Open Collections

UBC Theses and Dissertations

UBC Theses Logo

UBC Theses and Dissertations

Effect of three rations on blood metabolites in pregnant ewes Ross, James Pelter 1967

Your browser doesn't seem to have a PDF viewer, please download the PDF to view this item.

Item Metadata

Download

Media
831-UBC_1967_A4 R6.pdf [ 5.78MB ]
Metadata
JSON: 831-1.0104368.json
JSON-LD: 831-1.0104368-ld.json
RDF/XML (Pretty): 831-1.0104368-rdf.xml
RDF/JSON: 831-1.0104368-rdf.json
Turtle: 831-1.0104368-turtle.txt
N-Triples: 831-1.0104368-rdf-ntriples.txt
Original Record: 831-1.0104368-source.json
Full Text
831-1.0104368-fulltext.txt
Citation
831-1.0104368.ris

Full Text

THE EFFECT OF THREE RATIONS ON BLOOD METABOLITES IN PREGNANT EWES by JAMES PELTER ROSS B.S.A., University of British Columbia, 1964 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE IN AGRICULTURE in the Division of Animal Science We accept this thesis as conforming to the required standard THE UNIVERSITY OF BRITISH COLUMBIA January, 1967 In presenting t h i s t h e s i s i n p a r t i a l f u l f i l m e n t of the requirements f o r an advanced degree at the University" of B r i t i s h Columbia, I agree that r.he L i b r a r y shall, make i t f r e e l y a v a i l a b l e f o r reference and study,, I f u r t h e r agree that permission f o r extensive copying of t h i s t h e s i s f o r s c h o l a r l y purposes may be granted by the Head of my Department or by h i s representatives. I t i s understood that copying or p u b l i c a t i o n of t h i s t h e s i s f o r f i n a n c i a l gain s h a l l not be allowed without my w r i t t e n permission. The U n i v e r s i t y of B r i t i s h Columbia Vancouver 8, Canada ^ t e JtQgtch • £2^J3L2. - ABSTRACT The feeding of three different rations varying mainly in crude fat and crude fiber level at a high level of intake to pregnant and non-pregnant ewes had a number of effects on blood glucose, lactic acid, acetone plus acetoacetate, and total volatile fatty acid concentra-tions during the last three months of gestation for the pregnant animals (February, March, and April). There was no significant effect due to time on blood glucose level or on the level of blood ketone bodies, during the latter stages of pregnancy, for animals consuming high, medium or low roughage rations. The intramuscular injection of 1 mg./kg. body weight of growth hormone into pregnant sheep for 7 days during the last month of pregnancy, caused no significant effects on blood glucose, lactic acid, acetone plus acetoacetate, or total volatile fatty acids concentrations. ACKNOWLEDGEMENTS I am p a r t i c u l a r l y grateful to Dr. W. D. K i t t s , Professor of Animal Science, for h i s guidance and ins t r u c t i o n throughout the course of t h i s study. Sincere thanks are also extended to Dr. B. A. Eagles, Dean of the Faculty of Agriculture and Chairman of the Division of Animal Science for allowing the use of the necessary f a c i l i -t i e s for t h i s study. I also wish to express my gratitude to Mrs. J. Lit s k y for assistance i n laboratory work. Appreciation i s also expressed for the assistance of Miss 6. Wilson. LIST OF TABLES TABLE I II III IV i V VI VII VIII IX XI XII XIII xiv xv XVI XVII A XVII B XVII c XVIII Estimated requirements for glucose i Composition of ration 1 Composition of ration 2 Composition of ration 3 Summary of results for Part I A summary of proximate analysis data obtained for the three rations Proximate analysis of collected feces Feed consumed and feces excreted during digestibility trials Calculated digestibility of feed proximate fractions Comparison of results obtained by different sampling methods The effect of time after feeding on blood glucose and lactic acid concentration Recovery of glucose added to venous blood i Recovery of lactic acid added to venous blood i Recovery of acetone added to venous blood Recovery of volatile fatty acids added to venous blood Rate of distillation of volatile fatty acids Record of blood metabolites for animals on ration 1 Record of blood metabolites for animals on ration 2 Record of blood metabolites for animals on ration 3 Summary of results of growth hormone treatment PAGE 30 39 39 40 46 47 48 49 50 51 52 53 53 54 54 55 56 57 58 73 LIST OF FIGURES FIGURE PAGE I A Blood acetone plus acetoacetate concentrations of non-pregnant sheep for February, March and A p r i l 60 I B Blood acetone plus acetoacetate concentrations of pregnant sheep for February, March, and A p r i l 60 I I A Blood glucose concentrations of non-pregnant sheep for February, March and A p r i l 62 I I B Blood glucose concentrations of pregnant sheep for February, March and A p r i l 62 I I I A Blood t o t a l v o l a t i l e f a t t y acid concentrations of non-pregnant sheep for February, March and A p r i l 64 I I I B Blood t o t a l v o l a t i l e f a t t y acid concentrations of pregnant sheep for February, March and A p r i l 64 IV A Blood l a c t i c acid concentrations of non-pregnant sheep for February, March and A p r i l 66 IV B Blood l a c t i c acid concentrations of pregnant sheep for February, March and A p r i l 66 LIST OF APPENDICES APPENDIX PAGE I Record of Lamb Weights 95 II Record of Lambing 96 III Data Obtained in Growth Hormone Study 97 INTRODUCTION The metabolism of sheep and c a t t l e has been extensively studied i n the past two decades. The research into ruminant metabolism has been conducted mainly for the purpose of improving knowledge of the factors involved i n animal production. The majority of the studies which have been conducted have been for the purpose of elucidating the mechanisms involved i n the major ruminant metabolic disorder; that of ketosis. Considerable economic loss i s d i r e c t l y a ttributable to ketosis i n a l l liv e s t o c k producing areas. The extensive studies of ruminant metabolism have resulted i n an increased understanding of the significance of many of the factors important i n ketosis and also i n the accumulation of considerable informa-t i o n of value i n other aspects of livestock production. For example, a much more precise estimation of the nutrient requirements of sheep and c a t t l e has been made possible, p a r t i c u l a r l y for pregnant and la c t a t i n g animals, by studies carried out i n r e l a t i o n to ketosis. Although the precise mechanism or mechanisms involved i n the development of ketosis i n ruminants i s s t i l l obscure the use of radioisotopes has greatly f a c i l i t a t e d and accelerated the study of ruminant metabolism. In the past, ruminant ketosis was attributed to a glucose d e f i -ciency, but the numerous factors which might precipitate t h i s condition were not known. Gradually many of the factors involved i n ketosis became known. Although a carbohydrate deficiency i s s t i l l generally assumed to exist during ketosis i n the ruminant, the significance of t h i s deficiency i s s t i l l not f u l l y understood. A considerable amounts of work has therefore been done i n an attempt to determine the biochemical changes i n various tissues which accompany a carbohydrate deficiency. Although therapeutic measures have been quite successful i n the treatment of bovine ketosis, much less success has been obtained i n attempts to treat ovine ketosis. Even when i t has been possible to successfully treat ovine ketosis the cost has often been prohibitive. Considerable work has been done i n recent years which indicates a carbohydrate deficiency such as that occurring i n ketosis could be prevented by the feeding of certain types of rations. These include rations high i n concentrate^rations which have been f i n e l y ground,or rations i n which the concentrate portion of the rat i o n has been steam heated. These rations afford a r e l a t i v e l y high proportion of digestive end-products which give r i s e to glucose following absorption. In certain cases these rations have been shown to cause blood glucose values to be elevated above normal. This study was conducted to determine the effect of different rations on the changes i n blood metabolites which occur i n late-pregnant ewes. I t has been shown that normal late-pregnant ewes have blood changes which are similar to those seen i n ketotic late-pregnant ewes. However the changes seen i n the normal ewes are much less accentuated than the changes seen i n the ketotic ewes. The effect of the r a t i o n fed, on the changes i n normal late-pregnant ewes blood was studied i n an attempt to assess the prophylactic value of high concentrate rations. LITERATURE REVIEW 1. I. Introduction Pregnancy toxemia and acetonemia are metabolic disorders that have many features in common. Since these disturbances occur at different times in the reproductive process, i t appears reasonable to deal with them separately. Pregnancy toxemia usually occurs during the final weeks of the gestation period and most frequently is seen in ewes that are twin-pregnant. The symptoms of pregnancy toxemia have been well documented (16, 185, 177). There is some variation in the syndrome shown, and in many cases no visible characteristics are apparent during the early stages of the disease. When the disorder has become more advanced the symptoms may include inactivity, anorexia, faulty equilibrium, grinding of teefn, and a characteristic odor of acetone on the breath. If the disease is terminal as i t may be within 2 to 5 days of the first symptoms, the animal will go into a comatose state which is followed by death. Convulsions and blindness may also be present in the final stages. Most of the symptoms associated with pregnancy toxemia are typical of a generalized disorder of the central nervous system. Similar to pregnancy toxemia, acetonemia is characterized by symptoms of a generalized disorder of the central nervous system. Many of these symptoms are similar to those of pregnancy toxemia. For example, anorexia, incoordination, and an acetone odor on the breath are symptoms common to both disorders. Unlike pregnancy toxemia, however, acetonemia occurs post-partum, usually during the first 2 weeks of lactation. When acetonemia occurs there is a rapid loss of body weight and a marked drop in milk production. 2. A number of gross changes take place in the blood which are common to both pregnancy toxemia and acetonemia. Concomitant with these diseases are hypoglycemia and hyperketonemia, the extent of which is dependent on the severity of the disease (16). Both changes become more accentuated with increasing severity of the disease. Other changes which occur include acidosis with decreased plasma carbon dioxide transporting ability and elevated levels of non-protein nitrogen (16). These changes and others indicate that in both ketotic conditions there is an increased mobilization and catabolism of protein and fat. This is considered to occur in response to a carbohydrate Insufficiency (185). The changes in concentration of metabolites in the blood are reflected in elevated urine levels of ketone bodies and non-protein nitrogen (185). Similarly, the milk of a cow affected with the disease may be tainted with the flavor of ketone bodies. As a result of the elevated concentration of acetone in the blood, the odor of acetone may be detected on the breath of a ketotic animal. Post-mortem examination of tissues from animals which have died from pregnancy toxemia or acetonemia indicates changes in response to severe stress. The lesions found include atrophy of the anterior lobe of the pituitary gland, hypertrophy and patchy degeneration of the adrenal cortex, involution of the thymolymphatic system, acute involution of the pancreas, fatty infiltration of the liver, and very low levels of liver glycogen (16). 3. II. Main Theories Proposed The causes for acetonemia and pregnancy toxemia have not as yet been shown to be greatly different. The following is a general discussion of the theories proposed to explain these ketotic conditions. Although some workers have preferred to emphasize specific factors as responsible for ketosis (43, 192, 166, 173), others have emphasized the diverse number of causes that may be important with respect to the incidence of ketosis (99,185,21). As a result, the theories for ketosis have been numerous without any single theory gaining complete acceptance. Certain concepts of ketosis have gained at least general accept-ance. Many workers since the original work of Hupka (84), have demonstrated hypoglycemia during ketosis. It is generally accepted that hypoglycemia during ketosis results from a deficiency of carbohydrate (99, 62, 49, 173, 183). A number of concepts have been established which illustrate the rigorous carbohydrate economy of the adult ruminant (11, 162, 5). Very l i t t l e of the carbohydrate requirement of the ruminant has been shown to be met by the direct absorption of glucose from the gut. It is necessary, therefore, for the adult ruminamt to maintain its carbohydrate supply by gluconeogenesis from such possible glucose precursors as propionic acid, protein, or the glycerine portion of fat (15, 2, 30, 88). I t is also recognized that under certain conditions carbohydrate requirements may be increased considerably. This is true for any stress situation and is particularly important during the onset of lactation in the cow. The fetal drain of hexose is largely responsible for the increased glucose requirements of the pregnant ewe, while for the cow the lactational drain is responsible for the increased requirement. 4. If gluconeogenesis from glucose precursors plus glucose released from liver glycogen are unable to meet the Increased demand for glucose, or are unable to sustain an increased supply of glucose, then hypoglycemia results. If euglycemia is not maintained as in the case of hypoglycemia then an alternate supply of metabolites is made available to the body tissues. This alternate supply of metabolites is provided by the mobilization of body fat. The free fatty acids which are mobilized are taken in by muscle and liver tissue cells and oxidized for energy. Fatty acid catabolism in the liver gives rise to ketone bodies which cannot be utilized by liver tissue as rapidly as they are formed. Consequently the ketone bodies pass into the circulation and become available to the peripheral body tissues. The ketone bodies so provided are metabolized in the peripheral tissues and may provide a considerable portion of the animal's energy requirements under ketotic conditions (33, 53). Ketone bodies are normally present in the blood of the adult ruminant at levels significantly higher than for the monogastric (99), which indicates the contribution ketone bodies normally provide to the energy metabolism of the ruminant (33, 111). Although the concept of carbohydrate deficiency during ketosis has been generally accepted, there is a much less agreement on the manner in which this carbohydrate deficiency comes about or on the metabolic changes caused by a carbohydrate deficiency. Shaw et al.(177, 173) have suggested that a hypophyso-adrenal mechanism is responsible for the carbohydrate deficiency during ketosis. Basically the theory states that during periods of stress the pituitary-adrenal system may become exhausted resulting in reduced gluconeogenesis. When this occurs a marked glucose deficiency results in ketosis. This theory has also been advocated by Carlstrom (47). However, i f there is a pituitary-adrenal deficiency during ketosis, this 5. deficiency i s only a r e l a t i v e deficiency i n the sense that during ketosis the glucocorticoid output by the adrenals may be inadequate, but the output of glucocorticoids by the adrenals i s s t i l l much above normal. Lindner (111) has shown that blood C o r t i s o l levels are much above normal i n sheep with ketosis and Robertson et a l . (154), have shown blood corticosteroid levels to be elevated during ketosis i n c a t t l e . Shaw (173) has also attributed the development of ketosis to f a u l t y Carbohydrate metabolism but has shown normal glucose tolerance curves for ketotic cows. Kronfeld (99) found that glucose tolerance tests on ewes with pregnancy toxemia showed a greater u t i l i z a t i o n rate of glucose than normal. In recent years the effects of a carbohydrate deficiency on metabolism during ketosis has been considered largely i n r e l a t i o n to the t r i c a r b o x y l i c acid (TCA) cycle (15). The metabolite considered to be of pr i n c i p a l importance for the normal operation of the TCA cycle has been oxaloacetic acid (43, 15). Oxaloacetic acid can be used for glucose or protein synthesis and i t i s also the condensing partner of acetyl-coenzyme A required for the operation of the cycle. During l a t e pregnancy i n the ewe and early l a c t a t i o n i n the cow the heavy demands for glucose and i n t e r -mediates for i t s sythesis may res u l t i n a decrease i n TCA cycle operation. If a decrease i n TCA cycle operation occurs, then a decrease i n the oxidation of acetate to carbon dioxide and water would be expected. A reduction i n the rate of oxidation of acetyl-Co A by TCA reduces the rate of removal of acetyl-Co A formed by the catabolism of f a t t y acids; acetyl-Co A accumulates, condenses, and thereby increases the rate of formation of ketone bodies. Before ketosis w i l l occur the increased rate of ketone body formation by the mechanism described above must resu l t i n the formation of 6. ketone bodies at a rate greater than they can be utilized by the peripheral tissues (32, 62). Bach and Hibbitt (22) have suggested that bovine ketosis results because of an interference in the TCA cycle caused by a block in the conversion of pyruvate to citrate and other reactions involving oxidative decarboxylation. This conclusion was based on serum concentrations of these metabolites. They suggested that the utilization of coenzyme A for mammary lipogetaesis from acetate reduces the availability of coenzyme A for oxidative decarboxylation. In an attempt to support this theory, cysteamine was injected into cows and a 100% success of the treatment was reported. However other workers have been unable to confirm this finding (99). A number of criticisms have been made of the theory of reduced TCA cycle activity during ketosis (15). Since oxaloacetic acid is regen-erated at each turn of the TCA cycle, only catalytic amounts of oxaloacetic acid would be required for TCA cycle operation. Also, since extrahepatic ketone body oxidation is not impaired during ketosis a deficiency of oxaloacetic acid must occur in the liver. However, Ford (65) found no difference in t he oxaloacetic acid levels of liver tissue between ketotic 14 and normal sheep. The injection of acetate-C into one quarter of the udder of ketotic and normal cows showed that milk products derived from the TCA cycle were greater for the ketotic animal than for the normal one (103). Kronfeld et aL (103) have suggested a lipogenic block is respon-sible for the reduced synthesis of fatty acids from acetate in the mammary gland of the ketotic cow. This reduced fatty acid synthesis enhances the formation of ketone bodies from acetate. 