The Clinical-Chemical Interface of Medical Science: Its Development in This Century*

The Clinical-Chemical Interface of Medical Science: Its Development in this Century*

A. BAIRD HASTINGS, P h .D ., D .S c .

Department of Neurosciences, School of Medicine, University of California, San Diego, La Jolla, CA 92073

When I seriously sat down to prepare high state of the medical sciences. But each my remarks to you this evening, I was became involved in applying chemistry to aghast to see the title that I had given clinical problems through a fortuitous state Dr. Sunderman. Who am I to presume to of circumstances. speak on the “Development of the Clinical- Let me briefly introduce them to you. Chemical Interface of Medical-Science in First is Donald Van Slyke, born in 1883 this Century?” It’s a damn good title, but of Dutch descent, who left us in 1971. He I’m certainly not qualified to live up to it. worked hard in the lab and played tennis What I’d like to hear is a panel of friends throughout his life. He was unbeatable at such as Donald Van Slyke, Otto Folin, Wil either. liam Mansfield Clark and John P. Peters Second is Otto Folin, born in Sweden in gathered here to discuss the subject. So I 1867, who was head of biochemistry at invited them—and they accepted—or at Harvard from 1907 until his death in 1934. least let’s imagine that they have. Each in He invented many quantitative methods his way made technological and conceptual useful in clinical practice and was a much developments that have led to the present loved teacher. * Presented at the Association of Clinical Scien Third is William Mansfield Clark who tists’ Banquet, March 9, 1974, La Jolla, CA. lived from 1884 to 1964 and spent most of 213 214 HASTINGS his life as head of physiological chemistry Here he had intimate contact with a small at Johns Ilopkins. His contributions were group of clinical investigators and the re in acid-base and oxidation-reduction bio sponsibility of applying chemistry to the chemistry. He was a perfectionist in his solution of selected clinical problems and research and was highly respected by his here he remained until he reached the re students. tirement age of 65—after which he had a Finally, there was John P. Peters, always second productive career at the Brook- called Jack, born in 1887, who was Pro haven National Laboratory, dying while fessor of Medicine at Yale from 1921 until still active at age 88. He never really aged. his death in 1955. A scholar in medicine, One can say Van Slyke saw clinical who was feared and revered by his res chemistry develop; he contributed person idents and never compromised on prin ally to much of it. (As his first assistant ciple, Jack teamed up with Van to write from 1921-1926, I had the privilege of ob the classic two volume work Quantitative serving what was happening at the Clin- Clinical Chemistry. ical-Chemical Interface and of participat We’ll try to get a word from each of ing, at times, to a minor extent). them. This was the period when the chemical description of the body fluids in terms of mass and composition in health and disease was under investigation. Methods had to be devised for quantitative determination of individual constituents. Since existing chemical methods usually required large Donald D. samples of blood, they had to be modified Van Slyke and adapted to accommodate what were

1 8 8 3 - then regarded as small samples—meaning 1 9 7 1 one cubic centimeter of blood or other fluid. Quantitative measurement of the end products of the chemical reactions involved were limited to gravimetric, titrimetric, colorimetric and gasometric procedures. In his time, Van Slyke made use of all four— D o n a l d V a n S l y k e although he is best known for his ingenious Van Slyke had taken a Ph.D. in 1907 in use of gasometric techniques. organic chemistry under Moses Gomberg Each of our guests was young at the who discovered organic free radicals. He start of this century. Three, at least—Van had expected to follow in his father’s foot Slyke, Folin and Clark—were well equipped steps and become an agricultural chemist. to advance the chemical knowledge of the However, through a chance conversation day. The fourth, Peters, the M.D. of the between his father and P. A. Levene at a four, had the foresight to visualize a prom meeting of the American Chemical Society, ising future for the use of sound quantita he became an assistant to Levene at the tive chemical knowledge in the solution of newly-formed Rockefeller Institute for clinical problems and the ability to make Medical Research. Seven years later, he it stick. was selected by Dr. Simon Flexner, the What can they be said to have left us? director, to head the chemistry division of First and foremost their students. I’ll wager the Hospital of the Rockefeller Institute. that every one of you medical scientists in CLINICAL-CHEMICAL INTERFACE OF MEDICAL SCIENCE 2 1 5 this audience has come under the influence been exposed to Emil Fischer’s pioneer of one of these students—or if not directly work on amino-acids and proteins: we —then one of their student’s students. would try to study the fate of proteins in What is taken for granted today was a the body. Folin adapted the Nessler re completely blank wall in the early 1900’s. action for the determination of ammonia Let me enumerate some of the things, following Kjeldahl digestion of proteins to which today you would regard as