8. Whatever indulgence may be granted to those who have heretofore been the ignorant causes of so much misery, the time has come when the existence of a private pestilence in the sphere of a single physician should be looked upon, not as a misfortune, but a crime; and in the knowledge of such occurrences the duties of the practitioner to his profession should give way to his paramount obligations to society.
ADDITIONAL REFERENCES AND CASES.
Fifth Annual Report of the Registrar-General of England, 1843, Appendix. Letter from William Fair, Esq.—Several new series of cases are given in the letter of Mr. Storrs, contained in the appendix to this report. Mr. Storrs suggests precautions similar to those I have laid down, and these precautions are strongly enforced by Mr. Farr, who is, therefore, obnoxious to the same criticisms as myself.
Hall and Dexter, in Am. Journal of Med. Sc. for January, 1844.— Cases of puerperal fever seeming to originate in erysipelas.
Elkington, of Birmingham, in Provincial Med. Journal, cited in Am. Journ. Med. Sc. for April, 1844.—Six cases in less than a fortnight, seeming to originate in a case of erysipelas.
West's Reports, in Brit. and For. Med. Review for October, 1845, and January, 1847.—Affection of the arm, resembling malignant pustule, after removing the placenta of a patient who died from puerperal fever. Reference to cases at Wurzburg, as proving contagion, and to Keiller's cases in the Monthly Journal for February, 1846, as showing connection of puerperal fever and erysipelas.
Kneeland.—Contagiousness of Puerperal Fever. Am. Jour. Med. Sc., January, 1846. Also, Connection between Puerperal Fever Epidemic Erysipelas. Ibid., April, 1846.
Robert Storrs.-Contagious Effects of Puerperal Fever on the Male Subject; or on Persons not Child-bearing. (From Provincial Med. and Surg. Journal.) Am. Jour. Med. Sc., January, 1846. Numerous cases. See also Dr. Reid's case in same journal for April, 1846.
Routh's paper in Proc. of Royal Med. Chir. Soc., Am. Jour. Med. Sc., April, 1849, also in B. and F. Med. Chir. Review, April, 1850.
Hill, of Leuchars.—A Series of Cases Illustrating the Contagious Nature of Erysipelas and Puerperal Fever, and their Intimate Pathological Connection. (From Monthly Journal of Med. Sc.) Am. Jour. Med. Sc., July, 1850.
Skoda on the Causes of Puerperal Fever. (Peritonitis in rabbits, from inoculation with different morbid secretions.) Am. Jour. Med. Sc., October, 1850.
Arneth.—Paper read before the National Academy of Medicine. Annales d'Hygiene, Tome LXV. 2e Partie. ("Means of Disinfection proposed by M. Semmelweis." Semmelweiss.) Lotions of chloride of lime and use of nail-brush before admission to lying-in wards, Alleged sudden and great decrease of mortality from puerperal fever. Cause of disease attributed to inoculation with cadaveric matters.) See also Routh's paper, mentioned above.
Moir.—Remarks at a meeting of the Edinburgh Medico-chirurgical Society. Refers to cases of Dr. Kellie, of Leith. Sixteen in succession, all fatal. Also to several instances of individual pupils having had a succession of cases in various quarters of the town, while others, practising as extensively in the same localities, had none. Also to several special cases not mentioned elsewhere. Am. Jour. Med. Sc. for October, 1851. (From New Monthly Journal of Med. Science.)
Simpson.—Observations at a Meeting of the Edinburgh Obstetrical Society. (An "eminent gentleman," according to Dr. Meigs, whose "name is as well known in America as in (his) native land," Obstetrics, Phil., 1852, pp. 368, 375.) The student is referred to this paper for a valuable resume of many of the facts, and the necessary inferences, relating to this subject. Also for another series of cases, Mr. Sidey's, five or six in rapid succession. Dr. Simpson attended the dissection of two of Dr. Sidey's cases, and freely handled the diseased parts. His next four child-bed patients were affected with puerperal fever, and it was the first time he had seen it in practice. As Dr. Simpson is a gentleman (Dr. Meigs, as above), and as "a gentleman's hands are clean" (Dr. Meigs' sixth letter), it follows that a gentleman with clean hands may carry the disease. Am. Jour. Med. Sc., October, 1851.
Peddie.—The five or six cases of Dr. Sidey, followed by the four of Dr. Simpson, did not end the series. A practitioner in Leith having examined in Dr. Simpson's house, a portion of the uterus obtained from one of the patients, had immediately afterwards three fatal cases of puerperal fever. Dr. Peddie referred to two distinct series of consecutive cases in his own practice. He had since taken precautions, and not met with any such cases. Am. Jour. Med October, 1851.
Copland.—Considers it proved that puerperal fever may be propagated by the hands and the clothes, or either, of a third person, the bed-clothes or body-clothes of a patient. Mentions a new series of cases, one of which he saw, with the practitioner who had attended them. She was THE SIXTH he had had within a few days. ALL DIED. Dr. Copland insisted that contagion had caused these cases; advised precautionary measures, and the practitioner had no other cases for a considerable time. Considers it CRIMINAL, after the evidence adduced,—which be could have quadrupled,—and the weight of authority brought forward, for a practitioner to be the medium of transmitting contagion and death to his patients. Dr. Copland lays down rules similar to those suggested by myself, and is therefore entitled to the same epithet for so doing. Medical Dictionary, New York, 1853. Article, Puerperal States and Diseases.
If there is any appetite for facts so craving as to be yet unappeased,—lassata, necdum satiata,—more can be obtained. Dr. Hodge remarks that "the frequency and importance of this singular circumstance that the disease is occasionally more prevalent with one practitioner than another, has been exceedingly overrated." More than thirty strings of cases, more than two hundred and fifty sufferers from puerperal fever, more than one hundred and thirty deaths, appear as the results of a sparing estimate of such among the facts I have gleaned as could be numerically valued. These facts constitute, we may take it for granted, but a small fraction of those that have actually occurred. The number of them might be greater, but "'t is enough, 't will serve," in Mercutio's modest phrase, so far as frequency is concerned. For a just estimate of the importance of the singular circumstance, it might be proper to consult the languid survivors, the widowed husbands, and the motherless children, as well as "the unfortunate accoucheur."
ON THE ANTISEPTIC PRINCIPLE OF THE PRACTICE OF SURGERY BY JOSEPH LISTER
Joseph Lister was born at Upton, Essex, England, in 1827, and received Aw general education at the University of London. After graduation he studied medicine in London and Edinburgh, and became lecturer in surgery at the University in the latter city. Later he was professor of surgery at Glasgow, at Edinburgh, and at King's College Hospital, London, and surgeon to Queen Victoria. He was made a baronet in 1883; retired from teaching in 1893; and was raised to the peerage in 1897, with the title of Baron Lister.
