Significant Scots - Sir Alexander Fleming

It is the lone worker who makes the first advance in a subject but, as the world becomes more complicated, so we are less and less able to carry through anything to a successful conclusion without the collaboration of others.

fleming

The events and the stages leading to a great discovery are many and complex, Fleming had 'found' penicillin. He had demonstrated the bactericidal power and the non-toxicity of the substance in its crude state. He had suggested its use in the treatment of wounds infected by vulnerable microbes, and had published an account of the favourable results of his tests. He had tried to get the chemists to purify it. Obstacles and mishaps of every description had made it impossible for any of thdm to carry their attempts through to the end. Nevertheless, in 1935, the two men who together were to resolve the problem, were converging on Oxford from two points very far removed from each other on the earth's surface.

Dr Howard Florey, an Australian, was born in Adelaide in 1898. From childhood he showed a lively interest in science and, more especially, in chemistry. While still studying medicine, he married a fellow-student, Miss Ethel Reed. She wanted to be a practising physician, and he to give all his time to research. He was awarded a Rhodes Scholarship which took him to Oxford. There he studied physiology and later, at Cambridge, pathology. All the major subjects attracted him and in all branches of medicine he was successful, for he had a quick and vigorous mind combined with great force of character.

In 1925 the Rockefeller Foundation sent him to the United States where he worked in a variety of laboratories and made a number of friends, among them Dr A. N. Richards of the University of Pennsylvania who was destined to play a part in the story of penicillin. After his return to England in 1929 he learned about the work Fleming had been doing on lysozyme, and immediately showed interest in that astonishing substance which is present in human tears and human nails and can instantaneously dissolve certain microbes. The year 1935 saw him appointed to a Chair of Pathology at the Sir William Dunn School in Oxford. This institution was a model of its kind. It was situated at one side of the University Parks, was admirably equipped and employed far more research-workers than St Mary's Hospital, since there was a whole group of laboratories, all under one roof — pathological, biochemical and bacteriological. There Florey continued his researches into lysozyme. No one could have been better suited to direct and co-ordinate the work of a team of scientists, for he was competent in each of their special subjects. He entrusted Dr Roberts with the task of extracting pure lysozyme, which Roberts succeeded in doing in 1937, as did also another young chemist, Dr Abraham. Shortly after taking up his professorial appointment Florey invited Dr E. B. Chain to Oxford, there to direct the work of the biochemical department.

Chain was born in Berlin in 1906, of a Russian father and a German mother. Since his father was an industrialist dealing in chemical products, he had decided, while yet a boy, to make chemistry his profession. As a student at the University of Berlin he took an especial interest in the chemistry of the human body (biochemistry) and made physiology his second subject. He had already obtained his doctor's degree when the Nazis came to power in 1933. Chain was a Jew, and went to England where he worked first in London and then in Cambridge where the Director of the Institute of Biochemistry, Sir Frederick Gowland Hopkins, thought highly of him and took a great interest in his research-work. One day Sir Frederick turned up in the basement where Chain's laboratory was situated and asked him whether he would like to go to Oxford where the new Professor of Pathology, Florey, was looking for a biochemist.

Chain was delighted at the prospect. It had never occurred to him that he might get a post in England, and he was preparing to emigrate to Canada or Australia. He was at that time a young man of twenty-nine, with black hair, flashing eyes, and an exceptionally lively mind, very different from the English, who, however, got the impression that he was something of a genius — and not without reason. It was Sir Frederick Gowland Hopkins who, struck by his ability, had recommended him to Florey.

Chain went to see Florey, who explained to him the importance he attached in his department to biochemistry, since all pathological change rests upon a biochemical phenomenon. He promised Chain a perfectly free hand and the right to choose his own subject for research. At the same time, he suggested that he might do worse than investigate the behaviour of a bacteriolytic substance, lysozyme, which, said he, played a part in protecting the body against bacteria and, perhaps, gastric ulcers.