old hat the Dubosq colorimeter. Van Slyke took but for which they had to break ground: advantage of the reaction of nitrous acid with aliphatic amino acids with the evolu 1. The inorganic composition of the ex tion of gaseous nitrogen and devised a glass tra and intracellular fluids of the body. apparatus for its measurement. Both Folin 2. The transport of oxygen and C 02 and and Van Slyke used their respective meth the control of salt, water and acid-base bal ods to study the digestion of proteins, the ance. absorption of amino-acids and their metab 3. The metabolism of fats, carbohydrates olism in the tissues. Both greatly advanced and proteins by the several tissues. our knowledge. Folin was led to believe Subjects unknown to them in their early that all tissues were equally able to de- days were: aminate amino acids, whereas Van Slyke showed that the liver is the primary site of 1. The role played by vitamins and hor amino acid metabolism and urea formation. mones in metabolic control. When Van Slyke left Levene’s laboratory 2. That proteins were chemical indi for the Hospital of the Rockefeller Insti viduals. tute, he took his responsibility for aiding in 3. The composition and mode of action the study of disease through chemistry very of enzymes. seriously. Since F. M. Allen was then ap 4. Bioenergetics and biochemgenetics— plying the so-called starvation diet to the two subjects still under active investigation. treatment of diabetes, Van Slyke, with 5. Organelles of cells—their structure Glen Cullen, undertook to devise a method composition and functions. for the early detection of acidosis,-—an in 6. Membranes of all kinds and how evitable lethal concomitant of diabetes in transport is effected through them. the preinsulin days. This led to his sim 7. Immunochemistry. ple and adequately accurate gasometric 8. How the nervous system acts to method for plasma bicarbonate concen achieve a coordinated desirable metabolic trations. result. The method proved clinically useful in Included in this abbreviated list are all sorts of disease states involving acid- some topics that are essentially settled and base abnormalities and led Van Slyke, in others that are still under investigation and 1921, to his classic characterization of acid- obscure. base abnormalities which has stood the test For a moment, let us transport ourselves of time. back to the early 1900’s. What kind of H. “Tell us about your early lab days, Van.” questions would we first attempt to an swer? Remember, we’d have no isotopes, Van Slyke speaks: “First of all, I never set no chromatography, no sophisticated spec- out to be a clinical chemist. I was fasci tro-photometers—not even any glass elec nated by amino acid and protein chemistry trodes. and their metabolism in the body. During I expect we would do about what Van my seven years in Levene’s laboratory, I Slyke and Folin did, particularly if we had had the good fortune to literally dream up 2 1 6 HASTINGS a gasometric method for measuring amino gaseous state such as oxygen, C 02, CO and acid nitrogen. It was quite accurate and nitrogen. With the able help of Folch, with it Gus Meyer and I were able to show Sendroy and other fellows, gasometric that the liver is the primary organ for methods were developed for amino-nitro- amino acid metabolism and urea formation. gen, urea, lipids, sugar, lactic acid, chloride “After I transferred to the Hospital of the potassium and calcium. These proved use Rockefeller Institute to head its depart ful until they were supplanted by modem ment of chemistry, I was responsible for technology. chemical studies on the diseases under in “After I went to Brookhaven, I was able vestigation at the Hospital. At various to improve considerably the determination times these were diabetes, pneumonia and of the isotope C14 through adapting the nephritis. However, I never gave up my manometric apparatus for its measurement interest in and work on what might be of C140 2 in the gaseous state. In keeping called basic biochemical attacks on physio with the times, I also enjoyed, at Brook logical problems such as transport of oxy haven, microing the manometric apparatus gen and C02, control of aeid-base bal and methods for determinations on samples ance, the role of the kidney in ammonia in the microliter range instead of the milli formation and clearance of urea from the liter range. blood. “Of course, I didn’t do these things my “Nor did I ever completely stop work on self. Throughout the 42 years at the Insti amino acids. In 1920 while determining the tute and my 22 years at Brookhaven, I had hexone bases in a gelatin hydrolysate, I a constant procession of young associates encountered a discrepancy which led, (over 70 in all) in the laboratory who many years later, to the identification of stayed with me for a few years and then a new amino acid,—hydroxylysine. This moved on,—usually to professorships in the amino-acid has recently proved to be of academic world. Some had M.D. or Ph.D. great importance in membrane structure degrees when they arrived; a few received through its linkage with mucopolysac their Ph.D.’s while with me. Though I charides. never was officially a professor, I feel that “Although we tried to complete, to the I’ve done my share of teaching at the clin best of our methodology, each problem ical-chemical interface and would like to that we undertook before undertaking an be remembered for that quite