7. Trombropoulos and Kleiber (192) have attributed the llpogenlc block suggested by Kronfeld et a l . to a depressed rate of generation of reduced coenzymes necessary for f a t t y acid synthesis. The reason offered for the depression of reduced coenzyme generation was attributed to a decrease i n glucose oxidation by the pentose cycle. However, Kronfeld et al.(105) have since shown that i n the mammary glands of ketotic cows there i s a depression i n the levels of a l l forms of the coenzymes necessary for f a t t y acid synthesis, including both reduced and oxidized forms. Seekles (166) has shown a r e l a t i o n between the incidence of ketosis i n dairy cows and the lev e l of butyric acid i n certain forms of feedstuffs p a r t i c u l a r l y grass silage. Presumably the butyric acid i s metabolized by the rumen epithelium giving r i s e to appreciable quantities of ketone bodies (112, 29). The observations of Seekles have been confirmed by other workers (127, 3). Seekles speculated that when large amounts of butyric acid were absorbed, an impaired l i v e r oxidation resulted i n ketosis. However, the explanation of Seekles does not seem compatible with accepted pathways of ketone body u t i l i z a t i o n or with the carbohydrate deficiency theory of ketosis. /' / 8. III. Experimental Approach to Ketosis, There are many factors involved in the development of a ketotic condition in ruminant animals. The study of ketosis has been directed towards gaining an understanding of the importance of these various factors and towards making known^factors not yet recognized. The discussion which follows deals with much of the research that has been done in this regard. A. Spontaneous and Induced Ketosis. The etiology of ketosis would be greatly facilitated i f the experimenter could induce or simulate ketosis experimentally. Therefore a considerable amount of time has been devoted to the study of methods whereby ketosis may be induced. For the induction of ketosis to be successful, i t is necessary to establish metabolic conditions which precede the occurrence of this condition. Since a carbohydrate deficiency has been assumed to be a concomit-ant of ruminant ketosis, attempts to produce ketosis experimentally by starving have been made. Starvation eliminates.the source of carbohydrate precursors normally absorbed from the gut. Initially, glucose can be provided to the blood by a break-down of liver glycogen. Within a short period of time however, the liver glycogen stores become depleted. As a result, a l l of the carbohydrate requirement must be supplied by gluconeogenesis, particularly from protein. The mobilization of body fat is also a source of nutrients under these conditions. It is the produc-tion of ketone bodies during the catabolism of fat that is responsible for the hyperketonemia during starvation. Depending on the duration of starvation, and on the nutrition of the animal prior to starvation, there 9. will be varying degrees of hyperketonemla. Shaw (173) and Shaw et al,(177) have indicated that acetonemia cannot be induced early post-partum merely by fasting. Although fasting did produce marked hypoglycemia, hyperketonemla, low liver glycogen and fatty livers in more than 30 cases studied, there was an absence of the typical clinical symptoms of acetonemia. Shaw (173) terms the ketosis that occurs when an animal is consuming feed normally as primary or spontaneous ketosis and al l other forms as secondary ketosis. Bergmann (135) has pointed out that starvation of lactating cows reproduced some of the blood biochemical changes of acetonemia, but clinical symptoms were lacking. However, Bergmann (135) has indicated that starvation of late twin-pregnant ewes has been at least partially successful for inducing pregnancy toxemia in that, i f the disease occurred, the clinical symptoms and necropsy findings as well as the blood changes appeared indistinguishable from spontaneously occurring pregnancy toxemia. Weeth (197) induced ketosis in pregnant ewes by simulating a starving snowbound stress condition. Leng (110) found that the concentra-tion of yg -hydroxybutyrate in the blood was dependent on the rate of entry of /9 -hydroxbutyrate into the circulation both for pregnant sheep which had prolonged undernutrition followed by a short period of starvation and for ewes with pregnancy toxemia. This work suggests a metabolic similarity between pregnancy toxemia and starvation in the pregnant ewe. Fraser et al»(69) attempted to determine the effect of plane of nutrition on the severity and incidence of ketosis. Generally i t was found that the incidence and severity of ketosis was greatest with the 10. lower plane of nutrition. Other methods which have been used in an attempt to produce ketosis in ruminants have involved starvation combined with some additional treatment. Goetch (71) found that whereas fasting alone lowered blood glucose levels and increased blood ketones, treatment of fasted pregnant ewes with alloxan resulted in a much greater increase in blood ketones. Clinical signs of ketosis, however, were observed in the group that was fasted, but not in the group that was fasted and treated with alloxan. Procos (140) tested the effect of alloxan injected into Merino wethers and found that this led to a substantial increase in blood glucose and ketone body levels. Dye et aX, (57) also used alloxan in an attempt to produce ketosis in sheep. There was, however, only a mild hyperketonemia without clinical symptoms. Goetch (71) used a combination of fasting and phloridzin administered to pregnant and non-pregnant ewes. Ewes treated in this manner developed marked hypoglycemia within four hours of phloridzin administration and clinical signs were observed in one out of two animals so treated. A number of experiments were conducted in an attempt to produce both clinical and biochemical symptoms of bovine ketosis (184). The treatments imposed were phloridzin and phloridzin plus 800 units of insulin. All treatments were applied following periods of fasting which ranged from 24 to 96 hours. It was found that phloridzin did not cause sufficient stress to cause clinical symptoms of ketosis. There were only moderate changes in glycemia and ketonemia. When insulin injections were combined with phloridzin treatment there was a marked decrease in blood glucose but no clinical symptoms of ketosis were observed. 11. Kronfeld ( 9 6 ) has speculated that excessive endogenous growth hormone may"exaggerate a carbohydrate deficiency during periods of stress and thus cause ketosis. It has also been speculated that ruminants are selec ted for rapidity of body growth, fetal growth, and milk production, a l l of which are enhanced by anterior pituitary growth hormone. Therefore, the production of large amounts of endogenous growth hormone may be selected for indirectly. Kronfeld (100) injected crystalline growth hormone intramuscularly in doses of 50 mg, to 100 mg. for 4, 10, and 15 days into 3 sheep during the last month of pregnancy. The effect on blood glucose was an increase of about 12 mg/100 ml. for 5 days but then a decline to normal occurred despite the continuance of injections. Blood ketone bodies rose 3 to 7 mg/100 ml. after 3 to 5 days but about 5 days later the ketone body concentration became lower or erratic. Kronfeld speculates that normally pregnant sheep can compensate for excess growth hormone, so that excess endogenous growth hormone is unlikely to be of any major significance in pregnancy toxemia. Kronfeld (100, 9 9 ) also attempted to induce bovine ketosis during early lactation usimg growth hormone. In one experiment ( 9 9 ) , 500 mg, of growth hormone were Injected intramuscularly. The result was an increase in daily milk produetion from 53 lb. to 59 lb. and also ketonuria and hyperketonemia, but there was no other clinical symptoms of ketosis. In an attempt to produce the clinical symptoms of ketosis, 0.5 unit/kg. body weight of protamine zinc insulin was injected daily into lactating cows. However, there was no ketosis and only slight hypoglycemia. By increasing the dose to 1.0 trait/kg. body weight given twice daily, hypoglycemia and 12. clinical signs of ketosis were obtained but there was no hyperketonemla. In another experiment, Kronfeld (100) injected 1 mg./kg. body weight of growth hormone intramuscularly into 2 cows daily for 10 days, starting 2 weeks after parturition. In both animals plasma nonesterifled fatty acids and acetone plus acetoacetate level increased during the period of growth hormone administration and dropped when the treatment ceased. 14 By injecting acetate-G intracisternally, the incorporation of acetate into milk fat during growth hormone induced ketosis was measured. Similar to spontaneous ketosis the acetate uptake into milk fat was decreased during growth hormone induced ketosis. Although the clinical symptoms of ketosis could not be induced by this method, Kronfeld specula-ted that excessive growth hormone secretion may be of importance in the pathogenesis of spontaneously occurring bovine ketosis. Adler (1), in support of the observations of Seekles (166), found 27 clinical cases of ketosis in cows fed grass silage. Adler, however, found that the ketone body content of the silage, rather than the butyric acid content was high. This observation by Adler is in part supported by the work of Bergmann (33), who infused large quantities of acetoacetate intravenously to sheep thereby causing ketosis. Possibly the most successful method for inducing acetonemia in dairy cows is the method which has been used by Hibbitt (77) and by Baird (23). In both reports ketosis was produced in dairy cows under stall fed conditions by a combination of L-thyroxine injections and excessive protein feeding. L-thyroxine was given In 4 to 5 dfl»ses of 25 mg. to cows which had just reached maximum milk production. The ketosis produced by 13. this method, while possibly differing in some degree from primary spontaneous ketosis, appears to have many characteristics in common with the spontaneous condition. There was a drop in feed intake and a decline in milk yield with a rapid deterioration of body condition. There was also an increase in serum ketone body concentration to values seen during spontaneous ketosis. Baird has shown that during thyroxine induced ketosis the liver remains very ketogenic which is not considered to be the case in spontaneous ketosis (173, 98). Baird (23) has speculated that a possible explanation for the finding that perfused liver from cows suffering from spontaneous ketosis was not ketogenic is that this tissue may have been obtained from animals which have developed the condition some time previously. Possibly, when ketosis is prolonged the liver becomes moribund. Hibbitt (77) has also pointed out that the time of the thyroxine injection and the high protein levels in the ration are necessary for this method of induction to be successful. With respect to the induction of ketosis in dairy cows i t is interesting to consider the work of Reece (143). It was found that feeding 15 gm of thyroprotein to cows past their lactational peaks for various periods of time (3 to 8 mos.),caused an average increase in milk production of 24.8 per cent with an in i t i a l increase in milk fat. Reece also fed 15 gm of thyroprotein daily to 4 cows with ketosis and fotmd an increase in milk production and a removal of the ketotic condition. 14. B. The Effect of Ration and Digestive End Products on Blood Glucose and Ketone Bodies. Since certain of the digestive end-products have been shown to affect both glycemia and ketonemia, the extensive studies of such effects, have been carried out in relation to ketosis. Quantitatively the major end-products of digestion are the result of carbohydrate fermentation in the rumen. This gives rise to short chain volatile fatty acids, mainly acetic, propionic, and n-butyrlc acids. Small amounts of lactic and formic acids may also be absorbed from the rumen of animals fed certain rations (18). There has been some consideration given to propanediol which is produced in the rumen in small amounts (188). In simple-stomached animals, the most important carbohydrate metabolite is glucose. Glucose is readily absorbed from the digestive tracts of these animals. This is reflected by large post absorptive increases in glycemic levels. Glucose is probably an equally important metabolite in the metabolic processes within the ruminant body. Little glucose is normally absorbed from the digestive tract of the ruminant, yet i t can be demonstrated experimentally that glucose is readily absorbed from the intestinal tract. Schambye (16) has provided supporting evidence that only small amounts of glucose are absorbed from the alimentary tract of ruminants. By examining blood of the venous portal system and comparing i t with that of arterial blood i t was found that the level of glucose was never significantly different. Weller and Gray (16) found that when 148 gm. of starch was fed to sheep daily, only 7.8 gm. of the starch passed through the rumen. Allcroft (5) found that feeding high carbohydrate meals to ruminants caused small but variable changes in blood glucose level. It has also been generally accepted that monosaccharides present 15. in rumen fluid are acted on rapidly by rumen microorganisms giving rise to organic acids, thus preventing the absorption of any appreciable amount of carbohydrate. A number of concepts have been established pertaining to the metabolism of the volatile fatty acids following absorption (49,18). It has been fairly well established that the large amounts of acetate provided by rumen fermentation are utilized for energy purposes and for the synthesis of fat. Although acetate can be incorporated into glucose the net synthesis of glucose from acetate is not considered to occur within the body. Propionate, on the other hand, is a recognized glucose precursor and is considered to be important in ruminant gluconeogenesis. It has been estimated that the propionate available from rumen fermentation can, at least in theory, supply the glucose not accounted for by gluconeogenesis from protein (49). Butyric acid is s t i l l controversial with respect to its ability to contribute to the endogenous synthesis of glucose. There is as yet no adequately supported pathway of butyrate oxidation which would by-pass the formation of acetate, and yet a gluconeogenic effect of injected butyrate has been demonstrated in both lactating cows and sheep. It has been speculated by Armstrong (18) that the glucogenic effect may be the result of butyrate in some way sparing glucose. These general concepts of volative fatty acid metabolism have gained support from in vivo studies done by numerous workers. Goetch (72) orally administered sodium salts of acetic, propionic, and bu£yric acid to phloridzin treated non-pregnant ewes. Sodium acetate did mot alter the hypoglycemia and only slightly increased blood ketone levels. Sodium propionate rapidly elevated the blood glucose level and maintained the 1 6 . blood glucose level at normal or above normal for 24 hours post-treatment. Bipod ketonemla decreased, and the Intensity of the Induced glycosuria was* markedly Increased during the subsequent 48 hour period. Sodium butyrate administered in this way markedly increased blood ketone levels and significantly reduced blood glucose levels. There was also a depression of the urinary levels of glucose, urea nitrogen, and total urine nitrogen. Armstrong (19) infused volatile fatty acids; acetic, propionic and butyric, into the rumen of sheep for 48 to 72 hour periods, and the effects were measured on certain blood and urine constituents. The infusion of acetic acid caused a rise in blood total volatile fatty acids and ketone body concentration as well as a f a l l in glycemic level. The administration of propionic acid by rumen infusion for 48 hours resulted in a marked increase in blood sugar, a drop in blood ketonemla and a reduction in urinary nitrogen excretion. The administration of butyric acid caused hyperketonemia and a decrease in blood glycemia. Barcroft et aU (16) first demonstrated that the volatile fatty acids of ruminal fermentation are rapidly absorbed from this organ into the portal venous system. The metabolism of absorbed volatile fatty acids by ruminal epithelium was first indicated by Kiddle .et aL^ when i t was shown that the concentration of butyrate in blood draining the rumen was appreciably less than that expected on the basis of the decline in rumen concentration. This finding led to extensive studies on the metabolism of volatile fatty acids by ruminal epithelium. Pennington (131) conducted a number of in vitro studies with rumen epithelial tissue from sheep. One finding of importance relative to the problem of ketosis was that the uptake, of acetate and butyrate was 17. accompanied by ketone body production. Annison et atl. (13) stated that the branched chain fatty acids, valeric and isovaleric gave rise to ketone bodies when metabolized in the rumen epithelium. The significance of ketone body production from branched chain fatty acids is considered to be much less important than the production of ketone bodies from butyrate since the branched chain fatty acids are only produced in relatively .small amounts in the rumen. Propionate was found to diminish the amount of ketone bodies produced by the rumen epithelium. Similar results were found with OK rumen epithelium (11). Examination of portal and arterial blood taken simultaneously from normal sheep has indicated that ketone body levels are 1 to 2 mg/100 ml* higher in venous blood draining the rumen than in arterial blood supplying the rumen (16). Since portal blood flow is in the range of 1 to 2 liters per minute (16), i t is obvious that ketone body production by rumen epithelium may have considerable significance with respect to ketosis. It is also obvious that a considerable portion of the butyrate absorbed from the rumen is metabolized by the rumen wall. Seto (168) estimated that over half of the butyrate absorbed is converted to ketone bodies by the rumen epithelium. Hird (79) on the basis of isotope distribution, estimated that about three quarters of the butyrate carbon chains absorbed are converted to ketone bodies by the epithelium of the rumen and omasum. Jackson (85) has recently found that the in. vitro metabolism of palmitate, stearate, oleate, and5linoleate, the major constituents of ruminant depot fats, by rumen epithelium gives rise to the ketone, acetoacetate. The significance of this finding as related to ketosis is not as yet known. 