EXPERIMENTS BEARING ON THE FUNCTIONS OF THE LIVER IN THE METABOLISM OF FATS. I. BY H. S. RAPER. (From the Department of Pathological Chemistry, University of Toronto.) (Received for publication, January 27,1913.) Since the publication of Knoop’s experiments on the oxidation of phenyl substituted fatty acids in the animal body, a large amount of work has been directed to the object of determining the exact nature of the intermediate reactions involved in the breakdown of fatty acids to the ultimate end products, carbon dioxide and water. It would occupy too much space to give a com- plete summary of this work and would, further, be needless, since an excellent account of the work has recently been published else- where.’ It is sufficient to say that most of the results, so far obtained, support the hypothesis that in the catabolism of fatty acids the long chain of carbon atoms is gradually broken down by a successive series of /%oxidations involving the removal of two carbon atoms at a time. The first oxidation product is a P-ketonic acid or possibly a fl-hydroxy acid, though much of the evidence points to the conclusion that 8-hydroxy acids are produced by reduction of the ketonic acids. In some instances, the formation of o$-unsaturated acids has been observed, but whether they arise by direct oxidation or .indirectly, by the removal of the elements of water from a p-hydroxy acid, is not yet decided. Most of these exceedingly interesting results have been obtained by observations on the fate of phenyl or furfuryl derivatives of the lower fatty acids and it has been assumed that the same series of changes is the one which represents the mode of oxidation of the higher fatty acids when they are utilized as a source of energy in the animal body. This assumption receives support from the fact, estab- lished by Embden and his co-workers, that on perfusion of a sur- viving liver with blood containing caproic, octoic or decoic acids, 1 Dakin: Oxidalions and Reducliotis in the Animal Body, Longmans, Green and Company, 1912. 117 by guest on November 20, 2020 http://www.jbc.org/ Downloaded from
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EXPERIMENTS BEARING ON THE FUNCTIONS OF THE LIVER IN THE METABOLISM OF FATS. I.
BY H. S. RAPER.
(From the Department of Pathological Chemistry, University of Toronto.)
(Received for publication, January 27,1913.)
Since the publication of Knoop’s experiments on the oxidation of phenyl substituted fatty acids in the animal body, a large amount of work has been directed to the object of determining the exact nature of the intermediate reactions involved in the breakdown of fatty acids to the ultimate end products, carbon dioxide and water. It would occupy too much space to give a com- plete summary of this work and would, further, be needless, since an excellent account of the work has recently been published else- where.’ It is sufficient to say that most of the results, so far obtained, support the hypothesis that in the catabolism of fatty acids the long chain of carbon atoms is gradually broken down by a successive series of /%oxidations involving the removal of two carbon atoms at a time. The first oxidation product is a P-ketonic acid or possibly a fl-hydroxy acid, though much of the evidence points to the conclusion that 8-hydroxy acids are produced by reduction of the ketonic acids. In some instances, the formation of o$-unsaturated acids has been observed, but whether they arise by direct oxidation or .indirectly, by the removal of the elements of water from a p-hydroxy acid, is not yet decided. Most of these exceedingly interesting results have been obtained by observations on the fate of phenyl or furfuryl derivatives of the lower fatty acids and it has been assumed that the same series of changes is the one which represents the mode of oxidation of the higher fatty acids when they are utilized as a source of energy in the animal body. This assumption receives support from the fact, estab- lished by Embden and his co-workers, that on perfusion of a sur- viving liver with blood containing caproic, octoic or decoic acids,