as much as other, we made some oversights. One of for the publications from my laboratory.” these was our failure to recognize the pres H. “Well, Van, if I were to ask you what ence of carbonic anhydrase and of carb- gave you the greatest pleasure among the amino C 02 in red blood cells. Both of these many contributions you have made, what were within our reach during our experi would you say?” ments on C 0 2 transport by and distribution in blood, but we missed recognizing them. Van Slyke. “Working out the behavior of You were responsible for that, Baird.” blood as a physical-chemical system from the properties of its constituents, partic if. “Yes, Van. A little knowledge of thermo ularly hemoglobin. This required the deter dynamics is a dangerous thing.” mination of the concentrations of water Van Slyke. “I must confess some satisfac and inorganic constituents extra-cellularly tion over having the manometric gasomet and intra-cellularly, the concentrations and ric apparatus so readily adaptable to the distributions of oxygen and carbon-dioxide determination of a large number of sub in free and combined form and the titration stances other than those already in the curves of oxy and reduced hemoglobin. CLINICAL-CHEMICAL INTERFACE OF MEDICAL SCIENCE 217

From three known physical-chemical rela Chemistry and head of the department at tions ( electroneutrality, osmotic equilib the Harvard Medical School in 1907 where rium and the Gibbs-Donnan Law ), we he remained until his death in 1934. were able to account for the changes that Folin s early interests were centered occur in blood when its pH is changed or around protein metabolism and the devel its degree of oxygenation is altered. opment of quantitative methods applicable “Had it not been for the need for a more to such studies. The Dubosq Colorimeter, accurate method to measure blood gases in with which he had become familiar these studies, the manometric blood gas while in Germany in 1897, became his apparatus might not have been developed. favorite instrument for quantitation. He is “It also made me think through what was specially remembered for his methods and meant by buffer value in theoretical terms. metabolic studies of nitrogen, urea, ammo This was my best theoretical contribution.” nia, uric acid, creatinine, sugar and a sys tem of blood analysis developed with the H. “Thank you, Van.” late Professor Hsien Wu. He was a great and beloved teacher. In this connection, I cannot resist a digression. Shortly before I left the University of Chicago for Harvard in 1935, I received a telephone call from Professor Stiegliz ask ing me to come to his office. Since I barely Otto knew Professor Stiegliz, I wondered what Folin I had done. When I got there, he said: “Sit 1 8 6 7 - down, Mr. Hastings. I hear you are going 19 3 4 to Harvard. I want to tell you a story. A year or so after the University of Chicago opened in 1891, I was assigned a young graduate student. In time he came before the chemistry faculty for examination for his Ph.D. degree. The examination of my student was in the afternoon, following the O t t o F o l in examination of one of Professor Neff’s stu My next distinguished panelist is Otto dents in the morning. Professor Neff’s stu Folin. His entry into a career of medical dent had done brilliantly, answering all the chemistry differed greatly from that of Van questions put to him; but my student did Slyke,—though strangely enough their con miserably, answering hardly any of the tributions and interests often supplemented questions. However, since I was young and each others. this was my first graduate student, the Folin emigrated from Sweden to this faculty voted to pass him. Professor NefFs country in 1882 when he was 15 years old. student was never heard from again, but After undergraduate study at the Univer my student was named Otto Folin. Good sity of Minnesota, he earned a Ph.D. in day, Professor Hastings.” 1898 in organic chemistry while working H. “Professor Folin, what gave you the under Professor Julius Stieglitz at the then greatest pleasure of your contributions?” young University of Chicago. After eight years as a research chemist at the McLean Folin. “I suppose it was the night I de Hospital outside of Boston, Folin became livered my Harvey Lecture on Blood Anal Hamilton Kuhn Professor of Biological ysis in New York in 1920. Some years be 218 HASTINGS

fore, Hsien-Wu and I had introduced a Professor Benedict will also find it useful.’ number of methods for analyzing blood for I presented this so-called Folin-Wu tube clinically important constituents, among to him on the spot. I never heard any more them a method for determining blood criticism of the method from him.” sugar. The method was based on the reduc H. “Thank you, Professor Folin.” tion of cupric ion to cuprous oxide in an alkaline copper tartrate reagent, and then the reaction of the cuprous oxide with phospho molybdic acid, ending up with a blue solution of phospho-molybdous acid which was proportional to the glucose pres ent. Strict attention to temperature and time of heating was essential. My good William M. friend, Stanley Benedict, professor of bio Clark logical chemistry at Cornell Medical 1 8 8 4 - School, had taken exception in print to the 1 9 6 4 accuracy of our blood sugar method. His point was that reoxidation of the cuprous copper occurred erratically and much too readily. He had illustrated this with