, the radioactivity does ncrfe- persist sufficiently leng and with sufficient intensity to cause significant radiation damage to the tissues or cells under investigation or to any part of the experimental animal during the course of the experiment, otherwise the experiment is not truly physiological. UNIVERSITY OF IBADAN LIBRARY 55 The isotopic tracer method is one suited par excel­ lence for the study of biochemical reactions in the living cells. The major advantages of the method are: (a) The experiment can often be carried out under strictly physiological conditions on intact normal 'A animals. (b) With a few exceptions, the labelled compound has, for all purposes, the same biological properties and the same metabolic fate as the unlabelled compound. (c) The amount of isotope required is usually extre­ mely small, particularly with radioactive isotopes. (d) Laborious separation of radioactive compounds from tissues and tissue extracts is often unnecessary. (e) The precise origin of individual atoms in a compound produced by living tissues can oft*n be determined by isotope studies for instance the .N cf urea in blood or urine, the S or P atoms in proteins. There are some limitations, however, even though these are not of such a character as to reduce seriously the value of the isotopic tracer method as a general technique for agricultural research. These disadvantages are: (a) There is need for special technique and specialized equipment and these are usually expensive, for example Geiger counter, Mass spectrometer, Emis­ sion spectrometer. The compounds themselves for instance ”15 Cl or Na^ 32SO^ are expensive. UNIVERSITY OF IBADAN LIBRARY 56 (b) The method only involves following the label. The determination gives a measure of the amount of label in the sample analysed. It should not be automa­ tically concluded that the amount of isotope present in a particular tissue or cell gives a true measure of the concentration in that tissue of the substance originally administered to the animal. Also the isotope determinations give no direct information about the fate of any non-labelled parts of labelled molecules which have undergone disruption. These difficulties can often be overcome by the use of two or three different labels attached to different parts of the molecule of the compound studied, for example, Nitrogen and Sulphur in Ammonium -52S0̂ 4 (c) Radioactive isotopes may cause serious radiation damage to the tissues. This may apply to the tissues of the experimental animal or of the experimenter. « In the former case, the experiment may no longer be normal or physiological, and the results obtained may be largely due to a disordered metabolism of the irradiated or some indirectly affected tissues. UNIVERSITY OF IBADAN LIBRARY 57 (d ) Lack of a suitable isotope- There are few instances where it is impossible to find a suitable isotopic tracer for a biological investigation- Sometimes, however, a radioactive isotope which might otherwise be suitable has rate of decay of activity which is too short or too long for the experiment planned. In the former case, there would, be great difficulty in completing all the radioactive measurements before the labelled compound and its metabolites lose their radioactivity and in the latter case there will be a correspond­ ingly greater risk of radiation damage to the tissues. (e) Chemical non-identity of isotopes may not be strictly true. The physico-chemical differences between the isotope used as a label and the most abundant stable isotope of the same element may occasiona­ lly be sufficiently great to cause significant, quantitative differences between the metabolism of the labelled and unlabelled compound, for example, Heavy water D^O penetrates into red blood cells more slowly than does ordinary water. However, there is no evidence that these 'isotope effects' are normally of great magnitude in the complex biochemical systems of animal and plant organisms. UNIVERSITY OF IBADAN LIBRARY 5 8 Isotopic methods have been extensively used in the study of utilization of ruminal ammonia and blood urea by rumen micro-organisms. Mathison and Milligan (1971) used the isotopic tracer technique to determine the proportion of microbial protein derived from ruminal ammonia. 15NH^CI solution (2L/2^ hour) was continuously infused for periods of 120 - 216 hours into the rumen of sheep which were allowed to feed 2 out of every 10 minutes. These treatments achieved 'steady metabolic states' in the rumen in the period of the investigation. They found that 50 - 65% of bacterial N and 31 - 55% of protozoal N were derived from ruminal ammoniaj 60 - 92% of the daily N intake was transformed into ammonia, and 17 - 5^% of the ammonia formed was absorbed. The genera­ tion time of bacterial protein was found to be between 38 and hours. The investigators showed that increase in ruminal ammonia leads to a decrease in the conversion of protein into ammonia in the rumen,and was given by this relationship: Y = 123 - O.kk x where Y = Nitrogen converted into ammonia expressed as percentage of N intake. X = Concentration of ruminal ammonia (mg NH^ - N/litre). These results were similar to those of Pilgrim, Gray and Weller (1970). Nolan and Leng (1972) used isotopic dilution UNIVERSITY OF IBADAN LIBRARY 59 techniques with Ammonium sulphate, / • 15n_7 urea, and L 1 Ac_7 urea, and obtained similar results. They went further and showed that 59% of the dietary N was digested in the reticulo-rumen; 29$ of the digested N was utilized as amino acids and 71$ was degraded to ammonia. They also showed that urea was synthesized at the rate of l8.Ag N/day from 2.0g N/day of ammonia absorbed through the rumen epithelium and l6.Ag N/day apparently arising from deamination of amino acids and ammonia absorbed from tie lower digestive tract. They obtained similar results using continuous infusion and single injection techniques and therefore showed that both techniques were valid in isotopic dilution studies. The investigators also used this isotopic technique to determine body urea space. Landis (1968) using 35"S sulphate given intra-ruminally to lactating dairy goat showed that J>2 - A1$ of administered sulphur was utilized for synthesis of protein sulphur in the rumen when the ration contained ample amounts of protein but the value was 70$ when low protein ration was fed. Analysis of tissues of experimental animals showed that the rumen mucosa tissue protein had the greatest specific activity followed by the proteins of red bone marrow, liver, pancreas and kidney. This shows the sites of greatest utilization of UNIVERSITY OF IBADAN LIBRARY 6 0 administered sulphur. Milk was found to be strongly labelled, and the specific activity of individual amino acids of milk was similar to that of the amino acids of tissue proteins. This has brought the suggestion that the amino acid needed for the synthesis of body and milk proteins are drawn from a common amino acid pool. Piva and Silva (1968) used N - Diammonium phosphate to study the utilization of the com­ pound to produce milk and meat proteins. The nitrogen of the essential amino acids of ruminal bacteria and protozoa and milk of the sheep except tryptophan were significantly labelled. Only serine and oystine were found to be signi­ ficantly labelled of the amino acids of the tissues of the sheep. They also found that about 6% of administered N Diammoniura phosphate was metabolized in milk proteins. Land and Virtanen (1959) reported that feeding Ammonium nitrate to lactating coins resulted in the labelling of milk -within on& hour, and that of the milk amino acids, histidine and cystine were very weakly labelled. They interpreted the low content of 15N in histidine to be due to the incapability of the ruminal bacteria to synthesize the imidazole ring. They fouind that about 17% of administered 1bN was used for milk protein synthesis. Black, Egan, Anand and Chapman(1968) need c - 14 amino acids to show that amino acids play some role UNIVERSITY OF IBADAN LIBRARY 61 in gluconeogen^pis in lactating ruminants, that the process t is metabolically important for all animals, and becomes essential for survival when the body's glucose requirements exceed the alimentary supply. This may be so in ruminants where the glucose supply is always tenuous because the rumen micro-organisms rapidly ferment dietary carbohydrate conver­ ting it into short - chain fatty acids, leaving the animal very little glucose for absorption. Coccimano and Leng (196?) and Mugerwa and Conrad (1971) using intravenous infusion of C - 1't urea have calculated the urea pool size, rate of entry of urea into body urea pool, rate of degradation of urea in the rumen&adamount entering the body pool, that is q®graded* Theiri- results have shown the complexity of urea kinetics in the ru­ minant. The findings that have been described, using isotopic dilution techniques have shown how useful the method is and how promising it still is in biochemical and nutritional research. It has made possible the quantitative determination of the utilization or rate of transfer of metabolites in and between body compartments, the estimation of which could not otherwise have been possible without the use of isotopes. UNIVERSITY OF IBADAN LIBRARY 62 GENERAL OBJECTIVES The main objectives of those studios aro: 1. To determine the extent to which supplementation of a basal ration of hay (Cynodon nlenfuonsis/ Controsona pubescens) with protein concentrates affects production of ruminal metabolites of nitrogenous origin and blood urea levels, and to assess the efficiency of utilization of hay and concentrate supplements. 2. To determine the utilization of dietary nitrogen for the synthesis of microbial prote/in, blood urea and milk protein using /"" 15n J ammonium chloride, and the extent of recycling of blood uroa into the digestive tract of the sheep using f~ ̂̂ N_7 urea. 3. To estimate the digestibility, nitrogen retention, metabolic faecal nitrogen (MEN) f the endogenous urinary nitrogen (EM), the biological value of the,rations and the digestible crude protein requirement for maintenance. 4. To determine the digestibility of the rations in the stomach and intestines of the sheep using chromic oxide - impregnated paper as the marker and to show the reliability of the method for partitioning digestibility in the stomach and intestines of the sheep. UNIVERSITY OF IBADAN LIBRARY 6J CHAPTER TWO 2. RUMINAL AND BLOOD METABOLITES OF THE WEST AFRICAN DWARF WETHER SHEEP MAINTAINED ON BASAL HAY AND CONCENTRATE SUPPLEMENTS. 2.1 INTRODUCTION McDonald (1948) had shown tha-t dietary protein and non­ protein are degraded by the rumen mierobial population and that ammonia is the major end product of the degradation. The nitrogenous substances in the rumen are the feed and microbial protein, ammonia, amines, amides and amino acids. Sovoi’al investigators (Chalmers and Synge, 1954; Annison, 1956; Lewis, 1957; and Elliott and Topps, 1964) have shown that the levels of these metabolites in the rumen are dependent on the type of ration fed to the animal. In the present report, ruminal metabolites and blood urea were examined in four fistulated West African dwarf wether sheep maintained on basal hay and concentrate supplements to find the effect of varying levels of dietary protein on the ruminal and blood metabolites . 2.2 MATERIALS AND METHODS. 2.2.1 Animals and their management. Bight West African dwarf wether sheep, 1 - 1,5 years old and with live weights ranging from 13.2 to 26.3 kg were used. Four .cf the animals were fitted with permanent rumen cannulae. Each sheep UNIVERSITY OF IBADAN LIBRARY 64 was kept in a metabolism cage as described by Oyenuga (l96l). The animals were usually fed at 8.00 a.m. every day. The residues were collected at 8.00 a.m. every day, weighed, and stored for chemical analysis (in order to estimate nutrient intake). The animals had free access to salt licks and fresh clean water a,d lib. 2-2.2 Diets: There were six diets. The basal diet consisted of cynodon nlemfuensis/centrosema pubescens hay. The grass/legume mixture was cut on the field and left to dry for tiro days, after which the hay was packed and stored in the barn. There were five concentrates (c^ - C^) composed of cassava flour, groundnut meal, molasses and mineral mixture. The concentrates varied in the crude protein content of dry matter from 1.6 to 17.5^» The chemical composition of the concentrates and hay are shown in Table 2.1. RationA consisted of 1.0kg cf basal hay. Ration B consisted of 0.50kg of the basal hay and 0.50kg of concentrate C-,. Similarly, rations G, D, E and F consisted of 0.50kg of basal hay and 0.50kg respectively of concentrates C2> C^» and C^. 2.2.3 Plan of experiment. The experiment was divided into two trials. The first trial consisted of two periods and the second trial consisted of four periods in a 4 x 4 Latin square design. (Table 2.2), In the first trial, the eight animals were divided into two groups of four animal. Each group was made up of tiro fistulated and UNIVERSITY OF IBADAN LIBRARY <>5 TABLE 2.1 Components and. chemical composition of rations fed to the West African dwarf sheep. CONCENTRATES COMPONENTS C1 C2 s C4 C5 Grr»untlhut meal - 4.0 10.0 18.0 28.0 Cassava flour 92.5 88.5 82.5 74.5 64.5 Molasses 5.0 5.0 5.0 5*0 5.0 ♦Mineral mixture 2.5 2.5 2.5 ,*2*5 2.5 Total 100.0 100.0 100.0 100.0 100.0 ♦ 2.5kg of C2,A supplies the following: Vit. A (l.U) = 1.25 f f e ( s ) = 15 MVint. D ft _ 0.63 Cu II = 10(g) 40 Co ft = 0.75 Zn 30 MI g nit = 3.0= 5000 UNIVERSITY OF IBADAN LIBRARY 66 TABLE 2.2 Plan of experiment TRIAL 1 GROUP 1 GROUP 2 259, 179 173, 263 PERIOD 186, 268 184, 301 1 A B 2 B A TRIAL 2 : : GROUP 1 : GROUP 2 : GROUP 3 : GROUP 4 ; « * * 1 * * I PERIOD -259, 179'186, 268’ 173, 263* 184, 301 * « * - * * 4 D D F 3 E P G D 4 P C D E . . .u UNIVERSITY OF IBADAN LIBRARY 67 two intact animals. One group was put on a ration of hay (ration a ) and the other group was put on a ration of hay supplemented with cassava flour, molasses, and minerals (ration B), During the second trial, the eight animals were divided on a live weight basis, into four groups of two animals* Each group consisted of a fistulated and an intact animal, and each pair was allocated at random to each of rations C, D, E and F (fable 2.2). Each trial consisted of a 14-day preliminary period followed by a 6-day collection period. The animals were weighed at the beginning and at the end of each trial period. 2.2.4 Collection of faeces ..end urine. A day prior to collection, each animal was fitted with harnesses to which was attached a collection bag for the separation of urine and faeces. The removable trays in the metabolism cage permitted the urine to drain freely into the aluminium tray below it. This was sloped so as to allow easy drainage of urine to its tube at the centre. The tube led to a funnel placed at the mouth of a small plastic bucket below the cage in which urine was collected. The bucket contained 5 ml of 10/6 Mercuric chloride to prevent microbial breakdown of the nitrogenous components of the urine. Urine volvumes were measured immediately after collectxon and 10$ of the daily output was retained. The daily samples of urine for each animal were bulked and stored in a deep freezer at -5°C until required for analysis. UNIVERSITY OF IBADAN LIBRARY 68 A polythene tag was placed, in the collection tag to allow for easy collection of faeces. The bag was emptied daily before the morning feeding. Faeces were dried to constant weight in the forced- draught oven at 70°C for 48 hours. The daily dried faeces were bulked for each animal, milled with Christy Norris Hamner mill and stored in air-tight glass bottles until required for analysis. However, fresh samples of faeces stored in a deep freezer at -5 6 were used for the analysis of N of non-dietary origin. 2.2.5 Sampling of blood and rumen liquor Samples of rumen liquor and blood were collected during the last three days of the collection period. The animals were allowed to feed from 8.00 to 9.00 a.m., and then the feed was removed. Rumen samples were then taken at 10.00 a.m., 11.00 a.m. and 12.00 noon. Rumen samples were collected with the method of Alexander (1969) as modified by Mba and Olatunji (l97;j.). The sampling lasted 5 mins during which about 300 ml were obtained. The samples were stored at -5°C until required for analysis. Blood samples were collected from each fistulated animal at 1.00 p.m., using sterilised needles. The blood samples were obtained from the jugular vein and 5 ml of each blood sample was kept in a sample bottle containing some heparin. The blood was centrifuged at 2000 r.p.m. to obtain the plasma which was stored in the deep freezer at -5°C until required for analysis. UNIVERSITY OF IBADAN LIBRARY 69 2.2.6 Isolation of rumen bacteria and protozoa Ruminal digesta was strained through six layers of cotton cloth to remove plant debris. Bacteria and protozoa were then separated from the liquid fraction by differential centrifugation method (Blackburn and Hobson, I960). The microbial samples were freeze-dried at -20°C for five days. 2*2,7 analytical procedure. The N contents of feeds, faeces, and urine as well as crude fibre, other extracts and ash content of the feeds were determined according to A.0AC (1970) method, except that Markham’s (1942) semi-micro-Kjeldahl apparatus was used for the N determination. Nitrogen of non-dietary origin in faecal samples was determined according to the method of Van Boest and Wine (1967) as modified by Mason (1969). The rumen protein -N after precipitating with 10fo TCa and the total ruminal N were determined using Markham's (1942) semi-micro-kjeldahl method. Ammonia -N and blood urea -N were determined by the method of Fawcett and Scott (i960) as modified by Chaney and Marbach (1962). Amino acid composition of freeze-dried microbial samples was determined by the column chromatographic technique using the automatic Hitachi-Perkin-filmer amino acid analyser (model KLA. - 3B, Hitachi Ltd., Tokyo, Japan), after hydrolysis of 100 mg of each sample with 10 ml of UNIVERSITY OF IBADAN LIBRARY r 70 6N HC1 at 110°C for 24 hours. However, small quantities of 2 - t Hercaptoethanol (.0.5 ml per litre of 6N HCl) were added to the samples to improve the recovery of the amino acids, particularly methionine. The concentration of free -A- amino IT in the rumen liquor was determined by the method of Michel (i960). UNIVERSITY OF IBADAN LIBRARY r 71 2.3 R E S U L T S The concentrations of the various nitrogenous metabolites in the rumen and. blood are given in Table 2.3• Each value with its standard error is a mean for four fistulated animals. 2.3.1 Total ruminal nitrogen (mg/lOO ml) Total ruminal nitrogen, expressed in mg per 100 ml of rumen liquor (mg/lOO ml) for ration A was not significantly different from ration B (P> 0.05), the mean values being 40.8 ± 3*8 and 29.9 ± 2.3 for rations A and B respectively. The mean differences were highly significant (pk.,0.0l) for the rations in Trial 2. Ration P gave significantly higher levels of total ruminal nitrogen, (124.9 + 18.5 mg/lOO ml) than rations C, D and E the means of which xrere 44.8 + 6.4, 68.4 ± 5.4 and 78.2 + 2.2 respectively. The mean values of total ruminal nitrogen increased with increasing intake of dietary nitrogen, and also with increasing crude protein content of the ration. Total ruminal nitrogen was correlated with digestible crude protein intake (r = 0.56, P-C0.05). The highest value of total ruminal nitrogen occurred one hour after feeding and then declined; in some cases, no decline was observed after two hours. UNIVERSITY OF IBADAN LIBRARY 72 Table 2*3: Ruminal and blood notabolites+ the 7est Airican dwarf wether sheep maintained on basal hay and concentrate supplements Trial 1 Trial 2 Metabolites -1 T B C » D k E x « x . a ; a ‘ a 5 I DF TOTAL RUMINAL N (mg/lOOml) 40.8 ± 3.8 : 29.9 i 2.3 : 44.8 ± 6.4 ; 68.4 ± 5.4 • 78.2 + 2.2 - 124.9 ± 18.5 X ’ b RUMINAL PROTEIN - N (mg/lOOml) 29.4 ± 2.4 21.7 ± 1.5 : 31.5 ± 4.4 * 50.6 ± 2.8 ; 55^9 ± 1.8 ; 79.1 ± 9.8 RUMINAL NON-PROTEIN-N (mg/lOOml). b11.4 + 2.6 8.2 + 1.0 13.3 + 2.2 17.8 + 3.2 22.3 + 0.7 45.8 + 10.0 X y a a 5 "5 * RUMINAL AMMONIA-N(mg/ 100ml) 4.7 + 0.3 1.2 + 0.3 1.8 + 0.4 2.4 + 0.5 3.8 + 0.5 4.5 + 0.5 " i "rr d ’ *' ' "3 a b RUMIN.lL RESIDUAL-N (mg/lOOml) 6.8 + 2.7 7.1 i 0.9 11.6 + 1.9 15.2 ± 2.5 18.7 + 0.8 40.2 ± 9.1 5s" RUMINAL - AMINO-N (p ,mole/al) j 3>27+ 0>21 1#| + q .16 ; 2.31+ 0.44 ! 3-02+ 0.31 : 4.79+ 0.43 6.29+ 0.78 PERCENT PRO TEIN-N/ TOTAL N ; ?2>f ± 4#4 j 7f.x ± 1-5 ; 70.| ± 1.6 ; 72.5 + 2.6 ; 71.4 + 1.8 • 64.6 + 3.7 PERCENT RESIDUAL-N/NON PROTEIN-N) x a a a52.0 + 8.3 88.5 + 5.6 87.1 + 2.0 79.2 + 4.1 83.9 + 3.4 86.8 + 3*0 ammonia- n/ total- n X 11.7 ± 1-5 ?.13t 1.50 3.77± 0.92 3.91± 1.80 4.63± 1.93 4.39 ± 1.59 X y a a ab TT BLOOD UREA-N (mg/lOOml) 5.3 + 0.5 1.4 + 0.2 1.9 + 0.d 2.6 + 0.5 3-9 + 1.0 4.9 + 0.9 + EACH VALUE IS A MEAN FOR FOUR ANIMALS MEANS BEARING THE SAME SUPERSCRIPT IN THE ROW ARE ROT SI ©OTICANT (P > 0.05) UNIVERSITY OF IBADAN LIBRARY 73 2.3*2 Total Ruminal Protein Nitrogen (mg/lOO ml) The total ruminal protein nitrogen values of 29*4 + 2.4 and 21.7 + 1.5 mg/lOO ml respectively for rations A and S are shown in Table 2.3* Though the mean value was higher for ration A than for B, the difference was not significant (P>0,05). The mean differences in Trial 2 were highly significant (p<0.0l) with the mean values of 31.5 + 4.4, 50.6 + 2.8, 55.9 + 1.8 and 79*1 + 9.8 mg/lOO ml for rations C, D, B and F respectively. In sheep fed on concentrate-based rations, ruminal protein nitrogen increased with increasing crude protein intake and level of dietary crude protein. The highest concentrations of ruminal protein nitrogen occurred one or two hours after feeding. Individual observations were not significantly variable either within animals (p> $.05) or between periods (p > 0.05). The protein nitrogen expressed as percentage of total ruminal nitrogen were 72.7 + 4$ for ration A and 73*1 + 1.5^ for B. The mean differences were not significant (p > 0.05)« The mean value of 64.6 + 3.7/6 for ration F was lower than the mean values of 70.3 + 1.6, 72.5 + 2.6 and 71.4 + 1.8 for C, D and B respectively, although the the mean differences were not significant (p > 0.05).. Ruminal protein nitrogen as percentage of total ruminal nitrogen did not seem to be influenced by intake of nitrogen or by the levels of UNIVERSITY OF IBADAN LIBRARY 74 dietary crude protein but was remarkably constant for the rations especially from rations A to E. Differences between mean values were not significant either within the experimental animals (P.> 0.05) or between periods (P> 0.05). The lower value for ration F may be due to the fact that for this ration, the products of microbial breakdown of protein was not being as rapidly incorporated into microbial protein in ration F as in the other concentrate rations* 2.3.3 Non-Protein Nitrogen (mg/ioo ml) Supplementation of hay with concentrate depressed the level of non-protein nitrogen (NPN) from 11.4 + 2*6 to 8.2 + 1.0 but this depression was not significant (P>0.05). The ITPN concentration was higher for ration F (45.8 + 10.0) than for rations C, D and S which were 13*3 ± 2.2, 17.8 ± 3.2 and 22.3 ± 0.7 mg/lOO ml respectively, and the mean differences were significant (P <. 0.05). The NPN increased with increasing dietary nitrogen intake and dietary crude protein percentage of the rations. It is only at the highest nitrogen intake, and when the percentage crude protein in the ration was highest (ration F) that the levels of non-protein nitrogen was high enough to assume significance, due probably to a more rapid breakdown of dietary protein in the rumen than corresponding incorporation of non-protein nitrogenous materials to microbial protein. UNIVERSITY OF IBADAN LIBRARY J 75 2.3*4 Ruminal Ammonia Nitrogen (mg/lOO ml) Supplementation of hay with concentrate C-, (ration B) mainly cassava flour, very significantly depressed ruminal ammonia nitrogen from 4.7 ± 0.3 mg/lOO ml down to 1.2 + 0.3 ng/lOO ml (p O.Ol). The mean differences for ruminal ammonia nitrogen were also very highly significant for the rations in Trial 2 (P L. O.Ol) and these were 1.8 + 0.4, 2.4 + 0.5, 3*8 + 0.5 and 4.5 + 0.5 mg/lOO ml for rations C, D, E and F respectively. Ruminal ammonia nitrogen levels increased with increasing intake of dietary crude protein and also with increasing percentage of crude protein in the rations. Ruminal ammonia nitrogen was correlated with the level of non-protein nitrogen (r = 0.91» P ^ 0.05) when concentrate- based rations were fed, and the relationship can also be shown by the following regression equation Y = (0.086 + 0.015) X + 0.88 (r = 0.91) ..................... (2.1) where, Y is the concentration of ruminal ammonia nitrogen (mg/lOO ml)and X is the concentration of ruminal non-protein nitrogen (mg/lOO ml). Even though the mean differences were significant for treatments, low levels of ruminal ammonia were observed even with the ration highest in crude protein content and this shows that ammonia formed UNIVERSITY OF IBADAN LIBRARY 76 TABLE 2.4 The regression equations showing the relationship between ruminal and blood metabolites and nitrogen utilization in the West African dwarf sheep maintained on Lay and concentrate supplements. Y X REGRESSION EQUATION r 2.1 RAN NPN Y = (0.086 + 0.015) z + 0.88 0.91* 2.2 X.