In 1936 Chain, in company with a Rhodes Scholar named Epstein, attacked this problem, together with another — the biochemical action of snake-venom. The first thing to be decided was whether lysozyme was really, as Fleming said it was, an enzyme, that is to say, an element capable by its presence of encouraging certain reactions and breaking down certain molecules. If the answer to that question should be 'yes', then it must follow that there must be in the bacterial cell a substrate on which the product acted, since enzyme and substrate are as closely related as a lock and its key, which explains the specificity of the enzymes. The result was positive. Chain was able to extract from the famous yellow coccus (;micrococcus lysodeikticus) a substance (a polysaccharid which was broken up by lysozyme and, in this particular case, acted as the lock.

This piece of work having been carried through successfully, it seemed natural to go carefully through the 'literature of the subject', in other words, anything previously published, and to find out what had been done already in this field. Chain found close on two hundred papers dealing with antibacterial substances with a microbial origin, some of which had, like lysozyme, a lytic (or dissolving) action, while others killed this or that microbe in a different manner. Chain's attention was thus drawn to an immense field of study — microbial antagonism. Florey and he discussed the matter.

Of all the papers through which Chain had worked, the most interesting seemed to him to be the one written by Fleming in 1929 on penicillin. He learned from it of the existence of a substance 'with antibacterial properties rich in promise'. It had this superiority over lysozyme, that it could destroy dangerous microbes and in addition was, according to Fleming, completely lacking in toxicity. Continuing his reading, Chain found that a serious attempt had been made to extract and purify penicillin, which was more than could be said of the other substances, but that this had come to nothing.

He was ready to continue the attempt, but the necessary research would be pretty costly, and there was a shortage of funds. Walking one day with Florey through the lovely University Parks, he asked whether it might not be possible to get a few thousand dollars from the Rockefeller Foundation. The Medical Research Council had granted a few small sums, but these never amounted to more than fifty or a hundred pounds, which were almost worse than nothing. Florey forwarded the request and some time later informed Chain that if a programme of interesting biochemical research were submitted, the money would probably be forthcoming. Chain said that nothing would be easier and at once drew up for the Rockefeller Foundation a memorandum in which he suggested three subjects of study: snake-poison; the spreading factors; and bacterial antagonism. There was enough material in these to provide work for ten years.

Florey approved the memorandum and after a delay of a few months went in high spirits to tell Chain that a subsidy of five thousand dollars had been granted by the Foundation. This was marvellous news. The biochemical laboratory would no longer be prevented for want of a ridiculously small sum of money from buying the necessary equipment. Chain began his experiments on penicillin at the beginning of 1939. Then he went to Belgium for a holiday and, when he got back, war was just about to be declared.

Why had he started with penicillin? Florey and Chain had decided that three types of substance should be studied: an enzyme, pyocyanase; the antibacterial substances produced by the actinomycetes (which were later to give birth to such powerful antibiotics as streptomycin, etc.); and penicillin. The latter had many advantages over the others. It had already been studied under several aspects: it was known to be non-toxic and finally, though it could be neither conserved nor purified, it could at least be easily produced.

The Dunn School possessed a strain. How this came about must be explained. When Chain had arrived in Oxford in 1935, he had come across a colleague in one of the corridors carrying several Roux bottles containing a mould. At the time he had not paid much attention to this, but when he came to read Fleming's paper he realized that the culture he had seen in the hands of his colleague might perhaps have had something to do with penicillin.

He went and asked her whether this was so. She explained that she had been one of Dreyer's assistants: that Dreyer, Florey's predecessor, had been interested in the bacteriophages, forms of virus capable of destroying bacteria, and had thought that penicillin might be one of them. He had asked Fleming to send him a culture of penicillium and this Fleming, always pleased to find anyone interested in it, had done. Dreyer had soon seen that penicillin was not a phage, but had kept it with a view to further work. Chain, in his turn, asked for the culture.

At that time he knew nothing about moulds, and had to learn how to handle these extremely temperamental colonies. It seemed impossible to get any constant results. Sometimes the penicillium gave penicillin, sometimes it did not. The reason was that Fleming's strain had produced numerous mutations. Chain verified that the antibacterial substance was highly unstable, but this very instability aroused his curiosity. The chemists who before him had tried to extract pure penicillin had reached the conclusion that 'the substance vanished almost while you looked at it.' Chain set himself the task of finding the reason for this instability and attacked the problem by the very much gentler methods of the chemistry of enzymes, which he knew well.