1 Dakin: Oxidalions and Reducliotis in the Animal Body, Longmans, Green and Company, 1912.
aceto-acetic acid was produced, this being inferred from the increased amount of acetone obtained from the blood at the end of the experiment, whereas acids containing an odd number of carbon atoms gave no aceto-acetic acid. The excretion of the “acetone bodies” in the urine in human or experimental diabetes may also be accepted as evidence in favor of the above assump- tion, since in these conditions it is beyond doubt that fat is being used up in relatively large quantities as a source of energy. The fact that in such conditions, and especially in the experimental ones, the liver often contains a larger amount of fat than normal, has been taken to indicate that it is to this organ that the fat first comes when it is to be oxidized, since the other active organs of the body do not exhibit this fatty infiltration to anything like the same extent. When, however, we come to examine the nature of the fat in the normal liver, it is found to contain a much larger amount of unsaturated fats than that in the fat depots or, more correctly, the fat of the liver taken as a whole is more unsaturated than the connective tissue fat. This is also true of the other active organs, such as the kidney and heart, but the difference is not quite so marked. The exact meaning of the presence of these unsaturated acids is not yet clear. Does the presence of these highly unsaturated acids indicate that they represent a stage in the normal breakdown of fat in the liver, or does it represent a stage in the synthesis of fat or are the acids essential constituents of the protoplasm of the cell which enable it to carry out functions not directly associated with the catabolism of these acids them- selves?
With the knowledge at present at our disposal, it is not possible directly to answer any of these propositions, and it was with the object of trying to clear away some of the difficulties that the present work was started. We know so little about the synthesis of fats in the liver that the second possibility just mentioned cannot be put to the proof. If, however, we accept the view of Leathes, that fatty acids are synthesized by the condensation of acetalde- hyde, with coincident oxidation and reduction at the appropriate stage, then it is easy to understand how unsaturated acids could arise in such a process by removal of the elements of water from the hydroxy aldehydes or acids .formed, just as P-hydroxybutyric aldehyde or acid loses water and is converted into crotonic alde- hyde or acid as the case may be.
Some interesting experiments on the formation and distribution of the unsaturated acids in the liver have been carried out by Leathes and his co-workers.2 It has been shown that the fatty acids, contained in the lipoid substances of liver, are as a rule more unsaturated than those present as glycerides, so that it seems prob- able that some of the unsaturated acids are destined to be built up into phosphatides and it is possible that the liver may be spe- cially concerned in their manufacture. In a further series of experi- ments, in which animals (rats and cats) were fed with oils having a high iodine value, it was found on examination of the fat from the various organs that the liver fatty acids had a higher iodine value than that of the oil administered, no such change being observed in the fat from the other organs. The amount of fat, too, in the liver, appeared to be larger than normal in some cases and this also was true to some extent of the spleen. From these experi- ments it was concluded that “the liver takes up fat conveyed to it by the blood and changes the fatty acids in such a way as to increase their power of absorbing iodine. This may be interpreted as due either to the introduction of new unsaturated linkages, or to the transposition of existing ones from situations in which they are less liable to saturation by halogens to others in which they are more so.” Another possibility may be added, namely, that the liver has some selective affinity for thk highly unsaturated acids and takes them up in preference to the more saturated ones.
The work described in the present paper was undertaken w&h the object of determining definitely, if pbssible, whether saturated higher fatty acids, when taken up and metabolized in the liver, became unsaturated, as it was thought that this would tell us whether the phenomena observed by Leathes and Meyer-Wedell were due to a selective affinity of the liver for unsaturated acids. In order t.o accomplish this, it was necessary to have carried to the liver some saturated acid that could be separated subsequently from the acids normally present. in the liver and examined with respect to its power of combining with iodine.
2 Kennaway and Leathes: Proc. Roy. Sot. Med., 1909; Leathes and Meycr- Wedell: Proc. Physiol. Sot., 19OQ.
In view of Friedmann’s results3 on the fate of methylated amino-acids in the organism, which showed that the methyl- amino group in these acids apparently protected them from complete oxidation, it was decided to use phenylamino-stearic acid, C1sH3i-CH(NHCeHs)COOH, in the hope that, although the a-carbon atom might be immune from attack, the long chain of carbon atoms might still be open to oxidation. Moreover, phenylamino-stearic acid is insoluble in petroleum ether and can therefore be easily separated from the fatty acids normally present in the liver. The acid, however, was not easily absorbed, so this substance had to be rejected. Further, the small amount of acid absorbed was apparently oxidized for none could be found in the liver or connective tissues. This was confirmed by examining the urine for p-amino-phenol. After making the urine alkaline with sodium carbonate and extracting with ether, a residue was obtained from the ether which gave a very strong indophenol reaction. Enough was not obtained for further identification. A similar result was obtained when cu-phenylamino-hexoic acid was administered. In this case 4 grams of the acid given to a cat of 3.7 kilos proved fatal. The phenylamino group in these acids is therefore not immune from attack.