-AA NPN Y - (0.125 + 0,020) x + 0.87 0.95** 2.3 X - A A RAN Y = (1.401 ± 0.040) x -0.25 0.99** 2.4 BUN NR Y = (4.87 ± 0.14 ) x - 0.38 0.99*** 2.5 BUN ND Y = (4.44 ± 0.12 ) s - 0.32 0.99**5 *i 2.6, *1• BUN RAN * Y = (1.10 ± 0.02 ) x - 0.05 1 0.99** : RAN = Ruminal Ammonia nitrogen:, mg/lOOml NPN = Non-protein nitrogen, ng/lOOml X — AA = o<- Ajnino nitrogen p raole/ml BUN = Blood Urea nitrogen, mg/lOOml NR Nitrogen retained, g/day/w^g734 ND Nitrogen digested, ii/day/w^734 * _ Significant at 5% level (P< 0.05) *-* _ Significant at 1% level (p C o .o i) U IVERSITY OF IBADAN LIBRARY as a result of protein breakdown was efficiently fixed into microbial protein. The high correlation (r = 0.9l) between ruminal ammonia nitrogen and non-protein nitrogen shows that ammonia formation was very much dependent upon the nitrogen oonsumed that was converted into non-protein nitrogenous materials* There were no significant (P>0.05) variations within animals in their ruminal ammonia nitrogen concentration. The percentage total nitrogen that was ammonia -N was markedly depressed from 11.7 + 1.5/® for ration A down to 3*1 + 1*5$ for ration B/ (p<, O.Ol) by supplementation of hay with concentrate C^* In Trial 2, the mean values of the total nitrogen that was ammonia were not different (P> 0.05), being 3.77 + 0.92, 3*91 + 1.80, 4.63 + 1.993 and 4.39 + 1.59$ for rations C, D, E and F respectively. Mean values, however, greatly varied between periods and liithin experimental animals (P<0.0l). The mean value was lower in period one than in the other three periods of Trial 2. Mean values were not affected by nitrogen intake or percentage crude protein in the rations. The only ration where ammonia nitrogen formed an appreciable percentage of total ruminal nitrogen was ration ... Mean values for concentrate- based rations were low and seemed to be relatively constant during the first three hours after feeding. UNIVERSITY OF IBADAN LIBRARY 2.3.5 Ruminal Residual Nitrogen (mg/iQQ The ruminal residual nitrogen or non- ammonia non-protein nitrogen was not affected (p>0.05) by rations in Trial 1, the values being 6.8 + 2.7 and 7.1 + 0.9 for rations A and B respectively. In Trial 2, the mean differences were significant (P< 0.05). Ration F gave rise to higher concentration of residual nitrogen, 40.2 + 9.1 mg/lOO ml,, than rations C, D and E concentrations of which were 11.6 + 1.9, 15.2 + 2.5 and 18.7 + 0.8 mg/lOO mX respectively. The residual nitrogen concentration increased with increasing levels of dietary crude protein, with total nitrogen intake and also with the concentration of non-protein nitrogen. The concentration of residual nitrogen did not show much variation within animals, between periods or the first three hours after feeding even though the levels were consistently higher one hour after feeding. The residual nitrogen comprises amines, amino acids and some peptides (McDonald, 1948). Since the residual nitrogen is part of the non-protein nitrogenous materials, it is of interest to note that supplementation of hay with cassava flour increased the percentage of non-protein nitrogen less ammonia from 52.0 + 8.3^ to 88,5 + 5«6^* The mean differences were not significantly (P>0.05). For the protein-based rations in Trial 2, there were no significant variation in the percentage of residual nitrogen in the non-pr >tein fraction (P> 0.05) and the UNIVERSITY OF IBADAN LIBRARY 79 mean percentages were 87.1 + 2.0, 79.2 + 4.1, 83.9 + 3.4 and 86.8 + 3*8/ for rations C, D, E and F respectively. The percentage of non-protein nitrogen that was residual -N was not influenced hy nitrogen intake or level of dietary crude protein. The results showed that the residual -N formed the major fraction of the non-protein nitrogen of sheep fed hay/concentrate rations especially if the concentrates were rich in readily formentable carbohydrates. There were no variations in the mean values for experimental animals or periods (py 0.05). 2.3.6 Ruminal o4.~ amino nitrogen (/*■ mole/ml) The concentrations of ruminal c{- amino nitrogen (/mole/ml) was 3.27 + 0.21 and 1.61 + O.l6pmole/ml on rations A and B respectively and the mean differences were highly significant (P4.0,0l). The mean values were 2.31 + 0.44, 3.02 + 0.31, 4.79 + 0.45 and 6.29 + 0.78 for rations G, D, E and F respectively, and the mean differences were significant for the treatments (Pdl0.05). The levels of ai, - amino 17 increased with percentage crude protein in the ration, dietary nitrogen intake and levels of ruminal non-protein nitrogen. The concentration of ok - amino nitrogen was correlated to that of non­ protein nitrogen in the rumen of sheep fed on concentrate - based rations, relationship can be represented by the following regression equation: UNIVERSITY OF IBADAN LIBRARY 80 y = (0.125 + 0.020) X + 0 . 8 7 ........ ....... (2.2) where, Y is the concentration of bX,— amino nitrogen (U mole/ml) and. X is the concentration of non-protein nitrogen in the rumen (mg/lOO ml). Prom this equation, it is seen that an increase in the concentration of non-protein nitrogen will lead to a corresponding increase in the concentration of'X- amino acids, showing that$0- amino acids are derived from ruminal non-protein nitrogen. Since a similar relationship was obtained with ammonia and non-protein nitrogen, that increased concentration of non-protein nitrogen caused increase in ammonia nitrogen, it is expected that increase in ruminal ammonia will lead to increase i n ^ - amino nitrogen. This is in fact so, as shown by the regression equation showing the relationship between - amino nitrogen (y ) in / fmole/ml and ruminal ammonia nitrogen (x), in mg/lOO ml of rumen liquor of sheep maintained on concentrate based rations. Y = (1.401 + 0.040) X - 0.25. (r = 0.99, P C 0.01) .......................... (2.3) It was also observed that supplementation of hay with concentrate (C-j_) caused a very significant depression of amino nitrogen. Even on the ration richest in digestible crude protein, the c<- amino nitrogen concentration was still low (6.29 + 0.78 UNIVERSITY OF IBADAN LIBRARY J 81 H aole/ml). From this result, it is seen that the levels of o(- amino nitrogen were low on all the rations. There were no variations within experimental animals or between periods (P^ 0.05)• 2*3.7 Blood urea nitrogen (mg/100 ml) The animals on ration B had significantly lower levels of blood urea nitrogen (P^O.Ol) than those maintained on ration 2x (Table 2.5)• The mean values of blood urea nitrogen (mg/lOO ml) were 5*3 + 0.5, and 1.4 + 0.2 mg/lOO ml for rations A and B respectively. Thus, supplementation of hay with cassava flour - based concentrate caused a reduction in the concentration of blood urea nitrogen. In trial 2, the mean differences for treatments were significant (P< 0.05) and the mean concentrations were 1.9 + 0.3, 2.6 ± 0.5, 3*9 + 1.0 and 4*9 + 0.9 for C, D, a and F respectively. Blood urea nitrogen increased with increasing dietary nitrogen retained when concentrate-based rations were fed to sheep and the following equation shows the relationship: Y = (4.87 + 0,14) X - 0.38 .................. (2.4) (r = 0.99 P < O.Ol) where, Y is the mean value of blood urea nitrogen (mg/lOO ml) and X is the amount of nitrogen retained (g/day/wkg^* ) Blood urea nitrogen (y ) was also positively correlated with nitrogen digested, g/day/wkg °*7-54 with r Value of 0 .99 and the UNIVERSITY OF IBADAN LIBRARY R a t i o n s FIG. 2.1 LEVEL OF RUMINAL AMMDNIA N (°— o)AND BLOOD UREA N (a- - * ) IN THE SHEEP MAINTAINED ON HAY AND CONCENTRATE SUPPLEMENTS mg /1 00 ml UNIVERSITY OF IBADAN LIBRARY 83 relationship can be represented by the regression equation: I = (4.44 + 0.12) X - 0.32 ....... ............ (2.5) when concentrate-based rations were fed to the sheep. The regression equation was similar to the one obtained when blood urea was regressed on nitrogen retained and this shows that blood urea nitrogen was influenced by nitrogen retained as well as nitrogen digested. It was observed that with the experimental rations in trial 2, the highest concentration of blood urea nitrogen was still as low as 4.9 + 0.9mg/l00 nl. Observations of ruminal ammonia nitrogen and blood urea nitrogen showed that concentration of blood urea nitrogen (l) followed very closely the ruminal ammonia nitrogen concentration (x ) and the relationship can be represented by the following regression equation: I = (1.10 ± 0.02) X - 0.05 ................... (2.6) (r = 0.99, P < £ 0.0l). Ihu9..1 ow l-Qvels^af. xminataawoniavare ossGCiaie4;.udith lew-levels of blood *»ea«. -There wer.e:*ao_yariatiaa. •ftithin.* experimental, aaiiaala .(HXQ.Q5) but variations.between periods.were-significantly lower'in period 1 than in the other periods (P<0.05) of Trial 2 . 2.3*8 ilmino acid composition of ruminal bacteria j^xd protozoa: Table 2.5 shows the amino acid composition of ruminal bacteria and protozoa of sheep fed ration P. The results are expressed in g per I6g N and also in g/lOOg amino acids (g/lOOg Ui). From Table 2.5, UNIVERSITY OF IBADAN LIBRARY 84 table 2.5 Amino acid, composition of ruminal bacteria â'id protozoa for the West African dwarf aether sheep maintained on basal hay and concentrate supplements compared .with reported values B A C T E R I A P R 0 T 0 Z 0 AMINO ACIDS g/l6gN g/lOOg Amino acid g/lOOg iwino acid B B B* P P p * Lysine 8.51 12.60 8.83 8.53 10,40 12.98 Histidine 0.54 0.79 1.95 0.86 1.05 1.60 Arginine 3.14 4.65 5.30 3.73 4.55 3.70 ^spartic acid 8.66 12.81 12.00 8.01 9.77 13.53 Threonine 3.38 5.00 6.00 1 2,83 3-40 4.75 Serine 2.76 4.08 4.65 3.89 4.77 5.10 Glutamic acid 10.20 15.10 13.95 16.56 20.19 16.28 Proline 2.89 4.27 2.80 5.11 6.23 5.95 Glycine 3.54 5.23 5.88 2.80 3.42 4.10 Alanine 5.62 8.52 7.50 4.96 6.05 4.43 Valine 4.49 6.65 ; 5.20 \ 3.86 4.71 | 4.03 Methionine I 1 .5 2 ; 2.24 : 1.9 5 : 0.7 1: 0.87 : 1.63• I• soleucine “ 3.42 * 5.06 : 5.23 : 4.02; 4.90 ; 6.10 Leucine 4.64 6.87 7.73 9.96 12.14 j 7.45 Tyrosine 1.75 2.60 4.33 1.96 2.40 4.23 Phenylalanine 1.90 2.82 4.70 2.97 3.62 5.70 B*, P* - Rumen bacterial and Protozoal amino acid composition from Bergen, Purser and Cline (1968), Ration 4. UNIVERSITY OF IBADAN LIBRARY 85 tiie amiao«acid present in the least amount in ruminal bacteria is histidine (0.54g/l6gN) and followed by methionine (l.52g/l6gN). Glutamic acid, aspartic acid and lysine are present la- the greatest concentration of 10.20, 8.66 and 8.51g/l6gN respectively. Of the essential amino acids determined, lysine, leucine, valine and iseleucine were present in greatest concentrations of 8.51, 4.64, 4.49, 3»42g/l6gN respectively, while histidine, methionine and phenylalalanine were present in lowest concentrations of 0.54; 1.52, and 1.90g/l6gN respectively. The concentration of glutamic acid was very high in ruminal protozoal amino acid (l6.56g/l6gh). It accounts for about 20$ of the protozoal amino acids. Leucine, lysine and aspartic acid followed with concentrations of 9.96, 8.53, 8.01 g/l6gN respectively. Of the essential acino acids, leucine and lysine were present in high concentrations of 9.96 and 8.53 g/l6g IT respectively, and methionine, and histidine were present in low concentrations of 0.71 and 0.86 g/l6g N respectively. 2.4 DISCUSSION; The concentration of total ruminal nitrogen in the sheep maintained on hay (7.7$ crude protein) was 40.8 + 3*8 mg/lOO ml. This value is in very good agreement with the value of 40.6 mg/lOO ml obtained by .Elliott and Topps (1964) who maintained Persian wethers on UNIVERSITY OF IBADAN LIBRARY 86 a mixture of hay (8$ crude protein). Supplementation of hay with cassava flour-based concentrate significantly reduced (B-C0.05) the to.tal ruminal nitrogen to 29.9 + 2.5 mg/lOO ml. This value is jet however slightly lower than 34.4 mg/lOO ml obtained by Elliot and Topps (1964) for a cassava flour-based ration. They showed that the total ruminal nitrogen was correlated with the crude protein content of the diet. Their values of 49»9> 92.0 and 99.3 mg/lOO ml are higher than the present reported, values of 44.8, 68.4 and 78.2 mg/lOO ml in sheep maintained on rations G, D and E, similar to their low roughage rations. Both this present report and that of Elliott and Topps (1964) showed that maximum level of total ruminal nitrogen occurred about 1 hour after feeding. The unusually high variability associated with the N content of samples of the rumen liquor from sheep given low-roughage diets observed by Elliott and Topps (1964) was not observed in the present work as the mean differences were not significant within animals (P> 0.05) or between periods (p> 0.05). No sharp increases wore observed in the total nitrogen concentration even in the sheep receiving the suplement of the highest crude protein content between 1 and 2 hours after feeding, but there were sharp decreases three hours after feeding. The concentration of ruminal protein nitrogen, 29.4 + 2.4 and 21.7 i. 1.5 for rations A and B respectively were higher than 19.8 and UNIVERSITY OF IBADAN LIBRARY 87 l~.l mg/lOO ml obtained, for # cassava flour-based rations by Elliott and Topps (1964) but the values of 31.5 + 4.4, 50.6 + 2.8 and 55.9 + 1.8 for rations C, D and E respectively are comparable to 29.0, 59.1 and 59.3 obtained by these investigators. There seemed to be no difference in the percentage of N as protein in the rumen liquor which varied from 64.6$ with ration F to 73,1$ with ration B. These values are higher than 45 - 65$ obtained by Elliott and Topps (1964). They found that percentage of N as protein was negatively correlated with levels of ammonia. No such correlation was observed in the present investigation. Protein-N in the rumen may be derived from feed, bacteria or prcrfcoaoa. Weller, Gray and Pilgrim (1958) using Di-aminopimelie acid as marker for bacterial protein showed that bacterial protein formed 46$, protozoal 21$ and feed 26$ of ruminal nitrogen. Freitag et _q̂L. (1970) using the same indicator showed that the amount of bacterial protein in the rumen fluid was affected by the dietary nitrogen source. Th^jShowed that bacterial protein formed 99$ of the rumen fluid protein 7 hours after feeding urea-supplemented ration, the corresponding value for cotton seed meal - supplemented diet was 71$. The rations used in the present investigation contained groundnut meal as source of protein apart from that Bupplied by the basal hay. The presence of readily fermentable carbohydrates such as cassava flour and molasses enhanced rapid microbial growth. It is therefore reasonable to assume UNIVERSITY OF IBADAN LIBRARY 88 that a greater percentage of ruminal protein-N would be microbial proteins. The non-protein fraction (NPN) of rumen liquor is made up of amines, amino acids, ammonia and peptides (McDonald, 1948). The concentration of Mon-protein nitrogen was highest 1 or 2 hours after feeding and declined 3 hours after. Annison (1956) showed that casein and groundnut meal were rapdily degraded in the rumen with the formation of non-protein nitrogenous substances. The failure to observe a rapid increase in non-protein nitrogen was due to the presence of readily fermentable cassava flour in the rations, which is in line with the well established observation that the utilization of Non-protein nitrogen is improved when fermentable carbohydrates are also present. Only with ration F containing the highest level of crude protein was an appreciably lsLffa. level of non-protein nitrogen obtained. It may be that in this ration, the rate of. proteolysis of dietary protein was greater than the rate of assimilation of the non-protein substances formed. The supplementation of hay with cassava flour caused a very significant (P 4L O.Ol) depression in ruminal ammonia -N. This is in agreement with the findings of Chalmers and Synge (1954)*- that addition of starch greatly depressed ruminal ammonia production. When rations B to F were fed to the sheep, ruminal amonia concentration increased with increasing dietary nitrogen intake and percentage of crude protein UNIVERSITY OF IBADAN LIBRARY 89 in the rations. This is in agreement with the results of Elliott and Topps (1964). Ruminal ammonia concentration was highly correlated (r = 0.9l) with concentration of non-protein in rumen liquor, which shows that ammonia was obtained by hydrolysis of non-protein substances. Elliott and Topps's (1964) value of 5*9 mg/lOO ml. is higher than 4.7 mg/lOO ml obtained for ration A but their value of 1.3 mg/lOO ml was in very good agreement with present report of 1.2 mg/lOO ml for ration A. However, their values of 2.1, 5.2 and 9.7 mg/lOO ml are higher than the values of 1.8, 2.4 and 3*8 mg/lOO ml obtained in the present investigation. In any case, their rations contained less fermentable carbohydrate than those used in the present experiment. The highest level of ruminal ammonia nitrogen occurred 1 or 2 hours after feeding. No sharp decline was observed in their levels and this is attributable to efficient fixation of ruminal ammonia by ruminal microbial population (Chalmers and Synge, 1954). The high correlation (r = 0.99) between rumminal ammonia nitrogen and ruminal X - amino nitrogen indicates that both metabolites are dependent and were probably formed from the non-protein nitrogen fx*action of the rumen liquor. The relatively high levels of ammonia -N in the rumen of sheep maintained on only hay may be due to the fact that nitrogen in the UNIVERSITY OF IBADAN LIBRARY 90 form of urea and. mucoprotein was added to the rumen by the saliva and the degradation of these produced high levels of ammonia. This would tend to remain high as microbial protein synthesis would be restricted by a deficiency of available carbohydrate in the ration. The percentage of total nitrogen in the rumen liquor present in the ammoniacal form was very high in animals given hay (ll.7 ± 1.5) « and this was depressed to 3«1 + 1.5$ when cassava flour was given as supplement. For the concentrate-based rations, there were no differences between the rations even though the tendency was for the percentage to increase with increasing crude protein intake or as total nitrogen in rumen liquor increased. These observations agree with the results of Elliott and Topps (1964). Low levels of ammonia-N (4.39 ± 1.59) as percentage total nitrogen even on ration F showed that very little ammonia-N accummulated in the rumen which could subsequently be lost from the rumen; it indicates efficient utilization of the protein contents of the rations. Ruminal residual nitrogen, also known as the Non—protein non-ammonia nitrogen, comprises mainly peptides and low levels of -amino acids, and amines. Residual nitrogen was low on all rations except on ration F. The mean values for rations A and B, 6.8 and 7.1 mg/lOO ml, were lower than those obtained by Elliott and Topps (1964), which were 14.9 and 12.8 ng/100 ml respectively. Similarly, UNIVERSITY OF IBADAN LIBRARY 91 the values obtained for rations C, D and B which are 11.6, 15*2 and 18.7 were lower than 18.8, 27.7 and 30.3 mg/lOO ml obtained by the same investigators. Only on ration F (40.2 + 9»l) were high levels of residual nitrogen observed. The residual nitrogen as a percentage of non-protein nitrogen was lower (52.0 + 8.3) with ration A than with other rations (79 - Q&/°) although there were no significant differences (P> 0.05). This is due to the relatively high levels of ammonia in the rumen of hay-fed animals. Elliott and Topps (1964) and Moore and King (1958) showed that an increase in ammonia concentration in the rumen was accompanied by a decrease in residual nitrogen. The supplementation of hay with cassava flour significantly depressed the levels ofp<-amino nitrogen in the rumen. This is in agreement with the results of Chalmers and Synge (1954)» Annison (1956) and Leibholz (1969). For concentrate-based rations, the levels of o(-amino nitrogen increased with dietary nitrogen intake, percentage crude protein in the ration, and levels of ruminal total N and non-protein nitrogen. This is in agreement with the reports of Annison (1956) who also showed that though proteins were almost certainly converted into amino acids before degradation to ammonia, the concen­ tration of free amino acids was usually low presumably because of their rapid uptake or degradation. There was a high correlation between ruminal ammonia and 0^-amino nitrogen (r = 0.99)# and also between UNIVERSITY OF IBADAN LIBRARY 92 Nonprotein nitrogen and 3(-amino nitrogen (r = 0*95)* These results agree with those of iinnison (1956) who showed that increases in ammonia concentration followed similar increases in the concentration of o(-amino nitrogen. He also showed that the increase in the concentration cf free o^-amino N in the rumen immediately after feeding were largely due to the presence of free <>(— amino N and labile amide N in the feeds. The marked depression of ruminal ammonia observed when hay was supplemented with cassava flour was also observed in the case of blood urea nitrogen, and this is in agreement with the results of Lewis (1957) who found that changes in ruminal ammonia concentration resulted in similar changes in the blood urea levels. Increase in the concentration of blood urea was observed with increasing intake of dietary protein, and also with the increasing percentage of crude protein in rations B to F. Preston, Schnakenberg and Pfander (1965) obtained high correlation (r = 0.986) between nitrogen intake per metabolic size and blood urea nitrogen. Similarly, Wallace, Knox and Hyder (1970) obtained 0.92, 0.92 and 0.77 as the correlation coef- icients between blood urea and N intake, digestible N and retained N respectively. The value of r = 0.99 also obtained in the present experiment between blood urea and ruminal ammonia N is high and shows that the regression equation could be used to estimate the blood urea levels at varying concentrations of ruminal ammonia nitrogen UNIVERSITY OF IBADAN LIBRARY 93 for the rations used in the present experiment. Preston ej; al#(l965) suggested that blood urea nitrogen levels could be used to assess protein utilization in lambs. Certain difficulties, however, make this almost impossible. From their results, they showed that blood urea nitrogen in excess of 10 mg/lOO ml would indicate adequate protein intake in their ration. It is however, not correct to state that lower levels of blood urea nitrogen necessarily indicate poor nitrogen intake, for factors such as / breed, age of animal and percentage of readily fermentable energy in the rations influence blood urea nitrogen levels. However, when the ration is defined, their suggestion could be useful. In the present report, even at the maximum N intake of 1.55 g/day/w the level of blood urea nitrogen was still relatively low (4.9 mg/lOO ml). Blood urea levels can be used to assess utilization of dietary protein especially of herbage. High protein herbages are likely to give rise to high levels of ruminal ammonia and subsequently high blood urea levels, which in turn would increase urinary urea excretion (Coccimano and Lfcng, 1967). The low values of blood urea nitrogen in the present report would them be interpreted to mean that the dietary N were being efficiently utilized. This is supported by very low urinary excretion even on the ration of highest concentra­ tion of crude protein. The low levels of blood urea can also show UNIVERSITY OF IBADAN LIBRARY 94 that an appreciable amount of it was being recycled to the rumen and utilized there. The results of Weller jet al# (1958) and Freitafi\et alv(l970) showed that appreciable amoung of nitrogen presented at the abomasum is of microbial origin. The values ranged from 747° by Weller _et al. (1958) with a ration of hay to 99$ by Freitag et ĵ l. (1970) with urea-based ration. It is therefore essential to know the amino acid composition of microbial protein especially as dietary protein is being replaced by non-protein compounds in ruminant nutrition. The biological value of ruminal bacteria and protozoa wore 81 and 80$ respectively and true digestibility were 74 and 91$ for ruminal bacteri and protozoa respectively (McNaught et al, 1954). The present report has shown that of the essential amino acids determined, histidine and methionine were present in very low concentration in bacterial protein. This is similar to the report of Bergen, e_t al.(l968) who also obtained low levels of these two amino acids. However, the value of 1.95 g/lOO g amino acids obtained by these investigators for histidine is much higher than the present value of 0.79g/l00g amino acids obtained in this experiment. The lysine value of li.60g/l00g amino acid obtained in , , in the present studies is/jrery good agreement with that determined by Ahde King and Engel (1964) but much higher than that reported by Bergen £t al, (1968). The concentrations of tyrosine and phenylalanine UNIVERSITY OF IBADAN LIBRARY 95 reported in the present report were lower than those of Bergen e4 ,al* (1968) and Abdo et al.(l964)« Similarly for ruminal protozoal protein, the amino acids present in lowest concentration are methionine (0»87g/l00g AA^-and histidine (l.05 g/lOOg Amino acids) and were both lower than the result of Bergen et al (1964)- Prom these results, it appears that histidine and methionine present in least concentration might limit the utilization of microbial protein. The amino acid composition of microbial protein can only give an estimation of limiting amino acids but it can not -per se be assumed to limit the efficient utilization of dietary protein, Bergen e_t _gl. (1968a) therefore used the plasma amino acid score (PAA-S) method of McLaughlan (1964) and the restricted feeding regimen of rats with 10$ protein rations to determine the limiting amino acid of microbial protein. They found that for rumen protozoal protein, histidine was the limiting amino acid. The plasma levels of free histidine in rats fed protozoal protein-based diets for ten days were extremely low, indicating that this acid was most limiting. They found that the limiting amino acid in bacterial protein was oystine, whereas arginine and histidine were the next two least available amino acids. Purser (1970) showed that pepsin was ineffective in releasing arginine from protozoal and bacterial proteins, and this could account for its low concentration in the blood plasma of rats fed microbial proteins UNIVERSITY OF IBADAN LIBRARY 96 Land and Virtanen (1959) using labelled ammonium nitrate as major source of nitrogen for lactating goats observed that histidine was very weakly labelled of the amino acids of milk. They suggested that this may be due to the incapability of the ruminal bacteria to synthesize the imidazole ring. Cystino was also weakly labelled. Loosli and Harris (1945) suggested that the low level of methionine in microbial protein may be due to slow rate of synthesis in the rumen. Prom these results, it is likely that the limiting amino acids of microbial protein are histidine , oystine, methionine, and arginine. Though the present investigations are comparable to those of Bergen_et al. (1968), it must be emphasized that the method of pre­ paration of bacterial and protozoal specimens may have brought about the differences observed in the results. UNIVERSITY OF IBADAN LIBRARY CHAPTER THREE 3. ISOTOPIC STUDIES OFD WNAIRTF ROWGEETNH EMR ETSHABEOEPLISM IN THE WEST AFRICAN 3.1 I N T R O D U C T I O N The feeding of non-protein nitrogen (NPN) supplements to ruminants is based on the knowledge that ammonia is the major end - product of the degradation not only of the NPN but also of proteins in the rumen (McDonald, 19^+8). Ruminants have been maintained on diets in which the only source of N was either ammonium salts or urea (Loosli et a l . 