data showing what happened when you heated the solution longer and shorter times than we had prescribed. W i l l i a m M a n s f i e l d C l a r k “On the night of the lecture, I had re H. “Now, let’s hear what Bill Clark might quested that he sit in the front row directly have to say. I’ll let him speak for himself.” opposite the rostrum. I described the var ious methods that had been proposed for Clark. “Well, I never dreamed that I would measuring blood sugar on one cc samples. spend most of my scientific life in a medical Then I described our method and the crit school or do research that would be of icisms that had been leveled at it by my value to medicine. I wanted to be a first- friend Professor Benedict. Then, I said: ‘I rate physical chemist. That’s why I went did not say to heat the solution for eight to Hopkins and worked on osmotic pres minutes or fifteen minutes, but exactly ten sure measurements under Professor Morse. minutes.’ And I took up the other points That’s when I learned to be satisfied with of criticism in like manner. nothing less than the maximum accuracy “Then I said: ‘But I have devised a way attainable. I also learned to be a pretty to circumvent even those who do not good glass-blower. choose to follow directions,’ and I drew “My first job after getting my Ph.D. was from my pocket a standard reaction tube, as a research chemist in the dairy division with the bottom portion containing the re of the Department of Agriculture where actants drawn down so that it was con I was assigned to find out why there are nected with the upper portion by a four cm holes in Swiss cheese. This led me to the isthmus with only eight mm diameter. ‘This study of the metabolism of molds and mi cuts down reoxidation by air to an inconse croorganisms and their ability to produce quential amount. I find that Harvard med C 0 2 and other acids. I needed better meth ical students can now obtain accurate ods to measure pH and better understand values for blood sugar, and I hope that ing of acid-base equilibria in general. We CLINICAL-CHEMICAL INTERFACE OF MEDICAL SCIENCE 219 managed to design a rocking hydrogen “I never lost my love of glass-blowing, electrode which could measure the pH of which I would resort to for relaxation when protein containing fluids in a C 0 2 contain frustrated, or to golf where I could test the ing atmosphere and we synthesized a new ravages of time on my accuracy. To have series of acid-base indicators sulfaphtha- made three holes in one—one of them after leins, that covered a wide pH range. These I retired—was a special dividend in a were both widely used for years. happy life as a biochemist at the interface “In 1920 I put what was then known between medicine and chemistry.” about acid-base chemistry, the methodol H. “Professor Clark, what would you say ogy involved and the applications to biol were your major contributions to medi ogy, bacteriology and industry together in cine?” a book called the Determination of Hydro gen Ions. It must have met a need because Clark. “My graduate students and my med it went through three editions and four ical students. The former have contributed reprintings. much to today’s biochemical knowledge “Since the hydrogen electrode can be and the latter are practicing sound clinical thought of as measuring the potential be medicine. My book, Topics in Physical tween (atomic) hydrogen and hydrogen Chemistry, has been of some help to medi ions, it is a special case of an oxidation- cal investigators.” reduction potential. This led me and my H. “Thank you, Bill.” associates to a systematic study covering many years of reduction potentials of re- versibly oxidizable organic compounds with and without biological origin and im portance. The effect of pH changes and of coordination with ligands, which influ enced the potentials, led to the concept of an oxidation continuum, culminating in John P. what we now call the electron transport Peters chain. 1 8 8 9 - “I did part of this work at the Hygienic 1 9 5 5 Laboratory (now the NIH) of the Public Health Service between 1920 and 1926 and most of the work at Johns Hopkins where I was professor of physiological chemistry from 1927 to 1952. “Since I didn’t know much about bio chemistry as applied in medical schools, I J o h n P. P e t e r s took gross anatomy with the Hopkins med H. “Now, Jack, tell us how you got into ical students the first year I was there. This clinical chemistry. Was it in the year helped me, but it didn’t seem to help the 1920-21 when you were with Van, in his students learn what I considered they laboratory at the Hospital of the Institute.” should know. However, we both learned together and by the time I retired, I had Peters. “Well, in a way and it had some a great respect for medicine and the stu exciting moments. Franklin McLean came dents had a good idea of what biochemis breezing in from Peking and sold Van on try could do for medicine. taking on the job of proving whether L. J. 2 2 0 HASTINGS