18. Lactic acid has been shown to be produced in the rumen in signifi-cant amounts under certain conditions (173), particularly when there is a marked increase in the soluble carbohydrate of the ration, such as in ,changing from a diet of hay to grain. Feeding of large amounts of lactate caused a marked increase in blood lactate and glucose (173). Although i t has not been successfully demonstrated there is sufficient experimental evidence to indicate that certain rations may be effective in preventing and also in alleviating ketosis. Certain rations give rise to relatively large amounts of glucogenic end-products such as propionic acid (89, 174), whereas s t i l l other rations give rise to relative-ly large amounts of ketogenic end-products (89). Shaw (174) has shown that the molar proportions of the volatile fatty acids produced in the rumen can be regulated, to a remarkable extent by means of changes in the ration. In this regard, high concentrate and low roughage diets are found to produce relatively large amounts of propionic acid during rumen fermentation. However, Schmidt (163) found that"'3 different levels of grain feeding had l i t t l e affect on blood glucose or ketone body levels in lactating cows. All of the factors involved in affecting the ratio of rumen volatile fatty acids are not known. Similarly, the factors involved in ration effects on blood glucose and ketone bodies are not completely understood. Jorgensen et aj«(89), however, have shown elevated blood glucose levels and depressed blood ketone levels associated with high rumen propionic acid levels in lactating dairy cows. As pointed out by Shaw (174) and Jorgensen (89), when rumen propionic acid levels are high in lactating dairy cows there may be a marked decrease in fat content of the milk. Therefore, maintaining propionic acid levels high, as a prophylactic measure 19. against ketosis, may have certain disadvantages. Shaw (169) found that the greatest and most rapid decrease in the fat content of milk was obtained by a ration of finely ground hay and steam-heated corn. It appears as though more research is needed before the prophylactic value of adjust-ing rumen volatile fatty acid proportions can be assessed. Shaw (174) has stated that there is l i t t l e evidence to indicate a relation between the level of protein in a ration and the incidence of ketosis. Vigue (193) studied the rations of cows affected with ketosis and found that most had high calorie and low protein feeds. It was also found that there was frequently inadequate protein feeding after calving. Weeth (197) found that when sheep which had ketosis induced by starvation were placed on high protein feeds they recovered quickly. The recent work of Baird (23) and Hibbitt (77) on the induction of ketosis suggests that high protein feeding may be a stress factor significant in the etiology of ketosis. Although a fat condition is generally considered to be undesir-able in late pregnant ewes because of a greater propensity for ketosis (16, 53), Fraser (69) has found that over-feeding ewes to fatness did not increase the incidence of ketosis. The effect of grass silage on the incidence of ketosis has been considered in relation to the work of Seekles (166) and Adler (3). Adler found that the highest incidence of ketosis in dairy cattle was in late winter and early spring, which was related to the feeding of silo contents. In accord with this work,Potts (136) found that the ketone bodies in grass silage were highest in January and February. Seekles (166), however, attributed the ketosis resulting from the feeding of grass silage to the high level of butyric acid which i t contained. 20. Todd (191) fed a silage of grass and clover, which had been fermented at low temperatures and which contained large amounts of butyric acid, to lactating dairy cows. There was an increase in blood ketone body level of 5 out of 6 cows fed the silage and 2 of the cows become ketotic. There has been very l i t t l e evidence to suggest either, a mineral deficiency or a vitamin deficiency as being responsible for ketosis. Blackburn (40) tested high and low levels of mineral on the .incidence of ketosis but did not find any significant differences. Aspiotis (21) has speculated that trace element deficiencies and certain B-avltaminosis may be responsible for ketosis but evidence is lacking to corroborate these suggestions. C. The Effects of Hormones oh Ketosis 1. Insulin.' The mode of action of the hormone is well known (53, 62). Insulin acts to bring about a decrease in blood glucose concentration. It is considered to act on tissue cell membranes to facilitate the move-ment of glucose'into the body tissues. In muscle insulin acts to enhance glycogen formation, glucose oxidation and fat formation. It also acts to inhibit the release of non-esterifled fatty acids into the blood plasma. In liver insulin favors glucose uptake and lipogenesis. Due to the actions of insulin, this hormone is considered to be anti-ketogenic. It has been shown that insulin has slightly different effects in the ruminant than in the simple stomached animal (112). The rate of f a l l of blood glucose in ruminants following the injection of insulin is slower than the corresponding rate for non-ruminants. This may be 21. d i r e c t l y due, i n part, to a diminished action of i n s u l i n on muscle tissue. Also the symptoms of hypoglycemia following the i n j e c t i o n of i n s u l i n are delayed longer i n ruminants than i n non-ruminants. Comparatively large doses are required to produce i n s u l i n shock i n ruminants, and shock i s manifested only .when blood sugar levels between 10 and 20 mg.7-100 c"c. have been maintained for 5 to 8 hours (51). i ^ . " Since ketosis i n ruminants i s most frequently accompanied by hypoglycemia, t h i s , suggests insu l i n ' can Induce membrane changes. : Kronfeld (99) has speculated that since i n s u l i n favors an increase i n glucose metabolism and i s generally anti-ketogenic; hypoglycemia caused by i n s u l i n would probably hot favor the development of ketosis. However the a n t i -ketogenic potential of i n s u l i n may be reduced during carbohydrate deficiency. Bergmann (32) measured the anti-ketogenic action of i n s u l i n by measuring i t s effect on the rate of acetoacetate production and consequently the concentration of acetoacetate i n the blood plasma f e l l . Kronfeld (99) has measured the rate of glucose transfer from blood plasma to tissue c e l l s and has found that the transfer rate is not increased during ketosis. Yet, i f r e l a t i v e l y large amounts of i n s u l i n were injected the transfer rate was increased. Also, i n s u l i n can be used to induce hypoglycemia as well as a l l of the c l i n i c a l signs of ketosis except hyperket-onemla. These findings further suggest hypoinsulinism during ketosis. Because of t h i s i t seems that hypoinsulinism would favor the development of a ketotic condition i n the animal. 22. The major problem in determining changes in plasma insulin level associated with ketosis has been a methodological problem. A major advance has been the recent development of reliable immunochemical assay for plasma insulin. However the limit of sensitivity for this method corresponds very closely with the normal insulin level of the bovine. As a result hypoinsulinism is not detectable in the bovine. Kronfeld (99) has speculated that possibly a form of diabetes is a primary factor in the development of pregnancy toxemia. It was found that under certain conditions pregnancy toxemia will develop without hypoglycemia. It is interesting to note that Goetch (71) had less success inducing pregnancy toxemia in alloxan-diabetic ewes that were starved than in ewes that were simply starved. Shaw (173) has suggested that the level of blood glucose is not important in acetonemia because some cases of this disorder have been encountered in which there were normal blood glucose levels. These cases of acetonemia may in fact be diabetic conditions. The suggestion has been made that hyperinsulinism may be present during ketosis with a metabolic block acting as an antagonist to insulin, preventing the utilization of glucose by the tissue cells (99). The amounts of insulin-like activity extracted from plasma of non-pregnant cows and sheep generally indicated plasma-insulin levels of 100 to 500ymu/ml of plasma (50). It has'not been possible to show a correlation between plasma glucose and plasma-insulin levels (50). Leng (110) found that the plasma-insulin activity of 2 out of 4 pregnant toxemic sheep was very high. Calhoun (44) measured the serum insulin-like activity of normal, fasting and ketotic cows. It was found that the insulin level of ketotic cows 23. was lower theni normal and that fasting caused a decrease i n serum Insulin-l i k e a c t i v i t y . There has been some ind i c a t i o n of a hypoglycemic action of ketone bodies i n ruminants, both d i r e c t l y (32) and i n d i r e c t l y (191, 166). The i n vivo effects of hyperketonemia on carbohydrate metabolism have recently been characterized i n the dog (122, 122A, 115). I t was found that infusing ketone bodies into dogs caused a 60 per cent decrease i n hepatic glucose output and a 40 per cent decrease i n peripheral glucose u t i l i z a t i o n (122). The plasma mom-esterified f a t t y acid levels declined s i g n i f i c a n t l y when /S -hydroxybufcyric acid or acetoacetic acid were infused. Mebane (122) has suggested that the q u a l i t a t i v e s i m i l a r i t y between the effects of ketones and i n s u l i n not only i n hepatic glucose output and blood glucose levels but also on plasma non-esterifled f a t t y acid concentration indicates that ketones act by stimulating the secretaion of endogenous i n s u l i n . Maddison (115) found that infusions of /?-hydroxybutyric and acetoacetic acids resulted i n a 200 per cent increase i n the concentration of i n s u l i n i n the pancreatic venous blood. I f a similar mechanism operates i n the ruminant, i t may have considerable significance i n the etiology of ketosis. 2. ACTH (Adrenocorticotropic Hormone) and Glucocorticoids Although ACTH can cause an increase i n the rate of production of ketone bodies (28) t h i s ketogenic action i s normally much less s i g n i f i c a n t than i t s glucogenic action mediated by the glucocorticoids. C o r t i s o l , by stimulating gluconeogenesis from amino acids and i n h i b i t i n g t r i g l y c e r i d e uptake, has an anti-ketogenic influence on the l i v e r . C ortisol acts not only to stimulate gluconeogenesis from amins|acids but also has a catabolic action on protein. I t has been shown that glucocorticoids act on existing 24. enzyme systems to direct metabolites toward carbohydrate formation (142). Carlstrom (47) and Shaw (173) suggested that there is a reduced adrenal cortex function during acetonemia and consequently there is a decrease in gluconeogenesis and an increase in ketogenesis. Johnson ( 88 ) speculated that the conversion of protein to glycogen may be defective. However a defect seems unlikely since gluconeogenesis can be stimulated by exogenous glucocorticoids. The adrenal cortex will increase its produc-tion of glucocorticoids during conditions of stress such as that of ketosis. The increase in glucocorticoid production is stimulated by ACTH which causes hypertrophy and hyperplasia of the adrenal cortex. Dye (56) considered adrenal hypertrophy to become pathological in extreme cases, such as in severe acetonemia resulting in a fatal condition. Shaw (173) found that cows with ketosis generally required more glucocrticoids than normal animals on the basis of physiological response. He considered that this finding might explain a relative adrenal insufficiency in ketosis. However, no differences were found in response to cortisone acetate. An endocrine theory has been proposed in which a relative adrenal insufficiency conditioned by hypothyroidism is considered to occur during ketosis. It was found that the plasma corticosteroids were at a signifi-cantly higher l e v e l and the protein bound iodine level significantly lower in the ketotic than in the normal cow (154). Leng (110) found that there was no apparent relation between plasma C o r t i s o l concentration and blood glucose level in sheep. However, Lindner (111) found that C o r t i s o l levels in pregnancy toxemia were abnormally high. Kronfeld (99) has postulated that C o r t i s o l may inhibit glucose uptake by sensitive cells, such as brain 25. cells and the situation is somewhat like steroid diabetes. Yet cortisone administration exerts a beneficial effect on ketotic cows. It was also considered (99) that the adrenal corticoids were only relatively insufficient since plasma Cortisol and the urinary corticoid output were both increased during ketosis. 3. Growth Hormone Although certain actions of growth hormone on general metabolism have been established, the precise significance of these effects on the overall metabolism is poorly understood. This is particularly true in the adult animal. The injection of growth hormone into rats causes a low respiratory quotient presumably associated with a greater oxidation of fat. The suggestion has been made that growth hormone may have an effect on mobilization of fat and the release of non-esterifled fatty acids from adipose tissue (112). Growth hormone generally promotes synthetic processes especially protein synthesis, and reduces gluconeogenesis from amino acids (53). This action coupled with the depression in oxidative processes caused by this hormone might result in growth hormone being ketogenic. This has been verified in simple stomached animals since large amounts of exogenous growth hormone have been used to induce ketosis. A diabetogenic effect has been demonstrated for growth hormone (199). Under certain conditions, i t may cause cellular proliferation and / ^ - c e l l degranulation associated with an increased rate of insulin secretion. Growth hormone is antagonistic to insulin in that i t raises the level of blood glucose by inhibiting glucose oxidation (53). Despite this antagonistic effect to insulin growth hormone requires the presence of insulin to produce anabolic effects in the rat (112, 125). Because of this action of growth hormone on synthetic processes, i t is antagonistic to cortisone and Cortisol. Recently i t has 26. been demonstrated that hypoglycemia is a potent stimulus to the release of growth hormone in the human (156). Certain aspects of the actions of growth hormone have been considered in relation to ruminant metabolism and ketosis. It has been shown that ruminants depend on the lower chain fatty acids for much of their energy supply. Presumably these animals are adapted to the metabolism of acetate and must have a high capacity for oxidizing 2-carbon fragments. It has been speculated that the supply of acetate may be in-creased through the action of growth hormone, and this may be a means of meeting an increased metabolic demand (112). Possibly growth hormone tends to regulate the energy supply from fat in a similar manner to the regulation of carbohydrate metabolism by insulin. Lactating dairy cows were found to have an average of 50 per cent increase in plasma free fatty acids following the administration of growth hormone (204). Also it was found that growth hormone Increased the rate of acetate turnover. In the ruminant the sum of the actions of growth hormone may not always be ketogenic. This was found by Kronfeld in late pregnant sheep (96). However, Kronfeld demonstrated a definite ketogenic effect in lactating dairy cows, (100) but the magnitude of this ketogenic effect was not sufficient to induce acetonemia i n cows treated with growth hormone. In summary, growth hormone may have an important role in ruminant energy metabolism, but its significance to the etiology of ketosis in ruminants remains to be demonstrated. Insulin, ACTH and glucocorticoids, and growth hormones have been studied most extensively in relation to ruminant ketosis. However, the sparse and sometimes conflicting results which have been obtained make 27. it obvious that further study is needed before the significance of these hormones can be ascertained in relation to the problem of ketosis. Other hormones which may be of importance in the etiology of the disorder have received less attention than the hormones previously mentioned. These include such hormones as thyroxine, epinephrine, estradiol-17- ^  , estrone, progest-erone, glucagon, and possibly the mineral corticoids. Some of the generally recognized actions of these hormones would seem to implicate their involve-ment in the etiology of ketosis. 4. Epinephrine Hypoglycemia stimulates the adrenal medulla to increase its rate of epinephrine release which in turn causes the activation of a phosphorylase in the liver thus stimulating the breakdown of glycogen and the release of glucose into the circulation. During excitement epinephrine may be released from the adrenal medulla in such amounts as to cause marked increases in blood glucose. Epinephrine is known to enhance the breakdown of muscle glycogen. Shaw (176) has found that epinephrine in the normal cow stimulates AGTH release by the anterior pituitary. 