1 9 ^ 9 y Virtanen, 1966) , indicating that all the essential amino acids normally required by non-ruminants can be synthesized from ammonia by the ruminal micro-organisms. Isotopic methods have been used to determine the rate of production of ruminal ammonia, blood urea and bacterial and protozoal nitrogen in the sheep (Pilgrim et al., 1970; Mathison and Milligan, 1970; and Nolan and Leng, 1972). In the present report ,^^N_7ammonium chloride and £ 15nJ urea have been used to examine the rate of entry of ammonia into the ruminal ammonia pool, and urea into the body urea pool in sheep respectively. Estimates of the contribution of ruminal ammonia N to plasma urea and of plasma urea to ruminal ammonia pool, as well as utilization of ruminal ammonia by ruminal bacteria and protozoa were made- The utilization of infused ammonia for production of milk protein was also examined. UNIVERSITY OF IBADAN LIBRARY 98 3.2 MATERIALS AND METHODS 3.2.1 Animals and Their Management Four adult West African Dwarf wether sheep, 2 - 2.5 year old and weighing 19 - 35 kg, each fitted with a permanent rumen cannula, in addition to two intact lactating sheep of the same breed, were used in these studies. The animals were individually housed in metabolism cages (Oyenuga, 19 6 1). Five days before administration of isotope, the wethers were given their daily rations in equal portion at hourly intervals. 3.2.2 Diets; Ration A consisted of a high qualify Cynodon nlemfuensis/ Centrosema pubescens hay (15*3% crude protein) and 300g of this basal ration were given to each animal. Ration F consisted of 300g of hay and 150g of a groundnut - hased concentrate C_5 0 7 * 5 $ crude protein), and this was also supplied at hourly intervals, except for the lactating sheep where all the feed was supplied once. The groundnut cake-based concentrate is as already given in Charpter2, but the chemical composition of the basal hay is shown in Table 3«1» UNIVERSITY OF IBADAN LIBRARY 99 TABLE 3.1 Chemical Composition of the Basal Hay Fed to the West African- Dwarf Sheep % Crude Protein (CP) 15.3 t Crude Fibre (CF) 27.5 Ether extracts (EE) 1.2 Nitrogen-free extracts (NFE) 50.5 Ash 5-5 Total 100.0 3.2.3 Plan of Experiment On the day of the experiment, two sheep (Nos 210 and 273) received an aqueous solution (100ml) of ammonium chloride (250mg, 99% enriched with 15N) as a single infusion into the rumen. The other two sheep (Nos 186 and 259) were given a single infusion of / "hy urea (250 mg, 95% enriched with 15jj) into the blood. The two lactating sheep (Nos. 72 and 90) each received an aqueous solution (100 ml) of £ 1\ 7 ammonium chloride (200mg, 99% enriched with 'N). The solution was administered orally in two equal portions to each sheep at 8.00 a.m. and 2.00 p.m. UNIVERSITY OF IBADAN LIBRARY 100 3.2A Collection, of Faeces, Urine, Blood, Rumen and Milk Samples: Samples of 10 ml ruminal fluid were taken at two - hourly intervals and immediately transferred into a deep-freezer at - 5°C. Five samples of ruminal fluid were taken during the ten hour sampling period. Samples of 15ml of blood were placed in heparinized centrifuge tubes and centrifuged at 2000 rpm for 20 minutes. The plasma was stored in a deep freezer at-5°C. Urine from the wethers was collected in a collecting bottle containing 5ml of 10$ mercuric chloride solution to prevent microbial breakdown of the nitrogenous components of the urine. Urine from the ewes was collected by using bladder catheters and also stored at - 5°C. Faeces were collected from the wethers by means of harnesses to which were attached collection bags. Faeces from the ewes were collected from the cage floor at 8.00 a.m. in the morning. The ewes were hand-milked at 8.0 0 a.rm« The milk samples were stored in a deep - freezer at - 5°C, until required for analysis, 5.2.5 Separation of Bacteria and Protozoa in the rumen Liquor Bacteria and protozoa of ruminal fluid were separated by the differential centrifugation method of Blackburn and Hobson (i960). UNIVERSITY OF IBADAN LIBRARY 101 3.2.6 Analytical Procedures The preparation of ruminal ammonia and blood urea for isotopic analysis was according to Nolan and Leng (1972), except that distillation was carried out in Markhams’s (19^2) distillation apparatus. Frothing of samples was prevented by addition of sec-octyl alcohol as the anti-foamant. Nitrogen in the urine, faeces, milk and microbial samples was determined by the semi-micro kjeldahl procedures with NaOH used as alkaline reagent during steam distillation. •15 Samples were analyzed for enrichment of N by means of the mass spectrometer at IAEA, Vienna. 3.2.7 Theory Estimation of pool size, urea space and entry rate was by the method of Zilversmith (i960). Pool size (P) = »» fe- «* €T -U where, a = g of labelled material injected or infused, e^ = atom per cent excess of added material, e^ = atom per cent excess of isolated material. Ureagpace = Pool size (P)_____ Concentration of urea T i T t r Entry (g) = P __________ _ tyz x 1 • where ty2 is ihe time for half the enrichment to be lost from the sampled pool. UNIVERSITY OF IBADAN LIBRARY 102 3.3 RESULTS The recovery of„ 13.N, administered as ammonium chloride into the rumen or as urea into the blood stream is shojwn in Table 3*2* Low recovery of 15"'N injected into the rumen-occurred in the urine, and the label was not recovered in the faeces of the four wethers. The result showed that 5-296 and 6.9$ of administered dose of 'N were recovered in the urine of the wethers, and the values for the ewes were k.k% and if. 8$. Of this amount recovered, 63-3$ and 7 3-9$ were recovered during the first 2k hours in the wethers and ewes respectively. The mean recovery of 15N from the urine of the wethers r-15 -7 after intravenous administration of /_ "w_J urea was 30-5$ 1 and 89.896 of this was recovered during the first 2k hours of urination. None of the injected 15N was recovered in the faeces of the wethers. The mean recovery of 15N in the faeces of the ewes was 9.1$, and if3.996 of this was recovered during the first 2k hours of administration. The result showed that about 3-1$ of administered dose of 1"5/N was recovered in the milk protein, and of this i+1.9% was recovered during the first 2k hours after administration. The total amount of isotope recovered in the urine, faeces, and milk of the ewes was about 1?.2$ of administered dose. UNIVERSITY OF IBADAN LIBRARY 103 TABLE 3-2 The Recovery of 15N (Administered Either as Ammonium Chloride into the Rumen or Urea into the Blood) in the Faeces, Urine and Milk of Dwarf Sheep Animal Live Nitrogen 15 ^N recov ered in 15N Recovered xn Adm^iNnis­ N Recovered in " TOTAL No. Weight Intake Urine Adminis­ Faeces ($ Adminis- milk (as % 15 (g/day) tered tered) ter ed Administered) N recovered (mg) First Total First Total (as % adminis­ 2k hours % 2k hours % First teredTotal 2k hours 210 21.3 6.2 65.5 k.2 6.9 0 0 - - 6.9 273 25 10.1 63.5 3-3 5.2 0 0 - - 5.2 72 28.6 10.1 52. 2.9 *f.*f 6.5 8.5 1 .8 2 3.93 16.7 90 19-5 10.1 52.4 3.9 k.2 1.*f 9.6 0.79 3.17 17.6 186 27.6 6.2 116.7 30.8 32.3 0 0 - - 32.3 259 35-0 10.1 116.7 25.9 28.7 0 0 - - 28.7 - Enrichm ent w'.tti i5N Was z e ro ° U e x c e s s . — a Klethers usec=l C Ni® nr»rlk ) . UNIVERSITY OF IBADAN LIBRARY 10*+ The enrichment of ammonia N, blood urea N, bacterial N and protozoal N with time after a single injection of £ " ̂5N2. ammooium chloride into the rumen of sheep Nos* 210 and 273 is shown in Figures 3*1 and 3.2 respectively. The ratio of the area under the appearance curve for plasma r- 15 N u-rr e a , bacterial /_ _/ and protozoal ^N_/ to the area under the disappearan tion of urea N, ce curve forN N a m m o n i a , gave the propor­bacterial and protozoal N respectively contributed by ruminal ammonia (Table 3*3) • The results showed that ruminal ammonia N contributed 1*+.*>% of plasma urea N in sheep fed on hay, while the corres­ ponding value for that on hay/concentrNa te was 5*2%. In the Ns heep fed on hay, 33*2% of bacterial and 19*0$ of protozoal were derived from ruminal ammonia N in the period under investigation.N Ruminal ammoniN a N contributed 25*9% and 1*+.8% of bacterial and protozoal respectively in aaimalsN fed hay/concentrate ration. The enrichment of the protozoal with 15N was 57.2% of that of bacterial N in the sheep fed on hay, and hay/concentrate rations. The enrichment of blood urea N and ruminal ammonia N with time after a single injection of / ■ 15n_7 urea into the blood of the sheep Nos. 187 and 259 is shown in figures 3*3 and 3°*+ respectively. The ratio of the area under the appearance UNIVERSITY OF IBADA LIBRARY ' V ' ’ "/ Log Enrichment (Atoms % excess x10)UNIVERSITY OF IBADAN LIBRARYFIG.3-1 ENRICHMENT OF RUMINAL NH3- N (x-x), BLOOD UREA -N (»—o (No.210) BACTERIAL-N &— &), AND PROTOZOAL-N — °) AFTER INTRA RUMINAL INFUSION O F [15N] AMMONIA Log Enrichment (Atoms % excess x 100) : % P o i ^ a o N j c D o i s C D N j ( j » o N to -------1------- 1------- 1------- 1------- 1------- 1------- 1------- 1------- 1------- r c o t'o CD m > z c o 5 oC/} xm O N3z > m g ? i cn $x 3 c a > “0 2 — > Ko r ~ CDos > > T O OD > S ! m X I Cr _D8 O > i x m a > Kz 'O' > UNIVERSITY OF IBADAN LIBRARY FIG. 3-4 ENRICHMENT OF BLOOD UREA (°— °), AND RUMINAL (No.259) AMMONIA (x— x) AFTER INTRAVENOUS INJECTION OF [ 1 5 n ] UREA Log Enrichment (Atoms % excess x 10 ) UNIVERSITY OF IBADAN LIBRARY FIG. 3-4 ENRICHMENT OF BLOOD UREA(°— °),AND RUMINAL (No.259) AMMONIA (x— x) AFTER INTRAVENOUS INJECTION O F [ 1 5 n ] UREA Log Enrichment (Atoms % excess x 10 ) UNIVERSITY OF IBADAN LIBRARY 109 curve for ruminal ammonia to the area under the disappearance curve for blood urea N gave the proportion of ruminal ammonia N contributed by plasma urea N (Table 3-3)• The results showed that k 7 ,2% of ruminal ammonia - N of hay - fed animals was contributed by plasma urea - N, and the corres­ ponding value for sheep fed on hay/concentrate was 1 5*2%. The results presented in Table 3»3 showed that the fluid volumes of the rumen were 5°1L and 6,9 L for the two sheep used. The N pool size in the rumen were 1.03 and 1.25g respec­ tively. In the hay - fed animal, 8.6g of ammonia N entered the ruminal ammonia pool daily and was removed from it. The body urea pool size were estimated as 3«66g for the hay - fed sheep, and J . 8 k g for the sheep given concentrate - based ration. Body urea space were estimated as 17«^ and 21.16L for the animals used. When body urea space was expressed as per cent of body weight, values of 62,9% and 62.2% were obtained for the sheep. The amount of urea - N which entered the blood was estimated as 10.1g/day for the hay-fed animal, and S .k g/day for the animal on the concentrate - based ration. The amount of urea - N excreted per day were 2.8g and *t.7g for these animals (Table 3«3)« The values obtained when urea excretion (g/day) is subtracted from total entry (g/day) were values of urea degraded in the alimentary tract daily, and is the UNIVERSITY OF IBADAN LIBRARY 110 TABLE 3.3 Ammonia and Urea Metabolism in the West African Dwarf Sheep Estimated by Using Single Infusion of (15m ) Ammonium Chloride and Single Injection of (ig ) Urea into the Rumen and Blood Respectively A M M O N I A U R E A . No. 210 No. 273 No.186 No. 26®! (ration A) (ration F) (ration .A) (ration! Live weight (kg) 21.3 25.4 27.6 35.0 N intake (g/day) 6 . 2 1 0 .1 6 . 2 10*1 Fluid volume (l) 5.1 6.9 - - Ammonia concentration (mg N/100 ml) 2 0 .0 1 8 .0 - ! Plasma urea concentration (mg N/100ml) - - 21 .1 1 8 .2 Pool size (g) 1 .0 3 1 .2 5 3.66 3.84 Total entry (g N/day) 8.64 1 0 .3 2 10 .1 9.4 Urea excretion (g N/day) - - 2.8 4.7 Urea degradation (g N/day) - - 7.3 4.7 Body urea space (l) - - 17.4 21.2 Body urea space 62.2 (% body weight) - - 62.9 Microbial N from ruminal NH_5-N($) (a) Bacteria 33.2 25.9 - - (b) Protozoa 1 9 .0 14.8 - - Plasma urea from ruminal ammonia (%) 14.4 5.2 Ruminal ammonia contributed by plasma urea (,%) 47.2 15.2 - - • UNIVERSITY OF IBADAN LIBRARY 111 estimate of blood urea transferred to the digestive system. The amount of urea N from blood degraded in the digestive system were 7«3 g/day, and ^»7 g/day for the animals on the basal hay and the concentrate - based ration respectively. These were ^2*3$ an< ̂ of total urea entry per day, for the sheep maintained on the basal hay and the concentrate - based ration respectively. UNIVERSITY OF IBADAN LIBRARY 112 3A d i s c u s s i o n The validity of measuring urea and ammonia kinetics using a single dose of 15N in the form of urea or ammonium salt is based on the assumption that the system described is in "steady state", i.e. the compartment sizes and turn - over rates remained constant during the experimental periodo Under the hourly feeding conditions used, fairly constant concentrations of ammonia in ruminal fluid and urea in plasma were obtained, indicating that these conditions were largely fulfilled. The values of 5 .2% and 6.9% as the per cent of injected <1 £7 ammonium "N recovered in the urine of the wethers, and k.k% and ko8% in the ewes obtained in the present experiments were very low compared with values of J>8.6% observed by Piva and Silva (1968). Mathison and Milligan (1971)? Nolan and Leng (1972) have observed very low recovery of N in the urine of sheep and have siiggested that absorbed ammonia might not have been converted into urea in substantial amounts, and that the net retention of labelled N within the tissues of the sheep may not only indicate the extent of net utilization of ammonia, but could reflect exchange within the animal body as well. Nolan and Leng (1972) suggested that some of the absorbed ammonia could have entered nitrogenous compounds (such as amides or non-essential amino acids in the intestinal wall UNIVERSITY OF IBADAN LIBRARY 113 or liver), which are then incorporated into slowly equili­ brating pools of N in body protein. This is consistent with the reports of Piva and Silva (1968) who showed that adminisfe tered dose of 15N was incorporated into the muscle protein of the sheep. From the present report, 30.5% of 15N administered as urea through the blood was recovered in the urine, which suggest that administered urea was well utilized by the sheep. liugerwa and Conrad (1971) showed that from 5^ to 90% of injected urea was retained by the sheep. The present report with value of 6 9 .5% retention of injected urea could be taken to mean that injected urea was synthesized into ruminal microbial protein which was then digested and the products utilized for synthesis of tissue proteins of the sheep. However, the percentage of injected urea retained would be influenced by dietary nitrogen intake, it would be high at low N intake, and low at high N intake ( Coccimano and Leng, 196?; liugerwa dnd Conrad, 1971)« The present report showed that 9 . “)% of 15N administered to the lactating sheep was recovered in the faeces. This appears to be in good agreement with the value of 7 »9% obtained by Piva and Silva (1 9 6 8). UNIVERSITY OF IBADAN LIBRARY The fact that 15N was not recovered in the faeces of the wethers is very difficult to explain. This may be due to the fact that whereas it was applied in two doses to the ewes, it was applied only in one dose to the wethers, and the loss by absorption from the rumen would be less with the ewes than with the wethers, and a higher proportion of 15N would be incorporated into microbial protein in the ewes than in the wethers. The fact that absorbed ammonia may be utilized by a pathway other than the direct conversion into urea was demonstrated by the recovery of injected 15N in milk protein. In the present report, 3-1% of administered dose was recovered in milk protein and this is lower than the value of 5-6% obtained by Piva and Silva (1 9 6 8). However, Land and Virtanen (1 9 5 9) showed that the proportion of ammonia used for milk protein synthesis would be low with liberal protein feeding, and this may explain the observed low value in the present investi- gat ion. The total recovery of 15n in the ewes was 17.2% which showed that 8 2 .8% was retained in the tissues. The result of the present investigation showed that ruminal ammonia contributed 1^.^ and 5*2% of the plasma urea of sheep fed on the basal hay and concentrate - based rations respectively ten hours after infusion of 15 N into the rumen. UNIVERSITY OF IBADAN LIBRARY 115 Nolan and Leng (1972) obtained the values of 11% and 4-5% three hours after infusion and at time infinity respectively. The percentage of bacterial N derived from ruminal ammonia N were 33»2% and 25.9% for sheep fed hay and the concentrate - based ration respectively, ten hours after injection of N. Mathison and Milligan (197^) obtained the value of 50 - 65% as percentage of bacterial N derived from ruminal ammonia N after 144 hours of continuous infusion of 15N into the rumen. The percentage of protozoal N derived from ruminal ammonia were 1 9.0% and 14.8% for sheep fed on hay and con­ centrate - based ration respectively. The values obtained by Mathison and Milligan (1971) using continuous infusion were 31 - 55% after 144 hours of infusion. The present report with values of 1 9*0% and 14.8% after 10 hours showed that the ruminal protozoa were utilizing ruminal ammonia at comparably high rate. The enrichment of the protozoal fraction with was 57*2% of that of the baBterial fraction; the value obtained by Mathison and Milligan (1971) was 56 - 96%. These authors observed that as the concentration of ruminal ammonia increased, there was a decrease in the proportion of ammonia utilized by ruminal bacteria. In the present investigation, ruminal ammonia levels were high and this might contribute to the less UNIVERSITY OF IBADAN LIBRARY 116 than maximum conversion of ruminal ammonia N into bacterial protein. An appreciable quantity of blood urea was being taken to the rumen either via saliva or ruminal epithelium. This is shown by the enrichment of ruminal ammonia with 15N after intravenous injection of urea. The result showed that ten hours after injection, ^7*2$ and 1 5*2% of ruminal ammonia N were derived from blood urea in sheep fed on hay and the concentrate - based rations respectively. These values are higher than the value of 12$ obtained by Nolan and Leng (1972) and would appear that a larger quantity of blood urea was entering the rumen in the animals used for the present report. The total entry rate of ammonia N into the rumen were 8.6g and 10«3g/day in sheep fed on hay and protein concentrate - based rations respectively. The ammonia is removed either by absorption through the ruminal epithelium, or by incorpo­ ration into microbi&l N or by loss through the digestive tract. Nolan and Leng (1972) obtained values 1A.2 - 17«2g per day for sheep maintained on high quality hay. The body urea N pool size obtained in the present investigation were 3-7g and 3«8g for sheep maintained on hay and the protein rations respectively. These are in good agreement with the values of J>,6 to k .k g obtained by Nolan and Leng (1972). Coccimano and Leng (1967) obtained UNIVERSITY OF IBADAN LIBRARY 117 values of urea pool size ranging from 0.55 to 1 2«9 0g and showed that urea pool size increased with plasma urea concentration. The values of urea space obtained in the present inves­ tigation, were 1?.*f1 and 21.21, and these are in good agreement with the value of 18.7L obtained by Nolan and Leng (1972). Coccimano and Leng (1967) obtained values ranging from 6.3 to 27«OL. Urea space as per cent body weight were 62.9% and 6 2.2% for sheep fed on hay and mixed ration respectively, and these are within the range of the values of 55 - 66% obtained by Nolan and Leng (1972). The difference between total entry rate and total excretion rate of urea gave an estimate of the amount of urea degraded in the intestinal tract. The values were 7»3g/day and ^.7g/day in sheep fed on hay and the protein ration respectively, and urea degraded in the digestive tract as percentage of urea entering the body urea pool were 1 2 - 3 ^ and 50.0% for sheep fed on hay and protein rations respectively. Coccimano and Leng (1967) obtained values ranging from1.6 to 12.9g/day as urea degraded, and showed that the percentage of urea entering the body pool that is degraded decreased as dietary N increased, and this shows that in animals maintained on a low level of N, an appreciable amount of N can be supplied by blood urea. Nolan and Leng (1972) obtained the values 5«2 UNIVERSITY OF IBADAN LIBRARY 118 to 8.4gf/day as urea degradation rate in sheep maintained on high quality hay. The fact that much N can be brought to the digestive tract of the ruminant in the form of urea via saliva or ruminal and intestinal epithelia is of great importance to domestic ruminants in the tropics, which may at times have access only to roughage _■ of 3 - 5% crude protein content at the dry season of the year. The contribution of this blood urea would go a long way to making the ruminant livestock survive adverse climatic conditions particularly in the arid tropics where protein supply is at its lowest ebb. UNIVERSITY OF IBADAN LIBRARY CAHPTER POUR 4. THE UPTAKE AND DIGESTIBILITY OP DRY MATTER AND NITROGEN METABOLISM IN THE WEST AFRICAN DWARF NETHER SHEEP MAINTAINED ON HAY AND CONCENTRATE SUPPLEMENTS. 4. 1: INTRODUCTION Even though the potential for neat production by the West African dwarf sheep is realised, there has not been nuch investiga­ tion of the way in which this breed utilizes local forage and concentrates. As a result, the nutrient requirements for this class of livestock are entirely lacking. In the present report, the intake and digestibility of dry natter (DM) and the netabolisn of nitrogen (l'l) of basal hay and concentrate supplements fed to the Host African dwarf wether sheep were examined. Estimates of the crude protein requirement for maintenance as well as the metabolic faecal and endogenous urinary N were also made. 4.2: MATERIALS AND METHODS. The details of diets, animals and their management, plan of experiment, collection of faeces and urine, as well as analytical procedure are as previously described in Chapter 2. UNIVERSITY OF IBADAN LIBRARY 120 4.3: RESULTS The results of the digestibility of DM and N obtained with fistulated and intact sheep are presented in Table 4.1. Hie results for fistulated animals wore compared with those of intact animals in order to show if fistulntion .affected the digestibility of DM and N. The results show that fistulation had no effect on digestibility of DM and I? (pj> 0.05) and that digestive processes in fistula ted sheep closely approximated those in intact sheep* therefore the results of the two sets of animals were pooled for subsequent use in the present investigation. The mean DM intake^digestibility and IT metabolism for the West African dwarf wether sheep maintained on basal hay and concen­ trate supplements are summarized in Table 4.2, 4.3.1 The Dry natter intake. Supplementation of hay with concentrate C-j had not signifi­ cantly ( P j 0 .0 5) increased daily DII intake with the mean HI intake 0.734 of 49.4 ± 7.8,t and 62.4 i- 5 . intake per for rations A and B respectively. . Hie re wore no significant increases in DM intake in Trial 2 (P,) 0.05), with the means ranging from 61.2 + 3 .8 for rations C to 72.6 +3.3 g per kilogram metabolic size per day for ration. F. The DM intake increased with increasing levels of dietary crude protein and also with nitrogen intake. The DM intake was not affootod by the digestibility of DM even though there was some trends UNIVERSITY OF IBADAN LIBRARY 121 TABLE a . 1 Effect of runinal fistulation of the Eor.it African Dwarf 'Esther sheen, on digestibility of dry patter and nitrogen contents of basal hay and concentrate sunplenents T R E A T M E N T S A B C E F f! [ D | Fistulatec|1 Intact Fistulatod j Intact Fistulatod Intact j Fistula tod Intact Fistulatod Intact »Fistulato | Intact I _ __ ___________ Tj ..... r I i 57.0 t 72.7 64.5 75.2 73.3 67.8 69.3 69.4 65.2 66.9 | 76.8 DIGESTIBILITY | r —■ -5-5. -4---* ,T , , t1 OF 53.6 j 72.2 78.8 70.3 71.9 80.2 : 72.5 75.0 | 77.2 DRY MATTER 57.3 68.5 ! 79.4 (p e r c e n t) 57.9 56.7 -70.8 j 72.0 I 75.9 71.9 74.1 79.0 | 73.9 78.4 76.4 { 77.5 L. . . . ___ i_ — —J | j__________56.5 57.3 74.7 75.8. 80.8 . 74.1 76.1 76.3 77.9 76.7 76.4- 75.8-— MEAN ^57.2*0.3 55.7+0.7 71.7±1,2 71.1±2.3 77.8±1.5 f;74.1 ±1.3 74.1 ±2.1 75.4+1.5 ij 73.2+2.5 73.2+2.5 73.7+1.7 76.8+1.0_________ J __ _ . 56.7 57.1 63.4 63.6 59.7 58.6 62.5 58.3 jj 5 7 .4 | 61.4 72.0j ...... .. j. — 1 .. \ 72.4 j... DIGESTIBILITY 53.2 56.4 i 62.0 49.5 51.0 64.0 67.1 58.7 7 1 .6 ; 65.6 69.2_________ t . .......____ } i— 73._4__ NITROOGFE N ! 54.9 49.7 fI 56.1 63.8 55.6 t 74.2 ( ) i ; “ 71.0 73.1 62.4 73.5 70.7 j. -.... i p e r c e n t ' ___l59.4 I i : 59.7 ! .65.8 i .65.2 .59.0 .56.8 J- .65.9 .60.7 .69.5 I 70.4 .70.2 j. _ 67.5 MEAN 56.0+1.7 j55.7+2.0 | 61.3+2.0 1 60.5+3.3 56.3+1.9 |59.8+1.8 | 66.6+1.8 |62.7+3.0 | 65.2+3.0f 67.7+2.5 71.6+0.6 {70.8+1.6 _ _ _ _ A ----- i “ A— MEAN DIFFERENCES NOT SIGNIFICANT AT jfo LEVEL, BETWEEN FISTUT-ATED AND INTACT ANIMALS II.' THE PARAMETERS CONSIDERED. A ' UNIVERSITY OF IBADAN LIBRARY 122 TABLE 4.2 Dry natter intake, digestibility and N netabolisn for the ¥cst African dwarf wether sheep maintained on basal hay and concentrate supplements. _______ .... . .1. T R .E.. A . T M .E... N.... T...S.. . _ - __ . Trial 1 • Trial 2 | A B C D E P INTAKE OP DRY HATTER (g/day/wj^734 X X a49.4±7.6 62.4+5.5 6l.2a*3.8 67.8a+7.8 68.9 ±3.0 72.6 ±3.3 -—— - - INTAKE OP NITROGEN g/day/fc-^ X Q.0.62 + 0.27 0.49 + 0.11 0.8-2 + 0.07 0.91 + 0.28 1.28 + B 0.18 1.55 + 0.17 X y DIGESTIBILITY OP DRY MATTER OP 56.5 + 1.0 71.4 + 1.2 76.5 + 1.7 73.1 + 1.4 74.2 + 2.4 75.1 + 1.5 RATION, ($ I. ... ... ---- ----- --- r --- .. — DIGESTIBILITY OP DRY MATTER OP t CONCENTRATE (fo) - 85.5 + 4.6 | 90.3 + 2.2 83.0 +1.1 83.8 + 1.8 87.1 + 2.8 DIGESTIBILITY OP NITROGEN OP X x T a ¥ c RATION 55.9 + 1.0 6 1 . 2 + 1 . 