Casimir Funk and a Century of Vitamins

Casimir Funk, 1884-1967

One hundred years ago, in 1911, the Polish-born biochemist Casimir Funk published his first work on vitamins, titled “Experiments on the causation of Beri-Beri,” in the British Medical Journal, The Lancet.  Curiously, the word “vitamin,” coined by Funk, was missing from this paper as the Lister Institute in London, for which Funk was working at the time, did not accept the designation. It was not until 1912 that Funk was able to use his new word in a formal publication, the term appearing for the first time in the Journal of State Medicine.

And so began a new path of scientific inquiry into vitamins as constituents of food necessary to the maintenance of good health – an entire discipline whose start can be attributed in large part to Funk, the “father of vitamin therapy.” That said, even one-hundred years ago, it is apparent that Funk drew from what was already known about deficiency-related diseases as he charted his own experiments.

Before Funk’s time, it had already been established that certain foods acted like drugs in their capacity to prevent certain diseases. In the mid-1700s, Scottish physician James Lind conducted an experiment on sailors suffering from scurvy, a disease which is characterized by spots on the skin, spongy gums, and bleeding from mucous membranes. This ailment plagued many sailors who were at sea for longer than fruit could be preserved, and was suspected to be caused by a lack of fresh foods.

Lind tested this hypothesis by feeding one group of sailors two oranges and one lemon every day, while several other groups of sailors consumed a different diet that included garlic, mustard seed, cider and sea water. The two sailors who were given citrus fruits recovered from their symptoms of scurvy, while the others remained in the same condition or worsened. Lind came to the conclusion that citrus fruits were the cure for scurvy. Casimir Funk later discovered that diseases similar to scurvy, such as beriberi and pellagra, were likewise the physical manifestation of a nutritional deficiency.

Kazimierz Funk (“Casimir” is an anglicized form of his given name) was born in Warsaw, Poland, in 1884. He became interested in pathology and physiology at the age of fourteen, and at sixteen he went to Geneva, Switzerland to study the natural sciences – just the start of what would prove to be a peripatetic academic career. Funk later studied chemistry at Berne for three years, eventually focusing on organic chemistry under the supervision of Stanislaw Kostanecki.

His interests in the human body and chemistry eventually led to a program of research on “trace elements” in humans, carried out in 1904 at the Pasteur Institute with Gabriel Bertrand.  Funk’s trace elements work relied upon his ability to synthesize both organic bases and amino acids. Two years later he moved to Berlin, which was the world’s most scientifically vital city at the time. In Berlin, Funk and Nobel laureate Emil Fischer, a leading organic chemist of the period, undertook an analysis of amino acids – the building blocks of proteins – specifically focusing on the structures of cysteine and alanine.

Emil Fischer

At the same time that he and Fischer were analyzing amino acids, Funk accepted a position as a biochemist working for the municipal hospital in Wiesbaden. It was there that he came to the conclusion that foods could be divided into two categories: food that encouraged tumors and that which discouraged them. Funk also observed that “poor” proteins seemed to be poisonous to animals.  In an experiment conducted with Emil Abderhalden  – at the time one of Fischer’s assistants – one dog was fed horsemeat mixed with glucose and butter, while another dog was fed gliadin with glucose and butter. In fifteen days, the first dog gained 150 grams of protein while the second dog lost 450 grams. Based on this data, the American duo of Thomas Osborne and Lafayette Mendel showed that gliadin and edestin are “poor” proteins.

In 1910 Funk began the studies that led to his discovery of the vitamins when he traveled to London to work at the Lister Institute, and met its director, Charles Martin. Martin and Funk discussed the disease beriberi, which is found in populations of people who eat polished rice but not in those who eat the rice polishings. Beriberi was at one time fairly common among populations where rice is a staple, specifically in east Asia, and in its final stages caused paralysis and death.