Even before the work of Pasteur on fermentation and putrefaction, Lister had been convinced of the importance of scrupulous cleanliness and the usefulness of deodorants in the operating room; and when, through Pasteur's researches, he realised that the formation of PUS was due to bacteria, he proceeded to develop his antiseptic surgical methods. The immediate success of the new treatment led to its general adoption, with results of such beneficence as to make it rank as one of the great discoveries of the age.
ON THE ANTISEPTIC PRINCIPLE OF THE PRACTICE OF SURGERY (1867)
In the course of an extended investigation into the nature of inflammation, and the healthy and morbid conditions of the blood in relation to it, I arrived several years ago at the conclusion that the essential cause of suppuration in wounds is decomposition brought about by the influence of the atmosphere upon blood or serum retained within them, and, in the case of contused wounds, upon portions of tissue destroyed by the violence of the injury.
To prevent the occurrence of suppuration with all its attendant risks was an object manifestly desirable, but till lately apparently unattainable, since it seemed hopeless to attempt to exclude the oxygen which was universally regarded as the agent by which putrefaction was effected. But when it had been shown by the researches of Pasteur that the septic properties of the atmosphere depended not on the oxygen, or any gaseous constituent, but on minute organisms suspended in it, which owed their energy to their vitality, it occurred to me that decomposition in the injured part might be avoided without excluding the air, by applying as a dressing some material capable of destroying the life of the floating particles. Upon this principle I have based a practice of which I will now attempt to give a short account.
The material which I have employed is carbolic or phenic acid, a volatile organic compound, which appears to exercise a peculiarly destructive influence upon low forms of life, and hence is the most powerful antiseptic with which we are at present acquainted.
The first class of cases to which I applied it was that of compound fractures, in which the effects of decomposition in the injured part were especially striking and pernicious. The results have been such as to establish conclusively the great principle that all local inflammatory mischief and general febrile disturbances which follow severe injuries are due to the irritating and poisonous influence of decomposing blood or sloughs. For these evils are entirely avoided by the antiseptic treatment, so that limbs which would otherwise be unhesitatingly condemned to amputation may be retained, with confidence of the best results.
In conducting the treatment, the first object must be the destruction of any septic germs which may have been introduced into the wounds, either at the moment of the accident or during the time which has since elapsed. This is done by introducing the acid of full strength into all accessible recesses of the wound by means of a piece of rag held in dressing forceps and dipped into the liquid. [Footnote: The addition of a few drops of water to a considerable quantity of the acid, induces it to assume permanently the liquid form.] This I did not venture to do in the earlier cases; but experience has shown that the compound which carbolic acid forms with the blood, and also any portions of tissue killed by its caustic action, including even parts of the bone, are disposed of by absorption and organisation, provided they are afterwards kept from decomposing. We are thus enabled to employ the antiseptic treatment efficiently at a period after the occurrence of the injury at which it would otherwise probably fail. Thus I have now under my care, in Glasgow Infirmary, a boy who was admitted with compound fracture of the leg as late as eight and one-half hours after the accident, in whom, nevertheless, all local and constitutional disturbance was avoided by means of carbolic acid, and the bones were soundly united five weeks after his admission.
The next object to be kept in view is to guard effectually against the spreading of decomposition into the wound along the stream of blood and serum which oozes out during the first few days after the accident, when the acid originally applied has been washed out or dissipated by absorption and evaporation. This part of the treatment has been greatly improved during the past few weeks. The method which I have hitherto published (see Lancet for Mar. 16th, 23rd, 30th, and April 27th of the present year) consisted in the application of a piece of lint dipped in the acid, overlapping the sound skin to some extent and covered with a tin cap, which was daily raised in order to touch the surface of the lint with the antiseptic. This method certainly succeeded well with wounds of moderate size; and indeed I may say that in all the many cases of this kind which have been so treated by myself or my house-surgeons, not a single failure has occurred. When, however, the wound is very large, the flow of blood and serum is so profuse, especially during the first twenty-four hours, that the antiseptic application cannot prevent the spread of decomposition into the interior unless it overlaps the sound skin for a very considerable distance, and this was inadmissible by the method described above, on account of the extensive sloughing of the surface of the cutis which it would involve. This difficulty has, however, been overcome by employing a paste composed of common whiting (carbonate of lime), mixed with a solution of one part of carbolic acid in four parts of boiled linseed oil so as to form a firm putty. This application contains the acid in too dilute a form to excoriate the skin, which it may be made to cover to any extent that may be thought desirable, while its substance serves as a reservoir of the antiseptic material. So long as any discharge continues, the paste should be changed daily, and, in order to prevent the chance of mischief occurring during the process, a piece of rag dipped in the solution of carbolic acid in oil is put on next the skin, and maintained there permanently, care being taken to avoid raising it along with the putty. This rag is always kept in an antiseptic condition from contact with the paste above it, and destroys any germs which may fall upon it during the short time that should alone be allowed to pass in the changing of the dressing. The putty should be in a layer about a quarter of an inch thick, and may be advantageously applied rolled out between two pieces of thin calico, which maintain it in the form of a continuous sheet, which may be wrapped in a moment round the whole circumference of a limb if this be thought desirable, while the putty is prevented by the calico from sticking to the rag which is next the skin.[Footnote: In order to prevent evaporation of the acid, which passes readily through any organic tissue, such as oiled silk or gutta percha, it is well to cover the paste with a sheet of block tin. or tinfoil strengthened with adhesive plaster. The tin sheet lead used for lining tea chests will also answer the purpose, and may be obtained from any wholesale grocer.] When all discharge has ceased, the use of the paste is discontinued, but the original rag is left adhering to the skin till healing by scabbing is supposed to be complete. I have at present in the hospital a man with severe compound fracture of both bones of the left leg, caused by direct violence, who, after the cessation of the sanibus discharge under the use of the paste, without a drop of pus appearing, has been treated for the last two weeks exactly as if the fracture was a simple one. During this time the rag, adhering by means of a crust of inspissated blood collected beneath it, has continued perfectly dry, and it will be left untouched till the usual period for removing the splints in a simple fracture, when we may fairly expect to find a sound cicatrix beneath it. We cannot, however, always calculate on so perfect a result as this. More or less pus may appear after the lapse of the first week, and the larger the wound, the more likely this is to happen. And here I would desire earnestly to enforce the necessity of persevering with the antiseptic application in spite of the appearance of suppuration, so long as other symptoms are favorable. The surgeon is extremely apt to suppose that any suppuration is an indication that the antiseptic treatment has failed, and that poulticing or water dressing should be resorted to. But such a course would in many cases sacrifice a limb or a life. I cannot, however, expect my professional brethren to follow my advice blindly in such a matter, and therefore I feel it necessary to place before them, as shortly as I can, some pathological principles intimately connected, not only with the point we are immediately considering, but with the whole subject of this paper. If a perfectly healthy granulating sore be well washed and covered with a plate of clean metal, such as block tin, fitting its surface pretty accurately, and overlapping the surrounding skin an inch or so in every direction and retained in position by adhesive plaster and a bandage, it will be found, on removing it after twenty-four or forty-eight hours, that little or nothing that can be called pus is present, merely a little transparent fluid, while at the same time there is an entire absence of the unpleasant odour invariably perceived when water dressing is changed. Here the clean metallic surface presents no recesses like those of porous lint for the septic germs to develope in, the fluid exuding from the surface of the granulations has flowed away undecomposed, and the result is the absence of suppuration. This simple experiment illustrates the important fact that granulations have no inherent tendency to form pus, but do so only when subjected to preternatural stimulus. Further, it shows that the mere contact of a foreign body does not of itself stimulate granulations to suppurate; whereas the presence of decomposing organic matter does. These truths are even more strikingly exemplified by the fact that I have elsewhere recorded (Lancet, March 23rd, 1867), that a piece of dead bone free from decomposition may not only fail to induce the granulations around it to suppurate, but may actually be absorbed by them; whereas a bit of dead bone soaked with putrid pus infallibly induces suppuration in its vicinity.