7 | 63.5 + 0.6 69.4 + 1.6 70.0 + 1.8 76.3 + 1.8{%) ______ ~ ....... u .. “ _________ _ DIGESTIBILITY OF NITROGEN OP j a b * c CONCENTRATES. ($ - 75.7 + 5.9 | 68.7 + 1.9 78.6 + 2.4 80.6 + 1.3 88.6 + 2.1 . ... { NITROGEN DIGESTED, bo X x j a ab c g / d a y / w g ^ 0.35 + 0.05 0.30 + 0.03 ; 0.54+0.03 0.63 + 0.03 0.91 + 0.03 1.20 + 0.02 i *" i ~vzr~ c NITROGEN RETAINED. g/day/W0*754- x X r cl akg 0.29 + 0.05 0.20 + 0.02 i 0.48 + 0.04 0.59 + 0.09 0.86 + 0.08 1.11+ 0.09~ a ab" ' “ NJ'TUOGRJN RETENTION 7 "b T42.3 + 2.3 J 51.7 + 1.7 57.5 + 1.6 63.6 + 2.7 66.5 + 2.3 69.5 + 1.6 ROW UNIVERSITY OF IBADAN LIBRARY 123 The regression equations showing relationships between nutrient utilization in the West African dwarf sheep maintained on hay and concentrate Jupplenents. Nos y x ; REGRESSION EQUATION r 4.1 DMI MS Y = 0.28 + (0.040+0.002)X 0.50** i 4.4 log PI log ¥ Y = -1.055+(0.66840.026)X 0.97**!I 4.6 had foC-R Y = 48.95 + 0.39X 0«72* * ; / 7 DP log0 CP Y = 12.15 + (56.06+5.92)X 0.91** | DNp Cp-C Y = 62.91 + 1.43X 0.97** 4.9 UN Cp—C = X. Y = 51.67 + 1.35X-0.027X2 ^O-R = X, 4.10 DN/kg NI /k g Y = -5 .4 5 + (0.896+0.025)X 0.99** 4.11 DCp M. Y = -1.86+ (0.862+0.020)X 0 .8 6 "" y. ND NI Y = 0.163+ (0.863+0.045)X 0_ .^9 ̂9 -Xr* NR ND Y = -0 .014+(0 .939+0 .029)x 0.99** | n n - Dry Matter Intake, kg/day Ms - Metabolic Size, WleBg 4 - W *69'13t PI - Peed Intake, kg ¥ - Live-weight of animal, kg HID - Dry matter Digestibility,. per cent 5'cG—R — Percent Concentrate in ration DP - Percent Digestibility of Crude Protein of the ration - Percent Crude Protein in ration. DIIp - Percent Digestibility of Nitrogen of concentrate CP-C — Percent Crude Protein in concentrate, Dll/kg - Nitrogen Intake/eg Dry Matter Intake, DCp - Digestible Crude Protein, g ND - Nitrogen Digested, g/day/W^'J^ NI - Nitrogen Intake, g/dayW^'7^r NR - Nitrogen Retained, g/day/l^‘7 ^ ' * Correlation coefficient significant at (P <1 0.05) ** Correlation coefficient significant at (P754 FN - Faecal Nitrogen,• g/kg Dry Matter Intake UN - Urinary Nitrogen, g/day ^ ' P ' r Nitrogen Intake, g/day - + — MFN estimation for ration B. * - Correlation coefficient significant (p40.05) ** - Correlation coefficient highly significant (pcO.Ol) UN i f I IV 11 ... [ERSITY OF IBADAN LIBRARY o • CO t • *M25 125 with rations D, E and F of increasing DM intake with increasing DM digestibility. The effect of supplenontation of hay with the concentrates is shown in Table 4.4. The neon DM intake of hay was 49.3 + 7.3 . , 0.734 g/day/W^ . hlion the hay was supplemented with concentrate 01 , which consisted of nainly cassava flour, intake of hay foil to 0.734' , 0.734' 39.9 s / \ g t while 22.4g/wjJ.g, . of concentrate C-j was consumed. Table 4.4 shows that in Trial 2 with tho protein concentrates, there wore decreases in the intake of hay and increases in the intake of concentrates. Thus, 100 g of concentrate replaced 42,0 g of hay while 100g of concentrates C2 - O5 re placed 66 .8 g, 53.0 g, 54.6 ̂and 49.0 g of ha?/ respectively the neon being 53.1 + 8.0. Thus, 100 g of concentrates replaced 53.1 g of hay during the voluntary intake of hny/concontrato rations by the sheep. From Table 4.4 it is seen that the intake of concentrates increased with increasing levels of crude protein in the concentrates but the consumption of hay was almost equal for rations C, D, E and F which contained crude protein. Less of concentrate than O2- was taken b̂ r the sheep. The proportion of DM consuned which is concentrate increased with increasing crude protein in the concentrate, being 58.5$ for ration G and 62.9$ for ration F, but 35.9 for ration B. For rations G to F, the means differences of DM taken as concentrates were not significantly different (P>0 .0 5) It is thus seen from this 3duly that while supplementation of hay (f.7$ crude protein) with concentrates increased the total DM UNIVERSITY OF IBADAN LIBRARY 126 TABLE 4.4 Effect of supplementing basal hay with concentrates on Intake of hay fed to the Most African dwarf sheep Tried 1 Trial 2 A c i D P V--1“ “ “ 1 E ! TOTAL DRY-HATTER INTAKE , ,0.734-". 49.3 62.3 Ii 61.2 67.6 68.9 7 2 . 6g/day/Hj^ INTAKE OP' HAY g/day/ ̂ *734 49.3 39.9 25.4 28.4 25;7 2 6 . 9 — INTAKE OP CONCENTRATE _ 2 2 . 4 35.8 39.8 43.2 45.7 g / d a y A f ' J ^ ■ " ' 1 ____ INTAKE OP CONCENTRATE 1 'l AS PERCENT TOTAL INTAKE » 3 5 . 9 58.5 i 58.2 62.7 6 2 .9 I1— -— INTAKE OP HAY AS PERCENT ! TOTAL INTAKE 100 64.1 41.5 i1 41.8 37.3 37.1i *.“. •*_•. AMOUNT OP HAY REPLACED BY 100g OP CONCENTRATE 42.0 6 6 ,8 53.0 54.6 4 9 . 0 DRY MATTER I iJ------ UNIVERSITY OF IBADAN LIBRARY 127 intake, consumption of hay was reduced, and this reduction was greater for concentrates with higher levels of crude protein than for those of low crude protein content. At the sane tine, the level of consumption of c oncentrates increased with increasing levels of crude protein in the concentrates. The following regression equation relating DM intake (y) kg/day to the metabolic size of the sheep (x), was obtained: Y = 0.2C + (0.040 + 0.002) X, ................>(4.1 ) (r - 0,50 , P<0.01) Tile correlation coefficient was highly significant (P<0.01). This equation can be used to estimate the amount of food required for the sheep. For example, the live weight of sheep No. 263 was 2 3 .6 kg in the second trial and this is about 10.0 kg when converted to the metabolic 3ize. The animal then requires 0.68 kg DM according to the regression equation. This value is approximately 3^ of the live weight of the sheep. It was found that the equation of the type C where C is DM consumption in kilogram, and ¥ is the live weight in kilogram, a raid b being constants, could be used to estimate intake of DM by the sheep. Regression of log C over log ¥ gives *b* as the regression coefficient and ’a* as the constant. The equation above can be re-written as UNIVERSITY OF IBADAN LIBRARY 128 log C = log a + b log ¥ ......(4.3a) or I = A + b log X ......... (4.3b ) The equation of regression is Y = (0.668 + 0.26) X - 1.053 ........ (4.4) Hie antilog of the constant gives a value of 0.0885. Thus the equation is Y « 0.0885W0,668 ...................(4.5$ (r = 0.91 , P 4'0.05) The equation shows that DM consumption varies not with the 0.668 weight but with the metabolic size of the animal ( ¥ ^ ). This approximates to 0.67, the exapenont relating body weight to surface area. 4.3.2 Dietary IT intake; The mean N intake when expressed on a metabolic body size basis / 0.734 V*]cg , where, ¥ was the mean live weight over the period)N were not significant for rations A and B in Trial 1, but highly significant (P-CO.Ol) for rations C, D, E and P in Trial 2. Jntako of N incre ased with increasing levels of dietary crude protein, ranging from , 0.734 0.734 0.49 + 0.11 g/Wicg in ration B to 1.55 + 0.17 g / \ g in in ration P. The moan differences wore not significant for animals or periods (?.> 0.05). Supplementation of basal hay with cassava did not significantly (P_> 0.05) reduce N intake. In Trial 2, II intake increased with increasing digestibility of the dietary IT though not significantly (P> 0.05). UNIVERSITY OF IBADAN LIBRARY 129 4.3*3 The digestibility of dry natter: Hie supplcnentation of hay with cassava flour significantly (PCO.05) increased the DM digestibility of the ration. Hie DM digestibility for ration A was 56.5 + 1.0$, and was 71.4 + 1.2$ for ration B. Mean differences were not significant (P„> 0.05) for the rations used in Trial 2. There was a. slight increase in DM digesti­ bility wiih increasing levels of dietary crude protein especially with rations D, E and F. Fron Table 4.2 it is seen that the DM digestibility of ration B containing a concentrate of little crude protein content was just as high as those for the rations containing substantial anount of crude protein in the concentrates. The DM digestibility (y ) is related to the percent concentrate in the ration (x) by the equation Y = 0.39 X + 48.9 (r = 0 . 7 2 , P^0.05) ....... . ..(r.6) This shows that DM digestibility of these rations increased with increasing percentage of concentrate present in the rations. Fron this regression equation, the digestibility of the ration when no concentrate is present is 48.9$. This value is lower than 56.5$ obtained for DM digestibility of hay (ration a). On the other hand, when there is no hay in the ration, the digestibility of the concen­ trates alone is about 88$. This value agrees very well with 86,0$ obtained for the DM digestibility of the concentrates alone. Similarly, it can he shown that when the ration contains 63$ concentrate (Table 4.3 ration F), the DM digestibility is 73.5 and this is very UNIVERSITY OF IBADAN LIBRARY 150 close to 75.1 obtained for DH digestibility of ration P (Table 4.2). The neon differences of DM digestibility were not significant within the experimental animals. (p>0.05). 4.3.4 The digestibility of IT. The nonn differences were highly significant (PcO.01) for the treatments in 'Trial 2. The digestibility values ranged from 63.3 + 0.6 for ration C to 76.3 + 1 . 8 $ for ration P. There was / a linear increase in digestibility with increasing crude protein intake and percent dietary crude protein, and this relationship (with r = 0.91) was represented by the equation Y = (56.06 + 5.92) log X + 12.15 ............ (4.7) where, Y is the percent digestibility of crude protein of the ration and X is the percent crude protein of the ration. The relationship was .also represented by the equation Y = 62.91 + 1.43 X ........ ..... ....(4.8) (r = 0.97) where, Y is the percent digestibility of crude protein of concentrate and X is the percent crude protein of the concentrate. The digestibility of crude protein of the ration is not only influenced by the percent crude protein of the concentrate but also by the percentage of concentrate in the ration. Tills relationship wa.s represented by the equation Y = 51.67 + 1.33^ - 0 .027X3 (4.9) UNIVERSITY OF IBADAN LIBRARY 131 where, Y is the digestibility of dietary crude protein, X-| is the percent crude protein in the concentrate and X2 is the percent concentrate in the ration. From this equation, the digestibility of dietary crude protein increases with increasing crude protein content of the concentrate but decreases with increasing level of concentrate in the ration, which nay be due to increasing metabolic faecal N (M!®:) that accompany high levels of concentrates in the rations. Supplementation of hay with cassava flour did not significantly / (P> 0,05) increase the digestibility value for the N content of hay, tile mean values being 55.9 + 1.0 and 61.2 + 1.7 % for rations A and B respectively. The Truo Digestibility of IT. The truo digestibility of rations C, D, E and F used in Trial 2 has been determined by the regression methods and also by the Detergent method (Mason 1969). The regression of digestible nitrogen per kg 9 DM intake ( y ) on N intake per kg DM intake (x ) gives the regression equation: Y = (0.896 ± 0 ,025 )x - 3.45 ................................. (.4.10) (r = 0.99) The coefficient of X which is 0.896 gives the estimate of true digestibility of the rations. Thus, true digestibility of nitrogen in the rations is 89.6 fo. Regression of digestible crude UNIVERSITY OF IBADAN LIBRARY 132 protein ( y) on the percentage crude protein of the ration (x) given a regression equation: Y ss (0,862 ± 0.020) X - 1.86 ................... .(.4.11) (r = 0.86) 9 The coefficient of X gives an estimate of true digestibility and this is 0.862 or 86.2 c/o . when the value of the metabolic faecal nitrogen (MET?) is substracted from the total faecal nitrogen, the faecal nitrogen of dietary value was obtained. In table 4.5 is given the values of true digestibility for the experimental rations. For ration C, the total faecal nitrogen per kilogram DM intake was 4.75 ±0.68; and metabolic faecal nitrogen was 65.89 of this. Since the .apparent digestibility of ration C was 63.3 /4 the true digestibility value is given as 63.3 + 65.89 1° of /"lOO - 63. or 63.3 + (0.6589 X 36.7). and this giv* es the true digestibility to be 87.5 ' 9 Mien calculated in_this way, the true digestibility of rations C, D, E and F are. 87.5’» 92.9, 83,2 and 90.4 respectively and the moan value is 89.7 ± 2.1 # The value of true digestibility was also determined using the Detergent method. Those values are given in Table 4.6. It is possible to determine true digestibility since the proportion of faecal nitrogen derived from non-dietary origin is known. For instance, for sheep Ho. 186, tlis apparent digestibility of nitrogen UNIVERSITY OF IBADAN LIBRARY 133 True digestibility, “biolo-ricsl value -and not protoin utilisation values for the Nest African dwarf wo the r shoo? maintained on basal hay and concentrate supplements. — TOTAL METABOLIC . FAECAL | FAECAL NITROGEN APPARENT TRUE BIOLOGICAL I LET PROTEIN ! Periods RATI01 NITROGEN (g/kg DM intake) # MFN DIGESTIBILITY DIGESTIBILITY VALUE I UTILIZATION (g/kg D'K intake) MFN O T #TFN $ (t d x b v ) ■ NPTT 1 C 4.75 + 0.68 3.13 65.89 63.3 87.5 96.3 84.3 --- - “| .............! ■ * r 2 D 4.07 + 1.02 3.13 76.90 69.4 92.9 96.0 89.2 ■ 3 E 5.27 + 0.39 3.13 59-29 71.0 88.2 98.9 87.2 j ■--- 4 F 5.25 + 0.50 3.13 59.62 76.3 90.4 92.6 83.7 ' „ ■ i ( . jiM - IUIIL..* ML „ ' - - ! . — . _________ _____^MEAN 4.83 + 0.48 . * 65.42 ± 7.12 89.7 +2.1 95.9 + 2.2 86.9 + 2.2 | . . _ . .............. - ........... . . .............- _ ............................ ........................• _ _________ - . ______ - - - _ “ _ ____i A 5.51 + 0.14 3.13 56.80 55,9 80.9 85.7 69.3 j I » B 3.06 + 0.35 1.50 | 49.0 61.2 80.2 100.0 80.2 f ... . . “ .j MEAN 4.28 + 0.72 52.9 + 3.9 l 80.5 + 0.3 92.8 + 7.1 74.7 + 5.4 f --- ------------------------------------------------------I— _ — . . . . « , ■ • - — _ - UNIVERSITY OF IBADAN LIBRARY 134 of ration C is 64.1 %» This means that of ovory 100 g of nitrogen that is consumed, 64.1 g arc absorbed and 35.9 g are lost in the faeces. Since the non-dietary faecal nitrogen constitute 77.4$ of faecal nitrogen, (Table 4.6), it means that 0.774.X 35.9 g of the faecal nitrogen arc of non-dietary origin i.e. 27.8 g, The time digestibility of ration C is therefore 64.1 + 29.8 or 91.9 $ From these calculations, it would be soon that the time digestibility of rations A to F ranged from 89,3 a/° for ration A to 96.0 % for ration F, with the mean value of 92,9 +2,3 .The values of 89.6 io and 86.2^ from regression equations and 89.7 ± 2.1 and 92.9 + 2.3 obtained from non-dietary faecal nitrogen methods, would appear to be in very good agreement. The true digestibility of nitrogen of the rations nay therefore be estimatedas being between 86 and 93$. 4.3.5 Absorbed N Nitrogen digested per metabolic size, was not affected by supplementation of hay with concentrate , the mean values being 0.35 ± O.Oij and 0,30 + 0,03 for rations A and B respectively. In Trial 2, differences between means were very highly significant (P^O.OO^ for treatments, the values for nitrogen digested ranging from 0.54 + 0,03 on ration C to 1.20 + 0.02 |n ration F. Nitrogen digested per metabolic size increased with increasing levels of crude protein in the ration. It also increases with the nitrogen intake according to UNIVERSITY OF IBADAN LIBRARY the regression equation given: Y = (0 .0 6 3 + 0.045) X - 0.163, (4 .12) (r = 0.99) where, Y is the nitrogen digested per metabolic size per day, -and X is the nitrogen intake, also per metabolic size per day. The mean differences were not significant (P> 0.05) within experimental animals. It is of interest to note that nitrogen digested in rations A end B were 0.35 and 0.30 respectively even though the concentrate in ration B contained very little or no digestible nitrogen. 4.3.6 jfctained N; Nitrogen retained per 0.734r was the sane for the two rations, with the mean values of 0.29 + 0 .0 5 and 0.28 + 0.02 for rations A and B respectively. In Trial 2, the mean differences were significant (P< O.Ol) in nitrogen retained, ranging from 0.48 + 0.04 for ration G to 1.11 + 0.09 for ration F. Nitrogen retained increased as nitrogen- digested, and the following equation describes the relationship: Y = (0.939 + 0.029) X - 0.014 .................. <.(4.13) (r . 0 .9 9) where, Y is nitrogen retained per metabolic size and X is the N digested per metabolic size. The coefficient of X is 0.939 and this gives an index of biological value, and shows that the .animals were definitely utilizing the digested N efficiently. There were also UNIVERSITY OF IBADAN LIBRARY R a t i o n s FIG.4-1 THE RELATIONSHIP BETWEEN ABSORBED N AND RETAINED N (o— o) OF THE SHEEP MAINTAINED ON BASAL HAY AND CONCENTRATE SUPPLEMENTS UNIVERSITY OF IBADAN LIBRARY 137 , 0.734 very high correlations between N retained, Y (por ) and N intake, X (, per Wj°j--g.7 ^ K) wwh>i*ehrh. the r value being 0.91 in Trial 1 , and 0.98, 0.98, 0.97 and 0.99 for periods 1, 2, 3 and 4 respective! of Trial 2. Mien the results of the four periods in Trial 2 wore pooled, r value of 0.98 was obtained. Thus the values of retained II increased with increasing dietary It intake, and the relationship is represented by the following regression equations: Por Trial 1; Y = - 0.32 + (0.91 + 0.006) X, ...............(4.14 ) (r = 0.91) Por Trial 2; Period 1, Y = -0.20 + (0 .8 3 ± 0,03) X ............C4.15 ) Period 2, Y = -0.23 + (0.82 + 0.02) X ............ (4.16) Period 3, 7 = -0.07 + (0.75 + 0.02) X ............. 4.17 Period 4 , Y = -0.07 + (0.73 + 0.01) X ............. (4.18) pooled value, Y = -0.13 + (0.77 + 0.01) X ............^ 4-19) Fron these equations, the N intake at zero - K - balance were, for Trial 1, 0.35 , 0.734S / \ g , and 0.24, 0.2/., 0.093 and 0.096 respectively for periods 1, 2, 3 end 4 of Trial 2. 4.3.7 retention (/£) The percent retention of N of ration B was significantly higher than that of ration A (lA 0.05) with the mean value of 51.7 + 1.7 for ration B and 42.3 + 2.3 $ for ration A. In Trial 2, UNIVERSITY OF IBADAN LIBRARY FIG. A. 2 THE REGRESSION OF NITROGEN BALANCE ON NITROGEN INTAKE FOR THE SHEEP MAINTAINED ON HAY AND CONCENTRATE SUPPLEMENTS Nitrogen Balance(g/day/W|< UNIVERSITY OF IBADAN LIBRARY 139 the noon differences wore significant (P< 0.05) for treatments C to F in the N retention values, ranging from 57.5 +1,6 for ration C to 69.5 + 1.6 fo for ration F. The N retention followed the same pattern as the digestibility values. Nitrogen retention increased with increasing intake of N and also with increasing levels of dietary crude protein. The mean differences were not significant within animals (Ps 0.05). It is soon by the closeness of the values of N digestibility and N retention that almost all the II digested was retained, with very little loss of N in urine. Two factors may be responsible for this, one is_ the inclusion of cassava flour, a readily fermentable carbohydrate, and the second is th. fact that the animals were young and efficiently utilizing dietary IT. 4.3.8. The Metabolic faecal nitrogen Metabolic faecal nitrogen (MFN) which is the faecal N at zero N intake was obtained by two methods, the regression method and the detergent method (Mason, 1969). The regression of faecal II (y) ifi g /k g DM conoumod on II intake T = 3.13,+ (0.16 ± 0.02) X (4.20 ) (r « 0.62, P CO.01) Faecal II at zero II intake is given by this equation as 3.13g/kg DI' consumed, UNIVERSITY OF IBADAN LIBRARY FIG. 4-3 THE REGRESSION OF FAECAL NITROGEN ON NITROGEN INTAKE FOR THE SH EEP MAINTAINED ON HAY AND CONCENTRATE SUPPLEMENTS Faecal Nitrogen (g/kg/DM intake) UNIVERSITY OF IBADAN LIBRARY 141 TJhen the faecal N (y), expressed in g/kg DM consumed was regressed on percent crude protein in the ration (x), the following equation was obtained: Y = 3 .6 8 + (0.14 + 0.04) X ..................... (4 .21) (r = 0.75, P<0.05). which gives the value of M M as 3.68 g fkg DM consumed. Regression of digestible II per kg DM intal.ce (y ) on II intake per kg DM intake (x) gave equation 4.10 which gives the value of the M M as 3.45 g/feg PM consumed. Similarly, regression of digestible crude protein (y) on the percent crude protein of the ration (x) gave equation 4,11. The value of M M from the equation is 1.86 g crude protein per 100 g DM consumed or 2.98g I-IM/kg DM consumed. The values of M M obtained from regression equations are 3.15, 3.68, 3.45 and 2,98 g M M per kg DM consumed’, {jiving a mean value of 3.31 i 0.16g HM;kg DM consumed. It is possible to estim.nte the value of M M by the detergent method (Mason, 1969), as stated in Table 4.6. A knowledge of the faecal W per day, DM intake per day and the percent of faecal N that is non-dietary could allow the estimation of M M . For example, for ration C (Table 4.6), the faecal IT per day was 2640 mg and 77.4$ of it was of non-dietary origin that is 2080 mg is the M M . The DM y intake was 608.1 g per day. Therefore M M is 2080 mg per 608.1 g DM consumed or 340.20 ng/l00g DM consumed. The mean value of M M from UNIVERSITY OF IBADAN LIBRARY W EST AFRICAN DWARF WETHER SHEEP Faecal Nitrogen (g/kg/DM Intake ) UNIVERSITY OF IBADAN LIBRARY 143 Table 4.6 was 311.94 + 28,6 ng/lOOg DM consumed or 3.12 g/kg DM consumed. If the lowest two values (158.5 and 215.4) arc excluded fron the calculation of the noon, the Mill is 3.75 g /k g DM consumed. Tho value of the MFN obtained by the detergent method i3 in very good a.greenent with those obtained fron the regression equations and this indicates that the values of the IIM actually lie between 3*0 and 3.7 g/kg DM consumed for Most African dwarf wother sheep weighing 17 to 27 kg and maintained on hay/conqontrate rations. The porcontage of undigested nitrogen, microbial nitrogen, water-soluble nitrogen and non-dietary nitrogen in sheep faeces are also given in Table 4.6 About 1 g of faeces from each animal was .analyzed for the components of tho faeces. Table 4.6 shows that total nitrogen per unit weight of faeces is influenced by the content of nitrogen of the feeds, for while 1 g of faeces fron animals on ration A contained about 12.3 mg of nitrogen the corresponding amount of nitrogen in 1 g of faeces from animals on ration P was 22.4 mg (Table 4.6). This trend of increasing faecal nitrogen per g of faeces occurred from ration A to ration P except for ration D. Tho value of undigested dietary nitrogen recovered in the faeces ranged from 17.6 °/o for ration P to 24.3 % Tor ration A, the moan being 21.1 + 3.8. The value >f undigested dietary nitrogen (UDll) for ration P, 17.6 is lower than 24.3 obtained for tho UNIVERSITY OF IBADAN LIBRARY - 144 TABLE 4.6 The Porccntago of tile U n d ie s tod Hlcro 'b ia l N. Hater Soluble IT and Non-dietary Faecal IT in the Faeces o f tlic ~re s t Afr i c an dwarf wether. sloop n ain taiaed , on hay and concen trates — — 1 > SHEEP TOTAL [ UNDIGESTED MICROBIAL NON- HEN APPARENT TRUE FAECAL DRY-MATTERl METABOLIC M F N NOS. j RATIONS FAECAL } DIETARY & HATER - DIETARY NDFN DIGESTI­, SOLUBLE DIGESTI­ mg/ day INTAKE f a e c a l mg/lOOg ENDOGENOUS FAECAL 1 BILITY BILITY g/day N N N ng/day DM INTAKEN (MEN) j N (n d f n) $ I % j ] H 136 C 100$ 22.6 61.8 15.6 77.4 0.80 64.1 91.9 2640 608.1 2080 340.20 268 (15. Sag) (3.6) (9.6) (12.2) i I r 173 100$ 18.9 64.5 16.6 81.1 i 263 D (14.3) (2.8) (9.2) (11.6) 0.80 70.1 94.3 1690 423.1 1360 321.22 j I— _ _ 184 r E 100$ 19.9 63.9 16.2 ! 80.1 0.79 72.8 94.6 2655.8 506.0 2100 416.29 301 (17.1) (3.4) (11.o) (2.8) (13.3) y.mm. mtmmmmt 179 100$ 17.6 62.1 15.3 82.4 0.82 77.0 96.0 1190 629.3 980 158.45 * 259 (22.4) (3.9) (15.0) (3.4) (18.5)1 T 173 A 100$ 24.3 54.-4 21 ;4 75;7 ; 0.72 55.7 89.3 2460 443.4 1865 420.09 259 (12.3) (3.0) (6.7) (2.6) (9■.3) I 259 B 100$ 23.6 57.9 18.5 76.4 [ 0.76 63.6 91.4 1495 529.6 1140 215.41 473 (14.0) (3.3) (8.0) ( 2 ,6 ) (10.7)| MEAN N r 100 > 21.1 61.6 17.3 78.9 | 0.78 92.9 311.94if 1----- ....... . . J[ i =6.3 j =2.5 ! =0.05 \ 2 S i^j. ji 28.6 __ — {____________ UNIVERSITY OF IBADAN LIBRARY # 145 basal hay ration, 'this closeness in value suggests that the much lower digestibility of nitrogen of ration A than ration F is not so much duo to its indigestibility but to greater MFR in ration A than in ration F. In the estimation of UDN, there is the tendency of increasing percentage of UDN with decreasing level of crude protein in the ration and the relationship is illustrated as follows: Rations: A B C D E • F UDN 24.3 23.6 22.6 18.9 19.9 17.6 From this illustration, it may ho inferred that not onljr the apparent digestibility of crude protein but also the true digestibility increased with increasing levels of dietary crude protein. The percentage faecal nitrogen derived from microbial and endogenous nitrogen (MSN) ranged from 54.4 for ration A to 64.5 7° for ration D, with the mean value of 61.6 + 6 . 3 The value of MEN seemed to increase with increasing levels >.of dietary crude protein up to ration D. hater soluble nitrogen (WSU) accounted for 17.3 + 3.4 $ of faecal nitrogen, the value being highest for animals on ration A, 21.4 i° and lowest for animals on ration F, 15.3 The trend observed here is that of decreasing WSN with increasing levels of dietary nitrogen. The FSN nay be aminos, amides, some amino acids, ammonia, but mainly digestive juices. Non-dietary faecal nitrogen (iTDFN) is obtained by adding the values of microbial and endogenous nitrogen (lIEU) and water-soluble UNIVERSITY OF IBADAN LIBRARY 146 nitrogen (wsif), This is the faecal nitrogen of non-dietary origin. It is apparent that the terns Non-dietary faecal nitrogen (NDFN) and Metabolic faecal nitrogen (HI®) are synonymous. The former term (NDFN) i3 preferred since there is a tendency to regard MFN as a simple entity representing nitrogen of endogenous.origin. The values of NDH'I obtained ranged from 75.7 % for ration A to 82.4 % for ration F. The trend observed here is that of increasing concentration of NDFN with increasing dietary crude protein levels. The trend followed the sane pattern as that of MEN: The noan value of NDFN was 78.9 + 2.5 % The percentage of NDFN that is of MEN origin ranged fron 72./$ for ration A to 82.4 f° for ration F, with the neon value of 78.+ 5 % The trend observed hero is the increase in the percentage NDFN derived fron MEN fron ration A to ration F. Fron the results obtained, it nay be concluded that UDN, MSN and MSN accounted for 21. ] + 3 .8 %, 61.6 + 6 .3 f° and 17.3 + 3.4 % of tine faeces of sheep maintained on basal hay and concentrate rations. 4.3.9 Endogenous urinary nitrogen (SUN): Endogenous urinary nitrogen (eUN) is nitrogen excreted in urine at zero N intake. ‘The value of SUN was obtained b3r regression o f urinary IT (g/day/W^T^^ j on intake (g/day), X. The relation­ ship is given by the following equation: UNIVERSITY OF IBADAN LIBRARY 147 Y = (0.0096 ± 0.0011 ) X - 0.0238 ..................