The next experiments were of a different nature. Higher fatty acids substituted with bromine in the or-position were administered and an attempt was made to separate the bromo-acids from the tissue or organ fatty acids by converting the former into phenyl- amino acids-a change which is easily brought about by heating them with aniline. The free acid, however; even when mixed with meat, usually caused vomiting in cats, so this method of experiment was abandoned. In one case, in which a cat absorbed 7 grams of the acid in eighteen hours, none could be detected in the liver or connective tissues by means of aniline.
The next method chosen was the administration of cocoanut oil of which about 40 per cent of the fatty acids are volatile in steam; these acids moreover da not take up iodine. The method adopted was to separate the volatile acids by steam distillation from the liver of animals to which the oil had been administered and determine whether they had become unsaturated by estimating
3 Friedmann: Beitr. z. them. Physiol. u. Path., xi, p. 158.
the iodine absorption. Since the small amount of higher fatty acids, separated from the fatty acids of a normal liver by steam distillation, take up iodine, the method adopted had to be a differ- ential one. Some of the experiments indicated that the acids recovered in this way from the liver had acquired the property of absorbing iodine to a slight extent. The effect was not very marked and it does not appear to be sufficiently striking to prove the assumption that the liver desaturates these acids. It is possi- ble, however, that only the higher fatty acids, with 16, 18 or 20 carbon atoms, undergo this action to a marked extent, especially if the acids produced have to be built up into phosphatides.
The oil was administered to the animals by three methods, namely, by the mouth, by introducing soap solutions into the small intestine and by intravenous injections of fine emulsions of the oil. In the course of the experiments, some interesting results were obtained which have a bearing on the selective functions exercised by the liver in the mobilization of fat. Since cocoanut oil contains about 40 per cent of acids, volatile in steam, it was possible to estimate, roughly, the total amount of oil in the vari- ous tissues, after administration of a given amount of cocoanut oil to an animal. Many factors influence the rate at which vola- tile acids distil in steam so the results are only roughly quantitative. Controls have shown however that they are accurate enough to enable one to interpret them in the way that has been done.
When oil is given, mixed with food, it is not absorbed very rapidly and, after short periods of five to twelve hours, about 6 per cent of the oil absorbed is found in the liver. In the connective tissue fat its presence is just detectable. By infusion of cocoanut oil soaps into the intestine about 30 per cent of the absorbed fatty acids is found in the liver. The condition of the intestine at the end of these experiments was very abnormal, so too much stress cannot be laid upon the results. By intravenous injection of fine emulsions of the oil, none of the administered oil could be detected in the connective tissues, but from 25 to 60 per cent was found in the liver. This remarkable selective activity exhibited by the liver in the taking up of fat from the circulating blood is probably due in part to the effect of the anaesthetic. It was found, for instance, that a dog, which had cocoanut oil given by the mouth and lat,er half a grain of morphia followed by ether anaesthesia, had
twice as much of the oil in its liver as dogs similarly fed but receiv- ing no morphia and ether. The amount of oil in the liver was however, even then, not so great as in a dog to which the oil was given intravenously. Another factor which probably has an influ- ence on this selective action of the liver is the rate at which the oil enters the circulation. In the feeding experiments, the oil was absorbed by cats, rarely at a greater rate than 0.5 gram per hour, whereas in the intravenous injections it was given at the rate of 0.77 to 1.37 grams per hour.