5. Thyroxine Generally i t has been found that most hormones are dependent on an adequate level of thyroxine for their actions. This finding is consis-tant with the view that thyroxine is the primary regulator of metabolic processes in the body. Baird (23) and Hibbitt (77) induced a ketotic condition in dairy cows by means of exogenous thyroxine. Reece (143), on the other hand, successfully treated ketosis in dairy cows by feeding thyroprotein. Because of the relatively large amounts of thyroxine used in these works, i t is unlikely that such findings have much meaning in 2 8 . the etiology of the disease. Plasma protein bound iodine values in ketotic cows were found to be lower than for normal cows (154). Robertson (154) suggested that ketosis may be caused by a relative adrenal insufficiency. It was postulated that elevated glucocorticoid levels during stress conditions may cause hypothyroidism, and a failure in the utilization of glucocorticoids results. Blood leucocyte changes during ketosis have been recorded by a number of workers (173, 56, 47). The blood cell concentrations observed during ketosis have been quite variable. Despite the variability there has been some attempt to correlate these changes during ketosis with certain hormonal changes. Particularly with changes in plasma concentrations of thyroxine and glucocorticoids (56, 47). However, since the mechanisms Involved in the regulation of the concentrations of the various blood leucocytes are poorly understood, any conclusions reached appear to be only speculation. D. Other Metabolic Changes Accompanying Ketosis. In recent years a large amount of research has been directed at determining metabolic changes accompanying ketosis in the ruminant. Since normal metabolism has been only poorly understood, i t has been necessary to make a detailed study of normal metabolism for the purposes of comparison. These studies have attempted not only to determine concentrations of meta-bolites in different tissues, but also to determine the rates of utilization and formation of metabolites by means of radio-isotopes. Glucose because of its apparent central role in ketosis, is one such metabolite which has been extensively studied. Although l i t t l e glucose is absorbed from the gut of the ruminant,it has been found that i f glucose 29. turn-over rate, or the amount of glucose supplied or utilized per unit of time, is expressed in relation to surface area, then the values for the cow and the sheep are not greatly different from those of the dog or rat (112). This may be due to a dependence by the central nervous system on glucose .for metabolism. The glucose and oxygen uptake by the sheep brain was measured in vivo, and i t was estimated that i f a l l of the glucose was oxidized, this would account for the whole of the oxygen consumption by the brain (112) • It was not possible to demonstrate acetate uptake by the brain and therefore acetate is probably not an alternate to glucose for brain nutrient supply in the ruminant. These findings are in accord with the general concept that many organs including the heart and skeletal muscle can derive energy from fatty acids or ketone bodies as an alternative to glucose^whereas the nervous tissue and possibly blood cells have a primary requirement for glucose. In the pregnant ewe, the nervous tissue of the ewe receives competition for circulating glucose by the developing fetus, and this may, under certain conditions, jeopardize the l i f e of the ewe. A similar situation arises in early lactation in the cow. However, in this case, the mammary gland is in competition with the nervous system. In the latter case, the incidence of death is much lower, since a drop in milk production may result in restoring of carbohydrate balance. In the former case, If the carbohydrate imbalance is marked, only abortion will restore the carbohydrate balance and prevent death. The blood glucose concentration of the cow will drop slightly at the onset and during the early stages of lactation (83). Similarly, in pregnant ewes, particularly in twin-pregnant ewes there is a drop in blood glucose level in late pregnancy (148, 94). These changes in blood glucose 30. concentration presumably reflect an Increased utilization or demand for glucose. The daily glucose requirements for pregnant and non-pregnant sheep have been estimated using radio-isotope dilution studies and are shown in Table I (18). It appears that sheep weighing; from 50 to 60 kg. may utilize approximately 90 gm. of glucose per day, increasing to 150 or 160 gm. in the later stages of pregnancy. TABLE I Estimated Requirements for Glucose sm/dav gm/ Ks °!75/dav Sheep, non-pregnant (55 kg live weight; plasma glucose 55 mg/100 ml) 89 4.4 Sheep, pregnant (65 kg live weight; plasma glucose 53 mg/100 ml) 156 6.8 Jesse (87) and Shaw (173) have stated that the primary antiketo-genic factors are, high levels of liver glycogen and blood glucose. Shaw (173) has also indicated that rations providing high levels of rumen propionic acid may facilitate the maintenance of high levels of liver glycogen and blood glucose (174). It can be predicted from established metabolic reactions that propionate is convertible into carbohydrate in the animal body, however, the extent of this process in the ruminant is not definitely known. Ash (20) conducted infusion experiments with carboxyl-labelled propionate using sheep and found that very l i t t l e of the labelled carbon was incorporated into glucose. It was thus concluded that the hyperglycemia produced by propionic acid was not due to gluconeogenesis from this acid. An alternate explanation for the hyperglycemic effect of propionic acid was not offered. 31. Possibly propionic acid causes hyperglycemia by a glucose sparing effect. There has been a comparison of certain parameters of glucose metabolism between ketotic and normal ruminant animals. The pool size , and rate of turnover under various conditions have been measured i n some studies. Bergmann (30) studied non-pregnant, pregnant, and fasted pregnant ewes. The mean glucose pool sizes were 157 mg./kg. body weight for the fed non-pregnant ewes and 127 mg./kg. body weight for the fed pregnant ewes. Kronfeld (106) found that during pregnancy toxemia i n the ewe the glucose pool size and turnover rate became less than h a l f of normal non-pregnant ewes and less than a quarter of normal late pregnant ewes. Bergmann (30) found that the glucose turnover rates were 0.22 gm./hr./kg.0' body weight for non-pregnant ewes which had been fed, and 0.15 gm./hr./ 0 75 kS* * body weight for fasted non-pregnant ewes. The values for glucose turnover rates of normal ewes were for only 60 to 80 per cent of the pregnant ewes. Kronfeld (103) has made a comparison of normal and ketotic cows with respect to size and turnover rate of the glucose pool. I t was found that the size and turnover rate were about one-fifth greater i n the ketotic than i n the normal cow. The hypoglycemia which accompanies acetonemia was attributed to an expansion of glucose space. A lipogenic block theory of ketosis was discussed previously i n r e l a t i o n to reduced coenzymes necessary for fat synthesis (104). This work was supported by Trombropoulos (192) who correlated the decrease i n fat synthesis with a decrease i n glucose oxidation by the hexose monophosphate pathway. This work was i n turn supported by Armstrong (18). There has 32. been studies done indicating that glucose enters the TCA cycle i n abnormally large amounts as acetyl-Co A during ketosis i n the cow (37), whereas i n normal animals l i t t l e glucose carbon enters the t r i c a r b o x y l i c acid metabolic pathway by acetyl-Co A. The metabolism of the ketone bodies, mainly p> -hydroxybutyrate, acetoacetate, and acetone, has also been studied rather extensively. Bergmann (33) measured the turnover rate of acetoacetate i n ketotic and normal sheep. I t was found that i n normal sheep the turnover rate was 0.04 gm./hr/kg. body weight, however, i n ketotic sheep the turnover rate could be increased by as much as 10 f o l d . In both normal and ketotic ewes about 50 per cent of the acetoacetate used was oxidized to carbon dioxide. The per cent of the t o t a l respired carbon dioxide derived from acetoacetate metabolism increased from about 2 per cent i n normal ewes to 30 per cent during ketosis. Therefore the contribution of ketone bodies to energy metabolism increases greatly during ketosis. I t has also been shown that the rate of acetoacetate metabolism i s proportional to the plasma concentra-t i o n up to about 10 mg/100 ml above which the acetoacetate w i l l not be metabolized at an increased rate (32). As a r e s u l t , i t has been concluded that an overproduction of ketone bodies rather than an impaired u t i l i z a t i o n i s responsible for the elevated blood levels of ketone bodies during ketosis. Leng (110) studied the metabolism of /Q -hydroxybutyric acid i n ketotic and normal sheep and found that the rate of u t i l i z a t i o n was proportional to the plasma concentration up to 10 mg/100 ml. The c o n t r i -bution of p -hydroxybutyric acid to respired carbon dioxide increased from about 6 per cent i n the normal non-pregnant animal to a maximum of about 33. 20 per cent in the starved pregnant animal. The study of the metabolism of ketone bodies in the cow is rather incomplete. Black (37) has studied the metabolism of labelled /6 -hydroxy-butyrate and acetone in ketotic and normal cows. On the basis of the amount of the labelled carbon appearing in respired carbon dioxide and in milk products, in a given time period following ketone injection, i t was con-cluded that during ketosis the utilization of acetone and P -hydroxybutyrate is reduced. The metabolism of acetate has also been the subject of investi-gation in relation to the problem of ruminant ketosis. Ford (66) made a comparison of acetate metabolism between normal and ketotic twin-pregnant ewes. The acetate utilization rates were found to be 4.04 + 0.29 mg./min./ kg. body weight in the normal sheep and 2.07 + 0.11 mg./min./kg. body weight in the ketotic sheep. The determination was carried out 4 hours after feeding. Sabine (157) measured the utilization rate of acetate in normal sheep and found i t was about 5 mg./min./kg. body weight which compares favourably with that found by Ford. Ford (66) found the per cent expired carbon dioxide obtained from acetate was 13.4 + 1.48 for normal pregnant sheep and 8.75+0.83 in the ketotic pregnant sheep. The per cent of the utilized acetate which is oxidized to carbon dioxide was 1.14 + 0.68 for the normal pregnant sheep and 13.3 +2.1 for the ketotic pregnant sheep. Sabine (157) found the pool size and time of turnover for normal sheep to be 6.6 meq/sheep and 1.9 min. respectively. The pool size and turnover time of preg-nant ketotic sheep was not determined. Kronfeld (103) has studied acetate utilization by the mammary glands 34. 14 of normal and ketotic cows. It was found that of the total C dose injected intracisternally as acetate, 14 per cent occurred in the milk fat of normal cows within 48 hours, compared to only about 3 per cent in ketotic cows. The mean standardized specific activity of milk citrate, casein, lactose, plasma glucose, respired carbon dioxide and urine acetone was greater in the ketotic than in the normal cows. Their results suggested that the proportion of acetate metabolized by the TCA cycle is increased, while milk fat synthesis from acetate is impaired during bovine ketosis. This work led Kronfeld to propose a lipogenic block theory for ketosis which has been discussed. Luick (114) compared the fatty acid composition of milk fat between normal and ketotic cows. Although there was a decrease in the average molecular weight of the low molecular weight fatty acids in milk fat of ketotic animals, there was an apparent compensatory increase in the long chain fatty acids; palmitoleic, stearic, and especially oleic acid. Consequently i t was suggested that a specific enzymatic defect is unlikely in the fatty acid formation from acetate in the ketotic cow. A few metabolic relationships have been found in the study of ketosis which may be of importance to the etiology of the disorder. For example, for ketotic cows with a blood ketone body level below 45 to 50 mg./ 100 ml., a statistically significant depression of blood calcium has been found (73). It was suggested that the lowering of blood calcium levels was secondary to a reduced feed intake during ketosis. In experiments conducted with goats which had been made ketotic by fasting or fasting and phloridzin treatment, i t was found that the intravenous infusion of sodium 0-hydroxybutyrate caused a significant decrease in plasma non-esterified fatty acids (123). A direct feedback effect of ketone bodies on fat 35. mobilization was suggested, and the presence of i n s u l i n was considered to be of importance for maximum suppression of non-esterifled f a t t y acid release. A high degree of interchange of ketone bodies has been found i n the blood with f2> -hydroxybutyrate tending to be present i n the highest concentration (188). Various tissues are known to be ketogenic i n the ruminant, and yet the significance of each i n r e l a t i o n to ketosis has not been determined. Baird (23) compared the ketogenic potential of l i v e r and l a c t a t i n g mammary gland s l i c e s from normal and ketotic cows. Cows, which had been treated with thyroxine to induce ketosis, possessed l i v e r tissue of much higher ketogenic potential than normal cows. However cows which were spontaneous ketotic had non-ketogenlc l i v e r s . Mammary gland tissue from ketotic cows was only s l i g h t l y more ketogenic than mammary tissue from normal cows. The acetoacetate l e v e l i n l i v e r s of ketotic sheep did not d i f f e r from normal (65). Smith (178, 179) studied the metabolism of v o l a t i l e f a t t y acids by l i v e r and rumen from ketotic and normal sheep. The re s u l t s indicated that neither of these tissues are i n h i b i t e d i n the metabolism of v o l a t i l e f a t t y acids during ketosis. However, the ketotic l i v e r possessed a higher potential for acetoacetate production. The rumen epithelium from both normal and ketotic sheep exhibited a high potential for ketone body forma-ti o n i n the presence or absence of substrate. 36. IV. Therapy. The treatment of ketosis has been directed at increasing the supply of glucose to the body tissues. The success of t h i s approach has been the most impressive evidence i n support of a carbohydrate deficiency. The methods used to bring about an increase i n body glucose have been quite varied. The i n j e c t i o n of blood glucose was found to maintain blood glucose levels for only a short period of time (176). This has led to the i n j e c t i o n of glucose with adrenal c o r t i c o i d hormones. When 500 ml. of 50 per cent glucose and 50 units of ACTH were injected, the blood glucose le v e l was maintained for a much longer period of time than i f glucose was injected alone (176). The oral administration of large amounts of sugar or mollasses may also be of some use i n elevating blood glucose (87). Even though much of the carbohydrate administered i n t h i s way w i l l be fermented by rumen microorganisms, the r e s u l t w i l l give r i s e to high levels of propionic acid (174) which i s known to have a hyperglycemic effect (72). Sodium propionate given o r a l l y has also been demonstrated to be useful i n the maintenance of blood glucose levels (87,72,164). The use of sodium lactate as a therapeutic agent i n the treatment of ketosis has been studied. It was found that 1.5 to 3.0 l b . of l a c t i c acid as sodium lactate given o r a l l y to cows caused a 20 per cent increase i n blood glucose levels (173). Oxaloacetate and pyruvate given at 57 mg./kg. body weight were both found to be ef f e c t i v e i n r a i s i n g blood glucose and reducing ketonemia (129). Propylene glycol i s another o r a l l y administered glucose precursor which has been used with some success (16, 58). The i n j e c t i o n of 75 to 150 ml. of 20 per cent calcium gluconate has been p a r t i c u l a r l y useful i n treating cases of acetonemia complicated by parturient paresis (153). Chloral hydrate has been of some use i n the treatment of ketosis through i t s hyperglycemic 37. effect (53, 16). The mechanism of action of chloral hydrate i n t h i s respect i s not known. One explanation proposed i s that i t acts to enchance gluconeogenesis (53). Another explanation offered for i t s action i s that i t reduces the rate of oxidation of glucose (183). The use of ACTH and certain of the adrenal corticosteroids has i n many cases proven more effec t i v e than the use of glucose precursors. Cortisone and ACTH were used as effective treatments for over 150 cases of acetonemia, whereas sodium propionate was found to be much less useful (171). Cortisol was found to be more eff e c t i v e than cortisone acetate (70). Goetch (72) found that cortisone acetate was effective i n preventing the occurrence of ketosis but once the condition was present cortisone acetate would not reverse the hypoglycemia and hyperketonemia i n fasting ewes. The treatment of pregnancy toxemia has generally been less successsful than the treatment of acetonemia. 38. MATERIALS AND METHODS Part I1 Feeding T r i a l A. Materials 1. Animals Grade Rambouillet-Romney ewe-lambs, with an average i n i t i a l body weight of 58 l b . , were used i n t h i s experiment. 2. Feed There were 3 different rations employed i n the feeding t r i a l . The composition of these rations i s given i n Tables I I , I I I , and IV of Part I I . Mineral mix, cobalt-iodized s a l t , and water were provided free choice. B. Methods Thirty-three animals were divided into 3 groups of 11 animals per group. A different r a t i o n was fed to each of the 3 groups. The feed intake was recorded da i l y . The animals were weighed weekly. The feed e f f i c i e n c y of the 3 rations was determined both on a pounds of gain per pounds of feed basis, and also on the basis of t o t a l d i g estible nutrients per pound of gain. 39. Part I I D i g e s t i b i l i t y T r i a l s A. Materials 1. Animals Eighteen of the animals used i n the feeding t r i a l were used i n the d i g e s t i b i l i t y t r i a l s . 