Another instructive experiment is, to dress a granulating sore with some of the putty above described, overlapping the sound skin extensively; when we find, in the course of twenty-four hours, that pus has been produced by the sore, although the application has been perfectly antiseptic; and, indeed, the larger the amount of carbolic acid in the paste, the greater is the quantity of pus formed, provided we avoid such a proportion as would act as a caustic. The carbolic acid, though it prevents decomposition, induces suppuration—obviously by acting as a chemical stimulus; and we may safely infer that putrescent organic materials (which we know to be chemically acrid) operate in the same way.
In so far, then, carbolic acid and decomposing substances are alike; viz., that they induce suppuration by chemical stimulation, as distinguished from what may be termed simple inflammatory suppuration, such as that in which ordinary abscesses originate—where the pus appears to be formed in consequence of an excited action of the nerves, independently of any other stimulus. There is, however, this enormous difference between the effects of carbolic acid and those of decomposition; viz., that carbolic acid stimulates only the surface to which it is at first applied, and every drop of discharge that forms weakens the stimulant by diluting it; but decomposition is a self-propagating and self-aggravating poison, and, if it occur at the surface of a severely injured limb, it will spread into all its recesses so far as any extravasated blood or shreds of dead tissue may extend, and lying in those recesses, it will become from hour to hour more acrid, till it requires the energy of a caustic sufficient to destroy the vitality of any tissues naturally weak from inferior vascular supply, or weakened by the injury they sustained in the accident.
Hence it is easy to understand how, when a wound is very large, the crust beneath the rag may prove here and there insufficient to protect the raw surface from the stimulating influence of the carbolic acid in the putty; and the result will be first the conversion of the tissues so acted on into granulations, and subsequently the formation of more or less pus. This, however, will be merely superficial, and will not interfere with the absorption and organisation of extravasated blood or dead tissues in the interior. But, on the other hand, should decomposition set in before the internal parts have become securely consolidated, the most disastrous results may ensue.
I left behind me in Glasgow a boy, thirteen years of age, who, between three and four weeks previously, met with a most severe injury to the left arm, which he got entangled in a machine at a fair. There was a wound six inches long and three inches broad, and the skin was very extensively undermined beyond its limits, while the soft parts were generally so much lacerated that a pair of dressing forceps introduced at the wound and pushed directly inwards appeared beneath the skin at the opposite aspect of the limb. From this wound several tags of muscle were hanging, and among them was One consisting of about three inches of the triceps in almost Its entire thickness; while the lower fragment of the bone, which was broken high up, was protruding four inches and a half, stripped of muscle, the skin being tucked in under it. Without the assistance of the antiseptic treatment, I should certainly have thought of nothing else but amputation at the shoulder-joint; but, as the radial pulse could be felt and the fingers had sensation, I did not hesitate to try to save the limb and adopted the plan of treatment above described, wrapping the arm from the shoulder to below the elbow in the antiseptic application, the whole interior of the wound, together with the protruding bone, having previously been freely treated with strong carbolic acid. About the tenth day, the discharge, which up to that time had been only sanious and serous, showed a slight admixture of slimy pus; and this increased till (a few days before I left) it amounted to about three drachms in twenty-four hours. But the boy continued as he had been after the second day, free from unfavorable symptoms, with pulse, tongue, appetite, and sleep natural and strength increasing, while the limb remained as it had been from the first, free from swelling, redness, or pain. I. therefore, persevered with the antiseptic dressing; and, before I left, the discharge was already somewhat less, while the bone was becoming firm. I think it likely that, in that boy's case, I should have found merely a superficial sore had I taken off all the dressings at the end of the three weeks; though, considering the extent of the injury, I thought it prudent to let the month expire before disturbing the rag next the skin. But I feel sure that, if I had resorted to ordinary dressing when the pus first appeared, the progress of the case would have been exceedingly different.
The next class of cases to which I have applied the antiseptic treatment is that of abscesses. Here also the results have been extremely satisfactory, and in beautiful harmony with the pathological principles indicated above. The pyogenic membrane, like the granulations of a sore, which it resembles in nature, forms pus, not from any inherent disposition to do so, but only because it is subjected to some preternatural stimulation. In an ordinary abscess, whether acute or chronic, before it is opened the stimulus which maintains the suppuration is derived from the presence of pus pent up within the cavity. When a free opening is made in the ordinary way, this stimulus is got rid of, but the atmosphere gaining access to the contents, the potent stimulus of decomposition comes into operation, and pus is generated in greater abundance than before. But when the evacuation is effected on the antiseptic principle, the pyogenic membrane, freed from the influence of the former stimulus without the substitution of a new one, ceases to suppurate (like the granulations of a sore under metallic dressing), furnishing merely a trifling amount of clear serum, and, whether the opening be dependent or not, rapidly contracts and coalesces. At the same time any constitutional symptoms previously occasioned by the accumulation of the matter are got rid of without the slightest risk of the irritative fever or hectic hitherto so justly dreaded in dealing with large abscesses.