It was agreed that Florey should carry out biological tests on the substance when Chain had managed to isolate it and to elucidate its structure. Chain himself undertook to work on penicillin, and asked one of his colleagues, Mrs Schoental, to proceed with the study of the pyocyanase.

Chain thought that penicillin must be an unstable enzyme. It is a known fact that the enzymes in solution often lose their activity when concentrated by evaporation, because the inactivating substances axe concentrated at the same time as the enzyme and destroy it. But Chain had at his disposal a new method which had been developed since the time of the researches made by Ridley and Raistrick — lyophilization (freeze-drying), which was being much used in, for example, the conservation of blood-plasma.

The process is very simple and is based on the principle that when liquids become congealed in a vacuum they pass straight from the solid to the gaseous state. This phenomenon can be observed on the tops of high mountains, where ice is sublimated (transformed into vapour) without going through the intermediate stage of melting. Now if a liquid solution containing different substances becomes congealed, these substances, when solidified, cease to have any action on one another (<corpora non agunt nisi fluida). If, therefore, the liquid is eliminated by sublimation, the solid substances which form the dry residue retain their activity almost indefinitely. Here was a way in which penicillin could be saved.

By freeze-drying the culture-liquid, Chain obtained a brown powder which contained, together with penicillin, several impurities (proteins, salts), and consequently could not be used for purposes of injection. Could one, as his predecessors had hoped, extract penicillin by dissolving it in pure alcohol? He tried the experiment, but without success. This did not surprise him, because he believed it to be an enzyme, and, therefore, not soluble in alcohol. All the same, so as to leave no stone unturned, he tried again, this time with methyl-alcohol (or methanol) and to his amazement succeeded. Some part of the impurities was thus eliminated. Unfortunately, however, the volatile penicillin, when dissolved in methanol, once again became unstable. The cure for this was to dilute the solution with water, and then have further recourse to freeze-drying.

Now that he had in his possession a partially purified penicillin, Chain was eager to test it. Since Florey was just then much occupied with other research-work, he turned to a great Spanish surgeon, Joseph Trueta, who happened to be working at the Dunn School on the floor above, with a young English assistant, John Barnes. At Chain's request, Barnes injected thirty milligrams of concentrated penicillin into the vein of a mouse. To the delight of Chain and to the surprise of Trueta, who was watching the experiment, no toxic reaction occurred.

Florey, much interested, at once repeated the experiment on another mouse, with a dose of twenty milligrams, and again there was no toxic effect. He, too, was so completely surprised, that he feared he might have missed the vein and said to Chain: 'Let me have another dose^' — by no means an easy thing to do at that time. With the utmost difficulty Chain managed to get hold of the twenty milligrams required and once more Florey established the non-toxicity of the substance.

So, at last, the Oxford scientists had in their hands, in a concentrated state, stable and partially purified, a product which possessed the astonishing property of killing microbes but not the cells of the body. Florey asked Chain to enlist the help of Heatley, a young and inventive laboratory-worker who had just come back from Copenhagen, and together he and Chain perfected a practical method of purifying penicillin.

It would take too long to describe here the innumerable difficulties which they encountered. The essential points which emerged from their researches were the following: (i) the work must be done (a), at a low temperature, and (b) with a neutral pH; and (2) the neutral liquid solution must be freeze-dried in order to obtain a penicillin salt in powdered form. It is important to note that the method thus perfected was the one used by all the big industrial producers until 1946. Without freeze-drying it would have been impossible to manufacture penicillin on a large scale. Chain had been among the first to employ this process when he was studying the enzymes.

In order to measure the antibacterial power of penicillin, Heatley at first used Fleming's method (holes made in the agar of a Petri dish and filled with penicillin round which the vulnerable microbes disappeared). Then he substituted for the holes small glass or porcelain cylinders planted in the surface of the agar. His first experiments proved that the product, when partially purified, was a thousand times more active than the crude penicillin, and ten times more active than the most active of the sulphonamides. (When completely purified, penicillin turned out to be a thousand times more active than Chain's first samples and, therefore, a million times more so than Fleming's crude substance.)