2. Feed The 3 rations used i n the d i g e s t i b i l i t y t r i a l s were of the following composition: TABLE I I Ration 1 Constituent , ; lbs/Ton Rolled Barley 1635 Dehydrated grass meal 100 Fishmeal 90 Molasses 150 Tricophos (tricalcium phosphate) 15 Iodized s a l t 10 TABLE I I I Ration 2 ;  Constituent , lbs/Ton Rolled Barley 828 Dehydrated grass meal 1050 Fishmeal 45 Molasses 75 Tricophos (tricalcium phosphate) 7 Iodized s a l t 5 40. TABLE IV Ration 3 Constituent lbs./Ton Dehydrated grass meal 1000 Dehydrated alfalfa meal 1000 T Sv3. digestibility Cages During the digestibility trials animals were held in metabolism cages with a floor area of 3.5 f t . by 2.5 ft. Canvas sacking was used to support the animals and hold them in position during feces collection. The feces was collected separate from the urine by means of a screen directed downward towards a collecting tub. B. Methods For each digestibility t r i a l , 3 animals were selected at random from one of the 3 groups mentioned in Fart I. Six digestibility trials were carried out, 2 trials per pen, so that 6 different animals in each group were used. The feces was collected in large tubs covered with aluminum fo i l to prevent drying out of the feces prior to weighing. Since a l l of the animals used in the trials were ewes, the separation of feces from urine presented a problem which was resolved by the use of an angled screen onto which urine and feces would f a l l . The urine passed through the Screen while the feces rolled down the screen into the tubs described. The total feed intake and feces output was measured over a 48 hour period. The feces was collected and weighed 3 times daily. Feed and water were provided free choice. Proximate analysis was carried out on each of the 3 rations by methods outlined in the A.O.A.C. Methods of Analysis (17). In each of the digestibility trials proximate analysis of representative feces samples was conducted (17). The apparent 41. digestibility of the 3 rations was determined. Also the total digestible nutrients of these 3 rations was determined. Part III Hematological Studies of Pregnant and Non-pregnant Ewes A. Materials 1. Obtaining Blood The animals used for this study were pregnant and non-pregnant ewes. The pregnant animals had been bred during the last week of November. The blood obtained for the study was collected either in 10 ml or 35 ml hepa*inized test tubes. The test tubes were vacuum sealed, so that in combination with Vacutainer Needles, No. 1004, very rapid collection of blood was possible. In preliminary studies a cannula was used which could be placed in the jugular vein and used for repeated blood sampling. The cannula consisted of a Rochester Plastic Needle, 16 Ga. by 2.5 in., No. 503, which was sutured in position. 2. Analyzing the Blood The blood was analyzed for glucose,ace-tone plus acetoacetate lactic acid and total volatile fatty acids. B. Methods 1. Animal8 The animals used for this study consisted of 3 groups of 11 animal8 each. These animals had been used in Part I and in Part II as well. Each of the 3 rations which were studied in Part I and Part II were fed to one of the 3 groups free choice with water mineral mix and cobalt-iodized salt provided as well. 42. To obtain a sample of blood, 2 animals i n each group were placed i n large restraining pens situated within the main pen. This made i t possible to sample blood from 6 animals on the same day. The restraining pens prevented the animals held for sampling from becoming excited. These restraining pens had a f l o o r area of 5 f t . by 3 f t . which allowed the animals considerable freedom of movement. The animals were placed i n the restraining cages the day prior to sampling. Feed and water were available to the animals for the entire period i n which they were held i n the restraining pens. 2. Sampling Since excitement i s known to af f e c t certain of the metabolites studied to a marked degree, a considerable amount of time was spent i n an attempt to eliminate t h i s factor. F i r s t , animals were deliberately excited and then a sample of blood drawn and analyzed. Following t h i s , animals were sampled through a jugular cannula. The jugular cannula was placed i n position one or two days i n advance of sampling. The Rochester Needles used as cannulas are designed to be used i n combination with Luer-lok glass syringes which f a c i l i t a t e d sampling. The cannulas could be used repeatedly for up to 2 weeks provided a few drops of heparin solution was injected into the cannula following blood sampling. The cannula and syringe method of sampling did not appear to excite the animals. F i n a l l y , animals were sampled using vacuum tubes and vacutainer needles. Precautions were taken to avoid exciting the animal. I t was found that J>y the l a t t e r method animals could be approached and sampled i n from 40 to 60 seconds. The three methods of sampling were compared and the l a t t e r method was 43. found to be the most satisfactory. The procedure for sampling blood was standardized to prevent extraneous factors from af f e c t i n g the concentra-t i o n of c i r c u l a t i n g metabolites. The animals to be sampled were placed i n restraining cages, and the day following blood was drawn from a l l animals between 1:00 and 1:30 P.M. 3. Analysis Once the samples of blood were obtained they were rapi d l y deproteinated to prevent any metabolite concentration changes such as that due to g l y c o l y s i s by the blood c e l l s . The analysis of the blood was then carried out according to the following procedures: a. Acetone plus acetoacetate The blood concentration of acetone plus acetoacetate was determined by the method outlined by Behre and Benedict{25), i n which V concentration i s expressed as acetone. b. Glucose The glucose concentration of whole blood was determined using the method of Benedict (27). c. Lactic acid Lactic acid concentration of whole blood was determined by the method of Mendal and Goldsheider (124). A modification was found to be necessary. Calcium hydroxide was removed by f i r s t centrifuging the samples and then drawing the supernatant through a m i c r o t i I t e r i n g device. Yale Luer-lok syringes were used to draw the supernatant through a Swinny f i l t e r adapter (XX30 012 00), containing a M i l l i p o r e f i l t e r , Ha 0.45u, 13 mm. This procedure gave clear f i l t r a t e s . d. Total v o l a t i l e f a t t y acids The t o t a l V o l a t i l e f a t t y acids i n whole blood was determined by the method of Bensadoun (29), with some modification*. I t was found that following deproteinization, the a l k a l i n e f i l t r a t e s could be evaporated to dryness at! 30° C under vacuum using the Rot&vapor Model VB50GD. This method was much more rapid than freeze drying and did not reduce the accuracy of the procedure. Also, following d i s t i l l a t i o n the v o l a t i l e f a t t y acids were determined t i t r i m e t r i c a l l y using phenol red as the indicator. I t was found that bubbling with nitrogejn prior to t i t r a t i o n was not necessary provided a blank teas^titrated with every sample. The a c i d i t y of the blank was then subtracted from the a c i d i t y of the sample. The a b i l i t y to recover glutose,lactic acid, acetone, and v o l a t i l e f a t t y acids added to whole blood was determined. Since the procedures for glucose, l a c t i c acid and acetone showed a linear r e l a t i o n between op t i c a l density and concentration of the metabolite, a l l readings of o p t i c a l density were converted d i r e c t l y to concentration by means of a reference l i n e . Part T? The Effect of Growth Hormone on Blood Constituents A. Materials Growth hormone Raben-type was obtained from N u t r i t i o n a l Biochemicals Corporation, Cleveland, Ohio. Peanut o i l was used as a medium for growth hormone i n j e c t i o n . 45. B . Methods Growth hormone was injected at a le v e l of 1 mg./kg. body weight. These injections were given to 6 animals i n each group for 7 consecutive days prior to the f i n a l blood sampling of the animals. A comparison was made between the metabolite concentrations (mentioned i n Part III) of animals treated with growth hormone and those animals not treated with the hotmone. 46. RESULTS AND DISCUSSION Part I Feeding T r i a l The complete record of the weights of the animals used i n the feeding t r i a l over the experimental period are presented i n Appendix I. A summary of the data and calculations for the feeding t r i a l are presented i n Table V. TABLE V J Ration 39 184 1^4.7 88 35 195 1:5.6 70 27 194 1:7.2 59 *TDN or t o t a l digestible nutrients was determined for the rations from data obtained i n the d i g e s t i b i l i t y t r i a l of Part I I . TDN (%) = (protein (%) x d i g e s t i b i l i t y (%))+ (crude f i b e r (%) x d i g e s t i b i l i t y ( 7 . ) ) + 0&FE (%) x d i g e s t i b i l i t y (%)) + 2.25 (crude f a t (%) x d i g e s t i b i l i t y (%)). It was assumed that 1 l h . TDN = 2000 kc a l . of digestible energy. The r e s u l t s shown i n Table V indicate that r a t i o n 1 had the highest e f f i c i e n c y with one pound of gain to 4.7 pounds of feed consumed. Ration 2 was the next most e f f i c i e n t with an e f f i c i e n c y of 1 pound of gain to 5.6 pounds of feed, and r a t i o n 3 the least e f f i c i e n t with an ef f i c i e n c y of 1 pound of gain to 7.2 pounds of feed. The t o t a l digestible nutrients (TDN) were determined for each of the 3 rations on the basis of the d i g e s t i b i l i t y t r i a l s of Part I I . The TDN values for r a t i o n 1, ra t i o n 2, and r a t i o n 3 were 88, 70 and 59 per cent respectively,,and were found to correspond with the feed e f f i c i e n c y values as was expected. Summary of Results for Part I • Avg. Sain (lbs) TDN* T r i a l Avg. Total Total Feed (lbs) % Period (days) (gain (lbs.) Feed (lbs)  1 (barley) 63 2 (barley-grass) 63 3 (grass-alfalfa) 63 47. Part I I D i g e s t i b i l i t y T r i a l s A summary of the proximate fractions per cent determined for the 3 rations studied are given i n Table VI. TABLE VI A Summary of Proximate Analysis Data Obtained for the Three Rations Ration Dry Crude Matter Protein 1 (barley) 88.4 16.3 2 (barley-grass) 93.9 17.0 3 (grass-alfalfa) 92.7 17.8 Proximate Fractions (%) Crude Crude Fiber Fat Ash NFE 7.7 13.7 6.7 55.5 13.1 5.5 7.9 56.6 24.2 4.9 10.7 42.9 The crude protein l e v e l i n the 3 rations was not greatly d i f f e r e n t , the levels ranging from 16.3 per cent for r a t i o n 1 to 17.8 per cent for ratio n 3. The crude f i b e r and crude fat proximate fractions were quite different for the 3 rations. Crude f i b e r contributed 7.7, 13.1 and 24.2 per cent to the dry weight of rations 1, 2, and 3 respectively while crude fat contributed 13.7, 5.5 and 4.9 per cent for rations 1, 2, and 3 respec-t i v e l y . There was l i t t l e difference i n the ash content of the 3 rations, the levels ranging from 6.7 to 10.7 per cent. The nitrogen free extract (NFE). was about the same for r a t i o n 1 and 2 at 55.5 and 56.6 per cent while that for ration 3 was considerably lower at 42.9 per cent. The r e s u l t s of proximate analysis of feces are given i n Table VII. TABLE VII Proximate Analysis of Collected Feces Ration Animal Proximate Fraction (%) Tag Dry Crude Crude Crude No. Matter (%) Protein Fiber Fat Ash NFE 1 (barley) 233 31.6 13.8 27.1 4.5 14.6 40.0 234 26.4 12.2 26.5 4.5 13.3 43.5 226 32.0 14.1 30.5 4.3 13.1 38.0 235 42.7 14.2 29.7 5.6 13.1 37.3 2 (barley-grass) 215 42.8 20.0 23.6 6.4 10.2 39.9 169 34.6 17.4 26.9 7.5 10.6 39.9 207 . 34.3 17.7 25.5 6.1 14.1 36.7 216 35.1 17.4 25,6 6.4 14.0 36.7 3 (grass- 209 36.0 14.1 34.4 5.3 , 12.4 33.8 a l f a l f a ) 156 43.1 15.3 31.9 8.3 12.1 32.5 208 41.5 15.2 29.9 5.4 12.1 37.4 214 25.8 15.1 30.6 6.9 11.1 36.4 The reeord of feed intake and feces collected for each of the d i g e s t i b i l i t y t r i a l s appears i n Table VIII. TABLE VII I Feed Consumed and Feces Excreted During D i g e s t i b i l i t y T r i a l s Ration Animal Feces Feed Tag No. (l b . dry matter) ( l b . dry matter) 1 (barley) 233 1.4 4.8 234 1.1 4.7 226 0.9 4.2 235 1.6 5.2 2 (barley-grass) 169 1.9 4.8 215 1.2 3.8 207 0.9 3.2 216 1.7 5.7 3 (grass-alfalfa) 209 2.3 6.1 156 2.4 6.4 208 2.1 4.8 214 3.1 9.0 50. The apparent d i g e s t i b i l i t y of the proximate fractions i n the 3 rations was determined by means of the data for proximate analysis of the ra t i o n i n Table VI, the data for proximate analysis of the collected feces i n Table VII, and the data for feed consumed and feces excreted i n Table VIII. The r e s u l t s of these calculations are given i n Table IX. TABLE IX Calculated Apparent D i g e s t i b i l i t y of Feed Proximate Fractions Ration Proximate Fraction (%) Animal Dry Crude Crude Crude Tag No. Matter Protein Fiber Fat Ash NFE 1 (barley) 233 77.4 75.6 14.5 90.9 37.5 79.0 234 80.5 82.9 13.0 92.3 53.1 81.6 226 83.8 80.9 13.8 93.1 57.1 85.4 235 80.0 72.9 14.1 87.3 40.0 79.2 X 80.4 78.1 13.8 90.9 46.9 81.3 2 (barley- 215 68.5 63.1 44.0 61.9 60.0 77.7 grass) 169 66.7 65.2 31.4 52.4 51.6 76.5 207 68.9 70.4 42.9 70.6 48.0 81.8 216 40.2 69.1 42.7 64.5 46.7 80.8 X 61.1 67.0 40.3 62.4 51.6 79.2 3 (grass- 209 62.3 70.4 46.7 58.6 54.0 70.2 a l f a l f a ) 156 62.5 67.5 51.0 35.5 56.1 71.6 208 56.5 62.4 45.7 52.2 49.0 61.7 214 65.6 70.6 56.4 51.2 63.0 70.7 X 61.7 67.7 50.0 49.4 55.5 68.6 The crude protein d i g e s t i b i l i t y was 78.1 per cent for r a t i o n 1 as compared to 67.0 and 67.7 per cent for rations 2 and 3 respectively. The crude fat d i g e s t i b i l i t y was also greatest for rat i o n 1 at 90.9 per cent while that of rations 2 and 3 was 62.4 and 49.4 per cent respectively. The d i g e s t i b i l i t y of crude f i b e r was lowest for r a t i o n 1 at 13.8 per cent, intermediate for r a t i o n 2 at 40.3 per cent and highest for r a t i o n 3 at 50.0 per cent. The greater TDN value of rat i o n 1 over ration 2 and ra t i o n 3 was due mainly to a greater crude protein d i g e s t i b i l i t y and to the higher concentration and d i g e s t i b i l i t y of crude fat i n r a t i o n 1. 51. Fart I I I Hematological Studies of Pregnant and Non-pregnant Ewes A. Blood Sampling X \ \ Blood sampling was performed by three different methods described previously (see Methods). The effects of these methods on blood glucose and l a c t i c acid levels were determined. High blood glucose and l a c t i c acid concentrations are known to be associated with excitement. Normal blood glucose levels for sheep have been given as 35 to 60 mg./lOO ml. (53), and l a c t i c a c i d values as 9 to 16 mg./lOO ml.(53). During excitement l a c t i c acid levels r i s e very rapidly to 50 mg./lOO ml. or higher (5). The concentration of blood glucose and l a c t i c acid determined from blood obtained by the three different methods of sampling were compared and are shown i n Table X. TABLE X Comparison of Results Obtained by Different Sampling Methods Method Sample No. Glucose mg./lOO ml. Lactic Acid mg./lOO ml. Needle and syringe 1 2 X 75 84 80 50 44 47 Cannula and syringe 1 2 43 54 49 17 18 18 Vacuum tubes and needle 1 2 54 51 53 19 12 16 52. On the basis of the re s u l t s In Table IX, i t was assumed that the methods involving the cannula and syringe and vacuum tubes and needles, were best suited to minimize excitement as a factor i n t h i s study. The method involving vacuum tubes and needles was selected because i t was found to be the most expedient i n that i t f a c i l i t a t e d very rapid blood sampling. The effect of time a f t e r feeding on blood glucose and l a c t i c a cid was determined and the re s u l t s of t h i s study are given i n Table XI. TABLE XI The Effect of Time After Feeding on Blood Glucose and Lactic Acid Concentration Ration Animal Time Aft e r Glucose Lactic Acid Tag N O . Feeding (hr.) mg/100 ml ma/100 ml 1 (barley) 173 1.5 64 22 3.0 60 17 6.0 71 30 10.0 50 18 2 (barley-graai); 169 ! 1.5 58 37 3.0 58 30 6.0 66 25 10.0 56 36 3 (grass-alfalfa) 167 1.5 64 36 3.0 71 31 6.0 58 17 10.0 55 22 The data in Table X l suggested there was an effect of time a f t e r feeding on blood glucose c o n c e n t r a t i o n . A n effect of time after feeding on-blood l a c t i c acid l e v e l was not-suggested. To avoid an. ef f e c t of time af t e r feeding, on.blood glucose levels the samples of blood were taken from ewes which had free access to feed 10 hours prior to sampling. 53. The extent of recovery of glucose, l a c t i c acid, acetone plus acetoacetate and t o t a l v o l a t i l e f a t t y acids added to whole blood was determined. The re s u l t s of these determinations are shown i n Tables X I I , X I I I , XIV and XV. TABLE X I I Recovery of Glucose Added to Venous Blood Sample Glucose Recovered Added Control Sample Per cent mg/100 ml mg/100 ml 1 -4 72 40 43 112 100 80 80 148 95 2 — 80 • 40 39 120 100 80 83 164 104 X 100 TABLE X I I I Recovery of Lactic Acid Added to Venous Blood Sample Lactic Acid Recovered 3 Added Control Sample ^ Per cent mg./lOO mi. mg/100 ml mg/100 ml 1 50 20 20 70 100 " 4 0 39 104 110 2 -- 31 ... 20 22 52 "r."f:.: 105 40 40 76 112 X 107 54. TABLE XIV Recovery of Acetone Added to Venous Blood Sample Acetone Recovered Added Control Sample Per cent mg/100 ml ,_iL...... 1 3.2 --4.0 3.8 7.5 113 8.0 8.0 12.0 91 2 -- — 3.3 --4.0 4.2 7.8 93 8.0 7.8 12.1 113 X 105 . TABLE XV Recovery of V o l a t i l e Fatty Acids Added to Venous Blood Sample VFA • - - . • Recovered Added " Control Sample Per cent meq./lOO ml meq/100 ml meq./lOO ml 1 _ _ _ 0.075 __ 0.013 0.013 0.087 , 102 ~ 0.026 0.026 0.101 99 2 — 1.433 — 0.013 0.013 1.446 100 0.026 0.026 1.452 99 x too A l l of the methods used i n the blood analysis were tested and found to be adequate on the basis of recovery from whole blood of added metabolites. The data obtained i n these recovery t r i a l s i s given i n Tables XII, XIII, XtV, and XV. The rate of d i s t i l l a t i o n of v o l a t i l e f a t t y acids was determined and the r e s u l t s are shown i n Table XVI. 55. TABLE XVI Rate of D i s t i l l a t i o n of V o l a t i l e Fatty Acids Sample Sample Total VFA Collected Volume Volume Collected D i s t i l l e d (ml.) (ml.) (%) 1 25 25 70.5 2 25 50 88.2 3 25 75 95.3 4 25 100 97.4 Collecting 100 ml. of d i s t i l l a t e was therefore adequate assuring complete v o l a t i l e f a t t y acid d i s t i l l a t i o n . The data obtained i n the hematological studies of pregnant and non-pregnant ewes on each of the 3 rations are found i n Tables XVII A, XVII B, and XVII C. The record of lambing data i s recorded i n Appendix I I . 