In order that the treatment may be satisfactory, the abscess must be seen before it is opened. Then, except in very rare and peculiar cases [Footnote: As an instance of one of these exceptional cases, I may mention that of an abscess in the vicinity of the colon, and afterwords proved by post-mortem examination to have once communicated with it. Here the pus was extremely offensive when evacuated, and exhibited vibros under the microscope.], there are no septic organisms in the contents, so that it is needless to introduce carbolic acid into the interior. Indeed, such a procedure would be objectionable, as it would stimulate the pyogenic membrane to unnecessary suppuration. All that is requisite is to guard against the introduction of living atmospheric germs from without, at the same time that free opportunity is afforded for the escape of the discharge from within.
I have so lately given elsewhere a detailed account of the method by which this is effected (Lancet, July 27th, 1867), that I shall not enter into it at present further than to say that the means employed are the same as those described above for the superficial dressing of compound fractures; viz., a piece of rag dipped into the solution of carbolic add in oil to serve as an antiseptic curtain, under cover of which the abscess is evacuated by free incision, and the antiseptic paste to guard against decomposition occurring in the stream of pus that flows out beneath it; the dressing being changed daily until the sinus is closed.
The most remarkable results of this practice in a pathological point of view have been afforded by cases where the formation of pus depended on disease of bone. Here the abscesses, instead of forming exceptions to the general class in the obstinacy of the suppuration, have resembled the rest in yielding in a few days only a trifling discharge, and frequently the production of pus has ceased from the moment of the evacuation of the original contents. Hence it appears that caries, when no longer labouring as heretofore under the irritation of decomposing matter, ceases to be an opprobrium of surgery, and recovers like other inflammatory affections. In the publication before alluded to, I have mentioned the case of a middle-aged man with a psoas abscess depending in diseased bone, in whom the sinus finally closed after months of patient perseverance with the antiseptic treatment. Since that article was written I have had another instance of abscess equally gratifying, but the differing in the circumstance that the disease and the recovery were more rapid in their course. The patient was a blacksmith, who had suffered four and a half months before I saw him from symptoms of ulceration of cartilage in the left elbow. These had latterly increased in severity so as to deprive him entirely of his night's rest and of appetite. I found the region of the elbow greatly swollen, and on careful examination found a fluctuating point at the outer aspect of the articulation. I opened it on the antiseptic principle, the incision evidently penetrating to the joint, giving exit to a few drachms of pus. The medical gentleman under whose care he was (Dr. Macgregor, of Glasgow) supervised the daily dressing with the carbolic acid paste till the patient went to spend two or three weeks at the coast, when his wife was entrusted with it. Just two months after I opened the abscess, he called to show me the limb, stating that the discharge had been, for at least two weeks, as little as it was then, a trifling moisture upon the paste, such as might be accounted for by the little sore caused by the incision. On applying a probe guarded with an antiseptic rag, I found that the sinus was soundly closed, while the limb was free from swelling or tenderness; and, although he had not attempted to exercise it much, the joint could already be moved through a considerable angle. Here the antiseptic principle had effected the restoration of a joint, which, on any other known system of treatment, must have been excised.
Ordinary contused wounds are, of course, amenable to the same treatment as compound fractures, which are a complicated variety of them. I will content myself with mentioning a single instance of this class of cases. In April last, a volunteer was discharging a rifle when it burst, and blew back the thumb with its metacarpal bone, so that it could be bent back as on a hinge at the trapezial joint, which had evidently been opened, while all the soft parts between the metacarpal bones of the thumb and forefinger were torn through. I need not insist before my present audience on the ugly character of such an injury. My house- surgeon, Mr. Hector Cameron, applied carbolic acid to the whole raw surface, and completed the dressing as if for compound fracture. The hand remained free from pain, redness or swelling, and with the exception of a shallow groove, all the wound consolidated without a drop of matter, so that if it had been a clean cut, it would have been regarded as a good example of primary union. The small granulating surface soon healed, and at present a linear cicatrix alone tells of the injury he has sustained, while his thumb has all its movements and his hand a fine grasp.
If the severest forms of contused and lacerated wounds heal thus kindly under the antiseptic treatment, it is obvious that its application to simple incised wounds must be merely a matter of detail. I have devoted a good deal of attention to this class, but I have not as yet pleased myself altogether with any of the methods I have employed. I am, however, prepared to go so far as to say that a solution of carbolic acid in twenty parts of water, while a mild and cleanly application, may be relied on for destroying any septic germs that may fall upon the wound during the performance of an operation; and also that, for preventing the subsequent introduction of others, the paste above described, applied as for compound fractures, gives excellent results. Thus I have had a case of strangulated inguinal hernia in which it was necessary to take away half a pound of thickened omentum, heal without any deep-seated suppuration or any tenderness of the sac or any fever; and amputations, including one immediately below the knee, have remained absolutely free from constitutional symptoms.
Further, I have found that when the antiseptic treatment is efficiently conducted, ligatures may be safely cut short and left to be disposed of by absorption or otherwise. Should this particular branch of the subject yield all that it promises, should it turn out on further trial that when the knot is applied on the antiseptic principle, we may calculate as securely as if it were absent on the occurrence of healing without any deep- seated suppuration, the deligation of main arteries in their continuity will be deprived of the two dangers that now attend it, viz., those of secondary haemorrhage and an unhealthy state of the wound. Further, it seems not unlikely that the present objection to tying an artery in the immediate vicinity of a large branch may be done away with; and that even the innominate, which has lately been the subject of an ingenious experiment by one of the Dublin surgeons, on account of its well-known fatality under the ligature for secondary haemorrhage, may cease to have this unhappy character when the tissues in the vicinity of the thread, instead of becoming softened through the influence of an irritating decomposing substance, are left at liberty to consolidate firmly near an unoffending though foreign body.
It would carry me far beyond the limited time which, by the rules of the Association, is alone at my disposal, were I to enter into the various applications of the antiseptic principle in the several special departments of surgery.
There is, however, one point more that I cannot but advert to, viz., the influence of this mode of treatment upon the general healthiness of an hospital. Previously to its introduction the two large wards in which most of my cases of accident and of operation are treated were among the unhealthiest in the whole surgical division of the Glasgow Royal Infirmary, in consequence apparently of those wards being unfavorably placed with reference to the supply of fresh air; and I have felt ashamed when recording the results of my practice, to have so often to allude to hospital gangrene or pyaemia. It was interesting, though melancholy, to observe that whenever all or nearly all the beds contained cases with open sores, these grievous complications were pretty sure to show themselves; so that I came to welcome simple fractures, though in themselves of little interest either for myself or the students, because their presence diminished the proportion of open sores among the patients. But since the antiseptic treatment has been brought into full operation, and wounds and abscesses no longer poison the atmosphere with putrid exhalations, my wards, though in other respects under precisely the same circumstances as before, have completely changed their character; so that during the last nine months not a single instance of pysemia, hospital gangrene, or erysipelas has occurred in them.