Florey and his colleagues had tested the toxicity of penicillin given intravenously in a single injection. This Fleming had also done with his crude penicillin. But now, with most of the proteins removed and with the substance at last stable, the Oxford team could go farther. They proceeded to treat rats with an intramuscular injection of ten milligrams every three hours over a period of fifty-six hours. Once again there was no accident. They tried the action of their substance on the arterial pressure and the respiration of cats. They repeated Fleming's experiments on the leucocytes. 'From all these tests', they tell us, 'it was clear that this substance possessed qualities which made it suitable for trial as a chemotherapeutic agent.'

The moment had now come for the crucial test. It was made on May 25th, 1940, on three groups of mice infected, respectively, with staphylococci, streptococci and Clostridium septicum. Heatley remembers with emotion the night he spent in the laboratory observing the reactions of the animals, and his joy when he saw the controls die one after the other, while the treated mice survived. Next morning Florey and Chain went to ascertain the results. Chain's eyes still sparkle when he speaks of the occasion.

June 1940 — it was the time of the great German offensive ... and Dunkirk. Was England going to be invaded? The Oxford team had decided that, should there be an invasion, they must at all costs save the miraculous mould the importance of which could no longer be doubted. They soaked the linings of their clothes in the brown liquid. It would be enough if only one of them escaped, for he would have on his person spores enough with which to start new cultures. By the end of the month they had enough penicillin for a decisive experiment. This was carried out on July 1st, on fifty white mice. All of them were given a more than lethal injection of virulent streptococci — I c.c. Twenty-five served as controls. The other twenty-five were treated with penicillin every three hours over a period of two days and two nights. Florey and his assistant, Kent, slept in the laboratory and were aroused by an alarm-clock every two hours. At the end of sixteen hours the twenty-five control mice were dead. Twenty-four of the treated mice survived the experiment.

The results smacked of the miraculous. They were set down in sober black and white in a note published by the Lancet.^l The signatories were Florey, Chain and Heatley who had done the extracting and conducted the first tests on animals. To their names were added those of Jennings, Orr-Ewing, Sanders and Gardner, whose help Florey had enlisted for the purpose of studying the miraculous substance more thoroughly. Gardner supplied the bacteriological study, confirmed Fleming's results and added some microbes to the list of those on which penicillin had an effect, in particular, that of gas-gangrene which in time of war was of paramount importance.

These names accounted for all but one of the Oxford team. Fleming had never had so large a group of specialists working with him. For this discovery to come about, first and foremost the solitary worker had been necessary, and only after him, following on his heels, the team. 'The work of a team', Chain has written, ^eis important for the development of an idea already formulated, but I do not believe that a team has ever produced a new idea.' And Fleming: Tor the birth of something new, there has to be a happening. Newton saw an apple fall; James Watt watched a kettle boil; Roentgen fogged some photographic plates. And these people knew enough to translate ordinary happenings into something new ...'

When Fleming read in the Lancet the first communication made by the Oxford team, he had the happiest surprise of his life. He had always known, and had never tired of saying, that a day would come when penicillin would be concentrated and purified, and that then it would be possible to use it in the treatment of generalized infections. He had but one thought — to see his darling substance in its pure state.

He therefore went to Oxford to see Florey and Chain. The latter was taken completely by surprise: he had thought that Fleming was dead! 'He struck me', he says, 'as a man who had difficulty in expressing himself, though he gave the impression of being somebody with a very warm heart doing all he could to appear cold and distant.' The truth was that he was making a great effort to conceal his joy, for he had always made it a rule never to show his feelings. 'You have made something of my substance,' was all he said to Chain. Craddock who saw him on his return relates that, speaking of the Oxford team, he said: 'They have turned out to be the successful chemists I should have liked to have with me in 1929.'

Fleming to Florey, November 15th, 1940 Dear Florey,

I am sorry to have been so long in sending you cultures of the penicillium which did not produce much yellow colour. When I got back from visiting you, I planted out a large number of my old cultures in broth, and I have selected from these a number which, while producing a good yield of penicillin, did not appreciably colour the broth yellow. I am sending these on to you, and I hope you will find them useful.