Henderson was right or wrong about the faster than we could keep up with it,—at effect of oxygenating hemoglobin of differ least to both our satisfactions. The upshot ent pH’s on chloride and bicarbonate dis was that I brought out a second edition of tribution in the blood. It meant that Van Interpretations in 1946 without the chap had to invent a more sensitive apparatus ters on hemoglobin and acid-base balance for O2 and C 02 determinations and that and the inorganic ions. Van kept working I was responsible for the equilibration of on the revision of these and the methods blood with gases of different C 02 and 0 2 volume as long as he lived. tensions. The next year I went to Yale as “Though our collaboration was a fortu an associate professor of medicine, and nate one, it was difficult because he was there I was to remain for the rest of my accustomed, as a chemist, to be content life. It was where our book Quantitative with nothing less than proven accuracy, Clinical Chemistry was bom. whereas I was used to being confronted “The publishers, Williams and Wilkins, constantly with disease manifestations in asked me to write a modest handbook for patients,—no two of which were the same. clinicians describing useful chemical meth It made it difficult to satisfy both of us at ods and discussing their application to the same time on any one subject. How clinical problems. It was originally to be ever, the first edition seems to have been called “Quantitative Chemistry in Clinical worth doing.” Medicine.” I soon found that it was going H. “Now, Jack, what do you think the de to be a bigger job than I could handle velopment of biochemistry and other med alone, and I asked Van Slyke to join me ical sciences has done for medicine and the in writing it. He agreed and we made an treatment of disease in patients?” outline and divided up the writing of the first draft of the chapters between us. We Peters. “Well one thing is clear. It has made also agreed to exchange each chapter until it more expensive. It has also made the it met the satisfaction of both of us. This medical school curriculum longer and more may have improved the accuracy and com difficult. It has resulted in a generation of pleteness of the chapters, but it made pub clinicians who not only have more under lication of the book delayed until 1931; standing of the basis of many diseases, but each chapter turned out to be a mono it has given them the tools to improve di graph. It also had grown to two volumes,— agnoses, to make prognoses more accurate one of 1200 pages with 21 chapters called and to follow the effects of therapeutic pro Interpretations dealing only with the physi cedures. All the medical sciences have ological and clinical significance of these made this possible. substances for which quantitative methods “But when this is said, there remains still were available, and a second volume called the art of the physician to be reckoned Methods of about 1000 pages containing with,—the need of the individual patient those methods, described in detail, that had for that intuitive recognition by his physi been proven accurate and useful. Some cian of physical and mental symptoms and where along the line, the title was short behavior born of clinical experience and ened to Quantitative Clinical Chemistry. compassion. We can correct abnormal mass “It must have met a need because the and composition of body fluids; we can first edition sold out rather quickly here modify metabolism of fats, carbohydrates and abroad and the publishers soon asked and proteins by diet, drugs and hormones; us to prepare a second edition. God knows but we are still primarily dependent on the we tried, but medical research progressed wisdom of the body to heal itself. It is the CLINICAL-CHEMICAL INTERFACE OF MEDICAL SCIENCE 2 2 1 wise physician who knows when to stop closer the workers are drawn together, the tinkering.” clearer becomes their community of pur pose and the more significant to the wel H. “Thank you, Jack.” fare of mankind the upbuilding of natural knowledge.” In spite of the progress made in this cen So, ladies and gentlemen, you have heard tury toward an accurate computerized de snatches of what it was like to be groping scription of man as an organized hetero at the clinical-chemical interface fifty or geneous chemical system for the useful more years ago. I hope that my imagination transfer of chemical energy, the whole and voice will be forgiven. remains much bigger than the sum of its Was it a dawn that was breaking? Are parts. The medical scientist has magnifi we at high noon today? Or is today more cently enlarged the knowledge of man in likely another dawn? health and disease. The physician still I can think of no more fitting ending to stands in the position of its interpreter in this pleasant and profitable meeting of the the interest of the patient. But let us each Association of Clinical Scientists than to persevere to advance knowledge in that quote the Christmas thoughts from the manner for which he is best fitted. Each Sanscrit of my friend and contemporary, sound contribution will find its rightful the founding father of this organization, niche. F. William Sunderman, Sr.: As the late, wise William H. Welch so prophetically said in an address in 1907 on Look to this Day! It is Life. the Interdependence of Medicine with In its brief course lie all the Other Sciences of Nature, “It is well that Verities and Realities of Existence. the sciences of nature hold out attractions Yesterday is but a Dream, and to so many different types of mind, for the Tomorrow is only a Vision. edifice of science is built of material which But Today well lived must be drawn from many sources—the Makes every Yesterday a Dream of Happi deeper we can lay the foundations and ness; penetrate into the nature of things, the And every Tomorrow, a Vision of Hope.