As there appears to be no doubt regarding the cause of this change, the importance of the fact can hardly be exaggerated.
THE PHYSIOLOGICAL THEORY OF FERMENTATION BY LOUIS PASTEUR TRANSLATED BY F. FAULKNER AND D. C. ROBB AND REVISED
THE GERM THEORY AND ITS APPLICATIONS TO MEDICINE AND SURGERY BY MM. PASTEUR, JOURBERT, AND CHAMBERLAND TRANSLATED BY H. C. ERNST, M. D. PROFESSOR OF BACTERIOLOGY IN THE HARVARD MEDICAL SCHOOL
ON THE EXTENSION OF THE GERM THEORY TO THE ETIOLOGY OF CERTAIN COMMON DISEASES BY LOUIS PASTEUR TRANSLATED BY H. C. ERNST, M. D.
Louis Pasteur was born at Dole, Jura, France, December 27, 1822, and died near Saint-Cloud, September 28, 1895. His interest in science, and especially in chemistry, developed early, and by the time he was twenty-six he was professor of the physical sciences at Dijon. The most important academic positions held by him later were those as professor of chemistry at Strasburg, 1849; dean of the Faculty of Sciences at Lille, 1854; science director of the Ecole Normale Superieure, Paris, 1857; professor of geology, physics, and chemistry at the Ecole des Beaux Arts; Professor of chemistry at the Sorbonne, 1867. After 1875 he carried on his researches at the Pasteur Institute. He was a member of the Institute, and received many honors from learned societies at home and abroad.
In respect of the number and importance, practical as well as scientific, of his discoveries, Pasteur has hardly a rival in the history of science. He may be regarded as the founder of modern stereo-chemistry; and his discovery that living organisms are the cause of fermentation is the basis of the whole modern germ- theory of disease and of the antiseptic method of treatment. His investigations of the diseases of beer and wine; of pebrine, a disease affecting silk-worms; of anthrax, and of fowl cholera, were of immense commercial importance and led to conclusions which have revolutionised physiology, pathology, and therapeutics. By his studies in the culture of bacteria of attenuated virulence he extended widely the practise of inoculation with a milder form of various diseases, with a view to producing immunity.
The following papers present some of the most important of his contributions, and exemplify his extraordinary powers of lucid exposition and argument.
TO THE MEMORY OF MY FATHER FORMERLY A SOLDIER UNDER THE FIRST EMPIRE CHEVALIER OF THE LEGION OF HONOR
The longer I live, the better I understand the kindness of thy heart and the high quality of thy mind.
The efforts which I have devoted to these Studies, as well as those which preceded them, are the fruit of thy counsel and example.
Desiring to honor these filial remembrances, I dedicate this work to thy memory.
Our misfortunes inspired me with the idea of these researches. I undertook them immediately after the war of 1870, and have since continued them without interruption, with the determination of perfecting them, and thereby benefiting a branch of industry wherein we are undoubtedly surpassed by Germany.
I am convinced that I have found a precise, practical solution of the arduous problem which I proposed to myself—that of a process of manufacture, independent of season and locality, which should obviate the necessity of having recourse to the costly methods of cooling employed in existing processes, and at the same time secure the preservation of its products for any length of time.
These new studies are based on the same principles which guided me in my researches on wine, vinegar, and the silkworm disease— principles, the applications of which are practically unlimited. The etiology of contagious diseases may, perhaps, receive from them an unexpected light.
I need not hazard any prediction concerning the advantages likely to accrue to the brewing industry from the adoption of such a process of brewing as my study of the subject has enabled me to devise, and from an application of the novel facts upon which this process is founded. Time is the best appraiser of scientific work, and I am not unaware that an industrial discovery rarely produces all its fruit in the hands of its first inventor.
I began my researches at Clermont-Ferrand, in the laboratory, and with the help, of my friend M. Duclaux, professor of chemistry at the Faculty of Sciences of that town. I continued them in Paris, and afterwards at the great brewery of Tourtel Brothers, of Tantonville, which is admitted to be the first in France. I heartily thank these gentlemen for their extreme kindness. I owe also a public tribute of gratitude to M. Kuhn, a skillful brewer of Chamalieres, near Clermont-Ferrand, as well as to M. Velten of Marseilles, and to MM. de Tassigny, of Reims, who have placed at my disposal their establishments and their products, with the most praiseworthy eagerness.
Paris, June 1, 1879.
THE PHYSIOLOGICAL THEORY OF FERMENTATION
I. ON THE RELATIONS EXISTING BETWEEN OXYGEN AND YEAST
It is characteristic of science to reduce incessantly the number of unexplained phenomena. It is observed, for instance, that fleshy fruits are not liable to fermentation so long as their epidermis remains uninjured. On the other hand, they ferment very readily when they are piled up in heaps more or less open, and immersed in their saccharine juice. The mass becomes heated and swells; carbonic acid gas is disengaged, and the sugar disappears and is replaced by alcohol. Now, as to the question of the origin of these spontaneous phenomena, so remarkable in character as well as usefulness for man's service, modern knowledge has taught us that fermentation is the consequence of a development of vegetable cells the germs of which do not exist in the saccharine juices within fruits; that many varieties of these cellular plants exist, each giving rise to its own particular fermentation. The principal products of these various fermentations, although resembling each other in their nature, differ in their relative proportions and in the accessory substances that accompany them, a fact which alone is sufficient to account for wide differences in the quality and commercial value of alcoholic beverages.
Now that the discovery of ferments and their living nature, and our knowledge of their origin, may have solved the mystery of the spontaneous appearance of fermentations in natural saccharine juices, we may ask whether we must still regard the reactions that occur in these fermentations as phenomena inexplicable by the ordinary laws of chemistry. We can readily see that fermentations occupy a special place in the series of chemical and biological phenomena. What gives to fermentations certain exceptional characters of which we are only now beginning to suspect the causes, is the mode of life in the minute plants designated under the generic name of ferments, a mode of life which is essentially different from that in other vegetables, and from which result phenomena equally exceptional throughout the whole range of the chemistry of living beings.