56. TABLE XVIIA Record of Blood Metabolites for Animals on Ration 1 Date Animal Acetone Glucose Lactic V o l a t i l e Sampled Tag.No. (mg./lOO ml.) (mg./lOO ml .) Acid Fatty Acid (mg&60 ml. (meq./l.) February 'i 7 230 14.6 64 14 1.51 9 232 2.0 52 29 2.40 non-pregnant 14 228 5.5 67 17 2.14 16 235 5.5 64 20 2.69 X 6.9 61.8 20 2.19 7 213 "llw.6 67 40 1.74 9 234 4.0 50 23 0.97 12 173 7.0 70 17 2.87 pregnant 12 229 3.2 65 17 2.19 14 233 5.7 72 34 — 15 172 2.8 61 19 0.91 16 226 2.2 63 16 2.64 X 5.2 64.0 23.7 1.89 March 12 230 2.6 61 17 1.48 i 14 228 0.3 77 34 2.72 non-pregnant 16 235 2.2 70 16 1.10 23 232 2.0 57 20 1.23 X 1.8 66.3 21.8 1.63 7 173 2.8 73 40k 1.73 7 213 2.8 78 14 2.92 pregnant 9 233 ' 2.4 38 23 2.21 9 229 0.8 60 29 2.18 12 226 2.0 58 17 1.38 14 234 1.8 68 17 2.02 16 172 3.4 60 19 1.51 X 2.3 65.0 22.7 1.99 A p r i l : 12 232 0.2 50 14 1.05 ~ 14 235 2.2 58 7 15 226 1.8 58 14 __-16 230 0.0 68 14 1.16 "X 1.7 56 12.6 1.34 12 234 1.4 53 13 1.85 13 172 8.4 67 30 2.16 pregnant 13 173 0.4 62 54 1.83 14 213 0.8 55 23 16 233 2.2 61 17 1.11 16 229 4.0 68 15 1 2.9 61.0 25.3 1. 74 " 5 7 ' TABLE XVIIB Record of Blood Metabolites for Animals on Ration 2 Date Animal Acetone Glucose Lactic Volatile Sampled Tag No. (mg./lOO ml.) (mg./lOO ml.) Acid Fatty Acid (mR./100 ml.) (meq./l.) February 7 222 10,8 63 35 1.89 9 215 12.8 70 24 2.10 non-preg- 12 221 9.6 67 17 2.07 nant 12 206 6.8 70 15 2.30 14 224 0.0 53 14 2.21 15 216 2.4 69 44 0.55 X 7.1 .65.3 24.8 1.85 7 225 21.8 63 15 1.93 pregnant 9 218 7.0 53 25 2.64 14 207 0.0 59 11 1.99 16 220 3.4 51 12 3.09 16 169 2.5 62 15 2.44 X 6.9 57.6 15.6 2.42 March 7 224 2.8 66 29 1.49 12 222 0.8 57 14 2.02 non-preg- 14 206 2.6 65 11 3.00 nant 14 221 2.4 60 12 1.33 16 216 2.0 69 18 0.82 23 215 0.9 58 18 1.95 "•X . 1.9 62.5 17.0 1.77 7 225 3.2 45 15 1.97 pregnant 9 218 1.6 58 14 2.12 9 220 1.8 54 17 : 1.93 12 169 2.8 53 15 2.14 16 207 j 2.2 60 18 1.84 X 2.3 54.0 15.8 2.00 April 12 216 2.2 70 26 1.46 -13 222 3.4 57 22 13 206 2.2 63 23 1.59 non-preg- 14 215 3.6 51 32 1.44 nant 14 224 1.6 62 25 — 15 207 2.4 56 i6. 1.35 16 221 2.8 63 20 0.75 X 2.6 60.3 23.4 1.32 12 169 0.6 51 18 1.48 pregnant 15 225 2.8 53 14 1.20 15 218 7.2 52 23 16 220 2.4 58 18 1.42 t . X 3.3 53.5 18.3 1.37 58. TABLE XVIIC Record of Blood Metabolites for Animals on Ration 3 Date Sampled February pregnant March non-pregnant 23 pregnant A p r i l pregnant Animal Acetone Glucose Lactic ,, V o l a t i l e Tag No. (mg./lOO ml.) (mg./lOO ml.) Acid Fatty Acid (mg./I 00 m l .) (meq./l.] 16 167 3.0 67 14 2.00 7 156 16.1 80 23 2.27 7 214 18.3 86 53 — 9 212 5.9 57 30 2.44 9 219 5.9 64 30 2.12 12 208 0.5 75 14 2.99 12 210 0.5 63 16 2.59 14 211 0.0 65 15 3.00 14 209 5.2 76 45 2.97 15 223 2.8 58 14 2.35 16 X 3.6 52 14 2.63 •TL 6.4 67.4 23.4 2.60 14 167 0.3 70 14 — 212 2.4 59 14 0.82 X 1.4 64.5 14.0 0.82 7 219 1.6 60 21 1.09 7 210 1.8 62 18 0.70 9 223 0.6 62 17 2.40 9 156 0.8 64 27 1.93 12 209 0.8 53 18 2.40 14 211 1.8 65 15 0.44 16 214 3.7 53 29 2.00 16 208 4.0 69 54 0.82 23 X 0.8 58 13 "X 1.8 60.7 23.6 1.64 12 212 3.6 45 15 3.83 12 156 0.8 57 16 1.12 14 X 3.2 53 16 14 208 2.8 55 12 2.44 15 167 4.4 54 13 2.18 16 214 2.6 62 15 1.77 X 2.9 54.3 14.5 2.27 12 219 1.6 45 14 2.30 13 211 2.0 53 48 2.04 13 209 4.6 58 76 — 15 210 4.8 55 12 1.09 16 223 3.0 69 17 1.69 X 3.2 56.0 33.4 1.78 Figure IA Blood Acetone plus Acetoacetate Concentrations of Non-pregnant Sheep for February, March, and A p r i l . Figure IB Blood Acetone plus Acetoacetate Concentrations of Pregnant Sheep for February, March, and A p r i l . Graph 1A: Non-pregnant 01 Ration 1 Q Ration 2 A Ration 3 O o A A A • • © © A n A _ /rv - " . ' ^ t f ; A a IN - " p — — i — i — | — i — — - l — I — I — I — i — ' I 1 1 0 - i — i — i — i — i — i — i — i — i — i — i — i — ' 1 i 1 1 4 8 12 16 20 24 28 4 8 12 16 20 24 28 1 5 9 13 17 February "° n Ration 1 Q e Ration 2 A Ration 3 0 March / A P r i l Graph IB: Pregnant 15 10 © A D © A e © Q A A 0 . 0 ^ Q B J A 8 A B O • 6 © I I | I I I I I I I \ » I '1—I—I I 1 — 1 0 I I I I I I I I I ' I I I ' ' , . ,_, 4 8 12 16 20 24 28 *4 8 12 16 20 24 28 1 5 9 13 17 February _ March A p r i l Figure IIA Blood Glucose Concentration of Non-pregnant Sheep for February, March, and A p r i l . Figure IIB Blood Glucose Concentration of Pregnant Sheep for February, March,and A p r i l . 5 2 . 90 -85 ' 80 -§ 70 $65] I 601 o 3 « 55 50 45 0 Graph IIA: Non-pregnant Ration 1 Q Ration 2 A Ration 3 © A a O A A 1 0 © A B A © © © A • 8 *12 16' 20 24 28 4 8 12 16 20 24 28 1 5 9 13 17 Pebruary March < A p r i l 90 85 80 " 75 -g 70 e 65 S 60 o o 3 o 55 50 45 Graph IIB: Pregnant © © Ration 1 • Ration 2 A Ration 3 © © A O A B B © © a. • © © A El B © © © © © Q & 0 E A © © A \ i i | i i i i i I i i i I—i I i i—r—i—r—»—i i i I i i i 4 8 12 16 20 24 28 4 8 12 16 20 24 28 1 February March © B 1 3 a © A B© ~x—n—i" r ~ i ~ i 5 9 13 17 A p r i l Figure IIIA Blood Total V o l a t i l e Fatty Acid Concentrations of Non-pregnant Sheep for February, March, and A p r i l . Figure IIIB Blood Total V o l a t i l e Fatty Acid Concentrations of Pregnant Sheep for February, March, and A p r i l . 51 5" o-01 5 Graph IIIA: Non-pregnant Ration 1 a Ration 2 A Ration 3 0 • Q • • ca A ID © © © A A A A a , , ! ! i i i i — i — i — i — ( — i — i — i — i — i — i — i — i — i — < ' i — r — i i i i i i 1 i i 8 12 16 20 24 28 4 8 12 16 20 24 28 1 5 9 13 17 February March A p r i l 0 5 0 5 .0 .5 •P .5 Graph IIIBx Pregnant Ration 1 • Ration 2 A Ration 3 © • o A A • © © a © © © © © eta & A A A © T — i — i — i — i — r — i — i — i — i — i — i — i i i—i—i—i—i—i—«—i—i—i—Ti—i—i I — T T i r — » — i r 4 8 12 16 20 24 28 4 8 12 16 20 24 28 1 5 9 13 17 February March A p r i l Figure IVA Blood Lactic Acid Concentrations of N©m-pregnant sheep for February, March, and A p r i l . Figure XVB Blood Lactic Acid Concentrations of Pregnant Sheep for February, March, and A p r i l . 66. o 9 80 -70" 60 501 40 30 Graph IVA: Non-pregnant o (0 * 20 10 Ration 1 Q Ration 2 A Ration 3 O A ID • A a A A © A A 0 • A 8 ©A© February March April 80 1 70 1 60 8 50 J 40 u 0 30 u ,3 20 10 Ration 1 a Ration 2 A Ration 3© © 0 A ©13 O © Graph IVB: Pregnant © • © © • © © © © A • A © n I I I i - — i i i i — r — i — i — i — i — i — T T — r n — ' i I i ' • 1 1 1 ' ° ' ' 8 12 16 20 24 28 4 8 12 16V 20 24 28 1 5 9 13 17 February March April 67. Part III Discussion A study of the effect of time after feeding and excitement on the blood metabolites measured suggested that both of these factors need to be considered in the type of experiment described in Part III. Excitement was avoided by methods previously described. The effect of time after feeding was minimized by allowing animals free access to feed for at least 10 hours prior to sampling. All of the methods used in the blood analysis were tested and found to be adequate on the basis of repovery from whole blood of added metabolites. v The statistical analysis of the data was conducted to determine the effect of time of pregnancy, ration, and pregnancy on the blood concentration of glucose, lactic acid, total volatile fatty acids and ketone bodies. The data obtained in the hematological studies are shown in figures I, II, III, and IV. The data, as indicated on the graphs show considerable dispersion, the exact cause of which was, not determined. It is possible that there are normal fluctuations in the concentrations of these blood metabolites which range through rather wide limits. Due to the variability of the data and the sample numbers, there may have been treatment effects which were not found to be significant. Although there was considerable variability in the data, some significant effects due to the treatments imposed were detected. The data for ketone body readings expressed as acetone are shown in Figure IA and IB. The data were quite variable for all 3 rations and within pregnant and non-pregnant groups. On the basis of increased fetal competition for maternal blood glucose and the usual accompanying increase in fat catabolism, a slight increase in 68. ketone bodies was anticipated in the last month of pregnancy. [However, any changes in blood ketone body concentrations with time were not found to be significant. These results are in agreement with the results for i blood glucose, since there was no significant decrease in blood glucose with time. The blood glucose levels of the pregnant animals on ration 1 for the month of March (4th month pregnant) were found to be significantly higher (P=0.05) than for the pregnant animals on ration 2 fbr the same month. On the basis of work reported by Shaw et al. (169) and by Jorgensen et aJL. (89), i t had been expected that the high concentrate ration 1 would give higher blood glucose levels than would either ration 2 or ration 3. However, this was not conclusively demonstrated. Although in the one case mentioned, there was a significant difference between ration 1 and ration 2 for pregnant animals in March, ration 3 was not found to be significantly different from ration 1. There were no ration effects found to be significant in the months of February or April (3rd and 5th months pregnaae) or for the month of March with non-pregnant ewes. The decrease in rblood glucose which had been expected during late pregnancy on the basis of studies reported by Reid (148) and by Kronfeld (94) could not be shown to be significant. The decrease in blood glucose may have been more effectively demonstrated i f there had'been more ewes twin-pregnant. Reid (148) demonstrated that blood glucose levels decreased more markedly in twin-pregnant ewes than in ewes which were singly pregnant, during late pregnancy. Therefore, the comparison of the 3 rations studied on the basis of magnitude of glycemia decrease in late pregnancy could not be made. The data suggested a possible decrease in 69. blood glucose with tine in the non-pregnant ewes. There were no significant differences in blood glucose concentrations between pregnant and non-pregnant ewes. In summary, pregnant ewes fed. ration 1 had signi-ficantly higher blood glucose levels than pregnant ewes fed ration 2 during the month of .March (,4th month pregnant). (P=0.05). However, there were no other significant effects on blood glucose concentration due to ration, and no significant effects due t® time or to pregnancy. Althoughstandard texts of physiology, such as that by Dukes (53), state that normal blood glucose concentrations for sheep range between 35 and 60 mg/100 mL, of the 99 determinations of blood glucose concentrations made in the experiment discussed, the lowest reading obtained was 45 mg/ 100 ml. while the average of these determinations was 60 mg/100 ml. Therefore, the feeding of certain rations may be associated with blood glucose concentrations which extend considerably beyond the upper limit of 60 mg/100 ml. The results of the experiment of Part I I I corroborates the suggestions of Shaw et al,(175) in relation to high concentrate feeding or fine grinding of roughage. In conclusion, the results obtained did not confirm nor refute the hypothesis that the roughage level in a ration may affect the suitability of that ration for maintaining high blood glucose levels in late-pregnant ewes. It appears, however that the roughage level in the ration is not an important factor in maintaining blood glucose levels in late-pregnant ewes, when the rotughage portion of the diet is finely grotamd sad the animals are on a high level of feed intake, as was the case in this study. A significant increase in the blood lactic acid level was found for non-pregnant ewes on ration 2 for the month of April as compared to the month of March- (P=0.05). Although the data suggested other changes in blood lactic acid with time, such as an increase in blood lactic acid level for April over March for pregnant animals on ration 3 and a decrease from March to April for non-pregnant animals on ration 1, these changes were not found to be significant. Abnormally high blood lactic acid levels are sometimes associated with certain rapidly fermentable feedstuffs (11). The rations studied in the experiment described in Part III may have given rise to considerable lactic acid upon fermentation which may have in turn affected the lactic acid level in the blood. However the. blood lactic acid level is markedly affected by excitement and the level of muscle activity (5). Changes in blood glucose also occur with excitement or muscular activity, however the changes in blood glucose concentration occur over a period of a few minutes (53). This is in contrast to changes in blood lactic acid levels, under similar circum-stances, which occur almost instantaneously (5). Lactic acid which is absorbed from the rumen is metabolized by the rumen epithelium to a certain extent, sis well as by the liver so as to decrease the effect of high levels of rumen lactic acid on the level of lactic acid in the peripheral circulation (11). Therefore, i t is unlikely that ration effects on blood lactic acid level would be detected under the conditions of this type of study. The effect of excitement and muscle activity also are complicating factors in this type of study, so that obtaining normal resting levels of blood lactic acid concentrations was unlikely. The reason for a significant effect of time on the blood lactic acid levels of non-pregnant ewes on ration 2 between March and April is obscure. Possibly these ewes were excited to a greater extent during the final month of sampling as compared to the month of March. 71. The data obtained for total blood volatile fatty acid.levels are indicated in Figure IVA for the non-pregnant ewes and. in.FigureIVB for the pregnant ewes. The data for total blood volatile fatty acid levels were quite variable with small sample numbers. This would tend to obscure any effects on blood volatile fatty acids due to ration, time or pregnancy. However, two significant effects on blood volatile fatty acids due to ration were demonstrated. The blood volatile fatty acid levels associated with ration 3 were found to be significantly higher than the levels associated with ration 2 for pregnant animals in the month of April (5th month pregnant)] (P=0.05) Among non-pregnant animals in the month of March the volatile fatty acid concentrations in the blood were found to be significantly higher for ewes consuming- ration 2 as compared to ewes consuming ration 3 (JVO.IO). There were no significant effects on the level of blood volatile fatty acids due to pregnancy. There was a signfleant effect on blood volatile fatty acid levels for the pregnant ewes consuming ration 2 due to time (P-0.05). The total blood volatile fatty acid levels were significantly lower in April (5th month pregnant) as compared to February (3rd month pregnant). High concentrate rations which have been shown to be associated with relatively high levels of rumen volatile fatty acids as compared to high roughage rations (174). Therefore i t was expected that i f signfleant differences in blood volatile fatty acid levels occurred there would be signfleantly,higher blood volatile fatty_acid levels associated with ration 1 as compared to those associated with ration 3. However, the results obtained were somewhat different than expected. Since a l l three rations were finely ground it may be that a complicating factor was introduced. Changes in rumen volatile fatty acid production in the rumen are frequently associated with the fine grinding of the roughage in a feedstuff. This may account for the apparent discrepancy between the results obtained and current theories. A significant decrease in blood volatile fatty acid levels,.as was found for late-pregnant animals on ration 2, has been suggested by Reid (159), but has not been demonstrated. Since most of the energy requirements of the ruminant are met by the metabolism of short-chain fatty acids (18), and since energy requirements are greatly increased in late pregnancy a decrease in total blood volatile fatty acids in late pregnancy might be expected. Possibly the level of total volatile fatty acids in the blood is more sensitive to energy imbalances in the ruminant than is the level of blood glucose, since the late-pregnant ewes which had a signficant decrease in total blood volatile fatty acids did not have a significant decrease in blood glucose. The blood volatile fatty acid levels were more affected by the factors studied than were the other metabolites. 73. Part IV The Effect of Growth Hormone on Blood Constituents of Pregnant Ewes. The record of the data obtained from this study is found in Appen-dix III. The results are summarized in Table XVIII. TABLE XVIII Summary of Results of Growth Hormone Treatment. Blood Treated Pregnant Non-pregnant acetone 3.8 (mg/100 ml,) lactic acid 23 (mg/100 mL) glucose 61 (mg/100 ml.) volatile fatty 1.70 acids (meo/liter) 2.2 18 57 1.47 Non-treated Pregnant Non-pregnant 2.6 2.7 28 55 1.61 17 60 1.80 Since growth hormone enhances anabolic processes in the body and inhibits catabolic processes, exogenous growth hormone was injected into late pregnant ewes (5th month pregnant) for the purpose of increasing metabolic stress and accentuating changes in blood metabolites which normally occur in late pregnancy. The work of Kronfeld (100) in this regard has previously been discussed. Kronfeld found initial slight changes in blood glucose concentration when growth hormone was administered but when growth hormone administration was continued over a number of days the blood glucose level returned to normal. * The results of the experiment of Part IV showed that growth hormone injected at 1 mg/kg. body weight into late-pregnant ewes on a high level of feed intake caused no significant effects on the level of blood acetone plus acetoacetate, blood l a c t i c acid, blood glucose, or blood v o l a t i l e f a t t y acids. Therefore, i f large amounts of exogenous growth hormone have any profound effects on metabolism under the conditions described, then rapid compensation i s made for these effects so that at the end of 7 consecutive days of growth hormone i n j e c t i o n these effects are not detectable i n the blood. Possibly more extensive studies would be useful i n elucidating more information i n t h i s respect. However, abnormally large amounts of endogenous growth hormone secreted i n l a t e pregnancy would probably not be important i n changes i n the blood metabolites which were studied i n t h i s experiment. CONCLUSIONS 75. The following conclusions are based on the results of the experiments which are reported here. 1. The three rations studied varied mainly in crude fat and crude fiber levels resulting in high (ration 3) medium (ration2), and low (ration 1) roughage levels with corresponding TM values of 59, 70 and 88 per cent respectively. 