(4.22) (r = 0.79 , P^0.05) Prom this equation, the urinary N at zero N intake is the BUN giving a value of 0.0238 g/,d ay/Ŵ, 0g.,734 4.3.10 The Biological value of tlio rat ions.: The true digestibility (TD), Biological value (BV) and Net protein utilization (NPU) values for the rations are presented in Table 4.5. The BV was determined using the KPN value of 3.13 g/kg UK consumed, and 0.0238 g//d ay/aN k0.g7 34 as EUN. The equation used is: NI - (IN - MITl) - (UN - EON) = BV NT - (IN - MIN) (4.23 ) where, NI is the N intake, IN is the faecal N, UN is the urinary N, EUN is endogenous urinary N, and BV is the biological value of the ration. The BV for the basal hay ration was 85.7, while the BV for the protein-based rations ranged from 92.6 fo for ration F to 98.9 a for ration E with the mean BV of 95.9 + 2.2 % . Maximum 3V,(100.Oh0 was obtained with ration B, and minimum P/ (85.750 was obtained for the basal hay. The BV were also high for rations C (96.3 °/°) and D (96.0 t f ) , those containing 6.5 and 8.5 i° crude protein respectively. From the BV determinations, it is soon that the protein of hay and mixed rations are very well utilized by the growing sheep. UNIVERSITY OF IBADAN LIBRARY 148 It also shows that maximum BY was obtained with ration B“which supplied the least amount of cru.de protein \, .0.49g, /day/0%.g7 34). The BY of the basal hay (85.7 $ is almost as high as those of the mixed rations. 4.3.11 The coefficient of net utilization of dietary proteins (HfU) The coefficient of net utilization of dietary protein is the product of true digestibility (t d) and the biological value (BV) and is also known as the net protein utilization (iTFU). From Table 4.5 the coefficient of net utilization was calculated ass TD X BY 100 (4,23b) The value for ration B was 80.2 and the value for basal hay was 6 9 ,3 , but the values ranged from 83.7 to 89.2 for the protein-based rations. The ration with the highest level of crude protein (ration F, 14 crude protein) had the lowest vilue for the coefficient of net utilization of the protein-based rations. 'Hie mean value for protein-based rations was 86.1 +2.2 $. 4.3.12 The protein requirement for the maintenance of West African dwarf wether sheep. The digestible crude protein (DCP) required for maintenance is the amount of digestible crude protein intake required to keep the animal in zero IT - balance or IT equilibrium. UNIVERSITY OF IBADAN LIBRARY 149 The regression equations obtained when IT - balance, g/, dayA^0.g7 34*f was regressed on IT intake, g/day/w^73‘r f aro given in Table 4.7. The N intake at zero N-balance in Trial 1, with rations , ,0.734' A and B was 0.35 g/dayA^g , the crude portein intake at zero N balance was 2.20,g/dayA^0 .734• ® 1G ^oan digestibility of rations A and B was 58.5 f°, and the DCP required for maintenance was estimated as 1.28 g // d c/ a°*y75y4o r 1.73 g/dayAcg l iv e weight. In Trial 2, IT intake at zero N balance were 0.24, 0.28, 0.093 . 0.734 and 0.096 g/dayj.^^ for period 1 to 4, which are equivalent to 0.734 1.51, 1.75, 0.58 and 0.59 g crude protein/day/W^g * Since the mean digestibility coefficient of ration C, D, 3 and F was 70 f°, the DCP required for maintenance were estimated as 1.06, 1.23, 0.41 and 0.41 in periods 1, 2, 3 nod 4 respectively of Trial 2. Then these are expressed per kg live weight, the- valuco are 1.44, 1.70, 0.56 and 0.56 g/dajfcg live weight for periods 1, 2, 3 and 4 respectively. There was a gharp decrease (P< 0.05) in the value of the DCP required for maintenance in the 3rd and 4th period of Trial 2, therefore the moan value for periods 1 and 2, and also of periods 3 and 4 were used. The mean estimate of DCP required for maintenance in periods 1 and 2 of Trial 2 is 1.15 + 0.08 g/day/W^g or 1.57 + 0.13 g/dnyAg live"weight, while the mean value for periods 3 and 4 is 0.41 g// day/A°^*g7 34 or 0.56 g/day/kg live weight. The regression equation obtained when the four periods of Trial 2 were pooled gave an estimate of the mean DCP requirement for UNIVERSITY OF IBADAN LIBRARY 150 TABLE 4.7 Digoatiblo crude protoin rqquiroporit of tho West African dwarf shoop naintained on basal hay with concentrate supplononts TRIAL REGRESSION NITROGEN INTAKE AT Z0E R7O8/ j CRUDE PROTEIN INTAKE AT' DIGESTIBLE CRUDE PROTEIN DIGESTIBLE CRUDE PROTEIN Jj N - BALANCE (g/day/li^ ) j ZERO N BALANCE (g/daywg REQUIREMENT FOR KAINTE- REQUIREMENT FOR MAINTENANCE I HSSCE (e/irW * ______ ..... . ______ _ ___ (g/day /k g Livo Weight)! ri t 1 Y = -0.32 + 0.91 X 0.35 2.20 [ 1.28 1.73 ! - - - - - 2 (l) Y = -0.20 + 0.83X i 0.24- 1.51 1.06 1.44- f " "-Tl 1 r ............ .. _ * T"~ (ii) Y = -0.23 + 0.82X 0.28 1.75 1 .23 1.70 MEAN 0.26 1.63 — 1.- 15 . _ ______ (iii) Y = -0.07 + 0.75X 0.095 0.58 0.56 (iv) Y = -0.07 + 0.73X ] 0.096 0.59 0.41 0.56 MEAN | 0.095 0.41 0.56 I-IV Y = -0.13 + 0.77X j 0.170 J 0.59 1.06 0.74 1.01 f . . . . . . . _______ UNIVERSITY OF IBADAN LIBRARY V«J1 | 1---- 151 maintenance / , ° • 7 54is 0.74 g/day/Wkg or 1.01 g/day/lrg live weight. Prom the results, it is obvious tint the DC? requirement for maintenance was high in Trial 1 and in the first two periods of Trial 2 but sharply declined during the 5rd and 4th periods of Trial 2. .The stage of growth of the sheep will therefore indicate the DCP requirement for maintenance. A value of 0,74 g/day/W^h^ "r may be taken as the mean DCP requirement for maintenance over the experimental period, Tlic factorial method of the Agricultural Research council (1965) was also used to estimate the digestible crude protein (DCP) requirement for maintenance and growth (Table 4*8 )• The equations used were: DCP requirement for maintenance and growth (g/day) 100 * 6.25 (UE + C) HP ( 100BV~ BV" - 1) .(.4.24 ) where UE = Urinary endogenous loss (g/day) G = Retention of nitrogen in live weight gain, estimated as 2.5 i° of gain for sheep. BV =s Biological value of protein i MF = Metabolic faecal nitrogen estimated as lcD g/day, Where D is dry natter intake in kg/day and k is the metabolic faecal nitrogen, g ll/kg Dry natter intake. Tho estimates of the DCP requirement for maintenance and growth arc summarized in Table 4*8. The values of urinary rcndogaaouo# UNIVERSITY OF IBADAN LIBRARY The. ostiaate. _of the; digestible crude nrotein requironant for maintenance and -growth, of. the West African dwarf wether sheep maintained on ha:/- and concentrate supplpaent.3 by. the f notorial method. \ • Initial Pinal > Moan ! live I* Mean jI Moan Endogenouofl Metabolic j i Height of weight of| weight Mean Digestible weight jweight of|Dry natter jDigestible Digestible Digestible TRIALS i of Urinary N j paGcal Biologica] crudeanimals animals I anima ls change! animals ' intake | crude crude , crude (kg) (g/doy) | (s/day) value of protein ! protein (kg) I (kg) j ( i j < ^ > rations protein prortein require­ j requiremen t | requirement ment for requirement !i ! J i*i «t maintenancej? mSl+,iJUGoHralxlCG ^ formaintenance for growth i i (g/day) ; A/toy/w°; 3" (g/day) 1 « i and growth __ . _ _ |1 __ . 'r? | _ . _ _ ik_____________ (g/day ) “ 1 19.5+1.3 ! 19.6+1.4 19.5 2.5 ;! 8.9 1 0.48 0.21 1.50 6| 92.8* 2.17 0.24 2.57 0.40 ■ i “ !— . ... iI-.-.-.-.* . ' . ■J _____ _ ___________ 2 19.6—+1.4 20.5+1.3 20.0 11.3 9.0 0,59 C.21 1.84 I 95.9 1.33 0.20 3.58 1.75" .... 1 — m e a n 11 0.53+0.05 .0.21 1.67+0,17 94.3+1.5 2.00+0.17 0.22+0,02 3.07+0.50 1.08+0.6: --- — + --------1 U—• —* — 1— - * _____ _ , __ ; UNIVERSITY OF IBADAN LIBRARY 153 loss and metabolic faecal nitrogen used were 0.0238g/d, ayUj0C.g7 34 0.05), and this agrees with the reports of Elliott and Topps (1963) and of Crabtree and hiIlians (1971) that total DM and digestible energy intake increased when hay was supplononted with energy - rich concentrates. Dry natter intake in Trial 2 increased with increasing levels of dietary crude protein even though this was not significant (P>0.05). Elliott and Topps (1963) showed that voluntary intake of low protein feeds by sheep is closely related to the nitrogen content of the feed. In the present experiment, there were slight increases in DM intake with increasing DM digestibility of rations D, E and F, This relationship was not significant but agrees with the reports of Elliott and Topps (1963) who obtained a low correlation (r = 0.273> P,>0.05) between DM intake and digestibility. The value of 72.6 + / 1 0.' /13.3 g / d a .y / \ g obtained in this report as intake for ration F is close to 74.5 g/d, ay/tf. |0£.g7 34 obtained for a similar ration for sheep by Elliott and Topps (1963)- The supplementation of hay with concentrate (ration b) mainly cassava flour has resulted in the decrease in hay DM intake. When the concentrates containing crude protein were- fed in Trial 2, the intake of hay DM fell progressively as concentrate DM intake increased. This seems to contradict tlx; findings of HI ax ter and Waimnan (1963) that apatite of sheep for hays of low protein content is increaded by supplementation with concentrate feeds. Crabtree and UNIVERSITY OF IBADAN LIBRARY 156 Williams_(l971) found with hay of a higher digestibility of energy (60.7 f°) t that concentrate feeding reduced hay intake. In the present report, when the basal hay was supplemented with concentrates, 100g DM of , C2, C^, Cy and C 5 replaced 42.0, 66.8 , 55.0, 54.6 and 49«0 g respectively of hay DM., the mean value being 55.1 + 8.0. Blaster, Wainman ond Wilson (1961) showed that when concentrates wore added to high quality fodder, the HI consumption of fodder fell by slightly .loss than the* amount of DM consumed as concentrates, for instance, that 100g DM of concentrate replaced 79 g of fodder. They showed, however, that when concentrates were added to the ration of poor quality fodder, 100 g of concentrate DM replaced 47 g of poor quality hay. The value of 55.1 g obtained in the present investigation is slightly higher than 47 g obtained by Blaster ot ■.-.1. (1961). The intake of concentrate DM increased with increasing levels of crude protein in the concentrate. Similarly intake of concentrate as percentage total DM intake also increased with increasing crude protein content of the concentrate. Tile amount of DM intake by the sheep in the present investigation was estimated as 3$ of body weight or about 6 P> of the metabolic weight, a value used in several fedding experiments (Bergen et al,^ 1968). When equation 4.2, C = aWb UNIVERSITY OF IBADAN LIBRARY 157 is used to estinate DM intake, ’b* was obtained as 0.668 in the present report. This shows that DM consumption of these rations is related to the live weight of the sheep raised to the power of 0.668. Blaxter et al. (1961) showed that DM consumption is influenced by tile nature of the ration; consequently, the value of the exponent *b* will also be influenced by the ration. Blaxter et al. (1961) obtained 0.70 as the value of 1V for rations of hay. The present report with the value of 0.668 + 0.026 is not different from 0.66, the exponent which relates body weight to body surface area, and slightly lower than 0.734, also the exponent relating basal energy metabolism to body weight (Brody, 1945), The indication in the present report of a value of 'b! similar to 0.67 which confirms the reports of other investigators nay perhaps be attributed to the necessity for the sheep under tropical conditions to maintain homoeothorny by heat loss through the body surfo.ce and hence to a closer relationship between metabolism (in this case as indicated by intake) and body surface, than between intake and body weight (Butterworth, 1966). The West African dwarf wether sheep used consumed slightly less nitrogen in ration B (basal hay + Supplement ) than in ration A (basal hay); concentrate contained little protein (1.57/&) but the concentrate served mainly as a source of energy. Since intake of this concentrate did net improve the intake of hay, the UNIVERSITY OF IBADAN LIBRARY 158 pajor ■ source of dietary nitrogen, it is expected that reduction in hay intake with supplementation with concentrate C-| will lead to sone reduction in total intake of nitrogen. In Trial 2, nitrogen' intake increased with increasing level of c rude protein in the concentrate, which is in agreorient with the findings of Robinson and Forbes (1970) who used weaned lambs. From this, it is seen that lanbs are likely to take low mount of nitrogen in a ration of low crude protein content, which nay not be sufficient to maintain then in nitrogen equilibria^ In the present investigation, all the animals were in positive nitrogen balance indicating that nitrogen intake was adequate at least for maintenance since the animals did not,put on nuch weight. The fact that supplomentati/o■ i of hay (7.7 crude protein) with energy did not significantly reduce total nitrogen intake offers an advantage in animal feeding. In places where the crude protein level in herbage is just adequate (about 8 fo), supplementation of herbage with energy while removing the limitation to animal growth due to energy, would not introduce shortage of crude protein. It is likely that the animals would consume sufficient energy for their requirement hut not too much as to significantly reduce their intake of harbage, the major source of crude protein. This situation may not, however, apply if the crude protein content of herbage is very low (about 4/̂ ). In this case, nitrogen intake may be so low as to make nitrogen limiting to the growth of the anirils. UNIVERSITY OF IBADAN LIBRARY 159 Robinson and Forbes (1970) showed with lambs fed on so y a boon meal that the nitrogen intake per metabolic size were 0.53* 1.03, 1.60 and 1.81 g when the rations contained 7.3% , 13.3$, 19.8$ and 23.1 $ crude protein respectively which, in the present studies, the values of 0.84, 0.91, 1.28 and 1.55 g per Ityg*^rwere obtained. Hie HI digestibility of ration B was significantly (P<.0.0l) higher than that of ration A. This is in agreement with the results of Crabtree and Williams (1971) who showed higher DM digestibility of nixed rations than for hay alone. Hie DM digestibility of rations B to F were high, no doubt, due to the presence of concentrate supplements. The supplementation cf hay of low DM digestibilitjr with a concentrate of high DM digestibility is expected to lead to higher DM digestibility of the nixed ration. There was no marked effect of level of dietary ©ru.de protein on DM digestibility oven though slight CM digestibility increases wore ob&orvod with rations D, E and F. Robinson and Forbes (1970) had observed-linear increase in DM digestibility with increasing crude protein'intake. ' • In the present report, there was a. slight increase in Dli digestibility with increasing DM i: take especially for rations D, E and F. The present report that DM digestibility of a ration increases with increasing proportion of concentrate in the ration, is in agreement with the findings of Crabtree and Williams (1971). The lower digestibility of DM in period 1 than periods 2 to UNIVERSITY OF IBADAN LIBRARY 160 4 of Crial 2 reflects the higher intake of highly digestible concentrate fraction during periods 2 to 4 than in period 1 (.Robinson and Forbes (1970). Similarly, supplementation of the basal hay (7.7$ crude protein) with the concentrate (C-j) did not significantly affect its digestibility. Campbell, Sherrod and Ishizaki (1969) found that supplementation of Kikuyu grass (Pennisoturn Clandestinum). containing 5.5$ credo protein, with energy decreased its digestibi­ lity of nitrogen, and they concluded that the depression of apparent crude protein digestibility resulted from increased metabolic faecal nitrogen. Pick, itanernan ? Gowan, hoggins and Cornell (1975) reported improved digestibility of nitrogen of poor quality grass (3.28$ crude protein) with supplementation with energy in the form of corn meal, sucrose and starch, when energy-rich concentrates formed about 25$ of the rations hut that at higher levels of supplementation, energy did not affect digestibility of nitrogen contained in hay. Tile findings that supplementation of hay with energy-rich concentrates lead to increased metabolic faecal nitrogen would seen to depend on the quantities of the concentrates added to the basal forage and the quality of the forage. In ration B, cassava flour comprised about 33$ of the ration and it is not likely that at that level of supplementation metabolic faecal nitrogen had assumed any prominence. In fact, the slight increase in digestibility UNIVERSITY OF IBADAN LIBRARY 161 coefficient of hay nitrogen with energy supplementation in the present report nay he due to rapid multiplication of ruminal micr• o—organisms and hence slightly letter digestibility o»f the hay. It is however likely that with very poor quality hay supplenentation with energy would lead to increased netabolic faecal nitrogen mainly of microbial origin. In Trial 2, the digestibility of dietary nitrogen increased with increasing levels of dietary crude protein, and also with crude protein intake. This is in agreement with the reports of Robinson and Forbes (1970) and Andrews and 0rskov (1970a) who also reported increased digestibility of dietary crude protein with increasing levels of crude protein in the rations, and also with increasing intake of crude protein. This indicates decreasing quantitative importance of the netabolic faecal nitrogen with increasing crude protein content of the ration. The regression equation relating digestibility coefficient of nitrogen and percent crude protein in the rations (Eq. 4.7) would give higher values for corresponding dietary crude protein, than that of French, G lover and Duthie (1957) shown as follow: Y = 73.7 log X - 19.8 ..... . .......(4,25) Y = 70.3 lo g X - 14.9 ......... . . . . ( 4 , 2 6 ) for nixed food and herbage plus nixed feed respectively. Because of the costs and labour of conducting classical digestibility experi­ ments, attempts have been made to derive correlations between UNIVERSITY OF IBADAN LIBRARY 162 digestible nutrients available to the animal and the crude dietary components from which they are derived, Such regression equation as given in the present studies has several practical applications, one being the determination of the average digestibility of crude protein of rapidly growing grasses and herbages at different stages of development, another being the evaluation of average crude protein digestibility of single or compound feeds especially where adequate facilities are not available. It can also be used for computing maintenance find production rations. The digestibility coefficient of nitrogen is not dependent only on the crude protein content of concentrate but also on the percentage of concentrate in the ration. It has already been shown in Equation 4.9 that total digestibility of a ration increases with increase in percentage crude protein of supplemental concentrate but tends to decrease as the proportion of concentrate increases. The tendency for digestibility of nitrogen to decrease as percentage of concentrate increases is due to increased metabolic faecal losses that accompany supplementation with concentrate especially at low nitrogen intake. The lower digestibility of nitrogen in period 1 than in periods 2, 3 end 4 may be due to lower intake of the highly digestible concentrate fraction of the ration in period 1, compared with periods 2 to 4. Hie values of true digestibility obtained by regression and detergent methods are in very good agreement showing that the true UNIVERSITY OF IBADAN LIBRARY 163 digestibility of dietary nitrogen is between 86 and 92$. The value obtained from regression equation wore between 86' and 89$ and those from the detergent method were from 89 to 96%. The higher value obtained from the detergent method is as expected in view of the possibility of extracting fron tlx.- faeces other nitrogenous materials which are neither microbial nor endogenous, but dietary, and this would tend to increase the non-dietary faecal nitrogen and hence over-estimate the true digestibility. This may be particularly so in the case of water-soluble nitrogen. Some of the water-soluble nitrogen might in fact be of dietary origin but care assumed to be included in the non-dietary faocal fraction. Mason (1969) hao shown that the assumption that the process of digestion in the animal does not affect the extractability of the undigested dietary nitrogen in the faeces samples nay not bo strictl;? true. He used the detergent method and obtained the true digesti­ bility values of 91 - 92$ for ryegrass hay, end 98 - 99$ for soya bean meal-based ration and reported that the values were probably higher than the true values because some undigested dietary pigments were extracted by the procedures employed. The true digestibility value of 92.9 + 2.3 obtained in the present experiment is vers?- good agreement with Mason’s (1969) values of 91 to 92$ and Singh and Mahadevan’s (1970) value of 93.4 ± 1.9$ The amount of N absorbed by thm sheep did not differ with rations A and B, but increased with increasing intake of dietary UNIVERSITY OF IBADAN LIBRARY 164 crude protein. This is expected since the digestibility of the rations increased with increasing intake of nitrogen. The increase in ST absorbed with increasing N intake agrees with the reports of Stobo and Hoy (1973)• Since the value of the metabolic faecal N (HEll) is relatively constant por unit DM intake, it is expected that the preportion of N absorbed will increase with increasing dietary crude protein intake, . 0.734. T!ie N retained per metabolic size ) increased linearly with increasing IT intake and also with absorbed IT. About 93.9$ of absorbed N was retained by the sheep. This is to be expected since the sheep were young and laying down tissues by utilizing absorbed IT. The slope of the regression equation of retained IT with absorbed IT is 0.939 + 0.029, and has been termed 'the Nitrogen balance index of absorbed IT' (Allison, 1965) as it demonstrates the rate at which absorbed IT fills the protein stores of the animal body; consequently, this index is a function of the biological value of the dietary IT. The high N-bnlcnce index for the N content of the rations C, D, S and F indicates the very high efficiency wit:: which these animals utilized the protein of the concentrate-based rations. The IT balance index value of 0.959 was higher than the valuo obtained for casein (0 .65), and for urea (0 .83) but lower than the value of 1.05 obtained for gluten Tiy Deif, El-Shazly and Abeu Akkada (1968). UNIVERSITY OF IBADAN LIBRARY 165 Hie value of 0.77 obtained as the N balance index of intake II in the present studies was higher than 0.69 obtained for casein but less than 1.02 end 0.97 obtained for gluten and urea respectively by Deif, 31—Shazly and Abou Akkada (1968), again tending to indicate very efficient utilization of the IT contents of the rations. It is obvious fron Table 4.2 that at the hi^iest level of IT.intake, N retained was still increasing and the value of 1.11 +_ . , 0.734 0.09 g/day/I,JjCg could not bo the maximum IT retainable by the sheep. Black, Pearce and Tribe (1973) obtained the maximum value of IT retained per % g0. 734 for lambs weighing 20,8 kg 1.05 g/,d ay/¥k0.g7 34'. It is likely that, given rations higher in crude protein, the value of 1.11 g/dayAn0 * 734-+ obtained in tho present investigation night bo. . . , 0.734 excoodod. Black ot al.