The least reflection will suffice to convince us that the alcoholic ferments must possess the faculty of vegetating and performing their functions out of contact with air. Let us consider, for instance, the method of vintage practised in the Jura. The bunches are laid at the foot of the vine in a large tub, and the grapes there stripped from them. When the grapes, some of which are uninjured, others bruised, and all moistened by the juice issuing from the latter, fill the tub—where they form what is called the vintage—they are conveyed in barrels to large vessels fixed in cellars of a considerable depth. These vessels are not filled to more than three-quarters of their capacity. Fermentation soon takes place in them, and the carbonic acid gas finds escape through the bunghole, the diameter of which, in the case of the largest vessels, is not more than ten or twelve centimetres (about four inches). The wine is not drawn off before the end of two or three months. In this way it seems highly probable that the yeast which produces the wine under such conditions must have developed, to a great extent at least, out of contact with oxygen. No doubt oxygen is not entirely absent from the first; nay, its limited presence is even a necessity to the manifestation of the phenomena which follow. The grapes are stripped from the bunch in contact with air, and the must which drops from the wounded fruit takes a little of this gas into solution. This small quantity of air so introduced into the must, at the commencement of operations, plays a most indispensable part, it being from the presence of this that the spores of ferments which are spread over the surface of the grapes and the woody part of the bunches derive the power of starting their vital phenomena [Footnote: It has been marked in practice that fermentation is facilitated by leaving the grapes on the bunches. The reason of this has not yet been discovered. Still we have no doubt that it may be attributed, principally, to the fact that the interatices between the grapes, and the spaces between the bunch leaves throughout, considerably increase the volume of air placed at the service of the germs of ferment.]. This air, however, especially when the grapes have been stripped from the bunches, is in such small proportion, and that which is in contact with the liquid mass is so promptly expelled by the carbonic acid gas, which is evolved as soon as a little yeast has formed, that it will readily be admitted that most of the yeast is produced apart from the influence of oxygen, whether free or in solution. We shall revert to this fact, which is of great importance. At present we are only concerned in pointing out that, from the mere knowledge of the practices of certain localities, we are induced to believe that the cells of yeast, after they have developed from their spores, continue to live and multiply without the intervention of oxygen, and that the alcoholic ferments have a mode of life which is probably quite exceptional, since it is not generally met with in other species, vegetable or animal.
Another equally exceptional characteristic of yeast and fermentation in general consists in the small proportion which the yeast that forms bears to the sugar that decomposes. In all other known beings the weight of nutritive matter assimilated corresponds with the weight of food used up, any difference that may exist being comparatively small. The life of yeast is entirely different. For a certain weight of yeast formed, we may have ten times, twenty times, a hundred times as much sugar, or even more decomposed, as we shall experimentally prove by-and- bye; that is to say, that whilst the proportion varies in a precise manner, according to conditions which we shall have occasion to specify, it is also greatly out of proportion to the weight of the yeast. We repeat, the life of no other being, under its normal physiological conditions, can show anything similar. The alcoholic ferments, therefore, present themselves to us as plants which possess at least two singular properties: they can live without air, that is without oxygen, and they can cause decomposition to an amount which, though variable, yet, as estimated by weight of product formed, is out of all proportion to the weight of their own substance. These are facts of so great importance, and so intimately connected with the theory of fermentation, that it is indispensable to endeavour to establish them experimentally, with all the exactness of which they will admit.
The question before us is whether yeast is in reality an anaerobian [Footnote: Capable of living without free oxygen—a term invented by Pasteur.—En.] plant, and what quantities of sugar it may cause to ferment, under the various conditions under which we cause it to act.
The following experiments were undertaken to solve this double problem:—We took a double-necked flask, of three litres (five pints) capacity, one of the tubes being curved and forming an escape for the gas; the other one, on the right hand side (Fig. 1), being furnished with a glass tap. We filled this flask with pure yeast water, sweetened with 5 per cent, of sugar candy, the flask being so full that there was not the least trace of air remaining above the tap or in the escape tube; this artificial wort had, however, been itself aerated. The curved tube was plunged in a porcelain vessel full of mercury, resting on a firm support. In the small cylindrical funnel above the tap, the capacity of which was from 10 cc. to 15 cc. (about half a fluid ounce) we caused to ferment, at a temperature of 20 degrees or 25 degrees C. (about 75 degrees F.), five or six cubic centimetres of the saccharine liquid, by means of a trace of yeast, which multiplied rapidly, causing fermentation, and forming a slight deposit of yeast at the bottom of the funnel above the tap. We then opened the tap, and some of the liquid in the funnel entered the flask, carrying with it the small deposit of yeast, which was sufficient to impregnate the saccharine liquid contained in the flask. In this manner it is possible to introduce as small a quantity of yeast as we wish, a quantity the weight of which, we may say, is hardly appreciable. The yeast sown multiplies rapidly and produces fermentation, the carbonic gas from which is expelled into the mercury. In less than twelve days all the sugar had disappeared, and the fermentation had finished. There was a sensible deposit of yeast adhering to the sides of the flask; collected and dried it weighed 2.25 grammes (34 grains). It is evident that in this experiment the total amount of yeast formed, if it required oxygen to enable it to live, could not have absorbed, at most, more than the volume which was originally held in solution in the saccharine liquid, when that was exposed to the air before being introduced into the flask.
Some exact experiments conducted by M. Raulin in our laboratory have established the fact that saccharine worts, like water, soon become saturated when shaken briskly with an excess of air, and also that they always take into solution a little less air than saturated pure water contains under the same conditions of temperature and pressure. At a temperature of 25 degrees C. (77 degrees F.), therefore, if we adopt the coefficient of the solubility of oxygen in water given in Bunsen's tables, we find that 1 litre (1 3/4 pints) of water saturated with air contains 5.5 cc. (0.3 cubic inch) of oxygen. The three litres of yeast- water in the flask, supposing it to have been saturated, contains less than 16.5 cc. (1 cubic inch) of oxygen, or, in weight, less than 23 milligrammes (0.35 grains). This was the maximum amount of oxygen, supposing the greatest possible quantity to have been absorbed, that was required by the yeast formed in the fermentation of 150 grammes (4.8 Troy ounces) of sugar. We shall better understand the significance of this result later on. Let us repeat the foregoing experiment, but under altered conditions. Let us fill, as before, our flask with sweetened yeast-water, but let this first be boiled, so as to expel all the air it contains. To effect this we arrange our apparatus as represented in the accompanying sketch. (Fig 2.) We place our flask, A, on a tripod above a gas flame, and in place of the vessel of mercury substitute a porcelain dish, under which we can put a gas flame, and Which contains some fermentable, saccharine liquid, similar to that with which the flask is filled. We boil the liquid in the flask and that in the basin simultaneously, and then let them cool down together, so that as the liquid in the flask cools some of the liquid is sucked from the basin into the flask. From a trial experiment which we conducted, determining the quantity of oxygen that remained in solution in the liquid after cooling, according to M. Schutzenberger's valuable method, by means of hydrosulphite of soda [Footnote: NaHSO2, now called sodium hyposulphite.—D.C.R.], we found that the three litres in the flask, treated as we have described, contained less than one milligramme (0.015 grain) of oxygen. At the same time we conducted another experiment, by way of comparison (Fig. 3). We took a flask, B, of larger capacity than the former one, which we filled about half with the same volume as before of a saccharine liquid of identically the same composition. This liquid had been previously freed from alterative germs by boiling. In the funnel surmounting A, we put a few cubic centimetres of saccharine liquid in a state of fermentation, and when this small quantity of liquid was in full fermentation, and the yeast in it was young and vigorous, we opened the tap, closing it again immediately, so that a little of the liquid and yeast still remained in the funnel. By this means we caused the liquid in A to ferment. We also impregnated the liquid in B with some yeast taken from the funnel of A. We then replaced the porcelain dish in which the curved escape tube of A had been plunged, by a vessel filled with mercury. The following is a description of two of these comparative fermentations and the results they gave.