Another factor which would explain in part the divergence be- tween the results of the feeding experiments and the intravenous injections is the possibility that the lower fatty acids in cocoanut oil, for instance, capric and lauric acids, may be absorbed, not as glycerides but as sodium salts and may not display the same tend- ency to accumulate in the liver as the glycerides do. It is obvi- ous, when we consider the absorption of the sodium salts of the whole series of fatty acids, the lower members of which are ab- sorbed without further change and the higher members largely synthesized into glycerides before entering the circulation, that there must be a gradual transition from one method of absorption to the other. It is possible then that intermediate acids, such as capric and Iauric, reach the circulation partly as sodium salts, when their glycerides are introduced into the intestine; andsince these acids are among those which were used to localize the cocoa- nut oil in the tissues because of their volatility in steam, the amount of volatile acid obtained from any organ would only indi- cate the minimum amount of cocoanut oil in that organ, because the fat entering the organ might contain a smaller percentage of volatile acids as glycerides than the original cocoanut oil. This hypothesis was tested by administering cocoanut oil to a dog and collecting the lymph from the thoracic duct during absorption of the oil. It was found that the mean molecular weight of the acids present in the lymph as glycerides was much higher than that of the acids in the oil given and was still higher than that theoretically calculated allowing for the fat normally present in lymph. The divergence in results obtained in the feeding experiments and intra- venous injections may therefore be partly due to the fact that some of the lower acids from the cocoanut oil enter the circulation as sodium salts and consequently do not accumulate in the liver.
One other possibility remains to be considered, namely, that the fat taken up by the liver, when fine emulsions were introduced intravenously, was retained mechanically and the phenomenon was therefore purely one of obstruction. None of the experiments disprove this conclusively, as an emulsion has not yet been obtained so fine that a small fraction of the fat given does not remain in the lungs or spleen. This occurs with emulsions so fine that the major- ity of the fat particles are less than 2~ in diameter and certainly not more than 4,~~ The emulsions were injected into the external jugular vein so that all the fat had to pass the pulmonary capil- laries before entering the systemic circulation. In the earlier experiments, in which the emulsions were not so fine as in the later, a considerable amount of the fat remained in the lungs, the lung capillaries thus acting as filters and retaining the larger particles. Of the fat which finally entered the systemic circulation, none was found in the connective tissues, the intestinal mucous mem- brane or the kidneys, whereas the liver contained 30 to 40 per cent. With the finest emulsions, which resembled chyle in micro- scopic appearance, less than 2 per cent of the oil given was retained by the lungs and less than 3 per cent by the spleen, whereas 25 per cent was in the liver. It seems probable, therefore, that this rapid fatty infiltration of the liver by the injected cocoanut oil is a phenomenon identical with that observed in conditions of lipaemia, such as, for instance, occurs in dogs after excision of the pancreas. There seems to be every indication, therefore, when fat is brought rapidly into the circulation, that it is taken up selec- tively by the liver. It is hoped to make further use of this selec- tive function, in a manner similar to that described in the present series of experiments, in order to determine the changes which occur in fatty acids other than those of cocoanut oil when taken up by the liver.
EXPERIMENTAL PART.
The estimation of total fatty acids in the liver, when carried out, was done by Liebermann’s method of direct saponification, and all iodine values were determined by Wij’s method.
For the separation and estimation of the volatile acids, the organ was heated with about its own weight of 50 per cent caustic potash, until complete solution was accomplished, then alcohol was
added and the heating continued until saponification of the fat was complete. The liquid was transferred to a large flask, acidified with sulphuric acid and distilled in a current of steam; 2.5 liters of distillate were collected in every case. The distillate was made alkaline with caustic potash in order to dissolve the fatty acids and then evaporated to small bulk on the water bath. This resi- due was transferred quantitatively to a Liebermann flask and the fatty acids soluble in petroleum ether estimated in the usual way. From a portion of the fatty acids obtained, the small amount of unsaponifiable matter usually present was removed by converting the acids into soaps in 60 per cent alcohol and extracting the solu- tion vith petroleum ether. The purified fatty acids separated from the soap solution by acidification and extraction with petro- leum ether then served for the determination of the mean molecular weight.