I have been comparing the solid penicillin which I got from you with the sulphonamides and it seems to be, weight for weight, a great deal more potent than the most powerful of those on the ordinary septic microbes.

It only remains for your chemical colleagues to purify the active principle, and then synthesize it, and the sulphonamides will be completely beaten.

Yours sincerely

alexander fleming

Dr E. W. Todd to Fleming, August 23rd, 1940

London County Council, Public Health Dept, Belmont Laboratories, Stanley Road, Sutton, Surrey

My Dear Flem,

I was delighted to read in the Lancet this morning about penicillin. When can we start production? I am laboriously making gas-gangrene anti-toxin> and penicillin sounds much simpler.

I can claim to have been in the same room when you made the great discovery. Do you think that there is any chance that I might get a knighthood on that claim when you are raised to the Peerage?

Are you producing penicillin for therapeutic use?

Congratulations,

Yours ever

e. W. t.

The time had now come to try penicillin on human patients, but that would need great quantities of penicillin in as pure a state as possible. The substance to be dealt with was a madly temperamental mould and it was essential to move quickly. Heatley gave himself to the problem of extracting the product; Chain and Abraham concentrated on its purification. To describe here all their difficulties and disappointments would involve too many technical explanations. But it can and should be said that they showed admirable qualities of ingenuity and tenacity. The group met every day at tea-time, recorded their failures, deplored them, but never showed discouragement. The stake was well worth the trouble taken.

After innumerable washings, manipulations and filterings, they at last obtained a yellow powder which was a salt of barium containing about five hundred units of penicillin per milligram.²At first, the percentage of penicillin, for the same weight, had been half a unit. This was an excellent result. Next, the yellow pigment had to be precipitated. The final stage of the operation, the evaporation of the water in order to obtain a dry powder, still presented difficulties. The normal method of turning water into steam is to make it boil: but heat destroys penicillin. They had to have recourse to the other method, which consists of reducing the pressure above the water and so lowering the boiling-point. The use of a vacuum pump made it possible to evaporate the water at a very low temperature. The precious yellow powder remained at the bottom of the jar. It felt to the touch like cornflour. This penicillin was still only half pure. Nevertheless, when Florey made his bacteriological tests, he found that a thirty-millionth solution of this powder was enough to inhibit the development of the staphylococci.

The most convincing case would have been one of septicaemia. But this presented certain difficulties. On the one hand, the quantity of penicillin available did not permit the injection of a massive dose; on the other, the rapid excretion of the product would mean that it would remain in the body for an insufficient length of time. It was very quickly eliminated by the kidneys. No doubt it would be found in the urine and could be extracted and used again. But these operations would be lengthy, and the patient would have plenty of time in which to die. Giving the product by the mouth would be ineffective, because the gastric juices would destroy the penicillin as soon as it reached the stomach. What appeared to be desirable was the maintenance in the blood stream, by successive injections, of a quantity of the substance sufficient to allow the natural defence mechanisms of the body to destroy the microbes which, thanks to the penicillin, would have become far less numerous. The best method, therefore, would be frequent injections, or perhaps, even, intravenous drip.

The inadequate quantities on hand increased the natural anxiety always aroused by any brand-new experiment on a patient. There was the risk that a treatment might be begun which could not be continued. Florey went to see the directors of a great industry specializing in chemical products, told them that he had in his hands a substance which looked as though it would turn out to be a miraculous remedy, and asked, without concealing from them the difficulties of the enterprise, whether they were prepared to undertake its production on a large scale. After thinking the suggestion over, these industrial chemists refused. They are not to be blamed. Their factories were fully occupied by Government orders for war-material; the methods perfected, with the greatest difficulty, by the Oxford team, would have involved an enormous amount of work; and, last but not least, the concern would have run the risk, after equipping their factories at great expense, of seeing some research-worker suddenly solve the problem of making synthetic pencillin and so drastically reducing the cost of production.