The fermentable liquid was composed of yeast-water sweetened with 5 per cent, of sugar—candy; the ferment employed was sacchormyces pastorianus.
The impregnation took place on January 20th. The flasks were placed in an oven at 25 degrees (77 degrees F.).
FLASK A, WITHOUT AIR.
January 21st.—Fermentation commenced; a little frothy liquid issued from the escape tube and covered the mercury.
The following days, fermentation was active. Examining the yeast mixed with the froth that was expelled into the mercury by the evolution of carbonic acid gas, we find that it was very fine, young, and actively budding.
February 3rd.—Fermentation still continued, showing itself by a number of little bubbles rising from the bottom of the liquid, which had settled bright. The yeast was at the bottom in the form of a deposit.
February 7th.—Fermentation still continued, but very languidly.
February 9th.—A very languid fermentation still went on, discernible in little bubbles rising from the bottom of the flask.
FLASK B, WITH AIR.
January 21st.—A sensible development of yeast.
The following days, fermentation was active, and there was an abundant froth on the surface of the liquid.
February 1st.—All symptoms of fermentation had ceased.
As the fermentation in A would have continued a long time, being so very languid, and as that in B had been finished for several days, we brought to a close our two experiments on February 9th. To do this we poured off the liquids in A and B, collecting the yeasts on tared filters. Filtration was an easy matter, more especially in the case of A. Examining the yeasts under the microscope, immediately after decantation, we found that both of them remained very pure. The yeast in A was in little clusters, the globules of which were collected together, and appeared by their well-defined borders to be ready for an easy revival in contact with air.
As might have been expected, the liquid in flask B did not contain the least trace of sugar; that in the flask A still contained some, as was evident from the non-completion of fermentation, but not more than 4.6 grammes (71 grains). Now, as each flask originally contained three litres of liquid holding in solution 5 per cent of sugar, it follows that 150 grammes (2,310 grains) of sugar had fermented in the flask B, and 145.4 grammes (2,239.2 grains) in the flask A. The weights of yeast after drying at 100 degrees C. (212 degrees F.) were—
For the flask B, with air. ... ..1,970 grammes (30.4 grains). For the flask A, without air ... 1,368 grammes [Footnote: This appears to be a misprint for 1.638 grammes=25.3 grains.—D. C. R.].
The proportions were 1 of yeast to 76 of fermented sugar in the first case, and 1 of yeast to 89 of fermented sugar in the second.
From these facts the following consequences may be deduced:
1. The fermentable liquid (flask B), which since it had been in contact with air, necessarily held air in solution, although not to the point of saturation, inasmuch as it had been once boiled to free it from all foreign germs, furnished a weight of yeast sensibly greater than that yielded by the liquid which contained no air at all (flask A) or, at least, which could only have contained an exceedingly minute quantity.
2. This same slightly aerated fermentable liquid fermented much more rapidly than the other. In eight or ten days it contained no more sugar; while the other, after twenty days, still contained an appreciable quantity.
Is this last fact to be explained by the greater quantity of yeast formed in B? By no means. At first, when the air has access to the liquid, much yeast is formed and little sugar disappears, as we shall prove immediately; nevertheless the yeast formed in contact with the air is more active than the other. Fermentation is correlative first to the development of the globules, and then to the continued life of those globules once formed. The more oxygen these last globules have at their disposal during their formation, the more vigorous, transparent, and turgescent, and, as a consequence of this last quality, the more active they are in decomposing sugar. We shall hereafter revert to these facts.
3. In the airless flask the proportion of yeast to sugar was 1/59; it was only 1/79 in the flask which had air at first.
The proportion that the weight of yeast bears to the weight of the sugar is, therefore, variable, and this variation depends, to a certain extent, upon the presence of air and the possibility of oxygen being absorbed by the yeast. We shall presently show that yeast possesses the power of absorbing that gas and emitting carbonic acid, like ordinary fungi, that even oxygen may be reckoned amongst the number of food-stuffs that may be assimilated by this plant, and that this fixation of oxygen in yeast, as well as the oxidations resulting from it, have the most marked effect on the life of yeast, on the multiplication of its cells, and on their activity as ferments acting upon sugar, whether immediately or afterwards, apart from supplies of oxygen or air.
In the preceding experiment, conducted without the presence of air, there is one circumstance particularly worthy of notice. This experiment succeeds, that is to say, the yeast sown in the medium deprived of oxygen develops, only when this yeast is in a state of great vigour. We have already explained the meaning of this last expression. But we wish now to call attention to a very evident fact in connection with this point. We impregnate a fermentable liquid; yeast develops and fermentation appears. This lasts for several days and then ceases. Let us suppose that, from the day when fermentation first appears in the production of a minute froth, which gradually increases until it whitens the surface of the liquid, we take, every twenty-four hours, or at longer intervals, a trace of the yeast deposited on the bottom of the vessel and use it for starting fresh fermentations. Conducting these fermentations all under precisely the same conditions of temperature, character and volume of liquid, let us continue this for a prolonged time, even after the original fermentation is finished. We shall have no difficulty in seeing that the first signs of action in each of our series of second fermentations appear always later and later in proportion to the length of time that has elapsed from the commencement of the original fermentation. In other words, the time necessary for the development of the germs and the production of that amount of yeast sufficient to cause the first appearance of fermentation varies with the state of the impregnating cells, and is longer in proportion as the cells are further removed from the period of their formation. It is essential, in experiments of this kind, that the quantities of yeast successively taken should be as nearly as possible equal in weight or volume, since, celeris paribus, fermentations manifest themselves more quickly the larger the quantity of yeast employed in impregnation.
