Oct., 1914
T H E JOL'RS.1L O F I N D I ' S T R I A L A N D E S G I S E E R I S G C H E M I S T R Y
tions it is capable of causing local irritation of the mucous membranes with which it comes in contact. 111-Prolonged ozonization is capable of ridding, a t least in a measure, the atmosphere of food storage rooms of germ life, probably through rendering it an unsuitable medium for their support. IV-Ozone is a valuahle adjunct to ventilation, its function being the destruction of odor with consequent partial purification of the air; there is nothing either in the theory or in recorded experience to warrant its use for curtailing ventilation. V-Eggs may be preserved longer with the aid of ozone than under similar conditions without. VI-The analogy between laboratory tests and actual practical applications is often so obscure and replete with modifying factors that the extremest care must be exercised in applying the results of experimental observations t o practice. SPR.\Gl.F:
? d L E C T R l C XT'ORKS,
BLOOMFIELD, N. J.
THE IMPORTANCE OF ENZYMES AND ENZYME REACTIONS LN MEDICINE AND SURGERY' By \V. G . LYLE A K D P. A . KOBER
IT-e wish to present for your consideration this evening a brief review of the advances that have taken place in the methods of obtaining and identifying, for diagnostic purposes, some of the ferments present in the human body. These vary quantitatively so much, even under normal conditions, that only their absence, or marked diminution, is of importance in determining t o what extent the organ itself is diseased. We shall first discuss the digestive enzymes. Saliva contains a ferment called ptyalin, or better, salivary amylase. I t s activities are restricted t o the hydrolysis of the starches, converting them into maltose and a small amount of glucose. The ferment acts best in a slightly acid medium, strong mineral acids destroying it. Owing to the short time that food remains in the mouth, the amount of starch digested there is small. The ptyalin, after reaching the stomach, is destroyed by the hydrochloric acid present but many pieces of starch embedded in larger masses of food are protected from its action and may still undergo digestion, due t o the action of the saliva swallowed with them. It is estimated that this starch digestion in the stomach may proceed for a t least thirty minutes. In cases where the hydrochloric acid is low, or absent, pronounced digestion of starch will occur. An interesting fact observed by Warfield and Koelker is that saliva contains another ferment, erepsin, which is capable of splitting some of the peptids to the amino acid stage. The reason for the presence of this enzyme in the mouth is not clear; however, it is probably of no digestive importance, being derived from the cells which line the ducts of the glands, from the mucous membrane of the mouth, and from the bacteria present there. I n the stomach we find the strong proteolytic enzyme pepsin, acting in an acid medium. Its function seems t o be t o break down the coarser colloidal particles into a non-coagulable state, preparatory to their further digestion by the enzymes of the upper intestinal tract. Peptic digestion of proteids in the stomach does not proceed as far as the formation of peptids and amino acids; on this fact is based a diagnostic test for cancer, known as the glycyl-tryptophan reaction. Theoretically, in the normal stomach, the di-peptid, glycyl-tryptophan, cannot be broken up into glycine and tryptophan by pepsin, whereas, if cancer is present this will take place. The tryptophan thus liberated is tested for with bromine. The splitting of this peptid may be due to a n enzyme present in the carcinoma itself, or it may be brought about by the action of erepsin, a ferment widely distributed in the cells of the body, and undoubtedly secreted during the rapid proliferation and breaking down of the cancer cells. The practical difficulty with this test is that Presented a t the hTew York Section of the A. C . S., Chemists' Club, June 5 , 1914.