The only thing left for the Oxford chemists to do was once again to work with such means as they could find ready to hand. The task given by Florey to Heatley was to produce a hundred litres of culture a week and to extract the penicillin. A small quantity of the yellow powder was put aside in a refrigerator for emergencies at the beginning of February 1941. At that very moment a case turned up which, because it seemed desperate, justified the carrying through of a daring experiment. A policeman in Oxford was dying of septicaemia. It had started with a small infected scratch at the corner of the mouth. Then the whole blood-content of the body had been poisoned. The microbe in question was the staphylococcus aureus, which is vulnerable to penicillin. The patient had been treated unsuccessfully with sulphonamides. There were abscesses all over his body and his lungs had been affected. The doctors regarded his condition as hopeless. If penicillin resulted in a cure, it would be a shattering proof of its power.

On February 12th, 1941, an intravenous injection of a00 mg. of penicillin was given to the dying man and, thereafter, an injection of 100 mg. every three hours. At the end of twenty-four hours the improvement in his condition was startling. The wounds had ceased to suppurate. It was obvious that the patient, only the day before at the point of death, was now well on the way to recovery. While continuing the penicillin injections, the doctors gave him a blood-transfusion. Unfortunately, the tiny reserve of yellow powder was dwindling in a tragic fashion. It was possible to recover a certain amount from the man's urine and the improvement became still more marked. The patient felt better. He was eating. His temperature had dropped. Two facts in painful contrast to each other were now only too obvious. The penicillin treatment, if it could be maintained, would undoubtedly save his life, but it could not be continued long enough because there would not be sufficient penicillin available. Heatley worked devotedly, but he could only wait until the cultures had produced a fresh crop. He very soon had to stop the injections. The patient managed to hold on for a few days. But the microbes, being no longer attacked, got the upper hand and, on March 15th, the policeman died.

Florey now knew that if he had had enough penicillin, the man could have been saved. But he could not prove a hypothetical success. A transfusion had been given and it was open to sceptical critics to say that the improvement had been due to that. The first test, therefore, had partially failedw The yellow powder, fruit of so much hard work, had been used in vain. The Oxford team was saddened but not discouraged. When a new supply of penicillin was ready, three other cases were treated. All of them gave evidence of the immediately beneficial and spectacular action of the substance. Two of the cases were completely cured. The third, that of a child who had been brought out of a coma by penicillin, was getting much better, when death supervened as the result of the accidental rupture of a blood-vessel. But even severe judges could no longer doubt that medicine now possessed a new chemotherapeutic product of unparalleled strength, which was non-toxic. The first patient, when injected, had had a sharp rise in temperature and a rigor, but this was due to certain impurities still remaining in the drug, and was not repeated when penicillin had been completely purified.

Was it possible that on the strength of these first results the British Government would set on foot an immense effort to manufacture the miraculous remedy on an industrial scale? Florey very quickly realized that the answer to that question would be in the negative. England in 1941 was suffering under incessant bombing raids. It was conducting, or preparing, a war on all fronts. Urgent day-to-day necessities took priority over everything else. The men and women who were living under the constant threat of seeing their homes reduced to ruins on top of them could scarcely be expected to regard the war against microbes as a matter of essential importance. But Florey could measure the effects of a massive employment of penicillin and envisage the consequences for the wounded and for the war-effort as a whole.

The Oxford scientists approached nearly all the major manufacturers of chemical products. The reply in almost every case was the same: ^'Yes, Doctor, you have made a most important discovery, but the production of your substance on a commercial scale is impossible because your output is too small.' The treatment of a single case needed thousands of litres of cultures. The proposition was not a practical one. The obvious remedy for this state of affairs was an increase in output and the financing of a vast research programme. But the English factories, under the harsh conditions of war, were in no state to make the necessary effort. The only alternative was to turn to America.