(l973) also obtained the value of 1.46 g/day/I'Jlcg Since the weight range of the sheep used in the present report was 1 5 - 2 6 kg, the maxinun II-retained is more likely to ho closer to 1.05 than to 1.46 g/day/W0^.g7 34 . Therefore the value of 1.11 0,734g / d a y / \ g obtained in the present experiment would appear to have reached the maximum IT retention by this group of animals with nature weight of 20 kg. The fact that the mean difforonees within those animals were not significant (P^ 0 .0 5) indicates that N retained was almost tho sane within the weight range 15 - 26 kg of the animals used in tho present experiment. The percent II retention increased as the IT intake and digestibi­ lity of the ration. IT retention was also influenced by the stage of UNIVERSITY OF IBADAN LIBRARY 166 growth of the animals. In the present report, young dwarf sheep at their early maturity were used and nitrogen retention values were high, which shows that dietary IT was being utilized in the formation of tissues. Stobo and Roy (1973) reported curvilinear increase in IT retention with IT intake and this is in agreement with the report of Robinson and Forbes (1970). The supplementation of hay with concentrate (C^) mainly cassava flour, a readily digestible form of energy, increased IT retention. This is in agreement with the report of Fick ct al. (1973).that supplemental energy improved utilization of low quality hay. In the present experiment, supplementation of hay with concentrate (C^) decreased urinary IT and hence increased IT retention values. The values of metabolic faecal IT (I-IFN) obtained in the present report for lambs varied from 2.98 to 3.68 g /k g DM consumed. This is lower than the value of 5.0 g /k g DM consumed often quoted for ruminants (Maynard and Loosli, 1962). The values are however comparable to that of Dcif ot al„(l968) who reported MFI'T value of 3.58 g /k g DM consumed, and also to that of Elliott and Topps (1963), who obtained 3.66 DM consumed but higher than those of Black ot ajl* (1973), Walker and Fouchney (1964a) and Lofgreen and Kleiber (1953) who reported MFD values of 2.04, 2.90 and 2.70 g /k g DM intake respectively. These investigators with the lower MFI'T values usod young lambs or calves maintained on liquid diets. A wide variation had been obtained in M M values by some investigators. The estimate of Robinson and Forbos (1970) was UNIVERSITY OF IBADAN LIBRARY 167 6.5 g/fcg M intake and is higher than 5.0 g /k g DM intake while on (1969) using the detergent method obtained a range of 4.4 to 7.2 g /k g HI intake for the M M values. From this, it is evident that a number of factors influence excretion of metabolic faoool nitrogen. The composition of feeds influences excretion of metabolic faecal nitrogen. Most of the investigators who obtained low values for M M used highly digestible feed like solid or liquid milk (Black et al.? \ 975) and those who obtained higher values used hi^iljr fibrous rations (Mason, 1969). Schneider (1935) showed that the excretion of M M was influenced by body siao as well as by the level of feed intake, M M tending to increase with increasing live weight of animal. Blaxter and Wood (1951) showed that M M excretion increased on rations of low TO digestibility. Lofgreen and Kleiber (1953)» using the N:3r ratio to determine K M in young calves showed that M M increased with increasing calf weights. Walker and Faichacy (1964a) obtained a < mean of 2.90 M M / k g DM intake using protein-free diets. However, when nitrogen-free diets are given to the ruminant animals, thoir protein metabolism is substantially altered (Waterlow, 1968) and the results obtained nay not apply under normal feeding conditions Mason (1969) showed that the proportion of nitrogenous contents of faeces is influenced by the type of the rations. He found that when animals were maintained on rations of hay or other roughages, the percent of undigested dietary II (uDll) ranged UNIVERSITY OF IBADAN LIBRARY 168 from 11 - 2 8 increasing from 11 fo for dry grass to 28 fo for Oat straw. The estimated me,an value of 21.7 + 3.8 obtained in the present studies for hay/concentrate rations are well within the range obtained by Mason (1969). Since the growth of ruminal micro-organisms is enhanced by the presence of readily fermentable carbohydrates, it is expected that the microbial and endogenous fraction (TIEN) would respond to the levels of energy in the ration. The results of Mason (1969) showed higher percentage of MEN on barley ration than on dried grass, 73 f> and 60 fo respectively. The HEN fraction euisists of bacteria, protozoa and digestive juices and other N of endogenous origin. Water-soluble nitrogen is expected to be higher when rations which arc highly soluble are fed to animals. Tlius the highest percentage of WSN (56 fo) was obtained by Mason (1969) on a ration containing much glucose and sene urea, a synthetic diet the components of which are soluble. Tor conventional rations of hay and concentrates, the estimates of Mason (1969) ranged from 1 6 - 2 8 Tho values obtained in present experiment ranged from 15.3 to 21.4 with a mean, value of 17.3 ± 3.4 f° and are well within range of Mason's (1969) determinations. The groundnut meal used, though soluble in ruminal liquor, could easily said rapidly be incorporated into microbial cell components and therefore little of it will appear as part of WSN fraction. UNIVERSITY OF IBADAN LIBRARY 169 The mean value of Non-dietary faecal nitrogen (TOM) obtained in this present i' nvestigatio' n was 78.9 *+■» 2.5 f° and is *also well within the range 72 - 97 f° obtained by Mason (1969), who also showed that this fraction increased with increasing proportion of readily fermentable carbohydrates in the ration, lowest for oat straw (72/0 and highest for barley diets (97$). The percentage MEN in NDHT obtained in the present investi­ gation ranged from 72/’ to 82.4$ and the mean value of 78 + 5 $ also falls within the range, 66 - 81 $, obtained by Mason (1969) using conventional rations. The results obtained in the present report arc all comparable with that of Mason (1969) and show that for conventional type of rations, the proportions of UDN, MEN and WSN do not differ much from other reported values. It must be borne in mind that when detergents arc used in extraction, some undigested dietary fractions such as pigment could bo extracted (Mason, 1969), This would tend to lower the estimate of UDN and to increase that of NDFN. The estimate of endogenous urinary nitrogen (BUN) obtained in this report as-0.0238 g/day/W0^ g7 3^ is lower than the values of 0.170 g/day/Wkg^ r obtained by Walker and Faichney (1964) by giving 0.734 protein-free diets, and the value of 0.056 g/day/^kg obtained by Black et al. (1973) using lambs with live weights ranging from 7.8 to 30 kg. It is also less than 0.038 g/,d ay/lM 0k.g7 34 obtained by Singh and Mahadevan (1970) using adult rams. The value of UNIVERSITY OF IBADAN LIBRARY 170 endogenous urinary nitrogen (BUI!) nay not be constant under all conditions. The results of Ashworth raid Cowgill (1938) suggest that in rats., the mount of endogenous urinary nitrogen per Wv0g..7 24 rises slightly as live w&ght increase, but there is insufficient published infornation to identify a similar trend in lanbs. The low value of EUIT obtained in this work for tho West African dwarf sheep may bo an adaptation to subsistence on low quality forage, the condition prevalent in the tropics where natural grass species rapidly decline in nutritional value (Oyenuga, 1957). Mugerwa and Conrad (1971) have shown that urinary urea excretion is indicated by the levels of blood urea. It may be that in West African dwarf 3heep, blood urea nitrogen is preserved by re-cycling into the rumen thus decreasing the possibility of much loss via tho urine. The very low loss of nitrogen via urine nay be given in support of this suggestion. The biological value of ration B, 100$, was higher than that of basal hay (85.7$), while the biological value of protoin-based t rations was 95.9 ± 2.2 or 0.959 i 0.022 and these high values show that the dietary nitrogen was being wo11 utilised by the lanbs. The value is higher than 0.85 and 0.65 obtained for urea and casein respectively by Deif ot al. (l968). Hie high biological value of tho ration m y be due to very low excretion of urinary nitrogen, and this nay in turn, be caused by the presence of readily fermentable cabohydrate in the form of cassava flour in the rumen depressing UNIVERSITY OF IBADAN LIBRARY 171 ruminal ammonia production, blood urea and urinary urea levels. The sheep at this stage of growth were retaining a lot of nitrogen for tissue protein synthesis. The fact that maximum Biological value with the rations was attained on a ration supplying the least amount of crude protein and which then declined suggests that maximum W is obtained at minimum dietary N intake. The Biological value obtained in the present report is higher than that obtained by Singh and Mahadcvan (1970), a mean of 86.9 ± 8.68 fo. It is also higher than the value of 0.851 or 85.1 ‘f0 obtained b3̂ Stobo and Roy (1973) who also fed groundnut meal-based rations to ruminant calves. Hie Thomas-Mitchell concept of biological value as a practical neasure of the quality of a protein has two serious limitations. The first is the difficulty of obtaining a measure of the endogenous urinary, and to a less extent, the metabolic faecal nitrogen. In practice, the former is sometimes calculated on the basis of body size according to the equation mg EON = 146 W0 *75 .................... (4.27.) If the values for EON are to be determined experimentally, it is necessary to prepare a nitrogen-free diet of the type used by Walker and Faiehncy (1964a). The second difficulty arises from the fact that the levels of nitrogen fed modifies the calculated Biological value indepondentl3r UNIVERSITY OF IBADAN LIBRARY 172 of anino acid balance, because deamination appears to proceed somewhat according to the law of mass action (Crampton, 1956). Thus to obtain maximum Biological value, there must be a minimum of protein furnished. This is 3hown by the fact that ration B which supplied the least amount of IT had the highest biological value (100/6). This automatically means that production rations, including those for growth, where liberal protein feeding is necessary for maximum performance, show low biological values as compared to maintenance rations. Hence, biol#gioal values of individual foods will change according to the rations in which they are used. Biological values aro constants only if the proto in is used entirely for maintenance. In order to compare such- figures for different feeds, they must have been determined at the same protein levels of intake, and to standardise this such rations aro often adjusted to 10$ protein. Errors of the estimate of Biological value can be reduced by using the values of EU17 and MSTF determined during the experiment. It would be wrong to use equation 4.27 to estimate EU1I since the value obtained would be higher or lower than those of other investigators. Errors due to overestimation or underestimation of I!M and EUN were minimised in the present investigation. The finding of Barnes, Bates and Maack (1946) showed that age, class of animal and the production- involved would tend to influence the effective apparent biological value of a protein in addition to amino acid balance. UNIVERSITY OF IBADAN LIBRARY 173 Not protein utilization (NPU) takes into account the response of the aninal to the dietary protein intalce. The estimate reported here of 86.1 + 2.2 % is higher than the estimated value of 79.6 + 2.47 reported by Singh and Mahadevan (1970). This difference in the values of Net protein Utilization (lIPU) obtained in the present report and that of Singh and Mahadevan (1970) is due to a higher estimate of Biological value obtained in the present work (95.9 ± 2.2) than tint reported by those investigators (86,9 + 8.68). In determining the requirement of crude protein by the sheep, some assumptions have been made, and those have been summarised by Black ct_ ad. (1973) as follows (a) that measurement of nitrogen balance gives an accurate estimation of the nitrogen retained by the lambs (b) that the inevitable losses of nitrogen in urine and faeces can be determined by an extrapolation to zero protein intake of the results from animals given a wide range of protein intakes, and that (c) the inevitable losses of nitrogen per metabolic size, (d) the energy requirements for maintenance per metabolic size and (o) the efficiencies of utilization of metabolizable energy for maintenance and for production are constant over the range in live weight and diets covered by the experiment. With careful experimentation the common losses of N due to errors in urine collection and failure to consider dermal looses in- sheep range from only 1.2 to 2.6 % of the faecal and urinary output of N (Martin, 1966). Moreover, because If loss from the integument is UNIVERSITY OF IBADAN LIBRARY 174 a result of metabolism and is inevitable, it should be included * in an estimation of protein requirements. The endogenous losses of I'l in urine and faeces obtained by using H-frce diets are usually higher than when ample supply of IT is included in rations. However, the fact that values obtained by investigators using similar animals and rations agree closely, showed the reliability of extrapolation techniques. Wien animals with a narrow weight range are used, differences in the values of EON, MEN, and energy required for maintenance are not likely to be great. The value reported here of digestible protein requirement 0.734 for maintenance in periods 1 and 2 of Trial 2 was 1.15 g/dayA^kg or 1.57 g/dayA^g^^ live wei£ht, and for periods 5 and 4, the ̂ 0 734 value was 0.41 g/dayA^Cg or 0.56 g/day/kg live weight. A very sharp decline during periods 5 and 4 was obvious. This observation shows that protein requirements of growing animals change rapidly with age or live weight end agrees with the usual observations of some investigators (Black cat al.̂ 1973). The mean value for the experimental period was 0.74 g/d. ay/W, ^0g«,7 34 or 1.01 g/day/wg live weight. Black et al. (l973) found that the endogenous "T IT losses in lambs of 20 kg live weight was 0,20 g digestible n i t r o g e/n / d/a °y*/7N^j j o r 1.250 g digestible crude protein per metabolic size per day. The value of 1.250 is similar to 1.50 obtained in this present work during periods 1 and 2 but higher UNIVERSITY OF IBADAN LIBRARY 175 than 0.74 g/dayAg W°-75} the mean .value for periods 1 to 4. The present report with value of 1.150 is also similar to the value of 1.16 g/day/%. * obtained by Robinson and Forbes (1966) using non-pregnant ewes, slightly higher than the values of 0.875 and 0.893 g/daykg obtained by Singh and liahadevan (1970) using adult rans, maintained on groundnut meal-based rations, but very much lower than 3.6 g/day/fcg ¥ * recommended by I Brody (1945) # The present reported value is about 33$> of B r o d y * o recommendation. Elliot and Topps (1964) have shorn that the proportion of roughage to concentrate in the ration influenced maintenance requirement of sheep. They showed that the maintenance requirement increase with increasing roughage to concentrate in the ration. Elliott and Topps (1964) showed that the generally accepted standards for digestible N for maintainance appear to be excessive by a factor of 3 when applied to African cattle and sheep given diets adequate in energy. The present value of requirement is about 33% of Brody’s recommendation. This is therefore in agreement with the observation of Elliott and Topps (1964). The high efficiency of IT utilization of the rations fed to the hest African dwarf wether sheep is apparently associated with low endogenous losses and also high biological values of the protein. The rations used contained cassava flour, a readily fermentable source of energy and this reduced N losses due to deamination of protein in the rumen and subsequent loss of N in urine. The low protein requirement for the sheep used UNIVERSITY OF IBADAN LIBRARY 176 may also be due to the fact that an appreciable amount of nitrogen of urea was recycled to the rumen from the blood, and this would be efficiently fixed into ruminal microbial protein in the presence of readily fermentable sources of energy. Since in the sheep used, about 60fo of dry natter consumed was in the form of highly fermen­ table cassava flour, it is expected that any re-cycled urea would bo well utilized. Rosenthal and Allison (1951) have noted in monogastric animals, the protein-sparing action of energy-rich foods. Elliott and Topps (1953) suggested that the same or similar mechanism nay bo present in the ruminant, quite apart from the improvement in protein quality broughtabout by ruminal micro-organisms. They also reported that African cattle may through natural selection in a protoin-deficient environment, have evolved some physiological process for conserving IT under such stress conditions. The same is probably true of sheep and other ruminants under humid tropical onvii’onaonts UNIVERSITY OF IBADAN LIBRARY CHAPTER FIVE 5. DIGESTION AT DIFFERENT SITES OF THE ALIMENTARY TRACT OF THE 'NEST AFRICAN DWARF SHEEP 5.1 I N T R O D U C T I O N The digestion of feeds in the sheep may be divided into three stages, namely the fermentative processes that occur in the first three parts of the stomach (rumen, reticulum and omasum), hydrolytic digestion in the abomasum and small intes­ tine and finally a secondary fermentative stage in the large intestine. The present teport deals with sheep maintained on Cynodon nlemfuensis/Centrosema pubescens hay with or without protein and cassava flour supplements. The aim of the experi­ ment was to investigate the suitability of using chromic oxide- impregnated paper to determine the extent of digestion of feed in the different parts of the digestive tract of the West African Dwarf wether sheep. 5.2 MATERIALS AND METHODS 5.2.1 Animals and their Management Twelve West African dwarf wether sheep, 8 - 1 3 months old and weighing 13 to 20kg were used in this experiment. Each sheep was kept in a metabolism cage (Oyenuga, 1961). The animals were usually fed at 8.00 a.m. everyday. The residues which might be left over were collected, weighed and UNIVERSITY OF IBADAN LIBRARY 178 stored for chemical analysis (in order to determine nutrient intake). The animals had free access to salt licks and were weighed at the beginning of the experiment and just before they were slaughtered. 5-2.2 Experimental Rations The experimental rations used in this study are the same as those used in Chapter two, (Table 2.1). 5.2.3 Plan of Experiment The twelve wether sheep were randomised into six groups of two animals in each group, and to each group was assigned one of the six experimental rations. Four grams of chromic oxide - impregnated papers were given orally to each of the sheep with the aid of a balling gun just before feeds were offered every morning. There was a 14 - day preliminary period and a 6 - day collection period during which faecal samples were collected. 5.2.4 Collection of Faeces A day prior to collection, the animals were fitted with harnesses to which were attached collection bags. A polythene bag was placed in each collection bag to allow for easy coll­ ection of faeces. The bags were emptied daily just before the morning feeding. Faeces were dried to a constant weight in a forced - draught oven at 70°C for kS hours. The daily dried faeces were bulked for each animal, milled, and stored in air-tight glass bottles until required for analysis. UNIVERSITY OF IBADAN LIBRARY 179 5*2.5 Collection of Digesta from the Different sites of the digestive tract. After the faecal collection period, the animals were slaughtered, four in a day at 8.00 a.m. The viscera were removed as quickly as possible and ligatures were placed in the following places; the reticulo-omasal junction, the oraaso- abomasal junction, the pylorus, the proximal small intestine (the first metre), the distal small intestine (the last metre), the ileo - caecal junction, and the rectum. Digesta were removed from the sections, weighed and mixed, and an aliquot was taken and freeze-dried at - 20°C for 6 days. The digesta samples were milled and stored in air - tight sample bottles until required for analysis. 5.2.6 Analytical Procedure AOAC (1970) method was used to determine total N and ash in the digesta using Markham (19^2) semi-micro Kjeldahl apparatus for N determination. Chromic oxide in the digesta samples was determined by the method of Williams, David and Iismaa (1 9 6 2) using Parkin - Elmer Atomic absorption spectrophotometer (Model 290). 5*2.7 Estimation of Digestibility in the different sites of digestive system. Omasal samples were used to estimate digestion in the reticulo-rumaaand omasum; abomasal samples were used to estimate UNIVERSITY OF IBADAN LIBRARY 180 digestion in the stomach (rumen, reticulum, omasum and abomasum). Digesta sample from the terminal ileum was used to estimate digestion in the small intestine. Rectal samples were used to estimate total digestibility (Holmes et al_. , 1970. 5.3 R E S U L T S 5«3«1 Total Digestibility of Dry Hatter and Nitrogen The comparison of the total collection and chromic oxide methods for the determination of dry matter and N diges­ tibilities is shown in Table 5-1* The dry matter digestibility of the rations by the chromic oxide method ranged from 5 3 *3% with the basal hay, to 8 0.6% for ration F. The DM digestibility of the concentrate - supplemented rations were significantly higher than that of the basal hay (P < 0.01). There seemed to be increases in DM digestibility with increasing crude protein content of the ration, especially with rations E and F. The DM digestibility as determined by the total collection method ranged from ^k .2% for the basal hay to 82 .0% for ration F. The mean differences of DM digestibility by the chromic oxide and total collection methods were not significant (Pj>0.05)* The digestibility of N by the chromic oxide method ranged from 37 .8% with basal hay to 7^.8% with ration F. The digesti­ bility of N increased with increasing N intake and also with UNIVERSITY OF IBADAN LIBRARY 181 TABLE 3-1 A Comparison of the Total Collection and Chromic Oxide Methods*. For the Determination of Dry Hatter and N Digestibilities with the West African Dwarf sheep Maintained on Hay and Concentrate Supplements Nutrient Digestibility % Ration Nutrient Chromic Oxide Total MCeotlhlMethod o edction Dry matter 53.3 52.lv ^ A Nitrogen 37.3 *+0.7 B Dry matter 73.2 76.1 Nitrogen 31.3 k o . 6 C Dry matter 71.2 75.9 Nitrogen k k . 2 52.9 D Dry matter 71.1 7 6 . 2 Nitrogen 59.^ 6 6 . 8 E Dry matter 79.5 79.9 Nitrogen 6 6 . 0 7 0 . 0 F Dry matter 8o . 6 8 2 . 0 Nitrogen 7^.8 78.7 + Mean differences of digestibility values by the two methods not significant at(P> 0.05). UNIVERSITY OF IBADAN LIBRARY 182 increase in the crude protein content of the rations. The value of the digestibility of N by the total collection method ranged from kO.7% with basal hay to 78.7% with ration F, and this also shows that digestibility of N increased with level of dietary crude protein. The values of N digestibility by the total collection method were consistently higher than the values obtained by the chromic oxide method; however, mean differences were not significant (P^ 0 .0 5 ). 5»3»2 Chromic Oxide Recovery The chromic oxide recovery for the experimental animals is shown in Table 5-2. The value ranged from 8 ^ . 8 to 10^.5% with a mean value of 95«0 _+ 1.7%. Three of the experimental animals had a recovery of chromic oxide less than 90%» and three had a recovery greater than 100%. 5•3•3 Digestibility of dry matter at different sites of the digestive tract. The digestibility of dry matter at different sites of the digestive tract is shown in Tables 5«3»1 and 5«3*2 and total digestibility of the ration is obtained by adding together the digestibility in all the sections of the digestive tract (Table 5°3»1)» Table 5.3-1 shows that kk .6% out of a total digestibility of 5 3 *3% for dry matter, took place in the reticulo-rumen plus omasum with basal hay, and k 6 ,6% of a total digestibility of 7 3 *2% took place in the reticulo-rumen UNIVERSITY OF IBADAN LIBRARY 183 TABLE 5.2 Chromic Oxide Recovery for the West African Dwarf Wether Sheep Maintained on Basal Hay and Concentrate Supplements Sheep No. CR Intake (g) Faecal CR (g) % Recovery 31** 5 . 2 8 5 o02 95.08 336 5 . 2 8 5 . 1 2 96.97 3*+3 5 . 2 8 *+.66 8 8 . 2 6 399 5 . 2 8 *+.*+8 8*+.85 *+99 5 . 2 8 5.38 1 0 1 .8 9 510 5 . 2 8 *+.98 9*+.32 320 5 . 2 8 *+.62 8 7 . 5 0 513 5 . 2 8 5 . 2 2 18.86 518 5 . 2 8 5 . 3 2 100 .7 6 *+8*+ 5 . 2 8 5 . 5 2 10*+.55 358 5 . 2 8 *+.96 93.9*+ 570 5 . 2 8 *+.92 93.18 Mean 9 5 .0 1 + 1.7*+ UNIVERSITY OF IBADAN LIBRARY 184 TABLE 5-3.1 The percentage digestibility of dry matter taking place in the sections of the digestive tract of the West African dwarf wether sheep maintained on hay and concentrate supplements, using the chromic oxide ratio % % % % Total Ration Digesti­ Digesti­ Digesti­ Dige %sti­ Digesti­ Digesti­ bility in bility bility bility bility in bility in the raticu- in the in the in the the Caecun digestive lo-rume+ abomasum stomach small in­ and colon tract Omasum testine A if if. 6 13-3 57.9 0 -if.5 53.3 B 46.6 4.0 5 0 . 6 16.7 6 . 0 73.2 C if3.2 0 . 1 ^3-3 3.1 1 9 .8 7 1 . 2 D 6 5 . 8 1 1 . 0 54.8 -3.7 2 0 . 0 71.1 E 48.6 -8 . 0 if 0 . 6 1 6 . 2 2 2 . 7 79.5 F 47.5 7 . 5 55.0 9.0 16.6 8 0 . 6 Mean 49.4 1.0 50.if 7 : 7 13.* 71.5 SE 3 . 8 2.9 3.5 if.3 4.0 UNIVERSITY OF IBADAN LIBRARY 185 TABLE 5.3.2 The percentage digestible dry matter taking place in the sections of the digestive tract of the West African dwarf wether sheep Maintained on hay and concentrate supplements, using; chromic Oxide ratio % % % % % Ration Digestibi­ Digesti­ Digesti­ Digesti­ Digesti­ lity in the bility in bility in bility bility in reticulo- the the stomach in the the caecurr rume+ abomasum small and colon Omasum intestine A 8 5 . 6 2^.9 108.5 0 -8.5 B 65.5 k.O 69.5 22.9 7.6 C 60.7 0 * 1 6 0 . 8 1 1.if 22.8 D 9 1 . 0 -Ilf. if 7 6 . 6 -5.** 2 8 . 8 E 6 1 . 1 -10 .1 50.0 2 0. if 2 8 . 6 F 59.0 9*3 6 8 . 5 1 1 . 2 20.5 Mean 70.2 2.3 72.5 10 .1 17.if i; SE 5.6 5.8 8 .1 *U5 5.9 UNIVERSITY OF IBADAN LIBRARY 186 plus omasum with ration B. The corresponding values for rations C, D, E and F were 43.2, 65*8, 48.6 and 4-7.5% out of total digestibility of 7 1.2 , 7 1 .1 , 7 9 * 5 and 8 0.6% respectively. Table 5*3*2 shows the percentage of digestible dry matter taking place in the various sections of the digestive tract. The values in table 5.3*2 are obtained from Table 5*3.1 by dividing each value in Table 5»3«1 by the total digestibility value, for example, the percentage of digestible dry matter taking place in the reticulo-rumen and omasum with ration A is calculated as: 44.6 x 100 = 8 3.6% ............5.1 53.3 The results show that a mean of 72.5 +, 8.1% of digestible dry matter was obtained in the stomach. In the small intestine, the value was 10 .1 + 4.5%, and a mean of 17.4 _+ 5»9% took place in the caecum plus colon. 5.3.4 Digestibility of Organic Matter in the Various Sections of the Digestive Tract. The digestibility of organic matter is shown in Table 5.4.1 and 5*4.2. Table 5*4.1 shows that 44.6% of a total organic matter digestibility of 54.4% took place in the reti­ culo-rumen plus omasum with basal hay. The corresponding values were 48.4, 42.7, 6 5 .6 , 49.7 and 49.0% of total digestibility coefficients of 74.5, 72.3, 7 3 -8 , 8 0 . 6 and 8 2.0% with rations B, C, D, E and F respectively. UNIVERSITY OF IBADAN LIBRARY 187 TABLE 5.4.1 The percentage digestibility of organic matter taking place in the Sections of the digestivet/roafc tthe West African dwarf wether sheep maintained on hay and concentrate supplements, using the Chromic Oxide ratio °/o % % Digesti­ Digesti­ Digesti­ D^ges- TDVi g0e/0s- Total Digesti­ bility in bility bility tibili •tibilit bility in Ration the reti- in the in the ty in in the the diges­ culo-rumen4 absomasu n stomach the caecum Omasum small plus tive intes­ colon tract tine A 44.6 13.7 58.3 0.9 -4.8 54.4 B 48.4 2.4 5 0 . 8 18.4 6 .1 75.3 C 42.7 0.3 43.0 1 0 .6 18.7 72.3 D 6 5 . 6 -7.3 58.3 -3.5 19.1 73.9 E 4 9 . 7 -7.4 42.3 18 .1 20.2 8 0 . 6 F 4 9 . 0 8 . 9 57.9 8.4 15.7 8 2 . 0 Mean 5 0 . 0 1.8 51.8 8.8 12.4 73.0 SE 3 . 3 3.5 3.1 3.5 4.0 4.0 UNIVERSITY OF IBADAN LIBRARY 188 TABLE 5.4.2 The percentage digestible organic matter taking place in the Sections of the digestive tract of the 'West African dwarf wether sheep maintained on hay and concentrate supplements using the chromic oxide ratio % % % % % Digesti­ Digesti­ Digesti­ Digesti­ Digestibility Ration bility in bility in bility in bility in in the ca#curr the reti- the the the small and colon culo-rume+ abomasum stomach intestine omasum A 8 2 . 0 25 . 2 107.2 1 . 6 -8 . 8 B 6 % 6 2 . 2 6 7 . 8 24.3 7.9 C 59.1 0.3 59.4 14.7 25.9 D 8 7 . 0 -8 . 7 78.3 -4.9 2 6 . 6 E 6 1 . 6 -9.1 52.5 22.4 25.1 F 59.8 1 0 .8 7 0 . 6 10.2 1 9 . 2 Mean 6 9 . 2 3.5 72.7 1 1 . 4 15.9 SE 5.0 5.3 7.8 4.7 5.7 UNIVERSITY OF IBADAN LIBRARY 189 Table 5*4.2 shows that 82.0% of the total digestible O.M took place in the reticulo-rumen plus omasum with basal hay and the mean value with all the rations is 6 9 . 2 _+ 5*0%. Similarly, 3*5 _+ 5«3% of digestible OM was obtained for the abomasum, 11.4 _+ 4.7% for the small intestine and 16.0 + 5.7% for the large intestine. 