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there may be a regurgitation into the stomach of intestinal contents containing trypsin, which, of course, will split the peptid, and give the reaction, even if no carcinoma is present. For this reason the test has not proved satisfactory. Pepsin and hydrochloric acid are absent in a disease of the stomach known as .A4chyliaGastrica, in which the mucous membrane becomes atrophied, and the glands do not secrete the ferment. The absence of pepsin and hydrochloric acid is also found in many cases of cancer of the stomach, and is a factor of diagnostic importance. It is interesting to note that in some cases where these are not present in the gastric contents, patients may go for years with apparently little digestive disturbance, as the protein is probably digested by the intestinal enzymes, thus showing the comparative unimportance of peptic digestion. The pancreatic juice, which is poured out through a duct into the upper part of the small intestine known as the duodenum, continues the digestive process begun in the stomach, and breaks down the protein mass to the amino acid stage. Besides this proteolytic ferment called trypsin, the pancreas secretes a nuclease, an amylase, and a lipase. Owing to its deep retroperitoneal position, and consequent difficulty in gaining any information as to its condition by palpation and methods available for other organs, the study of its ferments has been utilized as a diagnostic help in diseased conditions, the supposition being that if they were absent, or greatly diminished, either the ducts through which these were discharged into the small intestine were obstructed, or the gland itself seriously diseased. Unfortunately, the amount of these ferments normally present is so variable, that unless they are completely absent or only traces of them found, great care must be taken in drawing conclusions as to the functional activity of the gland. However, cases have been reported by several observers where no ferments were found in the secretion of the upper intestine, and the diagnosis of an obstruction, or disease of the gland, was made, this afterwards being confirmed by operation or autopsy. Great advances have been made in the study of these enzymes by the use of the duodenal tube. The apparatus consists of a thin rubber tube, a quarter of a n inch in diameter, and 150 cm. in length. It is weighted at the end with a n olive-shaped metal ball, perforated on the sides. The patient swallows this tube on retiring, and is given a glass of milk in the morning. After an hour the contents are aspirated, and tested for the presence of ferments. This has a great advantage over the stool examination method, as we are able to obtain the ferments in a fairly pure condition. The usual test for trypsin is the one recommended by Oscar Gross. This consists of a solution of one-half gram of casein dissolved in a liter of sodium carbonate (solution 1-1000). Duodenal contents are added to this casein solution, incubated for about eight hours, and then tested with a I per cent acetic acid. If the mixture is digested, showing the presence of the ferment, the solution will be clear; if no digestion has taken place, a precipitate is thrown down. Other methods used for determining the presence of trypsin are the digestion of gelatin either in tubes or in plates, the digestion of fibrin, and of egg albumin. Amylase is tested for by a method introduced by Wohlgemuth. He uses six small test tubes into which are put 5 cc. of a I per cent soluble starch solution. Tube I serves as a control; t o the remaining five tubes are added o.oj, 0.25, 0.5 and 1.0 cc. of duodenal juice, diluted one-half with distilled water. The tubes are incubated for about thirty minutes and t o each is added a drop of -V)'IOiodin solution. The tube is selected as positive which shows an entire disappearance of all the blue color. Lipase is best determined by incubating a mixture of the duodenal juice with triacetin for a given length of time, then titrating the acid formed in the usual way. It is necessary in doing this to run blanks. As yet not a sufficient number of cases of pancreatic disease
T H E J O U R N A L OF I X D U S T R I A L A N D E N G I N E E R I N G C H E M I S T R Y have been investigated, and the results correlated to the findings a t operation and autopsy, to establish the value of these methods on a firm diagnostic footing. But it is likely that with the development of better technique they will prove of great aid to the surgeon and physician in diagnosing these obscure conditions. Before closing, we must mention some of the more recent clinical tests which are the direct result of enzyme investigations. I-A method for determining urea quantitatively in the urine and blood. 2-A test for the diagnosis of pregnancy. uREAsr2-~o~~owingthe discovery of Takeuchi that the soya bean contains a urease, a method based on this observation has been developed for estimating the amount of urea in the urine and in the blood. The procedure is briefly as follows: A given quantity of the extract, or powder, of the dried soya bean is added to the material to be tested, and the ammonia which is formed from the urea is estimated in the usual way. The advantage of the testcis that it is rapid and specific. Abderhalden’s test for pregnancy is based on the fact that foreign proteins circulating in the blood call out specific ferments which attack them. The test is as follows: A placenta, or after-birth, is ground up, washed free from all water-soluble material, and placed in a dialyzing bag with some serum from a suspected case. If the woman is pregnant, ferments specific for placental tissue are present in her serum. These will digest the placental tissue contained in the bag, and the dialysate will give the test for protein with ninhydrin, a reagent which gives a blue color with it. If she is not pregnant no ferments are present, and consequently the ninhydrin reaction is negative. A similar method is being tried for the diagnosis of cancer, using the cancer tissue instead of placenta, and dialyzing it with serum from the suspected case. The fact that proteins introduced into the circulation other than through the intestinal tract are split up by ferments is of great importance in the study of anaphylaxis and in diseases like hay fever, urticaria, asthma and peculiar disturbances in the nervous control of the circulation. It seems that proteins taken through the mouth are split up by the digestive juices in such a way as to deprive them of their specificity, and
Dudley breathed his last after a sudden stroke of paralysis. During recent years, Dr. Dudley had been in poor health and his many friends became much concerned about him, especially when two years ago he retired from active participation in his professional, educational and sociological work. Born April 16, 1859, in Covington, Kentucky, Dudley did not see the allotted three score years, but that part of his life following the attainment of his majority was full of hard work, enriched by good he did on every hand, and blessed with spontaneous affection from all those who enjoyed his friendship. Prepared in the public schools of Covington, he entered the .University of Cincinnati from which institution he was graduated in 1880 with the degree of Bachelor of Science. Last June he was the recipient of the degree of Doctor of Laws from his dma mater. Dudley early exhibited his interest in chemistry, coming as a student under the influence of F. W. Clarke, then professor of physics and chemistry in the University. As a junior in college1 he prepared a selenocyanate, an analogue of Buckton’s double sulfocyanate. While a senior under the direction of With F. W. Clarke, “Some Selenocyanates,” Be?., 11 (1878), 1325.