Funk knew of a disease in pigeons called polyneuritis that is equivalent to beriberi in humans – it occurred in pigeons that had been fed exclusively polished rice. It had been previously supposed by the English scientist Leonard Braddon, and later the Dutchman Christiaan Eijkman, that the endosperm of rice contains a poison, while the cortical layers – the “polishings” – contain the antidote. Funk, however, conducted preliminary experiments in which he administered a diet of various pure carbohydrates – such as starch, insulin, cane sugar and dextrin – and found that they all induced polyneuritis when administered alone. He came to the conclusion that there was no toxic agent at fault; rather, polyneuritis and beriberi were caused by a deficiency of some essential ingredient missing in polished rice.

From there, Funk performed a series of tests which fractured rice polishings into two sections, A and B. He gave one set of polyneuritis-stricken pigeons fraction A, and another set of pigeons fraction B. The pigeons which were given fraction A died, while the group that was given fraction B recovered. Funk further broke down fraction B, and discovered that very small “trace elements” of fraction B could cure pigeons of polyneuritis. He named this trace element a vitamin: “vita” meaning life and “amine” meaning a nitrogen-containing compound.  The word “vitamin” then, stands for “a life-sustaining compound containing nitrogen.” (Though as it turns out, Funk was mistaken about the “amine” part.) Funk named this first vitamin “B1,” now known as thiamine. He published his second paper on the vitamins, “On the Chemical Nature of the Substance which Cures Polyneuritis in Birds Induced by a Diet of Polished Rice,” in 1911.

Funk was sure that more than one substance like Vitamin B1 existed, and in his 1912 article for the Journal of State Medicine, he proposed the existence of at least four vitamins: one preventing beriberi (“antiberiberi”); one preventing scurvy (“antiscorbutic”); one preventing pellagra (“antipellagric”); and one preventing rickets (“antirachitic”). From there, Funk published a book, The Vitamines, in 1912, and later that year received a Beit Fellowship to continue his research.

For several years following the publication of his book, Funk served as director of the Hygiene Institute in Warsaw. At the Institute, he cured dementia symptoms in patients who suffering from pellagra by giving them vitamin B1 and adding yeast to their diet. Funk was correct in his supposition that vitamins are required for the proper metabolism of nervous tissues, and that the lack of vitamins causes the body to extract nutrients from its tissues, thus leading to weight loss as those vital resources are depleted. In 1922 Funk and Harry Dubin successfully created and marketed the first vitamin supplement containing vitamins A and D, found in cod liver oil. It was called “Oscodal” and was sold widely as a product used mostly in infant therapy.

Casimir Funk died in New York in 1967 at the age of 83.  His discovery of the vitamins is widely acknowledged as having catalyzed many more studies on and discoveries related to nutrition and health.  Among them was a 1928 project in which, after a number of efforts, physiologist Albert Szent-Györgyi was able to separate vitamin C from citrus fruits – the first instance of success in obtaining a pure vitamin. Within a few years, vitamin C (ascorbic acid) became recognized as a substance that greatly improved one’s health, and in the 1960s Linus Pauling began to take a special interest…

[Ed Note: Over the next four weeks we will mark the centenary of Casimir Funk’s discovery of the vitamins by examining a number of specific aspects of Pauling’s Vitamin C and the Common Cold.]

Advertisements

The Medical Research of Linus Pauling

By Tom Hager

[Ed Note:  In October 2010, Pauling biographer Tom Hager delivered a talk sponsored by the Oregon Health Sciences University which detailed and discussed the various contributions that Linus Pauling made to the medical sciences, including the controversy over his strong interest in orthomolecular medicine.   With the author’s permission, excerpts of this talk are being presented on the Pauling Blog over the next three posts.  The full text of Hager’s OHSU lecture is available here.  Those with an interest in learning more about Hager’s work, including his latest research on food issues and world hunger, are encouraged to visit his blog at http://thomashager.net.]

[Part 1 of 3]

Oil portrait of Linus Pauling, featuring a model of the alpha-helix in the foreground. 1951. Portrait by Leon Tadrick.

By 1939, at the age of 38, Linus Pauling was a full professor and head of the chemistry division at Caltech, as well as the father of four children (three sons, Linus, Jr., Peter, and Crellin; and a daughter, Linda).