Florey and Heatley left for the United States via Lisbon in June 1941, taking with them some strains of penicillium. The heat was intense and they were in a condition of the most extreme anxiety throughout the voyage, since the precious moulds could not stand high temperatures. In New York, Florey renewed acquaintance with an American friend who at once put him on the track of the man who could get things moving, Charles Thorn, who had identified the penicillium notatum, and was now Head of the Mycological Section of the Northern Regional Research Laboratory at Peoria, Illinois. This laboratory had been created to make research into the utilization of the organic by-products of agriculture which were polluting the rivers of the Middle West. The intention was to convert these waste products into usable fermentations, and the chemists there had been concentrating their efforts on producing gluconic acid by using the fermentation powers of moulds of the penicillium chrysogenum type. In this work they had been using, as their source of nitrogen, corn steep liquor, a by-product of the manufacture of starch from maize. This liquor had accumulated in the region to an embarrassing extent. The chemists had succeeded in making the gluconic acid production work by using a method of submerged fermentation.

Florey passed from scientist to scientist and finally reached Dr Coghill, who was Chief of the Division of Fermentation at Peoria. To him he explained his problem. It should be mentioned that the English scientists (and this is as true of Fleming as of Florey, of Chain as of Heatley) had done nothing to protect their discoveries by taking out a patent. In their eyes a substance which could render such services to mankind ought not to become a source of private profit. So extreme a degree of disinterestedness deserves to be stressed and applauded. They gave to the Americans all the results of their prolonged researches and all their methods of manufacture, asking in return only for penicillin so as to be able to continue their medical experiments.

Heatley stayed on in Peoria to take part in the work. The first objective was to increase output, which meant finding a favourable medium for the penicillium culture. The Americans suggested corn steep liquor with which they were familiar and which they had used as a medium for similar cultures. They quickly obtained an output twenty times higher than that of Oxford and this brought them within sight of a practical solution of the problem. It would be possible, at least for war purposes, to produce penicillin in quantity. The substitution of lactose for glucose still further improved the output.

Once again we are left wondering at the curious operations of Chance. If the Americans had not been embarrassed by an excessive accumulation of their steepiiig liquor, they would not have set up a laboratory at Peoria. Had it not existed, no one would have hit upon that particular culture medium — maize-liquor + lactose — and the commercial production of penicillin might have been indefinitely postponed. On the other hand, the arrival of the scientists from England had alone made it possible for this laboratory to do the work for which it had been set up, since the making of gluconic acid would not of itself have absorbed the immense quantities of corn steep liquor available, whereas the manufacture of penicillin would very soon make it extremely precious and increase its value a hundred times.

The new culture medium was not the only contribution made by Peoria, The mycologists attached to the laboratory were looking for mould-strains which would give a larger yield of penicillin, for it is a curious fact that all the cultures so far made in England and America were descended from the same spore, the one which had landed on Fleming's work-bench. Up till 1943, nothing better had been found, though a great many strains had been tried. It was most unlikely that this particular strain, which had not been deliberately chosen, should turn out to be the best. The American research-workers had enlisted the help of the Army authorities in getting specimens of moulds from all over the world, but none had proved to be usable. But the laboratory had also taken on a young woman whose function it was to go to market and buy every variety of mould she could find. She was very soon known as 'Mouldy Mary'. One day she returned from market with a mould of the penicillium chrysogenum type which she had found growing in a cantaloup melon which had gone bad. It showed itself to be remarkably productive. The application of genetic methods improved its yield still further, and most of the strains used today (after mutations) come from the rotten Peoria melon. The scientists, as so often happens, had on their doorstep what they had been searching for in vain all over the world.

While Heatley was working with the chemists at Peoria, Florey started on a pilgrimage which was to take him all over America and Canada, visiting numerous factories turning out chemical products and trying to interest industrialists in the production of penicillin on a really great scale. The situation in America seemed to be less difficult than it was in England. In early 1941 the country was still not yet at war. But it was receiving a considerable number of orders, and most of the industrial leaders whom Florey saw showed only moderate enthusiasm for an enterprise which they regarded as both uncertain and difficult. All the same, many of them expressed their good will and, when Florey left for England, he took with him promises from two firpls, each of which had undertaken to produce ten thousand Utr^s and to send the penicillin to Oxford for purposes of medical research. His last visit was to his friend Dr A. N. Richards whom he had known formerly at the University of Pennsylvania. Dr Richards had just been appointed President of the Committee of Medical Research. This important post would give him considerable influence and make it possible for him to interest the American Government in penicillin. The journey had been productive of great results.