For the volatile acids from the connective tissues, a fair sample of connective tissue from the subcutaneous tissues, the omentum and the subperitoneal fatty tissues was taken and, after saponifica- tion and liberation of the fatty acids, was distilled in steam, 2.5 liters of distillate being collected. A control, obtained by adding 1 gram of cocoanut oil to 80 grams of connective tissue fat and sub- sequent saponification and distillation in steam, showed that the actual weight of volatile acids obtained from the connective tissue fat gave very little quantitative indication of the amount of cocoa- nut oil present but the mean molecular weight of the volatile acids immediately showed that the presence of cocoanut oil in the con- nective tissues could be detected in this way.
For the experiments on cats, four control experiments were carried out. Three with the livers of normal cats without any addition, and one with a cat’s liver to which a gram of the oil had been added before saponification. Two controls were also carried out with dog’s liver on similar lines. The results are collected in table I.
It will be seen, on reference to this table, that from a normal cat’s liver, on an average, 109 mgm. of volatile acids soluble in petro- leum ether were obtained, with a mean molecular weight of about 250, and these absorbed 61.4 mgm. of iodine. When 1 gram of cocoanut oil was added to the liver and the subsequent treat- ment was the same, then the amount of volatile acids obtained
rose to 424 mgm. The iodine absorbed by these acids was 49.2 mgm. Similarly, with roughly the same amount of dog’s liver, 123 mgm. of volatile acids were obtained with a molecular weight of 255.? and absorbing 65.9 mgm. of iodine. When cocoanut oil was added to the same amount of the same liver, an increased yield of volatile acids was obtained with a corresponding decrease in their molecular weight and a slight diminution in the iodine absorption.
Feeding experiments.
In the experiment on cats, the oil was usually mixed with minced lean meat or with boiled codfish which contain less than 1 per cent of fat. When the’animal was killed, the contents of the alimentary tract were taken and the fat estimated, in order to gain some idea of the amount of oil absorbed. In the experiments with dogs, the oil itself was given by the mouth and was usually readily taken. As in the experiments with cats, the stomach and intestinal contents were worked up to find the amount of oil actu- ally absorbed. The results are collected in tables II and III.
On reference to table II, it will be seen that more volatile acids are obtained from the liver than in the control cats. In three out of the four experiments, the iodine absorption of the volatile acids is higher than in the controls, the most marked result in this direction being shown by the cat which had been receiving oil for nine days. In this experiment the iodine absorption was 50 per cent above the control average. In spite of this, the desatura- tion of the volatile acids can only have taken place to a slight extent, as in the most marked case, with aniodine absorption of 92 mgm., it only accounts for a rise in the iodine value of the volatile acids from 0 to 6. In all four cases the mean molecular weight of the volatile acids from the connective tissues is below the control, showing that some of the oil had been retained there. The result is most marked, as would be expected, in the cat which has received the largest amount of oil. In the other three cases, in which the animal was killed within twelve hours of administration of the oil, if allowance be made for the amount of volatile acids from a normal liver, and assuming that the amount of volatile acids obtained is equivalent to two and a half times its weight of oil, we find that between 5 and 6 per cent of the oil absorbed was in the liver.
In the experiments with dogs (table III) the presence of cocoa- nut oil in the liver is also revealed by the amount of volatile acids- obtained, but it is not quite as marked as in the experiments with cats. The case in which the most notable increase in volatile acids occurred is the one in which the dog was given morphia and ether subsequent to the administration of the oil. In only three, out of the five experiments, is the iodine absorption above the average for the controls, being highest in the dog which had been anaesthetized for four and one-half hours. Three grams of ure- thane and ether for only half an hour appears to produce little effect on the accumulation of oil in the liver.
Infusion of soap into the intestine.