5*3*5 Nitrogen Intake, Distribution and Absorption in the Various Sections of the Digestive tract of the Sheep Table 5*5 shows the intake, distribution and absorption of N in the stomach and intestine of the sheep* More N was recovered at the omasum than the total N intake. The values of N absorbed in the proximal small intestine were negative and this indicates that large amount of secretion of nitroge­ nous substances took place there. At the distal small intestine the absorption was much. The sum of N absorbed in the proximal and distal small intestine is the estimate of the net absorption of N in the small intestine, for instance, - 4.19 g/day of N was absorbed at ppoutlmal small intestine (4.19 g/day was secreted), and 4.99 g/day was absorbed at the distal small intestine, therefore ndt absorption in the small intestine per day is 4*99 + (-4.19g) or 0.80g N per day for the sheep maintained on ration A (Table 5»5)« The nitrogen absorbed in the small intestine as percentage of N intake was 61.6 _+ 22.6, and N UNIVERSITY OF IBADAN LIBRARY 190 TABLE 5 . 5 Nitrogen Intake, Distribution and Absorption at Different Sites of the Alimentary Canal of the West African Dwarf Sheep Maintanined on Hay and Concentrate Supplements N “ N — — '----------N N N n ---- u---- ----H---- N — TT— ----- n—N ----fl---- Ration Intake Passing Flowing Flow­ Flowing Flowing Absorbed Absorbed Absorbed absorbed Absor­ f^bsor- Absorbed Absorbed (g/day; through through ing through through in the in the in the in the bed in oed in in the in the omasum abomasum through the the reticulo- abomasum proximal distal the the small small per day (g/day) the pro­ distal return rume plus (g/day) small small whole Large intestine as (g/day) ximate small (g/day) omasum intestine intes­ small intes- as per cent small intes­ (g/day) (g/day) tine intes­ ;ine per cent of N intes­ tine (X) (g/day) tine !g/day) intake passing tine (g/day) (Y) (g/day) through (g/day) (X+Y) theabomasum per day A 5.59 3 . 8 8 2.41 6 . 6 0 1 .6 1 2.69 1.71 1.47 -6.9 4.99 0 . 8 0 -1 .0 8 14.3 33.2 B 3»61 4.64 4.68 7.89 1.78 2.65 -1 . 0 2 -0.04 -3 . 2 0 6 . 1 2 2 . 1 2 -0 . 8 2 80.9 62.3 C 3 . 6 2 3.72 3.4o 8 . 3 6 1.75 2.04 -0.78 0.33 -4.97 6 .6 1 1.64 -0.29 45.3 48.2 D 4.84 3-92 4.00 7.04 2.53 2.09 0.48 -0.07 -3.05 4.51 1.46 0.44 3 0 . 2 36.5 E 3.24 4.16 6.35 17'. 64 1 . 0 0 1 .1 1 -0 . 9 2 -2.19 -11.29 16.64 5.35 -0 .1 1 165 .1 84.3 F 12.03 1 2 . 0 7 7.27 6.67 3 . 2 2 3.09 -0.04 4.80 0 . 6 0 3.45 4.05 0 . 1 3 33.7 55.7 Mean 61.6 53.4 SE + 22.6 + 7.7 ____________ UNIVERSITY OF IBADAN LIBRARY 191 absorbed in the small intestine as percentage of N passing through the abomasum was 53«^ +, 7.7%. The correlation between N intake (X), in g/day, and the gain of N at abomasum (Y), g/day, was high, negative and significant (P<0.05). The regression equation is as follows Y = 0.76 x + 3-53 ...........5.2 (r = — 0.86)* From the equation, it can be shown that at zero N intake, 3.53 g/day of N could still flow into the abomasum from the ffeticulo-rumen and omasum. It can also be shown that at the N intake of ^.64 g/day, there would be no net gain of N from the rumen. At the intake of N higher than 4.6ilg per day, a net loss of N would be expected to occur in the rumen. The fact that the correlation was negative showed that a net gain of N in the stomach occurred only at low levels of N intake. D I S C U S S I O N The rations used in the present investigation were similar to that used for the experiments reported in Chapters 2 and *+. The values of the dry matter digestibility obtained in this report were therefore similar to the values obtained in Chapter k (Table 4.2); it shows that there has been no appreciable variation in the composition of the rations. UNIVERSITY OF IBADAN LIBRARY 192 In the present report, increasing digestibility coeffi­ cient of N was reported, with increasing levels of dietary crude protein; which is in agreement with the results of Andrews and Orskov (1970a). The digestibility coefficients of dry matter and N by the total collection and indicator methods showed no significant differences (P/>Q.05), and this shows the reliability of the chromic oxide paper method for deter­ mining total digestibility in the digestive tract. It must be borne in mind that if faecal samples were taken as often as possible, the differences in the digestibility values might be eliminated, and the fact that in the present report, the sample was taken once could have led to the little differences obtained since one sample might not be truly re­ presentative of faecal excretion for the day. The mean percentage recovery of chromic oxide for the animals was 95»0 _+ Virtually complete recovery of chromic oxide in the faeces of sheep has been reported by McRae and Armstrong (19&9), using chromic oxide - impregnated paper, by Putnam e_t al_. ̂ (1958) using chromic oxide in gelatin capsules and fed to cows, by Cowlishaw and Alder (1 9 6 3) using both chromic oxide - impregnated paper and the oxide with oil as the carrier. However, certain investigators have reported incomplete recovery of chromic oxide. Johnson et_ al_./ (196^) UNIVERSITY OF IBADAN LIBRARY 193 found 101.8# recovery with chromic oxide - impregnated paper but only 93*3# recovery when the powder was given. Pigden and Brisson (1956) obtained recoveries in the faeces of 101#. 9k% and 87# of the administered dose in three separate, - day trials each with four sheep, and Deinum, Immink and Dejis (1962) obtained recoveries of 97«5#i 9 8 .6# and 9 8.^# in experiments with cows given 50g of chromic oxide - impregnated paper daily and found traces of the oxide in the liver, lymph glands and kidneys. They suggested that some absorption of the marker might have occurred. The value of 95*1 + 1.7# obtained in the present report may therefore be taken to lie within the range obtained by several investigators. Estimates of chromic oxide recovery in the faeces would normally be less than 100# because most sources of error lead to losses of chromic oxide. Loss of faeces from the collection bag have been noted by several investigators including Carter, Bolin and Erickson (i9 6 0) and Beaut (1961). Losses may occur if the bags are not emptied often enough or if the harness is not correctly adjusted, particularly if the faeces have a high water content ( C 12# dry matter). Some of the faeces may stick to the bag. In the experiment reported, a polythene bag was fitted into the collection bag to prevent faecal loss. Carter ejt al. ̂ (1960) and Scaut (19 61) found that 5 - 7 days were sufficient for preliminary period. Bruce, Goodal, Kay, UNIVERSITY OF IBADAN LIBRARY 194 Phillipson and Vowles (19 6 6) showed that excretion of chromic oxide 'impregnated on to paper was irregular but fully recovered. In the present report 72«5% of digestible dry matter and 72.6% of digestible organic matter digestion took place in the stomach. These values are higher than the values of 4 3% obtained for hay by Balch (1957) , and 60% obtained for straw by Badawy et al . f (1958) but similar to the value of 71.3% obtained by Holmes e_t al• y (1970); however, these authors used (ignin as indigestible marker. Drennan _e_t al. ̂ (1970) using chromic oxide powder to estimate digestion in the stomach, obtained values ranging from - 7 to 36%. They suggested that the low values might be due to rapid or uneven passage of marker from the rumen. The present report shows that the digestibility values given by chromic oxide - impregnated paper are consistent. Most of the investigators who have determined digestibi­ lity of nutrients in the digestive tract of ruminants with chromic oxide as indicator have also used animals with re-entrant cannulae in the abomasum and terminal ileum. The results re­ ported in the present study also agree with their reports. Thus, Hogan and Phillipson (i9 6 0) found that of the total dry matter digested in the sheep, 70% took place in tie stomach, 11% in the small intestine and 19% in the large intestine. These values appear to be in very good agreement with the present report with values of 72.5 ± 8.1%, 10.1 _+ 4.5% and 17»4 ± 5*9% for UNIVERSITY OF IBADAN LIBRARY 195 the stomach, small intestine and large intestine respectively. Topps , Kay and Goodal (1968) showed that for animals fed with hay, 67% of the digestible dry matter disappeare in the stomach, 22% in the small intestine and 11% in the large intestine whereas for animals taking concentrate - based rations, the digestibility coefficients were 69%, 17%, and lk% in the stomach, small intestine and large intestine res­ pectively. The present results showed that of the total organic matter digested, 72 .6% took place in the stojnaoh, 11.4% took place in small intestine, and 16% took place in the large intestine. Bruce et al.^ (1 9 6 6) reported values of 68%, 20% and 12% of organic matter digested as taking place in stomach, small intestine and large intestine respec­ tively. Hogan, Connell and kills (1972) reported a value of 74% of the total digestibility of organic matter as occurring in the stomach when hay was fed to sheep. 0rskov, JSraser, McDonald and Smart (1974) also reported that 60% of the dietary organic matter disappeared in the rumen, 32% in the small intestine and 8% in the large intestine. In the present report, there was a net gain of nitrogen in the stomach of the sheep receiving low levels of nitrogen and a net loss in some receiving higher levels of nitrogen. There was a negative correlation (r = - 0.86) between the UNIVERSITY OF IBADAN LIBRARY 196 nitrogen intake and the gain of nitrogen in the stpmaah. This relationship indicates that at the zero nitrogen intake up to 3 -5 3g of nitrogen per day could still flow to the abomasum undoubtedly from microbial and salivary sources and other metabolic excretory nitrogen sources. Also from the regression equation, no net gain of nitrogen would take;* place at the intake of h .6 k g o f nitrogen per day. This is in agreement with the reports of Gray, Pilgrim and Weller (1958), Clarke, Ellinger and Phillipson (1 9 6 6), Nicholson and Sutton (1969)1 McRae (1970), Harris and Phillipson (1 9 6 2), Hogan and Weston (1 9 6 7), Topps at al . , (1 9 6 8), Kay, McLeod and Pavlicevic (1972) and jtfrskov et al.^ (197^)* These investigators reported that when sheep were consuming low dietary N more total N passed to the duodenum than were eaten daily in the feeds, whereas when these animals were consuming high dietary N, less total N passed to the duodenum than in the feeds. Gray et al.^ (1958) calculated that there was an apparent gain of N in the stomach when sheep consumed feeds containing less than 5g nitrogen daily. This is in agreement with the present report with value of k . 6 k g N per day. Harris and Phillipson (1 9 6 2) reported 50% more N per day, passing the abomasum of sheep than present in rations of low N UNIVERSITY OF IBADAN LIBRARY 197 The present results showed that a large quantity of nitrogen was secreted at the anterior small intestine and absorbed at the posterior portion. Badawy e_t al . j (1958) found that increase in the N was considerable at the proximal half of the small intestine while absorption took place in the terminal half. Ben-Ghedalia, Tagari and Bondi (197*0 reported that in the sections of the intestine from 1 - 15m posterior to the pylorus, the amounts of water, dry matter and total N decreased gradually as a result of absorption through the intestinal wall, that the region 7 - 15m from pylorus was more active with respect to the absorption of N whereas water, and dry matter were absorbed to a greater extent in the region 1 to 7m from the pyloruc.. The only part of the intestine in which substantial increases in water, dr̂ y matter and total N were found was the section immediately distal to the pylorus and these increases were caused by the inflow of bile, parcreatic and duodenal juices. These inves­ tigators found net increases beyond the entry of the common bile duct to be 2.7g protein - N and 2.0g non-protein - N per day, and that only very small changes occurred after 15m dis­ tant from the pylorus. From the present studies, the N absorbed in the small intestine as percentage of N intake was 61*6 _+ 22.6% and when expressed as percentage of N passing through the abomasum, the value was 53«*+ +. 7.7%» UNIVERSITY OF IBADAN LIBRARY 198 Even though there were individual differences between animals due to difficulty of sampling in the present exper­ iment, the results obtained are in reasonable agreement with those obtained with re-entrant cannulation for the purpose of partitioning digestibility in the various sections of the digestive tract. The general weakness of slaughter technique is that samples can only be obtaine&once from slaughtered animals, and since the samples might not be truly representative of the digesta flowing through the organ, considerable error due to sampling could be introduced if care were not exercised during the sampling. The great advantage of using animals with re-entrant cannulae over slaughter technique is that the former animals live long enough for repeated samplings to be carried out but the pre­ sent results showed that reliable results could be obtained using slaughter techniques. UNIVERSITY OF IBADAN LIBRARY 199 SUMMARY OF CONCLUSION The levels of ruminal metabolites of nitrogenous origin obtained in the present investigation were similar to the levels obtained by other investigators using similar rations, and this shows that the metabolism of forage and concentrate supplements in the West African dwarf sheep was similar to that of other breeds of sheep* The composition of the rations markedly influenced the concentrations of metabolites in the rumen* There was a tendency for total N, protein N, and non-protein N to increase with increasing levels of crude protein in the ration. Ruminal ammonia levels were low with all the rations used and this indicates that not much dietary N would be lost in the urine. Supplementation 0f the forage with cassava flour greatly depressed ruminal ammonia levels and this enables the rumen micro-organisms to convert ammonia - N into microbial protein. In the present study, low levels of ruminal ammonia could be interpreted to mean that dietary N was being very efficiently utilized. The levels of blood urea were low and increased as ruminal ammonia. It is known that urinary N excretion increases with increasing blood urea N, However, the low levels of blood urea obtained in the present investi­ gation was associated with low urinary N excretion, and high nitrogen retention. UNIVERSITY OF IBADAN LIBRARY 200 The ruminal micro-organisms have very low concen­ tration of methionine and histidine, and high lievels of lysine and leucine. Rations high in non-protein nitrogen, for example, urea-based rations may require methionine and histidine for efficient utilization by the micro-organisms. The results obtained when [_ N_J7 ammonium chloride and _15N _/_ urea were used to study the kinetics of ammonia and urea metabolism, were in very good agreement with those of other investigators (Coccimano and Leng, 1967; Mugerwa and Conrad, 1971; Nolan and Leng, 1972). The results however, showed that the West African dwarf sheep was able to recycle a substantial amount of blood urea to the digestive tract, which may be an adepatation for existence in areas where dietary N supply is often inadequate. The results obtained in the present report shows that fistulation has no effect on the digestibility of dry matter and nitrogen by the sheep. The intake of dry matter by the West African dwarf sheep was similar to that of other breeds of sheep when the results are expressed on the metabolic size basis. Dry matter intake was related to the body weight raised to the power of 0.668 kg The value of 0.668 is similar to 0.66, the exponent which relates body weight to body surface area, and this suggests that in the West African UNIVERSITY OF IBADAN LIBRARY 201 dwarf sheep, metabolism is related more to body area than to body weight. Butterworth (1966) suggested that this might be due to the necessity for the sheep under tropical con­ ditions to maintain homoeothermy by heat loss through the body surface.. The sheep used in the present investigation were utilizing dietary N very efficiently. Urinary N levels were low and N retention values were high especially with the concentrate - based rations, showing that dietary N is Y best utilized in the presence of readily fermentable sources of energy. In the present investigation, the readily ferina- table source of energy was in the form of cassava flour. The metabolic faecal nitrogen (MFN) values obtained in the present report ranged from 3*0 to 3*7g N/kg dry matter intake which is lower than the value of 5»0g/kg DM intake often quoted for ruminants (Maynard and Loosli, 1969) but is comparable to the values of 3 to 3*7 g/kg DM intake obtained by several investigators (Elliott and Topps, 1963; Deif et al., 1 9 6 8). However, the nature of the diets influence metabolic faecal N excretion (Mason, 1 9 6 9). Faecal analysis showed that for conventional type of ration, the percentage of non­ dietary faecal nitrogen (NBFN), microbial and endogenous nitrogen (MEN), and water-soluble nitrogen (WSN) do not vary appreciably. UNIVERSITY OF IBADAN LIBRARY 202 The low endogenous urinary N value (0.0238g/day/W^“^ ) obtained in the present report may be an adaptation of the tropical breeds of sheep for survival on forages of low N content. The biological values of the rations used were very high, showing that the sheep were utilizing dietary N efficiently. The value of 95«9 +, 2.2% obtained with protein- based rations is higher than the values 85 - 87% obtained for groundnut meal - based rations by singh and Mahadevan (1970), and Stobo and Roy (1973)* The value of the crude protein requirement for main­ tenance obtained in the present investigation, by the N - balance method was about 33% of Brody's (19^5) recommended value for a sheep of similar live weight. Elliott and Topps (196^) had observed that the generally accepted standards for digestible N for maintenance appear to be excessive by a factor of 3 when applied to African cattle and sheep given diets adequate in energy. The va'ue of digestible crude protein required for maintenance, obtained by the factorial method was much lower than the value obtained by N balance method. The low crude protein requirement for maintenance of the West African dwarf sheep may be an adaptation to life in areas where crude protein supply is often inadequate. UNIVERSITY OF IBADAN LIBRARY 203 The present report shows that chromic oxide is well recovered in the faeces but it is not very evenly distri­ buted in the digesta. The digestibility coefficient obtained could be subject to some error due to this uneven distribution. The variation in the concentration of chromic oxide between perio: 27. Blaxter, K.L., and WJ.. Wood, (1951). "The nutrition of the young Ayrshire calf. 1. The endogenous nitrogen and basal energy metabolism of the calf". Br. J . Nutr. jj: 11. UNIVERSITY OF IBADAN LIBRARY 208 Block, R .J e, J JL » Stekol and J.K. Loosli, (1957). "Synthesis of sulphur amino acids from inorganic sulphate by ruminants. 11. Synthesis of cysteine and methionine from sodium sulphate by the goat and by microorganisms from the rumen of the ewe” . Arch. Biochem. Biophys. 35: 353. 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"Effect of supplemental protein and energy levels on the utilization of Kikuyu grass (Pennisetum clandestinum) J. Agric. Sci. 22: 634. Carter, J.F., D.W. Bolin and D. Erickson, (i960). "Evaluation of forages". N,.Dakota agrio. exp, sta. Bull. No. 426. Chalmers, M.I. and R.L.M. Synge, (1954). "Ruminal ammonia forma­ tion in relation to protein requirement of the sheep - II Comparison of casein and herring - meal supplements" J. Agrio. Sci. 254. Chalupa, W., (1968). "Problems in feeding urea to ruminants." J. Rnim. Sci. 27: 207. Chaney, A.L. and E.P. Marbach, (1962). "Modified reagents for analysis of urea and ammonia." Clin. Chem. _8: 130. Chargaff and Sprinson,(l943) . oited by McLaren GJ1. (1964) in ’Symposium on Microbial digestion in Ruminants'. J. Aniia. Sci. 22: 377. Clarke Eileen M.W,, Gabrielle M. Ellinger and A.T. Phillipson,(l956). "The influence of diet on the nitrogenous components passing to the duodenum and through the lower ileum of sheep." Proc. Royal Soc. 166: Ser. 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"A new naturally occuring amino acid." Nature 165 : lb-. Wright, N.C., (1961). "Hunger: Can it be averted?" Brit. Ass. for Adv, Sci. London. Zilversmith, D.E,, (i960). "The design and analysis of isotope experiments." Amer. J. Med. 29: 832. Zuntz, N. (1891). Cited in 'Animal Nutrition' by Maynard L.A. and JJC. Loosli. (1969 ed.) 0 . 106. UNIVERSITY OF IBADAN LIBRARY A P P E N D I X TABLE At NITROGEN INTAKE, DIGESTIBILITY AND UTILIZATION IN THE NEST AFRICAN DWARF SHEBP. RATION - A N fo FAECAL - N ANIMAL | INTAKE FAECAL-N URINARY-N DIGESTED-N RETAINED-N DIGESTI­ RETEN­ g/kg D.M. NO. !A - (- g./ -d ay.) - . (g/ day) (g/day) (g/day) (g/day) BILITY TION CONSUMED 179 6.35 2.72 1.07 3.63 2.56 57.14 40.31 5.35 259 | 8.88 3.84 0.75 5.04 4.29 56.69 47.18 5.40 268 5.29 2.31 0.57 2.98 2.41 56.36 45.56 5.45 186 9.07 4.25 0.74 4.82 4.08 53.18 44.98 5.85 263 2.18 1.10 0.42 1.08 0.66 49.73 30.28 6 .3 0 173 5.79 | 2.61 0.45 3.18 2.73 54.89 47.15 5.63 301 2.65 1.07 0.49 1.58 1.09 59.69 41.13 5.04 184 2.91 1.18 ___ . _ “ 1.73 59.41 - 5.07 . ______________ ____________ A HAY 228 UNIVERSITY OF IBADAN LIBRARY TABLE 2. NITROGEN INTAKE, DIGESTIBILITY AND UTILIZATION IN THE WEST AFRICAN DWARF SHEEP. ........ FAECAL-N ANIMAL INTAKE-N f a e c a l-n TJEINAEY-N DIGESTED-N RETAINED-N DIGESTI­ RETEN­ g/kg D.M. (g/day) (g/day) (g/day) (g/day) (g/day) BILITY TION CONSUMED 179 3.98 1.45 0.04 2.53 2.49 63.57 62.56 2.84 259 3.83 1.40 0.11 2.43 2 .32 63.45 60.57 2.61 268 2.83 1.43 0.05 1.40 1.35 49.47 47.70 3.50 186 4.53 1.72 0.05 2.81 2.76 62.03 60.93 3.21 263 4.39 1.59 0.22 2.80 2.58 63.78 58.77 3.04 173 4.15 1.82 0.14 2.33 2.19 56.14 52.77 3.64 301 4.05 1.41 0.13 2.64 2.51 65.19 62.00 2.07 184 5.62 1.92 0.83 3.70 2.87 65.84 51.07 3.53 i i--------------- „ _____________________________ B 229 UNIVERSITY OF IBADAN LIBRARY TABLE 3 NITROGEN INTAKE, DIGESTIBILITY A!© UTILIZATION IN TEE WEST TRIAL 2 AFRICAN DWARF SHEEP. PERIOD 1 RATION I.D. NO j INTAKE-1T FAECAL-U URINARY-N DIG2STSD-N RETAINED-N ---D-I---$ANIIIAL GES --T--I-­-- REHTEN- FAECAL- (g/day) (g/day) (g/day) (g/day) (g/day) BILITY TION N g/kg.D.M C 179 5.98 2.28 0.20 3.70 3.50 61.87 58.52 4.70 C 259 7.41 2.92 0.07 4.49 4.42 60.59 59.65 4.38 D 268 9.41 3.67 0.25 5.74 5.49 61.00 58.34 4.90 D 186 6.71 2.33 0.09 4.38 4.29 65.27 63.93 4.28 l E 263 8.79 3.17 0.22 5.62 5.40 63.93 61.43 5.82 I ! E 173 11.59 3.84 1.10 7.75 6.65 66.87 57.38 5.37 F 301 12.28 3.22 0.18 9.06 8.88 73.78 72.31 5.22! F 184 12.49 3.22 0.27 9.27 9.00 74.22 12.06 5.74 5.05+0.55 UNIVERSITY OF IBADA LIBRARY TABLE 4 NITROGEN INTAKE, DIGESTIBILITY AND UTILIZATION IN THE WEST TRIAL 2 AFRICAN DWARF SHEEP. PERIOD 2 ------------ — RATION — I.D. NO. INTAEE-N FAECAL-N URINARY-N DIGSSTED-N RETAINED-N DIGESTI­ RETEN­ FAECAL-N ANIMAL (g/day) (g/day) (g/day) (g/day) (g/day) BILITY TION g/kg D.M D 179 10.55 3.47 0.89 7.08 6.19 67.11 58.67 5.30 D 259 11.31 3.19 0.55 8.12 7.58 71.79 67.02 4.57 E 268 13.19 3.61 0.93 9.58 8.65 72.63 65.58 5.58 E 186 12.34 2.82 0.66 9.52 8.86 77.15 71.80 4.88 P 263 13.00 2.67 1.33 10.33 9.00 79.46 69.23 4.36 P 173 13.07 3.48 1.76 9.59 7.83 73.37 59.91 5.24 C 301 6.00 2.04 0.12 3.96 3.96 3.84 66.00 3.931 C 184 7.10 2.37 1.11 ' 4.73 3.62 ' 66.62 51.00 6.12 __ - 5 .00+0.66 2J1 UNIVERSITY OF IBADAN LIBRARY TABLE 5 NITROGEN INTAKE, DIGESTIBILITY AND UTILIZATION IN THE WEST TRIAL 2 AFRICAN DWARF SHEEP. PERIOD 3 r1“ ------ i ---------j ! ration I.D. NO INTAKE-N FAECAL-N jURINARY-N DIGESTED-N RETAINED-N DIGESTI­ RETEN­ FAECAL-N ANIMAL (g/day) (g/day) j (g/day) (g/day) (g/day) BILITY TION g/kg D.M { CONSUMED i E 179 11.14 2.75 j 0.20 8.39 8.19 75.31 73.52 4.55 E 259 12.65 4.01 | 0.74 8.64 7.90 68.30 62.45 5.48 F | 268 15.53 3.41 0.59 12.12 11.53 78.04 74.24 5.72 P ! 186 j 15.09 3.41 1.00 11.68 10.68 77.40 1 0 ,1 1 4.95 « ! 1* c | 173 j 9.34 3.61 0.55 5.73 5.18 61.35 55.46 5.30j B j! 301 j 8.76 2.13 0.22 6.63 6.41 ! 75.68 | 73.17 3.53i 1 1 B ! 134 j 3.83 1.00 0.11 2.83 2.72- - - -- — * 73.89 | 71.01 1.96 4.5Q+1.16 232 UNIVERSITY OF IBADAN LIBRARYVfcO\CM TABLE 6 NITROGEN INTAKE, DIGESTIBILITY AND UTILISATION IN THE WEST TRIAL 2 AFRICAN DWARF SHEEP. PERIOD 4 7[— -"— — % 1° RATION I.D. NO. INTAKE FAECAL-N t URINARY-N DIGESTED—N RETAINED-N DIGESTI­ RETEN­ FAECAL-N ANIMAL (g/day) (g/day) (g/day) (g/day) (g/day) BILITY TION g/kg D.M. CONSUMED F 179 13.84 3.34 1.18 10.50 9.32 75.87 67.34 5.95 F 259 15.27 3.35 1.20 11.92 10.72 78.06 70.20 4.80 C 268 8.29 3.18 0.41 5.11 4.70 61.64 56.69 4.53 C 186 6,59 2,20 0.51 4.39 3.88 66.62 58.88 4 .2 8 ;1 D 263 6 ,7 2 2,02 0.63 4.70 4.07 69.94 60.57 4.75 » ■ 173 4.59 1.36 0.20 3.23 3.03 70.37 j 66.01 3.231 E 301 9.74 • 2.64 0.24 7.10 6.86 72.90 | 70.45 5.05 1 t 184 i 9.81 | 2.67 0 .3 2 7.14 6.82 7 2 . ! 69.52 5.4 E . - . . . . __ _______ 6_ _ 11 4.76+0.76 UNIVERSITY OF IBADAN LIBRARY TABLE 7 NITROGEN INTAKE, DIGESTIBILITY AND UTILIZATION IN THE WEST RATION A2 AFRICAN DWARF SHEEP. j -------------* i ANIMAL INTAKE-N FAECAL-N URINARY-N DIGESTED-N RETAINED-N DIGESTIBILITY RETENTION NO. (g/day) (g/day) (g/day) (g/day) (g/day) 179 3.19 1.49 0.75 1.70 0.95 53.43 29.78 259 5.10 2.61 1.30 2.49 1.19 48.74 23.33 ! 268 4.17 1.90 0.22 2.27 2.05 54.49 49.16 j 186 4.79 1.93 1.10 2.86 1.76 59.73 36.74 263 3.19 1.23 - 1.96 - 61.54 mm 173 4.79 2.15 0.22 2.64 2.42 55.13 50.52 301 3.19 J 1.44 0.49 1.75 1.26 55.00 39.50 184 2.55 1 1.12 0.30 1.43 1.13 55.92 44.31 i _ ________ A 234 UNIVERSITY OF IBAD N LIBRARY TABLE 8 RATION A RUMEN AND BLOOD METABOLITES OP THE WEST AFRICAN DWARF SHEEP RUMEN AMMONIA I BLOOD PROTEIN-N NON-PROTEJN-N NON-AMMONIA TOTAL-N I D mg/100 ml mg/lOO ml mg/lOO ml NON-PROTEIN-N mg/lOO NOS. 1 HR 2 HRS;! 3 HRS jmg/JoO |! 1 HR 2 HRS 3 HRS 1 HR 5 2 HRS 3 HRS 1 HR 2 HRS 3 HRS ml. . . . . . 179 i _ i 4.8 4.5 4.3 6.9 | 32.0 28.0 25.0 8.5 7.0 7.0 3.7 2.5 2.7 35.8 259 4.0 4.0 5.6 3.7 2.6 2.5 4.0 1 i ----------------------------------jj--------------------------------- 268 1J ...... \ .. _ , 4.6 4.2 4.3 7.5 36.0 3 2 .0 36.0 6.0 7.0 7.5 1.4 2.8 2.7 41.5 186 4.3 4.3 4.4 4.8 1 jj 5.6 | 4.5 4.0 4.2......... I _ . . ! 4.6 4.0 4.0 3.8 j 39.6 42.0 j 16.8 43.2 11.2 5.6 j38.6 12.8 1.6 61.6 173 | 5.4 5.4 6.0 4.8 1| 1 j 4.3 j 3.5 j 3.5 4.0 | 1 . . . . j ----3----0---1--- ------jr I :* i ; t <. _________ J --------------r 1 16.8 ! 