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they are not absorbed until this has taken place. This would point to some regulating mechanism for preventing the reabsorption of heterogeneous albumin. It is probable that the feeding of excessive amounts of foreign proteids is responsible for the condition known as hives, for the disorders following the eating of shell fish and strawberries, and probably for some of the digestive disturbances found in infants fed on cow’s milk. I n hay fever, the continued absorption of proteids present in pollen, produces, in sensitized individuals, the well known symptoms of the disease. Anaphylaxis, or in English, hypersensitiveness, is the opposite condition to protection. The word was coined to describe the peculiar effect which certain poisons possess of increasing, instead of diminishing, the sensitivity of the organism through their action. A hypersensitiveness occurs after the injection or absorption of a foreign proteid, and becomes manifest on the repetition of the injection. For example, if a guinea pig is injected with a given non-fatal amount of horse serum! and the injection repeated a t regular intervals, after a time the animal develops an immunity, and is able to tolerate a dose much larger than the original one. Whereas, if an interval of time is allowed to elapse, say ten to fourteen days after the first injection, the second dose will produce a train of symptoms, in severe cases, characterized by great restlessness, rapid and labored breathing, collapse and death. It is well known that some people are very susceptible to horse serum. This should be borne in mind if it is necessary to inject diphtheria antitoxin, as these very alarming and sometimes fatal anaphylactic phenomena occur, especially if t h e patient has had a previous, protective dose. In this connection it is interesting to note that some people are unable to ride behind a horse without attacks of sneezing, running of the eyes, etc. Some asthmatics who are sensitized to horse proteid have their attacks brought on in this manner. This brief r6sum6 suggests the great importance of ferment investigation in the practice of medicine and surgery. HARRIMAN RESEARCH LABORATORY ROOSEVELT HOSPITAL, NEW YORKCITY
OBITUARIES WILLIAM LOFLAND DUDLEY On the morning of Tuesday, September 8, 1914, en route from Clifton Springs Sanitarium to his home on the Campus of Vanderbilt University in Nashville, Tennessee, William Lofland
Val. 6 , N o .
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Professor Clarke, he isolated a new volatile alkaloid, “spigelina,” from Spigelia marilandica, or pink root.’ The youthful bachelor of science served as demonstrator of chemistry for one year in the Miami Medical College of Cincinnati during which time he worked out a modification of Bottger’s subnitrate test for sugar* and a new test for gallic acid.3 During the same time he published a work on t h e “Chemical Examination of Urine’I4 and a chart to be used by students in their study of urine. He a t once became professor of chemistry and toxicology, in which capacity he served that institution for six years, when he resigned to accept the Chair of Chemistry in Vanderbilt University in Nashville, Tennessee. In 1885, Miami conferred upon him the honorary degree of Doctor of Medicine. While a t Miami he carried out an elaborate investigation on tobacco smoke, the poisonous principle of which he determined to be carbon monoxide.6 He was prominent among the music-loving people of Cincinnati, possessing a rich voice which charmed many in solos and ensemble 1 “Preliminary Notice of a New Volatile Alkaloid,” J . A n . Chem. Soc.. 1 (1879). 286; and A m . Chem. J.. 1, 154-5. 2 “Laboratory Notes on a Modification of BBettger’s Test for Sugar,” A m . Ckem. J . , 2 (1880). 47. a “A New Test for Gallic Acid,” Am. Ckem. J., 1 (ISSO), 48. 4 Broadside, Cincinnati (1880). 5 “Poisonous Effects of Cigarette Smoking,” Medical N n v s (Philadel-. phia), 1883, p. 53.
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