The object of these experiments was to get rapid absorption and with this the possibility of a greater accumulation of the oil in the liver. It was hoped thereby to get more satisfactory evidence as to the desaturation of the saturated volatile acids than was obtained from the feeding experiments. In these experiments, cats were used. Urethane was given, followed by ether, and cannulae were inserted into the intestine just below the pylorus and just above the ileo-caecal valve, the lower cannula being iixed to a bu- rette. The solution to be administered was injected periodically through the upper cannula, and at the end of the experiment the intestinal contents were removed in order to determine the exact amount of fatty acid absorbed. It was found that whenever the fluid infused contained chiefly soaps, the intestine at the end of the experiment was very congested and the mucous membrane had partly desquamated so that the condition of the absorbing area had been very abnormal. On the other hand, when emulsions containing fatty acids or free hydrochloric acid were introduced, the intestine at the end of the experiment appeared quite normal and healthy. In spite of the numerous and varied mixtures intro- duced into the intestine, containing sometimes oil, sometimes free fatty acids or soaps, together with bile salts and glycerin, and although acid, neutral and alkaline fluids were tried, in only four of the experiments was enough absorption obtained to make it worth while searching for volatile acids in the liver. The results are collected in table IV.
The experiments show that the accumulation of the oil in the liver was much more marked than in the feeding experiments, the amount varying between 25 and 33 per cent of the fatty acid given. The iodine absorbed by the volatile acids was also higher than in the feeding experiments and still higher than in the controls. These experiments, then, indicate that desaturation of the volatile saturated acids from the oil has taken place, but, again, only to a slight extent.
Intravenous injection of emulsions.
The object of these experiments, as also that of the previous set, was to introduce fat rapidly into the circulation with the hope that more would be taken up by the liver and in consequence give a better chance for the detection of desaturation if it occurred. In all the experiments performed, the amount of fat taken up by the liver was greater than in the feeding experiments, but only slight evidence of desaturation was obtained. In the earlier experiments, the emulsions were relatively coarse, and rather less than half the oil given was retained, by the lungs. * These emulsions were made by shaking the oil with a 0.15per cehtemulsion of lecithin in normal saline. The emulsions usually contained about 4 per cent of oil. The amount of oil was estimated before each experiment so that the amount introduced could be accurately determined. It was discovered later that much finer emulsions could be made by using caseinogen instead of lecithin as emulsifying agent. These emul- sions were prepared as follows.
Four grams of casein were heated on the water bath in a porcelain dish with 25 cc. of normal saline and 4 cc. of decinorrpal sodium hydroxide until solution was practically complete; 8 grams of cocoanut oil were now added and the mixture rubbed up with a pestle until drops of oil were no longer visible. The dish was now removed from the water bath and the rubbing up continued until with gradual cooling the mass became gelatinous. If too much saline is used to dissolve the caseinogen the mass does not set when cold and the emulsion is not quite as fine as when the liquid sets to a jelly on cooling. In case the cold mixture is still creamy, it should be warmed up again until a little water has evaporated and the grinding continued until it is cold. An alkaline salt solution was now made containing 160 cc. of normal saline and 1 cc. of decinormal soda for each gram of oil used. The gelatinous mass was rubbed up in a mortar in small portions with a few cubic centimeters of the salt solution until dissolution was complete.
Finally the creamy solution thus obtained was diluted with the remainder of the salt solution, filtered through cotton wool and finally through filter paper. If the preparation of the emulsion has been successful, the whole of the fat should be in minute particles which show Brownian movement. No particles with a diameter greater than 4s should be present and these should be few in number. Almost the whole of the fat is usually in particles less than 2~ in diameter. On standing in the cold, practically no separa- tion takes place.
The method of administration was as follows:
Urethane and ether were used as anaesthetics and, in the case of dogs, mor- phine and ether. A cannula was inserted into. the external jugular vein and this was connected to the burette containing the emulsion so that the rate of entry of the fluid could be easily adjusted. The burette was sur- rounded by the outer tube of a Liebig’s condenser through which water was allowed to circulate at a temperature of 40” C. About 100 cc. of the emulsion was usually given in a couple of hours, the rate of administration of the oil being then about 2 grams per hour. The animal was kept alive for a further two to four hours, and then killed.