4.6 ! 4.4 8.0 | 22.4 29.2 | 14.0 14.0 14.0 5.6 7.2 13.4 1.2 33.1 184 8.0 ] 10.8 i 4.0 | 8.0 i ! 1 l1 | » i j 6.8 j 6.0 3.5 < 2.0 | i i \ i 1l .......... ............................ -----------------------L t __________________ i UNIVERSITY OF IBADAN LIBRARY oo in• TABLE 9 RATION B RUMEN AND BLOOD METABOLITES OF THE WEST AFRICAN DWARF SHEEP •rttMEN a m m o n i a BLOOD PROTEIN-N NON-PROTSIN-N NON-PROTEIN I D (mg/100 ml ) UREA (m g/100 m mg/100 1) (mg /100 ml 1 NON -AMMONIA-N TOTAL-N NuS J1. HTII PV 2 HRS 3 HRS 1 HR 2 HRS ) i 3 HRS 1 HR 2 HRS 3 HRS (ng/100 nl) ml ; l h r 2 HRS 3 HRS 179 0.1 ! 0.1 0.2 1.1 36.4 8.4 8.4 5.6 5.6 2.8 5.5 5.5 2.6 22.4 259 1.1 0.7 0.7 1.2 0.6 0.3 0.3 1.4 - - - - 268 0.6 0.6 0.6 0.5 25.2 25.2 14.0 16.8 8 .4 5.6 16.2 7.8 5.0 31.7 186 1.0 1.1 1.1 1.2 0.6 0.6 0 .5 1.2 265 0.6 0.7 0.6 0.9 16.8 16.8 30.8 8 .4 8.4 8.4 7.8 7.7 7.8 29.9 173 0.6 0.9 1.1 1.2 !i | 1.9 i 2.4 2.4 1.4 ! | I — — - — . i-....! ~ i i i 301 ! “ 2.4 2.4 j 2.0 33.6 30.6 14.0 | 11.2 8 .4 j 8 .4 7.2 6.0 6.0 35.54 . 0 1 ! 184 4.0; 1.1 | 2.0 i 1.2 i ii ____ 2 .8 ! 2.4 | 3.1 i i ! i __________! UNIVERSITY OF IBADAN LIBRARY TABLE 10 RATION C RUMEN AND BLOOD METABOLITES OP TEE NEST APRICAN DTJARP SHEEP RUMEN AMMONIA BLOOD PROTEIN-N NON-PROTEIN-N NON-AMI IONIA TOTAL-N PER­ I.D (mg/100 ml.) UREA I mg/100 ml) (mg/lOO ml) NON-PROTEIN (a v.) IOD NOS 1 HR 2 HRS i 3 HRS mg/100 ml 1 HR 2 HRS 3 HRS 1 HR 2 HRS 3 HRS 1 HR 2 HRS 3 HRS mg/lOQ ml. 179 0.6 0.6 0.7 1.2 25.2 16.8 12.0 11.2 11.2 3 .0 10.6 10.6 2.3 0.6 0.6 0.5 0.9 26.5 0 .7 0 .5 0.6 0.9 268 *+■ 4 186 i.8 2 .8 2.6 2.3 53.2 | 38.0 22.3 15.8 13.4 9.6 14.0 10.6 7.0 50.8 | 2.6 2.8 | 2.8 3 .0 ! 2.6 2.1 j 2 .7 CM 263 I 4- 173 ! 2 .0 2.1 32. 2.1 42 .0 ! 42.0 39.0! 22.4 30.8 8 .4 20.4 28.7 5.2 61.5 3.2 2.2 ! 1.6 i 2.8 1.3 | 1.2 1.3 )• 301 184 1.5 2.4 1 i 2 1.5 I 36.8 28.0 22.4 14.0 11.2 | 8.4 i 12.4 9.8 7.2 1.4 1.5 1.3 2.6 40.; 4 .0 2.6 1.3 2.0 UNIVERSITY OF IBADAN LIBRARY / TABLE 11 RATION D RUMEN AND BLOOD METABOLITES OP THE LUST AFRICAN DWARF SHEEP T------- PER­ ^ “ T p — ELUDE— ~I . D (mISg/O10m0 Umm — ~ PROTEIN-cr EJJ-K TOTAL, IOD i )NOS. 1 HR j 2 HRS 3 HRS mg/UlROEA ® S B 8 * nH M ® (av.) Oml 1 HR^J ^ H R S 1 HR 2 HRS 3 HRS (ng• /lOO m3 Dg/lOO 1 HR 2 HRS 3 HRS ml. 2 179 i. . . . . _ . . 2 259 5.2 3.6 2.4 3.6 30.8 50.4 50.4 j n . 2 8.4 8.4 6.8 6.9 7.4 53.2 4.4 1.5 1 .0 3.2 | 7.2 5.0 2.2 4.0 i--------- - - . . . . . . - . 1 268 j . r . 1 186 1.8 2.0 0.8 1.4 50.4 53.2 30.8 22.4 25.2 14.0 20.6 23.2 13.2 65.3 0.8 0.7 0.8 0.9 0.9 1 .0 1.2 1 .0 |___ 1— 4 263 ij - - - - - - - - - - - - - - - i i ' 4 173 4.8 1.8 1.8 3.4 58.8 53.2 50.4 28.4 23.5 10.6 23.6 21.7 8.8 79.0 J 4.8 3.2 1.6 2.6 1• 1 j f! j 4.0 2.6 1.4 4.6 J ! ! I 3 301! i j . . . . . . . . . . . j I ! J 3 184 1.6 2.8 2.4 2.3 56.0 72.8 j 50.4 22.4 j 30.8 8.4? 20.8 28.0 6.0 80.3 j 2.2 1.6 1.8 2.3 j ! j ! 1.8 !.5 j 1.4 2.1 1 j ! i 1- - - - - - - - - - - - --- - - - - - - - - - - - - - - - - 1 ----------------------------- L _ _ _ _ _ _ _ _ _ _ _ _ _ _ L u — - - - - - - - - - UNIVERSITY OF IBADAN LIBRARY TABLE 12 RATION E RUMEN AND BLOOD METABOLITES OP THE NEST AFRICAN DWARF SHEEP RUMEN AMMONIA BLOOD PROTEIN-N NON-PROTEIN-N NON-AMMONIA PER- I.D (ng/lOO nl) UREA (mg/lOO ml) (mg/lOO nl) NON-E TOTAL r o t e i n tn (av.) IOD NOS 1 HR 2 HRS 3 HRS ng/lOO 1 HR 2 HRS 3 HRS 1 HR 2 HRS 3 HRS nl 1 HR 2 HRS 3 HRS (mg/lOO nl 3 179 3 259 5.8 4.2 4.0 3.8 75.6 52.5 38.4 25.6 29.3 10.8 19.8 25.1 6.8 77.4 4.4 2.6 1.3 3.2 5.2 3.6 2.2 3.3 2 268 ----------- .. L L* . • 2 186 9.0 3.2 1.8 6.0 78.4 28.0 44.8 28.0 14.0 25.2 19.0 10.8 23.4 5.0 3.8 2.0 8.0 7.8 10.0 6.2 7.6 1 263 1 173 1.8 0.9 0.9 1.8 61.6 70.0 36.4 42.0 14.0 8.4 40.2 13.1 7.5 3.3 1.8 1.8 14. 77.5 4.2 3.6 3.2 1.0 4 30. 4 184 4.2 4.0 3.2 4.2 61.6 58.8 64.4 30.2 26.5 13 .8 26.0 22.5 10.6 6.0 5.9 2.0 3.8 85.1 3.6 3.8 1.6 3.2 — „ ___ -J ' —---- ---- - --- ________ 2 3 9 ■ UNIVERSITY OF IBADAN LIBRARY TABLE 13 RATION P RUMEN AND BLOOD METABOLITES OF THE WEST AFRICAN DWARF SHEEP RUM DN-NH^ BLOOD PROTEIN- N NON-PROTEI NON--n h 3 PER­ I.D (ns/ lOO ml ) UREA (mg/lO m1) (ng/lOO E NON-PIiOTEIN-f k1 I TOTAL IOD NOS 1 HR 2 HRS 3 HRS (mg/ 100 1 HR 2 HRS 3 HRS 1 HR 2 HRS (m 5/IOO El] L) Ell) 1 HR 2 HRS 3 HRS (mg/lOO ml.) 4 179 .... 4 259 6.4 5.6 6 .0 4.2 61 .6 7 0 .0 81.2 25.7 30.4 28.3 18.4 2 4 .8 22.3 99.1 6 .2 2 .2 1 .6 4.6 19.3 6 .0 3.4 2 .8 5.2 i 3 268 ;______ i______ 3 186 6 .8 6 .6 4.8 4.6 42.0 42.0 106.0 22.4 4 2 .0 9 8 .0 15 .6 33.4 93.2 117.5 5.8 3.6 2.4 4.4 35.4 4.8 3.0 1 .6 4.7 > 2 263 i 2 173 16.4 6.4 2 .6 8.4 5 6 .0 142.9 1 40 .0 28.0 98.0 ' 98.6 11.6 91.6 95.4 187.6 5.2 4.8 3.8 7.2 »>i 8 .2 5.3 2.4 6 .8 51 1 301 i 1 184 1.7 1.7 1 .6 1.9 72.8 6 7 .2 67.2 28.0 25.2 1 25.2- 26.3 23.5 2 3 .6 95.2 3.6 4.2 2.7 5.3 i J 240 UNIVERSITY OF IBADAN LIBRARY £3 ^ TABLE 14 TRIAL ONE PERIOD ONE -BY MATTER INTAKE, DIGESTIBILITY AND N UTILIZATION IN VEST AFRICAN DWARF SHEEP ANIMAL NO. WEIGHT 1 (kg) RATION N-INTAKI N-DIGEST]p N-RETAINED DIGESTIBILITY RETENTION d r y •MATTER g/ % % INTAKE DIGESTIBI­ g/day/Wkg^ LITY. % ! 179 15.88 A 0.85 I 0.48j 0.35 57.1 40.3 67.5 55.4 259 i 21.77 Ai 0.94 0.53 0.45 56.7 47.2 74.9 57.0 1 268 19.96 !a i! 0.60 0.34 0.27 56.4i 45.6 47.6 53.6 186 19.50 A |i 1.04 0.56 0.47 j 53.2 j 45.0 83.0 57.3 263 22.68 * i 0.48 0.31 0.28 63.8 58.8 57.0 72.0 173 26.31 B it 0.39 0.21 0.20 | 56.1 52.8 46.0 70.8 301 13.15 B 0.58 0.37 0.35 65.2 62.0 96.5 75.8 I 184 j 17.69 B 1 ( 0.6S 0.45 | 0.36 65.8— L. 66.7 74.7 | ------ 1 j !t 51.1 j --- -- „ i» UNIVERSITY OF IBADAN LIBRARY TABLE 15 TRIAL ONE DRY MATTER INTAKE, DIGESTIBILITY AND N UTILIZATION IN WEST AFRICAN DWARF SHEEP PERIOD TWO N-INTAKE N-DIGES'l N-RETAINED DIGESTI­ T-----------PED RETENTION DRY - ANIMAL WEIGHT RATION MATTER BILITY INTAKE DIGESTIBI­ NO. (kg) ... * % g/day/W^73 LITY % 179 15.88 B 0.53 0.33 0.33 62.6 62.6 67.8 64.5 259 21.77 B 0.40 0.25 0.24 63.4 60.6 56.6 72.7 268 19.96 B 0.31 0.16 0.15 49.5 47.7 45.9 72.2 18C 19-05 B 0.52 0.33 0.33 62.0 62.0 62.4 68.5 263 20.86 A 0.23 0,12 0.07 49.7 30,3 17.9 56.7 j 173 26.31 A 0.53 0.29 0.25 54.9 47.1 42 .6 57.9 301 j 14.51 ! A 1 0.41 0.24 0.17 i 59.7 41.1 32.4 57.3 184 j 17.69 A 0.36 0.21 i Jf ii 59.4 28.6i 56.5 242 UNIVERSITY OF IBADAN LIBRARY TABLE 16 2ND TRIAL DRY MATTER INTAKE, DIGESTIBILITY AND N UTILIZATION IN WEST AFRICAN DWARF SHEEP 1ST PERIOD T------ i ANIMAL |1 WEIGHT RATION N-INTAKE N-DIGESTEI N-RHTAINED DIGESTI­ RETEN­ DRY- HATTER NO. (kg) 3 BILITY TION INTAKE DIGESTI­ g/day/W°g g/day/W^73 BILITY % 7o 1° 179 15.42 C 0.81 0.50 0.47 61.9 58.5 65.8 73.3 259 21.77 C 0.78 0.47 C.47 60.6 59.6 70.4 75.2 268 20.86 D 1.02 0.62 0.60 61.0 58.3 81.5 69.3 186 18.60 D 0.79 0.52 0.51 65.3 63.9 64.4 67.8 265 21 .32 E 0.94 0.60 0.58 63.9 61.4 58.3 65.2 173 i 26.31 E 1.07 0.72 0.61 66.9 57.4 65.7 69.4 501 |5 14.51 F 1.74 1.29 1.26 73.8 72.3 87.5 76.8 184 j 17.69 F 1.53 1 1.14 1.10 74.2 76.2 69.0 66.1_____i,_ _______ UNIVERSITY OF IBADAN LIBRARY TABLE 17 2ND TRIAL DRY MATTER INTAKE, DIGESTIBILITY AND N UTILIZATION IN WEST AFRICAN DWARF SHEEP 2ND PERIOD — - ANIMAL WEIGHT RATION N-INTAKE N-DIGESTED N-RETAINED D IG EST I­ RETENTION DRY - MATTER • NO. 1I B IL IT Y INTAKE D IG EST I- | i ! 1° 1° g/day/W^73 B IL IT Y ! 1° j ) 179 14.51 D 1.50 1.01 0.88 67.1 58.7 92.9 71.9 259 21.32 D 1.21 0.87 0.81 71.8 67.0 74.7 70.3 268 19.96 E 1.48 1.07 0.97 72.6 65.6 72.7 72.5 186 17.24 E 1.54 1.19 1.11 77.1 71.8 72.4 80.2 [ 263 23.59 F 1.29 1.02 0.89 79.5 69.2 60.9 77.2 i 173 24.95 F 1.25 0.92 0.75 73.4 59.9 63.4 75.0 1 0.61 0 .58 66.0 64.0 78.8 1 30.1 13.15 , C 0.91 79.1 184 ! 15.42 | C 0.96 0.64 0.49 i 66.6 51.0 52.5 79.4 i | 1! ... 244 UNIVERSITY OF IBADAN LIBRARY TABLE 18 ?. 0 TRIAL DRY HATTER INTAKE, DIGESTIBILITY AND N UTILIZATION IN WEST AFRICAN DWARF SHEEP 3RD PERIOD — ANIMAL WEIGHT RATION N-INTAKE N-DIGESTHD N-RETAINSD DIGESTI­ RETEN­ DRY --MATTER NO. (kg) BILITY TION INTAKE DIGESTIBI­ i g/day/W^ 13 g/day/W®*73 LITY 1° % f> 179 16.78 E 1.42 1.07 1.04 75.3 73.5 77.0 78.4 259 22.68 E 1.29 0.88 0.81 68.3 62.4 74.9 73.9 268 20.41 F 1.72 1.34 1.27 78.0 74.2 80.0 77.5 186 19-05 F 1.76 1.36 1.24 77.4 70.8 80.1 76.4 263 22.22 C 0.34 36.7 71.9 173 26.76 C 0.85 0 .5 2 0.48 61.3 55.5 71.7 75.9 ! 301 16.33 D 1.19 0.89 0.87 75-7 73.2 81.9 79.0 184 17.24 D 0.48 0.35 0.34 73.9 i 71.0 j 63.9 74.1! 245 UNIVERSITY OF IBADAN LIBRARY TABLE 19 2ND TRIAL DRY MATTER INTAKE, DIGESTIBILITY AND N UTILIZATION IN WEST AFRICAN DWARF SHEEP 4TH PERIOD — — ANIMAL WEIGHT RATION N-INTAKE N-DIGESTED N-RETAINED DIGESTIBI­ RETEN­J DRY-MAT1! PER LITY TION INTAKE DIGESTI­ BIOLO­NO. (kg) £j/day/W^73 % fo g/day/W^75 BILITY GICAL VALUE 1° 179 17.24 F 1.73 1.31 1.17 75.9 67.3 70.3 75.8 91.92 259 23.59 F 1.52 1.18 1.03 78.1 70.2 69.4 76.4 93.19 268 22.68 C 0.85 0.52 0.48 61.6 56.7 71.8 74.1 97.54 186 19.50 C 0.75 0.50 0.44 66.6 58.9 58.9 80.8 95.00 265 21.32 D 0.72 0.50 0.44 69.9 60.6 45.6 76.3 93.20 173 26.31 D 0.42 0.29 0.28 70.4 66.0 38.7 76.1 98.70 301 16.33 E 1.27 0.92 0.89 72.9 70.4 68.1 76.7 99.31 184 16.78 E 1.25 0.91 0.87 72.8 69.5 62.4 77.9 98.50 1 L - ____________ J ---------.------------------- ------------------i UNIVERSITY OF IBADAN LIBRARY TABLE 20 1ST TRIAL CONCENTRATION OP RUMEN AND BLOOD METABOLITES (mg/lOOnl) IN TEE DUST ANSI CAN D’ .ARP SHEEP 1st PERIOD ' ANIMAL NO. RATION TOTAL-N PROTEIN-N NON-PROTEIN-N AMMONIA-N NON-AMMONIA RESIDUAL tA-AMINO-N BLOODN N (Uaele/nl) UREA-N 259 A 35.8 28.3 7.5 4.51 31.3 3.0 3.1 6.9 186 A 41.5 34.6 6.9 4.4 37.1 2.5 3.6 7.5 173 B 29.9 21.5 8.4 0.6 29.3 7.8 1.8 0.9 184. B 35.5 26.1 9.4 2.9 3 2 .6 6.5 1.1 2.0 LST I’RIAL. 2ND PERIOD 259 B 22.4 17.7 4.7 0.1 22.3 4.6 1.8 1.1 186 B 31.7 21.5 10.2 0.6 31.1 9.6 1.7 0.5 173 52.8 32.8 20.0 4.2 48.6 15.8 3.6 3.8 ?I 184 jt A 33.1 21.9 11.2 5.3 27.8 i 5.9 2.7| : 247 UNIVERSITY OF IBADAN LIBRARY •COo TABLE 21 21® TRIAL CONCENTRATION OP RUMEN AND BLOOD METABOLITES (ng/lOOml) IN THE WEST AFRICAN DWARF SHEEP PERIOD 1 248 UNIVERSITY OF IBADAN LIBRARY TABLE 22 2ND TRIAL CONCENTRATION OF RUMEN AID BLOOD METABOLITES (ng/lOOnl) IN THE WEST AFRICAN DWARF SHEEP PERIOD 3 ANIMAL NO. RATION TOTAL-N PROTEIU-N NON-PROTEIN-N AMIIONIA-N RESIDUAL-N r<-AMIN0-N BLOOD UREA (u mole/ml) N 259 E 77.4 55.5 21.9 4.7 17.2 5.65 3.4 186 F 117.5 63.3 54.2 6.1 48.1 5.85 4.6 173 C 61.5 41.0 20.5 2.4 18.1 3.50 1.9 184 D 80.3 59.7 20.5 2.3 18.3 2.25 2.2 2ND TRIAL PERI 01) 4 259 F 99.1 70.9 28.2 6.0 22.1 4.50 4.7 186 C 50.8 37.8 13.0 2.4 10.5 1.50 v 2.7 173 D 74.9 54.1 20.8 2.8 18.0 2.70 3.5 184 E 85.1 61.6 23.5 3.8 19.7 3.50 3.7 - - i ■ 249 UNIVERSITY OF IBADAN LIBRARY - 250 - TABLE 23 DRY MATTER INTAKE AND DIGESTIBILITY OF RATIONS BY THE WEST AFRICAN DWARF SHEEP HAY (i) RATION A I.D NO. INTAKE b*v\ FAECAL £ A i° ANIMAL (g/day) (g/day) DIGESTIBILITY 179 508.0 226.8 55.35 259 710.6 305.4 57.02 268 423.4 196.6 53.57 186 725.8 310.0 57.29 263 174.6 75.6 56.70 173 463.4 195.0 57.92 301 212.4 90.7 57.30 2 32 .8 101.3 . 5„ 6...4.9 i— n- m41 HAY II 179 255.2 102.1 59.99 259 408.2 141.8 65.26 268 333.4 153.1 54.08 186 383.3 131.5 65.69 263 255.2 88.5 65.32 173 383.3 139.5 63.61 301 255.2 79.4 68.89 184 204.1 64.6 68.35 _________,________ ____ 00 UNIVERSITY OF IBADAN LIBRARY 251 - TABLE ?4 DRY MATTER INTAKE AND DIGESTIBILITY OF RATIONS BY THE WEST AFRICAN DWARF SHEEP HAY AND CASSAVA. (RATION B) ANIMAL NO. INTAKE FAECAL (g/day) (g/day) DIGESTIBILITY 179 510.3 181.4 64.45 259 536.4 146.3 72.75 268 408.2 113.4 72 .22 186 536.4 169.0 68.49 263 5 2 2 .8 146.3 72 .02 173 500.1 146.3 70.75 301 680.4 164.4 75.84 184 543.2 137.2 74.74 UNIVERSITY OF IBADAN LIBRARY -• 252 - TABLE 25 DRY MATTER INTAKE AND DIGESTIBILITY OP RATIONS BY THE WEST AFRICAN DWARF SHEEP PERIOD 11 RATIONS ID. NO. g j lay g/day % ANIMAL INTAKE FAECAL DIGESTIBILITY C 179 484.8 129.5 73.29 c 259 667.2 165.4 75.21 D 268 748.4 229.6 69.32 D 186 544.5 175.5 67.79 E 263 544.5 189.5 65.19 E 175 714.4 218.3 69.44 F 501 616.6 143.2 76.78 F 184 561.3 143.2 66.08 PERIOD IT D 179 654.9 184.3 71.86 D 259 697 = 4 207.4 70.26 E 268 646.4 177.7 72.51 E 186 578.3 114.3 80.23 F 263 612.4 139.9 7 7 .1 6 F 173 663.4 165.8 75.00 C 301 518.8 110.1 78.78 C 184 387.0 79.8 79.36 l___________ UNIVERSITY OF IBADAN LIBRARY - 253 - TABLE 26 DRY MATTER INTAKE AND DIGESTIBILITY OP RATIONS BY THE TOST AFRICAN DWARF SHEEP PERIOD III RATIONS ID. NO. g/day g/day % ANIMAL INTAKE FAECAL DIGESTIBILITY E 179 603.9 130.4 78.40 E 259 731.4 190.9 73.90 F 268 722.9 162.5 77.52 F 186 688.9 162.5 76.41 C 263 353.0 99.2 71.89 C 173 680.4 164.0 75.90 D 301 603.9 126.6 79.03 D 184 510.3 132.3 74.07 PERIOD IV F 179 561.3 136.1 75.76 F 259 697.4 164.4 76.42 * 268 701.7 181.4 74.14 C 186 514.6 98.8 80.80 D 263 425.2 100.6 76.33 D 173 421.0 100.6 76.09 E 301 523.0 121.9 76.69 E 184 489.0 108.2 77.87 ____________ . ... UNIVERSITY OF IBADAN LIBRARY TABLE 27 THE DIGESTIBILITY OF THE NITROGEN OF THE CONCENTRATE SUPPLEMENTS OF THE FEED BY THE WEST AFRICAN DWARF SHEEP C 0 N C E N I R A T E , 3 ANIMAL NO. C1 cC2 °3 °4 °5 179 69.3 67.6 73.1 84.1 83.0 259 83.5 64.1 82.5 77.5 93.3 268 44.5 65.5 64.9 82.2 94.7 186 79.6 78.0 81.7 87.2 94.0 263 90.5 — 77.9 79.0 94.4 173 87.7 65.7 85.9 76.4 84.9 301 74.7 71.4 83.3 79.8 82.7 184 * 68.8 79.3 78.4 81.6 TOTAL 529.8 481.1 628.6 644.6 708.6 MEAN 75.7 68.7 78.6 80.6 88.6 i SE !! 5.9 1.9 2.4 1.3 2.1 * Value greater than lOOfo, excluded from estimation of mean. UNIVERSITY OF IBADAN LIBRARY - 255 TABLE 28 THE DIGESTIBILITY OP TEE DRY MATTER OP THE CONCENTRATE SUPPLEMENTS OP THE PEED BY THE WEST AFRICAN DWARF SHEEP. C 0 N C E N T R A T E S I.D. NOS. °1 C2 C4 C5 179 72.5 94.7 81.8 89.4 83.4 259 * 91.0 79.6 87.3 88.5 268 85.9 88.9 82.5 83.6 96.0 186 90.6 97.7 82.5 83.6 96.0 263 * 79.2 84.0 74.2 87.4 173 93.1 87.9 83.9 78.5 85.5 301 * 97.0 89.4 86.7 89.0 184 * 86.2 80.5 86.9 70.9 TOTAL 342.1 722 .6 663.9 670.2 690.5 MEAN 85.5 90.3 83.0 83.8 87.1 SE 4.6 2.2 1.1 1.8 2.8 * Values greater than 100%, excluded from estimation of mean. UNIVERSITY OF IBADAN LIBRARY - 256 - TABLE 29 DRY MATTER INTAKE (g/day/W^73) The Test of the Differences of the means of dry matter intake hy sheep fed rations C, D, E and F. A N I M A L S PERIOD 1 2 3 4 SIM MEAN 1 C: 68.08 D: 72.96 E: 62.00 F: 78.20 281.24 70.3 2 D; 83.82 E: 72.54 F: 62:14 Cr 65.79 284.29 71.1 3 Es 75.94 F: 80.04 C: 49.21 D: 72.40 277.59 69.4 4 F: 69.82 C: 61.85 D: 42.13 E: 65.23 239.03 59.8 SUM 297.66 287.39 215.48 281.62 1082.15 MEAN 74.4 71.8 53.9 70.4i i SUMMARY BY TREATMENT C D E P* SUM 244.93 271.31 275.71 290.20 1082.15 MEAN 61.2 67.8 68.9 72.6 ANOVA SOURCES df SS MS F Tab. F.05 TOTAL 15 1771.12 PERIOD 3 336.54 112.18 5.44* ANIMALS 3 1043.45 347.82 16.88** 4.76 TREATMENT 3 267.43 89.14 4.32 ERROR 6 123.70 20.61 Differences of means significant for round (P0.05). TABLE 47 BLOOD UREA-N (mg/lOO ml) The Testing of the differences of the means of the blood urea levels (mg/lOO ml) of sheep fed rations C, D, E and F. AROVA SOURCES df SS MS F Tab. F.05 TOTAL 15 56.11 ANIMALS 3 3.48 1.16 1.22 PERIOD 3 25.64 8.55 9.00* 4.76 TREATMENT 3 21.27 7.09 7.46* ERROR 6 5.72 0.95 __ _____«J i Differences of means significant for both period and Treatment (P<_0.05). UNIVERSITY OF IBADAN LIBRARY - 267 “ ■TABLE 48 DUNCAN MULTIPLE RANGE FOR TESTING DIFFERENCES OF MEANS (l) Digestibility of nitrogen (ft). Treatment being Range between means No. of means L S R compared F and C 76.5 - 65.5 = 15.0 4 4.6 F and D 76.5 - 69.4 = 6.9 5 4.5 F and E 76.5 - 71.0 = 5.5 2 4.4 E and C 71.0 - 65.5 = 7.7 5 4.5 E and D 71.0 - 69.4 = 1.6 2 4.4 D and C 69.4 - 65.5 = 6.1 2 4.4 Treatments: F E D C Mean 76.5 71.0 69.4 65.3 (il) Nitrogen intake (d/day/Wk^g*’^^) F and C 1.55 - 0.84 = 0.71 4 0.35 F and D 1.55 - 0.91 = 0.64 5 0.35 F and E 1.55 - 1.28 = 0.27 2 0.34 E and C 1.28 - 0.84 = 0.44 3 0.35 E and D 1.28 - 0.91 = 0.57 2 0.34 D and C 0.91 - 0.84 = 0.07 2 0.34 Treatments: F E D C Means: 1 55 1.28 0.91 0.84 Means joined by the same under-line not significant (P> 0.05) UNIVERSITY OF IBADAN LIBRARY 268 TABLE 48 CONTD. (ill) Nitrogen digested (g/day/wj^^) J U.._.u L , L . » . - ¥ Treatment being compared Range between means No. of means L S R P and C 1.20 - 0.54 = 0.66 4 0.28 P and D 1.20 ~ 0.65 = 0.57 5 0.27 P and E 1.20 - 0.91 = 0.29 2 0.26 E and C 0.91 - 0.54 = 0.37 3 0.27 E and D 0.91 - 0.63 = 0.28 2 0.26 D and C 0.63 - 0.54 = 0.09 2 0.26 Treatments: F E D C Means: 1.20 0.91 0.63 0.54 (IV) Nitrogen retention (fo) P and C 69.5 - 57.5 = 12.0 4 7.6 P and D 69.5 - 63.6 = 5.9 3 7.5 P and E 69.5 - 66.5 = 5.0 2 7.2 E and C 66.5 - 57.5 = 9.0 3 7.5 E and C 66.5 - 57.5 = 2.9 2 7.2 D and C 63.6 - 57.5 = 6.1 2 7.2 Treatments: F E D C Means: 69. 5 66.5 63.6 57.5 Means joined by same under--line not significant (P> 0.05) UNIVERSITY OF IBADAN LIBRARY ~ 269 - TABLE 48 CONTI) Treatments: F E D C Means: 1.11 0.86 0.59 0.48 (Vi) Digestibility of the N of concentrates Cc5 and C„2 53.5 - 68.3 = 20.2 4 6.7 C,5- and C,3 88.5 - 78.7 = 9.8 3 6.6 C3,- and C.4 88.5 - 80.6 = 7.9 2 6.3 and 80.6 - 68.3 = 12.3 3 6.6 C.4 and C,3 80.6 - 78.7 = 1.9 2 6.3 and Cg 78.7 - 68.3 = 10.4 2 6.3 Treatments: 5 4 ° 3 C2 Means: 88.5 80.6 78.7 68.3 Means joined by same under-line not significant (P^ 0.05) (VIl) Total ruminal N (mg/lOO ml) F and C 128.9 - 44.8 = 30.1 4 44.3 F and D 124.9 - 68.4 = 56.5 3 43.6 F and E 124.9 - 78.2 = 46.7 2 42.1 E and C 78.2 - 44.8 = 33.4 3 43.6 E and D ! 78.2 - 68.4 = 9.8 2 42.1 D and C I 68.4 - 44.8 = 23.6 2 42.1 Treatments: F E D C Means: 124.9 78.2 68,4 44.8 Means joined by same under-line not significant (P^0.05) UNIVERSITY OF IBADAN LIBRARY - 2 7 0 - TABLE 48 CONTD. (VIIl) Ruminal protein N (mg/lOO ml). — Treatments being compared Range between means No. of means L S R P and C 79.1 - 51.5 = 47.6 4 25.4 P and D 79.1 - 50.6 = 28.5 5 2 5 .0 P and E 79.1 - 55.9 = 25 .2 2 24.2 E and C 55.9 - 51.5 = 24.4 5 25.0 E and D 55.9 - 50.6 = 5.5 2 24.2 D and C 50.6 - 51.5 = 21.1 2 24.2 Treatments: F E D C Means: 79.1 55.9 50.6 51.5 (IX) Ruminal non-protein N (mg/lOO ml). F and C 45.8 - 15.5 = 52.5 4 25.0 P and D 45.8 - 17.8 = 28.0 5 22.6 F and E 45.8 - 22.5 = 25.5 2 21.9 E and C 22.5 - 15.5 = 9.0 5 22.6 E and D 22.5 - 17.8 = 4.5 2 21.9 D and C 17.8 - 15.5 = 4.5 2 21.9 • K - , Treatments: P E D C Means: 45.8 22.5 17.8 15.5 Means Joined by same under-line not significant (P>0;05). UNIVERSITY OF IBADAN LIBRARY 271 TABLE 48 CCNTD. ( x ) Ruminal ammonia N (mg/lOO ml) Treatments being compared Range between means No. of means L.S R F and C 4.50 - 1.80 = 2.70 4 0.87 F and D 4.50 - 2.37 = 2.13 3 0.86 F and E 4.50 - 3.83 = 0.67 2 0.83 E and C 3.83 - 1.80 = 2.03 3 0.86 E and D 3.83 - 2.37 - 1.46 2 0.83 D and C 2.37 - 1.80 = 0.57 2 0.83 Treatments: F E D C Means: 4.50 3.83 21.37 1.80 (Xl) Blood urea N (mg/lOO ml) F and C 4.88 - 1.90 = 2.90 4 2.07 F and D 4.88 - 2.60 = 2.28 3 2.04 F and E 4.88 - 3.92 = 0.96 2 1.97 F and C 3.92 - 1.90 = 2.02 3 2.04 E and D 3.92 - 2.60 = 1.32 2 1.97 D and C 2.60 - 1.90 = 0.70 2 1.97 - - Treatments s F E D C Means; 4.88 3.92 2.60 1.90 Means joined By same under-line not significant (P>0.05). UNIVERSITY OF IBADAN LIBRARY - 272 - TABLE 48 CONTD (XIl) Ruminal residual N (mg/100 ml) Treatments being compared Range between means No. of means L S R P and C 40.2 - 11.6 = 28.6 4 20.9 P and D 40.2 - 15.2 = 2 5 .0 3 20.6 F and E 40.2 - 18.7 = 21.5 2 19.9 E and C 18.7 - 11.6 = 7.1 3 20.6 E and D 18.7 - 15.2 = 3.5 2 19.9 D and C 15.2 - 11.6 = 3.6 2 19.9 Treatments: F D D C Means• 40.2 18.7 15.2 11.6 (XIIl) - amino N (u mole/ml) P and C 6.29 - 2.31 = 3.98 4 2.62 P and D 6.29 - 3.02 = 3.27 3 2.58 F and E 6.29 - 4.79 = 1.50 2 2.49 E and C 4.79 - 2.31 = 2.48 3 2.58 E and D 4.79 - 3.02 = 1.77 2 2.49 D and C 3.02 - 2.31 = 0.7} 2 2.49 Treatments: P E D C Means: 6.29 4.79 3.02 2.31 Means joined by same under-line not significant (P^0.05) UNIVERSITY OF IBADAN LIBRARY - 273 - TABLE 49 DRY MATTER INTAKE. PER CENT CONCENTRATE IN RATION AIR) PER CENT. CRUDE PROTEIN OF THE RATIONS No. Ratio Total Intake Concentrate fo Concentrate f> C.P. (g/d) 179 C 484.8 221.1 45.61 6.49 259 C 667.2 357.2 53.54 6.30 265 D 748.4 407.7 54.47 8.41 186 D 544.3 229 .6 42.19 8.25 263 E 544.3 263.7 48.44 10.31 173 E 714.4 399.7 55.95 10.72 301 F 616.6 378.5 61.38 14.02 184 F 561.3 374.2 66.66 14.57 — 1------ ' 2ND PERIOD 179 D 654.9 409.3 62.34 8.51 259 D 697.4 408.3 58.54 8.46 268 E 646 .4 408.3 63.16 11.11 186 E 578.3 408.2 70.59 11.51 263 F 612.4 408.2 66.66 14.57 173 F 663.4 408.3 61.54 14.04 301 C 518.8 280.7 54.10 6.29 1184 C 387.0 297.7 76.92 ___5_._6_9_ ! Per cent Concentrate in the Ration. Mean values: lTION MEAN VALUE B 35.9 + 14.1 c 58.5 ± 9.2 D 58.2 + 12.8 E 62.7 + 7.6 F 62.9 ± 5.0 UNIVERSITY OF IBADAN LIBRARY 274 TABLE 49 DRY MATTER INTAKE. PER CENT CONCENTRATE (CONTINUED) [Nos. Ratio Total Intake Concentrate fo Concentrate fo C.P j : - . (g/a) 3RD PERIOD 179 E 603.9 408.3 67.61 11.35 259 E 731.4 408.2 55.81 10.72 268 F 722.9 408.2 56.47 13.52 186 P 688.9 408.2 59.26 13.80 263 C 353.0 238.2 67.47 5.94 ! 173 C 680.4 408.2 60.00 6.141 301 D 603.9 408.3 67.61 8.58: {510 D 374.3 374.2 73.33 8.6. 6. (i I 4TH PERIOD t 179 P 561.3 408.2 72.72 15.18! 259 P 697.4 408.3 58.54 13.73i 268 C 701.7 408.2 58.18 6.19 j 186 C 514.6 268.0 52.07 6.34 263 D 425.2 306.1 72.00 8.63 173 D 421.0 148.8 35.35 8.15 301 E 523.0 344.4 65.85 11.25 184 E 489.0 344.4 70.43 11.51. fo Concentrate in the Rations. RATION B ------------- Animal Nos. Ration j Total D ^ Intake Concentrate fo Concentrate 179 B 510.3 272 .1 653.33 259 B 536.4 134.1 2 5 .0 0 268 B 408.2 236.5 57.94 186 B 536.4 178.8 33.33 263 B 522.8 180.5 34.52 j 173 B 500.1 183.0 36.59 | 301 B 680.4 249.0 36.59 184 ____L _ _ 543.2 54.7 10.07 UNIVERSITY OF IBADAN LIBRARY - 275 - TABLE 50 H E M L/C LIVE WEIGHT OF THE WEST AFRICA! DWARF SHEEP DURING THE DIGESTION EXPERIMENTS TRIAL ONE Period Animal Nos. Weight (kg) kg 179 15.88 7.53 259 21.77 9.48 268 19.96 8.89 1 186 19.50 8.74 263 22.68 9.76 173 26.31 10.88 301 13.15 6.56 184 17.69 8.14 179 15.88 7.53 259 21.77 9.48 2 268 19.96 8.89 186 19.05 8.60 263 20.86 9.18 173 26.31 10.88 301 14.51 7.05 184 17.69 8.14 UNIVERSITY OF IBADAN LIBRARY - 276 - TABLE 50 MEAL LIVEWEIGHT OF THE ANIMALS (KG) TRIAL WO Period Animal Nos. Weight (Kg) nJ.734 ! kg 179 15.42 7.37 259 21.77 9.48 268 20.86 9.18 1, 106 18.60 8.45 263 21.32 9.33 173 26.31 10.88 301 14.51 7.05 184 17.69 8.14 179 14.51 7.05 259 21.32 9.33 268 19.96 8.89 2 186 17.24 7.99 263 23.59 10.05 173 24.95 10.47 301 13.15 6.56 184 15.42 7.37 UNIVERSITY OF IBADAN LIBRARY 277 - TABLE 50 M E M LIVE WEIGHT OF THE ANIMALS (Kg) ? — Period Animal Nos. Weight (W) Kg. ¥0.734kg ! 179 16.78 7.84 259 22.68 9.77 268 20.41 9.04 3 186 19.05 8.60 263 22.22 9.62 173 26.76 11.02 184 17*24 7.99 179 17.24 7.99 259 23.59 10.05 268 22.68 9.77 4 186 19.50 8.74 263 21.32 9.33 173 26.31 301 16.33 7.68 184 16.78 7.84 ; UNIVERSITY OF IBADAN LIBRARY TABLE 51 INTAKEOUTPUT AND APPARENT DIGESTIBILITY OF NITROGEN AND NITROGEN RETENTION OF SHEEP 9g/day) MAINTAINED ON BASAL HAY AND CONCENTRATE SUPPLEMENTS Animal Ration Intake-N Faecal-N g/day Urinary-N Digestibility Retained-N Milk-N 186 A 6.2 2.3 2.8 62.9 1.1 210 A 6.2 1.9 1.6 69.3 2.7 259 F 10.1 2.0 4.7 79.2 3.4 273 P 10.1 2.5 4.4 75.2 3.2 72 P 10.1 2.5 4.4 75.2 3.2 2.1 90 P 10.1 2.0 5.4 79.2 2.7 0.8 l ........... - ......... 278 UNIVERSITY OF IBADAN LIBRARY - 279 - TABLE 52 ENRICHMENT (E) OF RUMINAL AMMONIA. BLOOD UREA. BACTERIA AND PROTOZOA AFTER ADMINISTRATION OF 15n int o tee h d m e n or bl o o d of w e t h e r she ep No No No No 210 273 186 259 atoms EXCESS OP 15n \ 2.577 2.473 0 .322 0.075 j RUMINAL 1.291 1.290 0.288 0.060 AMMONIA 0.442 0.440 0.261 0.050 0.278 0.275 0.259 0.043 0.234 0.235 0.196 0.041 0.677 0.323 2.647 1.905 BLOOD 0.568 0.251 1.939 1.269 UREA 0.476 0.130 1.600 0.829 0.386 0.128 1.357 0.692 0.324 0.086 1.220 0.509 0.161 0.105 0.620 0.665 0.080 0.055 0.069 0.056 URINE 0.043 0.056 0.456 0.436 0.398 0.331 BACTERIA 0.336 0.275 0.275 0.247 0.251 0.243 0.240 0.229 0.220 0.210 PROTOZOA 0.209 0.201 0.190 0.189 1 0.174 0.165 j------------ UNIVERSITY OF IBADAN LIBRARY - 280 TABLE 53 ENRICHMENT (e ) OF URINE,, FAECES AND MILK AFTER ADMINISTRATION OF 15 N INTO THE RUMEN OF THE EWES MAINTAINED ON BASAL HAY AND CONCENTRATE SUPPLE­ MENTS , Day No. 90 A T O M ^ E X C E !3 S O F 15N 1 0.189 0.188 2 0.071 0.084 URINE 3 0.039 0.047 1 0.158 0.065 2 0.040 0.070 FAECES 3 0.078 1 0.032 0.047 MILK 2 0.037 0.053 0.043 0.032 0.030 MILK SECRETION IN DICTATING SHEEP No. 72 D A Y JI----------- 1 2 6. . . . . . . . . . . 3 4 5 7 Milk (ml) 330 330 150 220 240 200 150 gN 2.98 2.98 1.35 1.99 2.17 1.81 1.35 No. 90 Milk (mj) 100 60 60 60 70 70 60 gN 0.88 0.84 0.74 0.86 0.71 0.98 0.61 UNIVERSITY OF IBADAN LIBRARY CM C~- • O - 281 - TABLE 54 PER CENT N. CM AND CR IN DIGESTA OF THE WEST AFRICA!? DWARF SHEEP Proxina Distal Sheep No. Feed Omasum Ahomasun Snail Snail Re-ctum! Intes­ Intes­ t i n e t i n e fo N 1.23 1.54 1.26 2.17 0.84 1.27 336 foOM 92.50 93.40 92.40 89.60 89.60 90.40 foCR 0.097 0.175 0.230 0.149 0.230 0.208 fo N 0.76 1.89 1.19 1.82 1.12 1.96 314 f OM 93.50 90.30 92.40 88.60 90.00 83.80 fo CR 0.072 0.175 0.145 0.145 0.215 0.218 fo N 1.04 1.82 1.05 2.45 1.05 2.11 343 fo CM 92.77 93.90 94.20 88.60 89.90 89.40 fo CR 0.104 0.215 0.185 0.190 0.223 0.358 fo N 1.32 2.38 1.61 1.40 1.26 1.33 399 f OM 9.304 95.30 88.60 88.00 88.00 90.80 fo CR 0.086 0.160 0.160 0.177 0.145 0.254 — f> N 1.67 1.67 1.40 2.03 1.19 2.32 499 f OM 93.01 91.80 77.30 84.60 • 88.60 88.40 fo CR 0.072 0.074 0.043 0.066 0.120 0.296 fo N 1.98 3.78 : 2.66 2.31 1.47 2.38 510 fo OM 93.32 90.59 j 87.30 j 86.00 87.50 86.60 fo CR 0.072 9.137 0.160 | 0.152 0.200 0.370 1 UNIVERSITY OF IBADAN LIBRARY - 282 - TABLE 54 PER CENT IT. OM AND CR IN DIGESTA OF THE WEST AFRICAN DWARD SHEEP (CONTINUED) — n 1 Proxima DistalSheep No. Peed Cnasun Abonasun Snail Snail Rect-uia; Intes­ Intes­ ji 1........ tine tine fo N 1.23 1.61 1.82 3.01 1.54 1.47 | 320 f OM 92.50 92.30 92.50 89.70 89.90 92.60 j f° CR 0.097 0.043 0.043 0.045 0.300 0.150 fo N 0.79 2.10 3.57 4.06 1.20 2.00 ! 513 f CM 93.44 89.90 89.20 84.70 85.00 89.00 ■ f CR 0.135 0.205 0.274 0.175 0.420 0.654 |• f N 1.01 2.03 2.66 3.64 1.00 1.86 ! 518 f OM 92.82 93.60 93.00 87.90 87.70 89.00 f CR 0.156 0.240 0.274 0.145 0.310 0.546 f N 1.27 4.55 4.20 5.18 1.40 1.82 484 f OM 92.73 87.20 82.20 87.20 83.30 90.00 f CR 0.190 1.300 0.520 0.215 0.496 0.755 f N 1.40 1.16 4.62 5.60 1.00 2.33 • 358 fo 014 92.69 90.80 90.00 85.00 84.90 87.60 f CR 0.190 0.370 0.320 0.240 0.440 0.926 , fo N 2.15 2.87 2.10 4.06 2.03 2.78 i 570 fo OM 93.51 93.30 69.50 87.80 88.00 89.00 | fo CR ̂- -.. - - •' _ 0.06_7_ 0.024 0.024 0.010 0.430 0.314 !J UNIVERSITY OF IBADAN LIBRARY