He was also beginning to turn his considerable talents toward understanding the complicated molecules inside the human body. He started with proteins.

The Molecules of Life

Determining the structure of proteins at this time was a gigantic problem. Most were difficult to purify, easily degraded, and hard to characterize. Proteins appeared to be not only gigantic, comprising hundreds or thousands of atoms – much too large to solve directly with x-ray crystallography – but also relatively fragile, losing their function (denaturing) after even slight heating or mechanical disturbance. No one at the time was even sure that they were distinct molecules – one popular theory held that proteins formed amorphous colloids, gels that did not lend themselves to molecular study.

Studying them at the molecular level seemed an impossible task with the tools available in the late 1930s. But Pauling took on the challenge. He started with the building blocks of proteins, the amino acids, and directed his growing lab team toward pinning down their precise structures. Then he set himself to figuring out how they formed protein molecules, often building models out of wood, wire, and paper.

He based his approach in part on the ideas of the German biochemist Emil Fischer. Like Fischer, Pauling came to believe that proteins were long molecular chains of amino acids linked end-to-end. Working with Alfred Mirsky in the mid-1930s, Pauling discovered that the denaturing of proteins resulted from breaking weak bonds, called hydrogen bonds, that pinned these chains into specific shapes. Between the early 1930s and early 1950s he made a string of important discoveries about hemoglobin, antibodies (including the most sophisticated work at the time into the structural relationship between antibody and antigen), enzymes, and other proteins.

Foldable paper model of the alpha-helix protein structure published in the Japanese journal Chemical Field, 1954.

In May 1951, he put everything he knew into a celebrated series of seven papers detailing the structures of a number of proteins at the level of individual atoms, including the structure of the single most important basic form of protein, the alpha helix (a hydrogen-bonded helical chain that is a structural component of almost every protein). It was an astounding breakthrough, and it opened the door for an understanding of biology at the molecular level. Within two years, Watson and Crick had used his approach to decipher the structure of DNA.

Biological Specificity

But structure was not everything. Pauling realized that life resulted not from individual molecules, but from the interactions between them. How did organisms make offspring that carried their specific characteristics? How did enzymes recognize and bind precisely to specific substrate molecules? How did antibodies recognize and bind to specific antigens? How did proteins, these flexible, delicate, complex molecules, have the exquisite ability to recognize and interact with target molecules?

It all fell under the heading of biological specificity at the molecular level. Pauling directed much of his attention here during through the 1940s, performing a great deal of careful work on the binding of antigens to antibodies.

Drawings of antibodies and antigens made by Linus Pauling in the 1940s.

His findings were surprising. Pauling demonstrated that the precise binding of antigen to antibody was accomplished not by typical chemical means – that is, through covalent or ionic bonds — but solely through shape. Antibodies recognized and bound to antigens because one fit the other, as a glove fits a hand. Their shapes were complementary. When the fit was tight, the surfaces of antibody and antigen came into very close contact, making possible the formation of many weak links that operated at close quarters and were considered relatively unimportant in traditional chemistry — van der Waals’ forces, hydrogen bonds, and so forth. To work, the fit had to be incredibly precise. Even a single atom out of place could significantly affect the binding.

Having demonstrated the importance of complementary structure with antibodies, Pauling extended his idea to other biological systems, including the interaction of enzymes with substrates, odors with olfactory receptors, and to the possibility of complementary structure in genes.

Pauling’s idea that biological specificity was due in great part to complementary “fitting” of large molecules to one another proved to be essential in the development of molecular biology. His research now formed a coherent arc, from his early work on the chemical bond as a determinant of molecular structure, through the structures of large molecules (first inorganic substances, then biomolecules), to the interactions between large biomolecules.

He carried out much of this research during World War II, when he also worked on synthetic plasma substitutes and a fruitless search for ways to produce artificial antibodies.

He had already earned a place among the nation’s leading researchers in the medical applications of chemistry. But his greatest triumph was still to come.