The results of these experiments are collected in table V. With the exception of the last experiment, the method of determining the amount of oil in the liver was the same as in the previous experi- ments, that is, by distillation in steam. In the last experiment, the amount of oil going to the liver was determined by excising a lobe of the liver previous to the injection of the emulsion and 1 hen comparing the amount of fat in this lobe with the amount in the rest of the liver at the end of the experiment. The result is approxi- mately the same as obtained by the distillation method and serves to confirm the results obtained by it. In this case the spleen was examined for the presence of cocoanut oil by comparing the mean molecular weight of the acids from a normal spleen with that of the acids found in the stileen after the injection of the oil, the mean molecular weight of the acids in the oil being 212. In this case it was foupd that 0.09 gram of the oil was present in the spleen. It will be seen, from the tabulated results, that there is marked evi- dence that the fat accumulates in the liver, but again the phenome- non of desaturation is only slightly indicated, the mean iodine absorption of the volatile acids being 71.9 mgm. as opposed t,o 61.4 mgm. in the controls.
In conclusion, two experiments were performed on dogs in which, two hours after a meal of cocoanut oil, an attempt to collect the
lymph flowing from the thoracic duct was made. The object of these was to determine whether the fat in the chyle had the same composition as that administered, or, if of different composition, whether the difference was caused by a failure of all the volatile acids to undergo the glyceride synthesis. In one of the experi- ments, although 56 cc. of lymph were collected, it contained so little fat in addition to what one might normally expect to be pres- ent in lymph, that no deductions in reference to the point in ques- tion could be made. In a second experiment only 7.5 cc. of lymph were coilected as the animal died owing to a misadventure wit.h the anaesthetic. The lymph however contained 5.37 per cent of fat and was sufficient for the purpose of the experiment. On examination of the fatty acids obtained from the chyle fat, the following results were obtained: Mean molecular weight, 236; iodine value, 19.1. The iodine value of the oil given was 7.7 and the mean molecular weight of its fatty acids, 212. Assuming the iodine value of the fat normally present in lymph to be 90, and its molecular weight 284, which are approximately what one would expect, then if enough of the normal fat were present to raise the iodine value from 7.7 to 19.1 the mean molecular weight would only be 222, whereas it was found to be 236. It thus appears as if the lower fatty acids in cocoanut oil do not completely undergo the glyceride synthesis during absorption.
SUMMARY.
1. Cocoanut oil administered to cats or dogs by the mouth can be detected in the liver in five or six hours. The amount present after times varying from five to twelve hours does not exceed 6 per cent of that absorbed.
2. If cats be anaesthetized (urethane and ether), and a solution of cocoanut oil soaps containing glycerin and bile salts be run into the small intestine, then about 30 per cent of t,he absorbed fatty acid is found in the liver.
3. When cocoanut oil is given to cats or dogs intravenously in the form of a very fine emulsion, containing about 4 per cent of the oil, then from 25 to 60 per cent of the oil which enters the systemic circulation is found in the liver.
4. It is probable that the greater retention of the oil by the liver, when it is administered in the form of soap or a fine emulsion, is
partly due to the anaesthetic and partly to the rapidity of adminis- tration.
5. When cocoanut oil is being absorbed, the fat in the chylecon- tains fatty acids with an average higher molecular weight than those in the oil administered. It is probable, therefore, that the lower fatty acids in the oil are partly absorbed as sodium salts.
6. The volatile acids obtained from the liver in the above sets of experiments absorbed more iodine than the volatile acids from normal livers. The increase was not great but it probably indi- cates that saturated fatty acids containing 10, 12 or 14 carbon atoms may become unsaturated in the liver.
I wish to express my thanks to Dr. V. E. Henderson for assist- ance in the thoracic duct experiments and to Professor Leathes for much valuable help and criticism.