2. The blood glucose concentrations were significantly higher for pregnant ewes fed ration 1 than for pregnant ewes fed ration 2 during the fourth month ©f pregnancy (March) (P=0.05). 3. A significant increase in the blood lactic acid levels was found for non-pregnant ewes on ration 2 for the month of April over the month of March (P«0.05). 4. The blood volatile fatty acid levels associated with ration 3 were found to be significantly higher than the levels associated with ration 2 for pregnant animals during the fifth month (April) of pregnancy.(P=0.05). 5. Among non-pregnant animals in the month of March, the volatile fatty acid concentratioas in the blood were found to be significantly higher for ewes consuming ration 2 than for ewes consuming ration 3 (P=0.10). 6. The total volatile fatty acid levels for pregnant ewes fed ration 2 were found to be significantly higher in the third month of pregssaacy (February) than in the fifth month of pregnancy (April) (P=0.05). 7. The intramuscular injection of 1 mg/kg. body weight of growth hormone into pregnant sheep for 7 days during the last month of pregnancy caused no significant effects on blood glucose, l a c t i c acid, acetone plus acetoacetate, or t o t a l v o l a t i l e f a t t y acids concentrations. 8 . The blood glucose levels which were obtained i n t h i s study were generally higher than those given i n standard textbooks. REFERENCES 1. Adler, J. H. Some aspects of bovine ketosis. Diss. Abstr., 17s 2044, 1957. 2. Adler, J. H., and Dye, J, A, On the mechanism of ketogenesis in ruminants. Cornell Vet., 46: 58, 1956. 3. Adler, J. H., Robert!, S. J. and Dye, J. A. Fiwther observations on silage as a possible etiological factor in bovine ketosis. Amer. Jour. Vet. Res. 19: 314, 1958. 4. Alexaad@r, G. S., and Zieve, L. Ketogenesis and hyperketonamia, Amer. Jour. Med. 107: . 514, 1961. 5. Allcroft, W. Mo, and Strand, R. Studies on the lactic acid, sugar and inorganic phosphorus of the -blood o£ ruminants. ii©chem. Jour. 27: 512, 1933. 6. Allison, M . J. Nutrition of rumen bacteria. In Physiology of Digestion in the Ruminant. Bffltterworth^Iac", Washington, D. C. 1965. 7. Allyn, 1. J, A contribution to a better understanding of ketosis in dairy cows. J@ur. Amer. Vet. Med. Ass. 126: 129, 1955. ' 8. Amb®, K. Reto, K., Skaurai,Y., and Bmezu, M. Studies on ketosis in ruminants II. Principal site ©f ketone body formation. J«p. Jour. Vet. Sci. .23: 265, 1961. 9. Ann!son, E. F.. Absorption from the ruminant stomach. In Physiology of Digestion in the Ruminant. Butterworth Inc., Washington, D. C. 1965. 10. Annisoa, E. F, H i l l , K„ J., and Lewis, D. Studies on the p@rtal blood ©f sheep. II. Absorption of ;ile fatty acids from the rumen of the sheep, lem. Jour. 66: 592, 1957. 11. Annis@iffi, E.. F., and Lewis', D... Metabolism in the Rumen. men and Company L t d ., London, 1959. 12. Annis®ffl,^ .E. f., and Lindsay, B. 3- Acetate utilization in'sheep. shem. Jour. 78| 777 , 1 9 6 I . 13. Annison, E. P., and Pennington, R. J. The metabolism of short chain fatty acids in the sheep. III. Formic, n-valeric, and some branched-chains. Biochem. Jour. 57: 685, 1954. 14. Annison, E. F., and White, R. W. Glucose utilization in sheep. Biochem. Jour. 80: 163, 1961. 15. Anonymous. Clinical Biochemistry of Domestic Animals. Academic Press, New Yorki1963. 16. Anonymous. The Merck Veterinary Manual 2nd Edition. Merck and Co. Inc. Rahway, N.J., 1961. 17. Anonymous Official Methods of Analysis of the Association of Official Agricultural Chemists. 9th edition. Association of Official Agricultural Chemists. Washington, D.C. I960. 18. Armstrong, D.G. Carbohydrate metabolism in ruminants and energy supply. In Physiology of Digestion in the Ruminant. Butterworth Inc., Washington, D. C., 1965. 19. Armstrong, D. G., and Blaxter, K. L. The heat increment of volatile fatty acids in fasting sheep. Brit. Jour. Nutr. 11: 247, 1957. 20. Ash, R. W. Pennington, R. J. and Reid, R. S. The effect of short chain fatty acids on blood glucose concentration in sheep. Biochem. Jour. 90: 353, 1964. 21. Aspiotis, N. Physiopathologic mechanism produc-ing ketosis of milk cows. \ Rec. Med. Vet. Ecole Alfort. 136: 989, 1960. 22. Bach, S. S., and Hibbitt, K. G. Biochemical aspects of bovine ketosis. Biochem. Jour. 72: 87, 1959. 23. Baird, O.D. Kstogenesis in slices of liver and lactating mammary gland from normal and ketotic cows. Biochem. Biophys. ACTA. I l l : 339, 1965. 24. Ballard, F.J., and Oliver, L.T. citrbohydrate metabolism in liver from fetal and neonatal sheep. Biochem. Jour. 95: 191, 1965. 25. Behre, J. A., and Benedict, S, R. A colorimetric method for the determination of acetone bodies in blood. Jour. Biol. Chem. 70: 487, 1926. 26. Bell, F. R. and Jones, E. R. Glucose tolerance in the bovine. Jour. Comp. Path, and Therap. 55: 117, 1945. 27. Benedict, S. R. A colorimetric method for the deter-mination of glucose in blood. Jour. Biol. 92: 141, 1931. 28. Bennet, L. L., Kreiss, R. E. Gfeoh Mao Li and Evans, H. M. Production of Ketosis by the growth hormone and adrenocorticotropic hormone. Amer. Jour. Phys. 152: 210, 1948. 29. Bensadoun, A. Direct estimation of the absorption of volatile fatty acids from the gastrointestinal tract of ruminants. Ph. D. Thesis, Cornell laiversity Library. Ithaca, New York, 1960. 30. Bergsaaan, E. 1. Quantitative aspects of glucose in pregnant and non-pregnant sheep. Amer. Jour. Phys. 204: 147, 1963. 31. Bergmann, E. N., and Kon, K. Acetoacetate turnover and oxidation rates in ovine pregnancy ketosis. Amer. Jour. Phys. 206: 449, 1964. 32. Bergmann, E. N., and Kon, K. Factors affecting aceto-acetate production rates by normal and ketotic pregnant sheep. Amer. Jour. Phys. 206: 453, 1964. 33. Bergmann, E. S3., Kon, K., and Katz, M. I. Quantitative measurements of acetoacetate metabolism and oxidation in sheep. Amer. Jour. Phys. 205: 658, 1963. 34. Bergmann, E. N*, Kon, K., and Katz, M. L. Acetoacetate metabolism rates in ovine ketosis. In: 47th Annual Meeting of the Federation of American Societies for Experimental Biology. 1963. Federation Proc. 22: 575, 1963. 35. Bergmann, E. N., and Sellers, A. F. Comparison of fasting ketosis in pregnant and non-pregnant guinea.pigs. Amer. Jour. Phys, 198: 1083, 1960. 36. Bergmann, E. N. Sellers, A. P., and Spurrell, F. A. Metabolism of - labelled acetone, acetate and palmitate in fasted)pregnant and non-pregnant guinea pigs. Amer. Jour. Phys. 198: 1087, 1960 37. Black, A. L. Metabolic change In the utilization of glucose carbon in the TCA cycle in bovine ketosis. In: 47th Annual Meeting of the Federation of American Societies for Experi-mental Biology. Federation Proc. 22: '611, 1953. 38. Black, A. L. and Luick, J. R. Metabolism of ketone bodies in lactating cows. Int. Cong. Biochem. 6: 714, 1964. 39. Black, A. L., and Luick, J. R, The metabolism of ketone bodies in normal and ketotic cows and their utilization for milk synthesis. In Radioisotopes in Animal Nutrition and Physiology. Proceedings of Symposium, Prague, November 1964. International Atomic Energy Agency, Vienna, 1965. 40. Blackburn, P. S. Castle, M. E. Brysdale, A. D. and Strachan, N. H. The effect of feeding a high or a low mineral concentrate on the incidence of ketosis in dairy cows. Brit. Vet. Jour. 117: 158, 1961. 41. Blaxter, K. L. Energy utilization in the ruminant. In Digestive Physiology and Nutrition of the Ruminant. Butterworth and Co., London, 1961. 42. Bloom, 1. Stetten, M. R. Stetten, D., Jr. Evaluation of catabolic pathways of glucose in mammalian systems. Jour. Biol. Chem. 204: 681, 1953. 43. Bressler, R. The biochemistry of ketosis. In: Some aspects of metabolic diseases in man and animals, i. New York Acad. Sci. 104: 735, 1962. 44. Calhoun, M. C,, Fleeger, J. L., and Richards, C. R. Serum Insulin like activity of normal fasting and ketotic cattle. Jour. Dairy Sci. 45: 421, 1962. 45. Campbell, J., and Best, C. H. Physiological aspects of ketosis. Metabolism 5: 95, 1956. 46. Card, C. S., and Schultz, L. H. Ttoe effect of ration on the volatile fatty acids produced in the rumen. Jour. Dairy Sci. 36: 599, 1953. 47. Carlstrom, B. Deficiency diseases particularly acetonemia in cattle. Vet. Rec. 62: 718, 1950. 48. Cook, R. M. Brown, R. E.. and Bavis, C. L. The incorporation of C^ from.carboxyl labelled volatile fatty acids into goat liver glycogen. Federation Proc. 22: 611, 1963. 49. Cornelius, C. E. Small, M., and Kleiber, M. C 1 4 studies on ketogenicity of metabolites in lactetlng dairy cows. Proc. Soc. Exptl. Biol. and Med. 172, 1957. 50. Cunningham, N.F. The insulin-like action of extracts of bovine and ovine blood plasma. Jour. Endocrinology. 25: 43, 1962. 51. Cutler, J. T. Studies on the carbohydrate metabolism of the goat. Jour. Biol. Chem. 106: 653, 1934. 52. Drummond, 6 . I., and Stern, J. R. Enzymes of ketone body metabolism. II. Properties of an acetoacetate-synthesizing enzyme prepared from ox liver. Jonnr. Biol. Chem. 235: 318, 1960. 53. Dukes, 1. H. The Physiology of Domestic Animals: 7th Edition. Comstock Publishing Ass., Ithaca, New fork, 1955. 54. Dye, J. A. The effects of the administration of acetic, propionic, butyric acids upon the blood glucose and ketone body levels of goats. Cornell Vet. 45: 273, 1955. 55. Dye, J.-A. and Chidsey, J. L . Ketone body—total carbohydrate utilization ratios and their relation to the problem of ketosis. Amer. Jour. Phys. 127: 745, 1939. 56. Dye, J. A., Roberts, S. J., Blampied, N., and Fincher, M. G. The use of cortisone in the treatment of ketosis in dairy cows. Cornell Vet. 43: 128, 1953. 57. Dye, J. A., amd Woodward, B, A. Alloxan diabetes in the sheep. Federation Proc. 6:99, 1947. 58. Emery, R. S< Burg, N., and Brown, L. D. Borderline ketosis: detection, oeerarrence and prevention. An. Sci. 22: 1119, 1963. 59. Emery, R. S., and Williams, J. A. Incidence of ketosis, other diseases and some post-parturn reproductive ailments in normal and triiodo-thyronine- treated cows. Jour. Dairy Sci. 47: 879, 1964. 60. Emery, R. S., Burg, M. Brown, L.D., and Blank, 6. N. Detection occurrence and prophylactic treatment of borderline ketosis with propylene glycol feeding. Jour. Dairy Sci. 47: 1074, 1964. 61. Engel, F. L. A consideration of the roles of the adrenal cortex and stress in the regulation of protein metabolism. Rec. Prog, in Horm. Res. 6s 277, 1951. 62. Engel, F. L. Amatruda. Hormonal aspects of ketosis. Iras Some aspects of metabolic diseases in man and animals. Ann. New York Acad. Sci. 104: 753, 1963. 63. Engel, F. L. Schiller, S., and Pesntz, E. I. Studies ©n tine nature of .the protein catabolic response to adrenal cortical extract. Endocrinology 44: 458, 1949. 64. Flux, B..-S., Folley, S. J., and Rowland, S. J. The effect of adrenocorticotropic hormone.on the yield and composition of the milk of the cow. Jotmv Endocrinology 10: 333, 1954. 65. Ford, E.~ J. H. Oxaloacetate levels in normal and ketotic pregnant sheep. Res. Vet. Sci. 4: 408, 1963. 66. Ford, E. J. H. Acetate utilization and carbon dioxide production in normal and ketotic sheep pregnant with twins. Res. Vet. Sci. 5s 161, 1964. 67. Forenbacher, S., and Srebocan. On the part played by the liver in the transformation of tricarbonic acids, particularly pyruvic, oc -ketoglutaric and lactic acid in ketosis of milch cows. ¥eterimarski ARH. 33: 1, 1963. 68. ForenbacMer, S., Zdenka, P., and Sherbocan, V . On glycolysis in the blood in ketosis in milch cows with special reference to the exogenous and endogenous gluconeogenesis. Vet. Archives. 32: 215, 1962. 69. Fraser, A. H. H., Godden, w"., §»®®k, L.C. and Thomson, W. The influence of diet up©n ketonemia in pregnant ewns. Jour. Phys. 94: 346, 1938. 70. Gessert, R. A., Shaw, J. C,' a©d Cmung, A. C. Studies sis in dairy cattle. Jour. Amer. Vet. 127: 215, 1955. 71. ©oetseh, ©. 1. Studies on the production and treatment of experimental ketosis of ruminants. Diss. Abstr. 17s 2364, 1957. 72. Goetseh, 6. 1., and Pri'tchard, W. R. -Effects of oral administration of short chain fatty acids on certain blood aad urine.constituents of fasted phlorizin-treated ewes. Amer. Jour. Vet. Res. 19: 637, 1958. 73. Halse, K., and Weiert, V. Blood calcium In bovine ketosis. Amer. Jour. Vet. Res. 19s 575, 1958. 74. Hetziolos, B. C. and Shaw, J. C., Bovine ketosis: An histologic study. II. The thyroid glands. Zentralbl. Veterinarmed. Reihe A. 10: 163, 1957. 75. Heath, T. J., and Morris, B. The absorption of fat in sheep and lambs. Quart. Jour. Exptl. Phys. 47: 157, 1962. 76. Heuter, F. 6. The oxidative metabolic pattern of normal and ketotic cow liver as studied with l - C 1 4 labelled Gj —>- G5 aliphatic acids. Diss. Abstr. 21: 58, 1959. 77. Hibbit, K. 6. Experiments on the induction of a ketosis in the dairy cow. Vet. Record. 76: 738, 1964. 78. Hird, F. J. R. Ketone body synthesis in relation to age in lambs. Biochem. J©ur. 93: 423, 1964. 79. Hird, F. J. R. and Symons, R. H. The mode of formation of ketone bodies from butyrate by tissue from the rumen and omasum of the sheep. Biochim. et Biphys. Acta. 46: 457, 1961. 80. Hird, F. J. R. and Symons, R. H. The mechanism of ketone-body formation from butyrate in rat liver. Biochem. Jour. 84: 212, 1962. 81. Hodgson, R. E. Riddel1, W. H. and Hughes, J. S. Factors Influencing the blood sugar level of dairy cattle. Jour. Agr. Res. 44: 357, 1932. 82. Hoflund, S., and Hedstrom, H. Disturbances in ruminant digestion as a predisposing factor in acetonemia. LI Vet. 38: 405, 1948. 83. Horroeks, D., and Patterson, J. Y. F. Some observations on glucose, ketone bodies and volatile fatty acids in the blood of dairy cattle. Jour. Comp. Path, amd Ther. 67: 33, 1957. 84. Hupka, E. Bie Azetonamie der Binder Duet, tieraztl w®ehseher. Ruminant Digestion. 36 No. 98, 1928. 85. Jackson, H..D., and Taylor, H. C. Formation of ketone bodies from long chain fatty acids in ruminant epithelium and liver from ketotic sheep. Arch. Biochem. Biophys. 105: 575, 1964. 86. Jarrett, 6. I., Jones, G. B., and Potter, B. J. Changes in glucose utilization during the development of the lamb. Biochem. Jour. 90: 189, 1964. 87. Jesae, S . Blood sugar and liver glycogen in ketosis of the ruminant. Jour. Amej Vet. Hed. Ass. 124: 341, 1954. 88. Johnson, R . B. The relative rates of absorption of the volatile fatty acids from the rumen and their relationship to ketosis. Cornell Vet. 41: 115, 1951. 89. Jorgensen, I. A., and Shultz, L. H. Ration effects on rumen acids, kefeogeaesis and milk compo-sition. Jour. Dairy Sei. 46: 437, 1963. 90. Kennedy, W. L., Anderson, A. K. Bechdel, S. I. Shigley, J. F. Studies on the composition of bovine blood as influenced by gestation, lactation and age. Jonnr. Dairy Sci. 22: 251% 1939. 91. Klussendork, R. 0, Ketosis confusion clearing. Jour. Amer. Vet. Med. Ass. 127: 328, 1955. 92. Knodt, C. B., Shaw, J. C., and White, G. C. The effect of stall and pasture feeding on blood ketone bodies aad urine ketone bodies. J©ur. Dairy Sci. 25: 837, 1942. 93. Krebs, 1. A. The biochemical lesion in ketosis. Arch. Internal Med. 107: 51, 1961. 94. Kronfeld, D. S. A comparison of normal concentra-tions of reducing sugar volatile fatty acids and ketone bodies in the blood of lambs, preg-nant ewes and non-pregnant adult ewes. Aust. Jour. Agr. Res. 8: 202, 1957. 95. Kronfeld, D. S. The effects ©ra blood sugar and ketone bodies of butyrate, acetate and ft -hydroxybutyrate infused into sheep. Aust. Jorar. Exptl. Biol. and Med. Sei. 35: 257, 1957. 96. Kronfeld, D. S. Growth hormone administration to pregnant Cornell let. 4 7 : 255,1957. 97. Kronfeld, B. S. The fetal drains hexose in ovine pregnancy toxemia. Cornell •••Vet. 48: 394, 1958. 98. Kronfeld, D. S. Metabolic aspects ©f ruminant ketosis. Amer. Jour. Vet; Res. 22: 496, 1961. 99. Kronfeld, D. S. Ruminant ketosis: A speculative approach. In Some aspects of metabolic diseases in man and animals. Ann. New York Acad. Sci. 104: 799, 1963. 100. Kronfeld, D, S. Growth hormone- induced ketosis in the cow. JOur. Dairy Sci. 48s 342, 1964. 101. Kronfeld, D. S. The enzymatic regulation of ketogenesis. Nord. Vet. Med. 17: 182, 1965. 102. Kronfeld, D. S., and Brown, G. H. Proceedings of the International Conference on the Use of Radioisotopes in Animal Biology and Medical Sciences, Mexico City, Academic Press, New Y®rk, New York, 2&7&1962. 103. Kronfeld, 1. S., and Kleiber, M. Mammary ketogenesis in the cow. Jour. Appl. Phys. 14: 1033, 1959. 104. Kronfeld, D. S. Kleiber, M., and Lucas J. M. Acetate metabolism in bovine ketosis. Jour. Appl. Phys. 14: 1029, 1959. 105. Kronfeld, D. S. and Raggi, F. Pyridine nucleotide coenzyme concentrations in mammary biopsy samples from ketotic cows. Federation Proc. 22: 471, 1963. 106. Kronfeld, D. S. and Simesen, M. 6. Glucose biokinetics in ovine pregnancy toxemia. Vet. 51: 478, 1964. 107. Kronfeld, D. S. Tombropoulos, and Kleiber, M. Glucose biokinetics in normal and ketotic cows. Appl. Phys. 14: 1026,, 1959. 108. Kumar S., Lakshmanan, S., and S&aw, J. C. /©-hydroxybutyrate metabolism; of the perfused bovine udder.,; Jour. Biol. Chem. 234: 754, 1959. 109. Leng, R. A. The metabolism of D (-)-/3 -hydroxybutyrate in sheep. Biochem, Jour. 90: 464, 1964. 110. Leng, R. A. ketone body metabolism in normal and underfed pregnant sheep and in pregancy toxemia. Res.'Vet. Sci. 6: 433, 1965. 111. Lindner, H. R. Blood cortiso^ in the sheep: Normal concentration and changes in ketosis of preg-nancy. Nature. 184: 1645, 1959. 112. Lindsay, D. S. Endocrine control of metabolism in ruminants. In Digestive Physiology and Nutrition of the Ruminant. Butterworth and 60., London, 1961. 113. Lindsay, B. B., and Ford, H. J. Acetate utilization and the turnover of citric acid-cycle components in pregnant sheep. Biochem. Jour. 90: 24, 1964. 