Sickle-Cell Anemia

Toward the end of World War II, Pauling’s reputation was great enough to earn him an invitation to join a national committee that was brainstorming the best structures for postwar medical research. This committee’s work led to the foundation of the National Institutes of Health.

Pauling was the only non-physician asked to join the committee.

At a dinner with other members one night, talk turned to a rare blood disorder called sickle-cell anemia. One of his dinner companions described how red blood cells in the victims were twisted into sickle shapes instead of discs. The distortion appeared to hinder the blood cells’ transport through capillaries, resulting in joint pain, blood clots, and death. The disease primarily affected Africans and African Americans. What caught Pauling’s attention most, however, was one odd fact: Sickled cells appeared most often in venous blood, rather than in the more oxygenated blood found in the arteries.

Pastel drawing of sickled Hemoglobin cells, 1964. Drawing by Roger Hayward.

He thought about this during the next few days. From his previous work with blood, he knew that red cells were little more than bags stuffed with hemoglobin. He had also shown that hemoglobin changed its shape slightly when it was oxygenated. If the red cells were changing shape, perhaps it was because the hemoglobin was altered in some way. What if the hemoglobin molecules in sickle-cell patients were changed in some way that made them clump, stick to one another, as antigens stick to antibodies? Perhaps something had changed that made the hemoglobin molecules complementary in shape. Perhaps adding oxygen reduced the stickiness by changing the molecules’ shape.

He presented his ideas as a research problem to Harvey Itano, a young physician who was then working on his Ph.D. in Pauling’s laboratory. Itano, later joined by postdoctoral fellow John Singer, worked for a year trying to see if sickle-cell hemoglobin was shaped differently from normal hemoglobin. They found no detectable differences in any of the tests they devised. But they kept at it. Finally, in 1949, using an exquisitely sensitive new technique called electrophoresis that separated molecules by their electric charge, they found their answer: Sickle-cell hemoglobin carried more positive charges on its surface.

This was an astounding discovery. A slight change in the electrical charge of a single type of molecule in the body could spell the difference between life and death. Never before had the cause of a disease been traced to a molecule. This discovery – to which Pauling attached the memorable title “molecular disease” – received widespread attention. Itano and Singer’s followup work demonstrated the pattern of inheritance for the disease, firmly wedding molecular medicine to genetics.

Medical Chemistry

It was a great triumph – there was talk of a Nobel Prize in Medicine or Physiology for Pauling – and it led Pauling to make greater efforts in the medical field. He encouraged M.D./Ph.D. candidates, hired physicians to work in his laboratory, and began focusing his own research on medical problems, including developing a new theory of anesthesia.

He was ahead of his time. An example of what the atmosphere was like: Pauling noted that as he went around in the late 1940s seeking funds for a comprehensive marriage of biology and chemistry to attack medical problems, people at funding agencies were telling him that they found the term “medical chemistry” to be “a disturbing description.”

In the late 1950s, Pauling extended his concept of molecular disease to the brain. After reading about phenylketonuria (PKU) – a condition in which a mental defect can be caused by the body’s inability to metabolize an amino acid, phenylalanine, leading to a buildup of that substance and others in the blood and urine – Pauling theorized that the problem might be caused by a defect in an enzyme needed to break down phenylalanine. PKU, in other words, might be another molecular disease. Now interested in the possibility that there might exist a range of molecular mental defects, Pauling visited a local mental hospital, saw other patients whose diseases seemed hereditary, and decided to seek support for an investigation into the molecular basis of mental disease. The Ford Foundation in 1956 awarded him $450,000 for five years’ work – a vindication of Pauling’s approach and a tribute to his reputation. The grant, however, yielded little in the way of immediate results, with much of the funding going toward testing his (ultimately found to be mistaken) theory of anesthesia.

The long-term results were more significant. Pauling’s immersion in the field, thanks to the Ford grant, led him to read widely in psychiatry and general health, always on the lookout for another molecular disease that might lend itself to new therapy. By the mid-1960s he was coalescing his findings into another overarching theory, this one combining much of what he knew about chemistry and health. He called his new idea “orthomolecular” medicine.