114. Luick, J. R., and Smith, L. M, Fatty acid synthesis dwiag fasting bovine ket©sis. Jour. Dairy Sci. 4 6 : 1251, 1963. 115. Maddis®a, L. L., Mebane, B, 1. and Lochner. The hypoglycemic action of ketones; II. Evidence for a stimulatory feedback of ketones on the pamereatie beta cells. Jorar. Clin. Invest. 43: 408, 1964. 116. Marshak, R. R. The pathological physiology of bovine ketosis. Jour. Amer. Vet. Med. Ass. 132: 328, 1958. 117. Mayes, P. A . Level of liver glycogen in ketosis. 182: 324, 1958. 118. Mayes, P. A . A caloric deficiency hypothesis of Ls. Met. Clin. Exptl. 11: 781, 1962. 119. Mayes, P. A. Role of the pituitary gland in ketogenesis. Nature. 194: 939, 1962. 120. McGandless, E. L., and Dye, J. A. Physiological changes in intermediary metabolism of various species of ruraiaamts incident to functional development of the rumen. Amer. Jour. Phys. 162: 434, 1950. 121. McKay, E. M. Barnes, R. H. Game, H.O., and Wick, A.W. ketogenic activity of acetic acid. Jour. 135: 157, 1940. 122. Mebane, D., and Madison, L. L. a ® hypoglycemic effefi ©f ketome bodies. Jorar. Clin. lav. 41: 1383, 1962. 122A.Mebane, B., and Madison, L. L. Clinical and experimental hypoglycemic action of ketoaes. I. Effects of ketasaes ©a hepatic gl«c@s® output and peripheral glucose utilisation. Jowr. Lab. and Clin. Med. 63: 177, 1964. 123. Memahaa, L. A., Sehultas, L. H. and !®ekstira. Relation-ship of ketone body metab©lism and carbohydrate utilisation to fat mobilization in the ruminant. Jossr. Dairy Sci. 49: 957, 1966. 124. Mendel, B., and Geldscheider, I. The colorimetric estimation of lactic acid in blood. Biochem Z. 164: 163, 1925. 125. Milman, A. E., De Meor, P. and Luhens, F. D. W. Relation of purified pituitary growth hormene and insulin in regulation of nitrogen balance. Amer. Jour. Phys. 38j 331, 1948. 126. Moir, R. J., and Harris, L. E. Ruminant flora studies in the sheep. X. Influence of nitrogen intake upon ruminal function. Jour. Nutr. 77s 285, 1962. 127. Outhouse, J. B. Influence of diet on ketosis in sheep. Diss. Abstr. 17: 463, 1957. 128. Parry, H. B., and Taylor, W. H. Renal function in sheep during normal and toxemic pregnancies. Jour. Phys. 131: 383, 1956. 129. Pauline, H. 0.,and Reber, E. F. The effect of oxalacetie and pyruvic acid on hypoglycemia and ketonemla in pregnant ewes. Cornell Vet. 47: 131, 1957. 130. Pennington, R. J. The metabolism of short chain fatty acids in the sheep. I. Fatty acid utilization and ketone body production by ruminal epithelium and ®ther tissues. Biochem. Jour. 51: 251, 1952. 131. Pennington, R. J., and Pfander, W. H. The metabolism of short chain fatty acids in the sheep. Biochem. Jour. 65: 109, 1957. 132. Pennington, R. J. and Sutherland, T. M. Ketone-body production from various substrates by sheep-rumen epithelium. Biochem. Jour. 63s 353, 1956. 133. Pennington, R. J., and Sutherland, T. M. The metabolism ©f short chain fatty acids in the sheep to the pathway of propionate in the rumen epithelial tissue. Biochem. Jour. 63: 603, 1956. 134. Phillipson, A. T. The fatty acids present in the rumen of lambs fed flaked maize. Brit. Jour. Nutr. 6: 199, 1942. 135. Phillipson, A. T. and McAmally, R. A. Studies on the fate of carbohydrate in the rumen of sheep. Jour. Exptl. Biol.19: 199, 1942. 136. Potts, R. B., and Kesler, E. M. Effect of grass silage on milk flavours and milk acetone bodies. Jour. Dairy Sci. 40: 1466, 1957. 137. Procos, J. Ovine ketosis. I . The normal ketone body values. Onderstepoort Jour. Vet. Res. 28: 557, 1961. 138. Procos, J. Ovine ketosis. II. The effect of pregnancy on the blood ketone body levels of well-fed ewes. Onderstepoort Jour. Vet. Res. 29: 107, 1962. 139. Procos, J. Ovine ketosis I I I . The effect of starvation on the blood sugar and ketone body levels of wethers. Onderstepoort Jour. Vet. Res. 29: 259, 1962. 140. Procos, J. Ovine ketosis I f . The effect of alloxan. Onderstepoort Jour. Vet. Res. 30: 161, 1963. 141. Randle, P. J. Regulation of glucose uptake by muscle. 7. Effects of fatty acids, ketone bodies and pyruvate, and of alloxan diabetes, starvation, hypophysectomy and adrenalectomy, on the concentra-tions of hexose phosphates, nucleotides and inorganic phosphate in perfused rat heart. Biochem. Jour. 93: 641, 1964. 142. Ray, P. D . Foster, D. 0. and Lardy, H. A. Mode of action of glucocorticoids. Jour. Biol. them. 239: 3396, 1964. 143. Reece, R. P. The influence of thyroprotein in the ration of dairy cattle. Jour. Dairy Sci. 30: 574, 1947. 144. Reid, R. L. Studies on the carbohydrate metabolism of sheep. I. The range of blood sugar values under several conditions. Austr. Jour. Agric. Res. 1: 182, 1950. 145. Reid, R. L. Studies on the carbohydrate metabolism of sheep. I I . The uptake by the tissues of glucose and acetic acid from the peripheral circulation. Aust. Jour. Agr. Res. Is 322, 1950. 146. Reid, R. L. Studies on the carbohydrate metabolism of sheep. I X . Metabolic effects of glucose and glycerol in undernourished pregnant ewes and in ewes with pregnancy toxemia. Aust. Jour. Agr. Res. l i t 42, 1960. 147. Reid, R. L. Studies on the carbohydrate metabolism of sheep. X I . The role of adrenals in;ovine pregnancy toxemia. Aust. Jour. Agr. Res. 11: 364, 1960. 148. Reid, R. L. Energy requirements of ewes in late pregnancy. In Digestive Physiology and Nutrition of the Ruminant. Butterworth and Co., London, 1961. 149. 8*1 d, R. L. Studies era the carbohydrate metabolism of sheep. XVI. Partition of ketone bodies in blood, tissues and urine. Aust. Jour. Agr. Res. 13: 307, 1962. 150. Reid, R. L. Studies on the carbohydrate metabolism of sheep. XIX. The metabolism of glucose free fatty acids and ketones after feeding and during fsating or radern®urishment of non-pregnant, pregnant and lactating ewes. Aust. Jour. Agr. Res. 13: 1124, 1962. 151. Reid, R. L. and Hinks, N. T. Studies on the carbohy-drate metabolism of sheep. XVII. Feed require-ments- and voluntary feed intake in late pregnancy with; particular reference to prevention of hyp©glyeemiaran3 hyper ketonemla. Aust. Jour. Agr. Res. 13: 1092, 1962. 152. Reid, R. L. and Hinks, H. T. Studies on the carbohy-drate metabolism of sheep. XVIII. The metabolism of glucose, free fatty acids, ketones and amino acids in late pregnancy and lactation. Aust. Jour. Agr. Res. 13: 1112, 1962. 153. Roberts, S. J. Ketosis parturient paresis complex. Jour. Amer. Vet. Med. Ass. 124: 368, 1954. 154. Robertson, W. G., Lennon, H. D., Bailey, W. W., and Mixner, J. P. Interrelationships among plasma 17-hydroxycortieosteroid levels, plasma protein bound iodine levels, and ketosis in dairy cattle. Jour. Dairy Sei. 40: 732, 1957. 155. Roderick, Lj-M., Harshfietd, G . S.£and Hawn, M.C, The. pathogenesis of ketosis: Pregnancy disease of sheep. Jour. Amer. Vet. Med. Ass. 90: 41, 1937. 156. Roth, J. <§lick,"S.. M. Yalow, R. S. and Berson, S. A. Hyp@glyeemia: A poteat stimulus to secretion of-growth, hormone. Science. 140:-. 987, 1964. 157. Sabine, J. R., and Johnson, B.C.-Acetate Metabolism in the ruminant. Jour. Biol. Chem. 239: 89, 1964. j 158. Sampson, J»» and Boley, L. E. Studies on the''total. ket®ne.:boddLes, .sugar., and calcium of the blood of nom^pregant, ndn-lactatiag ewes. Jour. Vet. Med. Ass. 96: 480, 1940. 159. Sampson, J., and Hayden, C. E. The acid-base balance in cows and. ewes during, and after.pregnancy-with special reference to milk fever and acetonemia. Jour. Amer. Vet. Med. Ass. 86: 13, 1935. 160. Sargent, F., Johnson, R. E. Bobbins, E., and Sawyer, The effect,of environment and other factors on nutritional ketosis. Qraart. Jour. Expt. Phys. 43: 345, 1958. 161. Scarisbrick, R., and Storie Pagh, P.D. The passage of acetate across the placenta and its uptake by ths foetal sheep. Brit. Vet. Jour. 113: 328, 1957. 162. Schamhye, P.» and Phillipsom, A. T. V o l a t i l e fatty acids in the portal blood of sheep. Nature.' '•' 164: 1094, 1949. [ V 163. Schmidt,, ®. 1., and Sehulta,, <S„ H. The effect of three levels of grain feeding during'the dry period on the incidence of ketosis, severity, of udder edema and subsequent milk production of dairy cows. Jour. Dairy Sci. 42: 170, 1959. 164. Schultz, L. H. Treatment of ketosis. in dairy cows '  with sodiran propionate. Cornell Vet. 42: 148, 1952. 165. Schttltz, L. 1., and Smith, V. R. Experimental ' alteration of the sugar and ketone levels i n the blood of ruminants. Jour. Dairy Sci. 34: 1191, 1951. 166. Seekles, L. A. Gastrointestinal autointoxication i n cattle and horses. Brit. Vet. Jour. 104: 238, 1948. 167. Seekles,, L. A. A note on ketosis treatment with assB8®iHii« lactate. Vet. Ree. 63: 494, 1951. 168. Seto, KatsuOs, Tsaneyuki, Tsuda, and Motoyosi. The meehamism of the ketone body production with rwsea epithelium. I. Bae ketone body prodwetlon from volatile fatty acids with rneem epithelium and the influence - of substances concerned with the tricarboxylic acid cycle on their production. Tohoku Lc. Res. 6: 91„ 1955. 169,. -Shaw, J. C. Nutritional physiology of the rumen., (general Reports. Pg. 29 s 1961. 170. Shaw, J. €. Studies on ketosis in dairy cattle. V. The development of ketosis. Jour. Dairy Sci. 26s 1079, 1943. 171. Shaw, J. G. The present status of the ketosis problem. Flour and Feed. 5^! 30, 1954. 172. Shaw, J. C. Studies on the etiology and treatment of ketosis in dairy cows. II. Studies on the corticoids in the treatment of ketosis. Proc. Amer. Vet. Med. Ass. :80, 1954 173. Shaw, J. C. Ketosis in dairy cattle. A review. Jour. Dairy Sci. 39; 402, 1956. 174. Shaw, J. C. Nutritional physiology of the rumen. In the VHIth. International Congress of Animal Production, Hamburg Germany, 1961. 175. Shaw, J. C. Ensor, W. L. Tellechea, H. F., and Lee, S. D. Relation of diet to rumen volatile fatty acids, digestibility, efficiency of gain and degree of unsaturation of body fat in steers. Jour. Nut. '71s 203, 1960. 176. Shaw, J. C. Oessert, R. A. and Chung, A. C. Studies on the etiology and treatment of ketosis in dairy cows. I. Etiological considerations in bovine ketosis. Proc. Amer. Vet. Med. Ass., page 78, 1954. 1 177. Shaw, J* C., Hatziolos, D.V.M., and Leffel, E. C. An approach to the etiology of ketosis in dairy cows. Jour. Amer. Vet. Med. Ass. 117s 103, 1950. 178. Smith, E. E. Oxidation of volatile fatty acids as related to ketosis in ruminants. Diss. Abstr. 21s 42, 1960. 179. Smith, E. E., Goetch, G. D., and Jackson, H. D. Oxidation of volatile fatty acids by rumen epithelium and by liver from ketotic sheep. Arch. Biochem. and Biophys. 95s 256, 1961. 180. Smith, G. S., Dunbar, R. S., McLaren, G. A., Anderson, G. C, and Welch, J. A. Measurement of the adaptation response to urea-nitrogen utilization in the ruminant. Jour. Nut. 70s 20, 1960. 181. Smith, R. M., and Osbourne, W. S. Metabolism of propionate by homogenates of normal sheep liver. Nature. 192s 868, 1961. 182. Sosteda, Clinical and biochemical studies on the basis of ketone bodies in cattle. I. Blood ketone bodies in dairy cattle. Jap. Jour. Vet. Res. 4s 47, 1956. 183. Sporri, H. Pathogenesis and Therapy of Ketosis of ruminants. Schweiz. Arch. Tierheilk. 100: 347, 1958. 184. Staubus, J. R. The effect of fasting, phloridzin, Insulin and butyrate on bovine energy metabolism as related to ketosis. Diss. Abstr. 20: 2997, 1960. 185. Stevenson, D. E. and Wilson, A. A. Metabolic Disorders of Domestic Animals. F. A. Davis Co., Philadelphia, Pennsylvania, 1963. 186. Strand, R., Anderson, W. and All croft, W. M. Further studies of the lactic acid, sugar and inorganic phosphates of the blood of ruminants following adrenalectomy or after intravenous injection of insulin. Biochem. Jour. 28: 642, 1934. 187. Swift, R. W., Bxatzler, J. W., James, W. H., Tillman, A.D., and Meek, D. C. The effect of dietary fat in utilization of the energy and protein of rations by sheep. Jour. An. Sci. 7: 475, 1950. 188. Thin, G., Paver, C. H., and Robertson, A. The metabolism of ketone bodies in the ruminant. Jour. Comp. Path, and Ther. 69: 45, 1959. 189. Thin, C., and Robertson, A. _he estimation of acetone bodies. Biochem. Jour. 51: 218, 1952. 190. Thin, C., and Robertson, A. Biochem. aspects of rmiassmt ketosis. Jo<asr. Comp. Path, and Ther. 63: 184, 1953. 191. Todd, J. R. An experiment on bovine ketosis using two different types of silage. Brit. Vet. Jour. 114: 414, 1958. 192. Trombr©poul©s, E. G., and Klelber, M. The metabolism of specifically labelled glucose in bovine ketosis. a. Jour. 80: 414, 1961, 193. Vigue, R. F. Evaluation of eeaae concepts of bovine ketosis from a practitioners standpoint. Jour. Amer. Vet. Med. As®. 127: 101, 1955. 194. Vigue, R. F. The use of adrenal corticosteroids in dairy cattle. Can. Vet. Jour. 4: 137, 1963. 195. Wakil, S. I.„ and Bressler, R. Fatty acid metabolism and ketone body formation. ,Im:'- Lipid metabolism Clin. Expt. 11: 742, 1962. 196. Wateya K. Studies on the action of adrenalin on f a t t y acid metabolism, especially on ketone formation. Med. Jour. Osaka Univ. 6: 397, 1955. 197. Weeth, H. J. Effects of a simulated snowbound stress condition on ewes. Jour. An. S c i . 18s 694, 1959. 198. Weinhouse, S. Me des, G., and Floyd, N. F. Fatty acid metabolism. The mechanism of ketone body synthesis from f a t t y acids with isotropic carbon as the tracer. Jour. B i o l . Chem. 155s 143, 1944. 199. West, E. S., and Todd, W, R. Textbook of Biochemistry. 3rd e d i t i o n . The MacMillan Co., New York, 19(62. 200. White, R. R., C h r i s t i a n , K. R., and Williams, V. J. Blood chemistry and-hematology i n sheep on decreasing l e v e l s of feed intake followed by starvation. New Zealand Jour. S c i . Tech. 38s 440, 1956. 201. Wieland, 0., Matschinsky, F, L o f f l e r G., and Muller, U. The dependence of l i v e r ketone body formation on the reduced dlphosphopyridine nucleotide l e v e l . Biochem. et Biophys. Aeta. 53s 412, 1961. 202. Wieland, 0., and Weiss, L. Increase i n l i v e r acetyl coenzyme A during ketosis, Biochem. et Biophys. Res. Comm. 10s333, 1963. 203. Williams, R. H. Ketosis, Arch. Int. Med. 107s 69, 1961. 204. Williams, W. ~F. Lee, S. D., Hsad, H. H. and Lynch, J. Growth hormone effects on bovine plasma f a t t y acid concentration and metabolism. Jour. Dairy S c i . 46s 1405, 1963. 205. Young, F. ©. Growth hormone and experimental diabetes Jour. C l i n . End. l i s 531, 1951. '''' 94. APPENDICES I 95. APPENDIX I Record of Lamb Weights Animal Dated (Day/Month) Weights (lbs)  Ration Tag No. 18/6 25/6 2/7 9/7 16/7 23/7 6/8 20/8 27/8 1 (barley) 226 49 49 56 58 66 70 73 80 89 228 56 55 64 68 73 77 83 96 98 229 48 52 60 62 67 72 81 99 100 230 55 59 67 70 79 82 99 102 106 231 48 50 50 52 61 64 71 80 84 232 47 49 55 60 67 72 77 84 90 233 57 64 70 74 75 86 96 109 111 234 50 52 60 68 74 76 83 91 97 235 62 68 73 74 83 86 93 102 108 173 83 89 98 100 104 112 , 109 127 136 172 84 88 97 98 105 107 l i 7 126 136 x'~ 58.8 62.2 68.9 72.1 78.5 82.2 89.3 I 99.6 104.8 2 (barley-grass) 216 218 52 50 60 55 66 62 64 64, 62 77 68 73 79 75 86 83 95 88 220 64 64 72 70 77 82 87 90 94 221 52 59 65 68 74 78 78 90 96 222 43 44 45 47 52 56 59 65 72 224 54 59 66 66 71 73 74 72 89 225 55 59 64 64 70 74 75 77 84 206 60 70 76 78 82 89 91 105 108 207 52 56 64 66 74 78 85 87 93 215 57 55 57 58 60 65 66 75 84 169 93 92 102 102 109 116 121 134 140 X 57.4 61.2 67.2 67.9 73.4 78.8 82.0 88.7 95.8 3 (grass- 208 49 50 51 56 62 63 63 69 71 a l f a l f a ) 209 49 48 52 51 50 56 58 66 72 210 55 60 60 61 70 71 75 76 83 212 44 48 49 50 54 59 62 66 70 213 45 49 52 54 60 58 60 62 66 214 61 65 66 69 76 81 77 82 91 219 52 56 58 64 70 74 82 87 89 223 52 56 61 64 72 75 79 85 88 X 50 54 58 63 71 73 81 94 100 167 79 84 85 84 92 94 99 102 108 156 99 104 106 106 118 118 126 136 141 211 54 58 62 63 68 69 68 72 74 X 57.4 61.0 63.3 65.4 71.9 74.3 "1 1-Ki-U. -77.5 83.1 87.8 96. APPENDIX I I Record of Lambing Ration Ewe No. Date of B i r t h Lamb No. Weight at " (day/mo./year) B i r t h (lb.) 1 (barley) 226 9/4/66 Aborted 233 28/4/66 549 8.5 173 28/4/66 546 8.0 229 28/4/66 Died 234 29/4/66 547 9.5 172 30/4/66 536 9.0 213 12/5/66 542 6.5 2 (barley- 216 15/2/66 Aborted grass) 207* 7/4/66 Aborted 169 12/4/66 405 9.0 225 4/5/66 533 10.0 220 29/5/66 544 7_5_ 3 ( g r a s s - a l f a l - 212 23/2/66 Aborted fa) 214 10/4/66 402 10.5 208 10/4/66 Died X* 10/4/66 401 7.5 10/4/66 404 9.0 156 11/4/66 403 10.5 210 11/4/66 Died 209 24/4/66 406 9.0 211 29/4/66 Died 223 29/4/66 537 10.0 219 30/4/66 J538 11.0 *twin lambs 97. APPENDIX I I I Data Obtained i n Growth Hormone Study Treated Acetone (mg./lOO ml.) Lactic Acid (mg./lOO ml.) Glucose (mg./lOO ml.) VFA (meq./l.) pregnant 211 2.0 48 53 2.04 223 3.0 17 69 1.69 225 2.8 14 53 1.20 220 4.4 18 58 1.42 172 8.4 30 67 2.16 229 4.0 15 68 X 3.8 23 61 1.70 non-pregnant 156 0.8 16 57 1.12 X 3.2 16 53 2.18 167 4.4 13 54 — 216 f 2.2 26 70 1.46 206 2.2 23 63 1.59 215 3.6 32 51 1.44 232 0.2 14 50 1.05 235 2.2 7 58 226 1.8 14 58 X 2.2 18 57 1.47 98. Data Obtained i n Growth Hormone Study Controls Acetone Lactic Acid Glucose VFA meq/1. No. Pregnant 219 1.6 14 45 2.30 209 4.6 76 58 - - -210 4.8 12 55 1.09 169 0.6 18 51 1.48 218 7.2 23 52 _ _ _ 234 1.4 13 53 1.85 173 0.4 54 62 1.83 213 0.8 23 55 233 2.2 17 61 1.11 236 x » 2.6 250 x - 27.7 492 x 5 5 9. 66 x - 1.61 non-preg- 212 3.6 15 45 3.83 nant 214 2.6 15 62 1.77 208 2.8 12 55 2.44 o,/ 222 3.4 22 57 — 224 1.6 25 62 — 207 2.4 16 56 1.35 221 2.8 20 63 0.75 228 2.6 14 48 1.81 230 14 68 1.16 218 x - 2.72 153 x =17.0 536 x » 60 13.11 'x-1.87 

Cite

Citation Scheme:

        

Citations by CSL (citeproc-js)

Usage Statistics

Share

Embed

Customize your widget with the following options, then copy and paste the code below into the HTML of your page to embed this item in your website.
                        
                            <div id="ubcOpenCollectionsWidgetDisplay">
                            <script id="ubcOpenCollectionsWidget"
                            src="{[{embed.src}]}"
                            data-item="{[{embed.item}]}"
                            data-collection="{[{embed.collection}]}"
                            data-metadata="{[{embed.showMetadata}]}"
                            data-width="{[{embed.width}]}"
                            async >
                            </script>
                            </div>
                        
                    
IIIF logo Our image viewer uses the IIIF 2.0 standard. To load this item in other compatible viewers, use this url:
http://iiif.library.ubc.ca/presentation/dsp.831.1-0104368/manifest

Comment

Related Items