If we compare under the microscope the appearance and character of the successive quantities of yeast taken, we shall see plainly that the structure of the cells undergoes a progressive change. The first sample which we take, quite at the beginning of the original fermentation, generally gives us cells rather larger than those later on, and possessing a remarkable tenderness. Their walls are exceedingly thin, the consistency and softness of their protoplasm is akin to fluidity, and their granular contents appear in the form of scarcely visible spots. The borders of the cells soon become more marked, a proof that their walls undergo a thickening; their protoplasm also becomes denser, and the granulations more distinct. Cells of the same organ, in the states of infancy and old age, should not differ more than the cells of which we are speaking, taken in their extreme states. The progressive changes in the cells, after they have acquired their normal form and volume, clearly demonstrate the existence of a chemical work of a remarkable intensity, during which their weight increases, although in volume they undergo no sensible change, a fact that we have often characterized as "the continued life of cells already formed." We may call this work a process of maturation on the part of the cells, almost the same that we see going on in the case of adult beings in general, which continue to live for a long time, even after they have become incapable of reproduction, and long after their volume has become permanently fixed.
This being so, it is evident, we repeat, that, to multiply in a fermentable medium, quite out of contact with oxygen, the cells of yeast must be extremely young, full of life and health, and still under the influence of the vital activity which they owe to the free oxygen which has served to form them, and which they have perhaps stored up for a time. When older, they reproduce themselves with much difficulty when deprived of air, and gradually become more languid; and if they do multiply, it is in strange and monstrous forms. A little older still, they remain absolutely inert in a medium deprived of free oxygen. This is not because they are dead; for in general they may be revived in a marvellous manner in the same liquid if it has been first aerated before they are sown. It would not surprise us to learn that at this point certain preconceived ideas suggest themselves to the mind of an attentive reader on the subject of the causes that may serve to account for such strange phenomena in the life of these beings which our ignorance hides under the expressions of YOUTH and AGE; this, however, is a subject which we cannot pause to consider here.
At this point we must observe—for it is a matter of great importance—that in the operations of the brewer there is always a time when the yeasts are in this state of vigorous youth of which we have been speaking, acquired under the influence of free oxygen, since all the worts and the yeasts of commerce are necessarily manipulated in contact with air, and so impregnated more or less with oxygen. The yeast immediately seizes upon this gas and acquires a state of freshness and activity, which permits it to live afterwards out of contact with air, and to act as a ferment. Thus, in ordinary brewery practice, we find the yeast already formed in abundance even before the earliest external signs of fermentation have made their appearance. In this first phase of its existence, yeast lives chiefly like an ordinary fungus.
From the same circumstances it is clear that the brewer's fermentations may, speaking quite strictly, last for an indefinite time, in consequence of the unceasing supply of fresh wort, and from the fact, moreover, that the exterior air is constantly being introduced during the work, and that the air contained in the fresh worts keeps up the vital activity of the yeast, as the act of breathing keeps up the vigour and life of cells in all living beings. If the air could not renew itself in any way, the vital activity which the cells originally received, under its influence, would become more and more exhausted, and the fermentation eventually come to an end.
We may recount one of the results obtained in other experiments similar to the last, in which, however, we employed yeast which was still older than that used for our experiment with flask A (Fig. 2), and moreover took still greater precautions to prevent the presence of air. Instead of leaving the flask, as well as the dish, to cool slowly, after having expelled all air by boiling, we permitted the liquid in the dish to continue boiling whilst the flask was being cooled by artificial means; the end of the escape tube was then taken out of the still boiling dish and plunged into the mercury trough. In impregnating the liquid, instead of employing the contents of the small cylindrical funnel whilst still in a state of fermentation, we waited until this was finished. Under these conditions, fermentation was still going on in our flask, after a lapse of three months. We stopped it and found that 0.255 gramme (3.9 grains) of yeast had been formed, and that 45 grammes (693 grains) of sugar had fermented, the ratio between the weights of yeast and sugar being thus 0.255 divided by 45 = 1 divided by 176. In this experiment the yeast developed with much difficulty, by reason of the conditions to which it had been subjected. In appearance the cells varied much, some were to be found large, elongated, and of tubular aspect, some seemed very old and were extremely granular, whilst others were more transparent. All of them might be considered abnormal cells.
In such experiments we encounter another difficulty. If the yeast sown in the non-aerated fermentable liquid is in the least degree impure, especially if we use sweetened yeast-water, we may be sure that alcoholic fermentation will soon cease, if, indeed, it ever commences, and that accessory fermentations will go on. The vibrios of butyric fermentation, for instance, will propagate with remarkable facility under these circumstances. Clearly then, the purity of the yeast at the moment of impregnation, and the purity of the liquid in the funnel, are conditions indispensable to success.
To secure the latter of these conditions, we close the funnel, as shown in FIG. 2, by means of a cork pierced with two holes, through one of which a short tube passes, to which a short length of india-rubber tubing provided with a glass stopper is attached; through the other hole a thin curved tube is passed. Thus fitted, the funnel can answer the same purposes as our double-necked flasks. A few cubic centimetres of sweetened yeast-water are put in it and boiled, so that the steam may destroy any germs adhering to the sides; and when cold the liquid is impregnated by means of a trace of pure yeast, introduced through the glass- stoppered tube. If these precautions are neglected, it is scarcely possible to secure a successful fermentation in our flasks, because the yeast sown is immediately held in check by a development of anaerobian vibrios. For greater security, we may add to the fermentable liquid, at the moment when it is prepared, a very small quantity of tartaric acid, which will prevent the development of butyric vibrios.
The variation of the ratio between the weight of the yeast and that of the sugar decomposed by it now claims special attention. Side by side with the experiments which we have just described, we conducted a third lot by means of the flask C (Fig. 4), holding 4.7 litres (8 1/2 pints), and fitted up like the usual two-necked flasks, with the object of freeing the fermentable liquid from foreign germs, by boiling it to begin with, so that we might carry on our work under conditions of purity. The volume of yeast-water (containing 5 per cent. of sugar) was only 200 cc. (7 fl. oz.), and consequently, taking into account the capacity of the flask, It formed but a very thin layer at the bottom. On the day after impregnation the deposit of yeast was already considerable, and forty-eight hours afterwards the fermentation was completed. On the third day we collected the yeast after having analyzed the gas contained in the flask. This analysis was easily accomplished by placing the flask in a hot-water bath, whilst the end of the curved tube was plunged under a cylinder of mercury. The gas contained 41.4 per cent. of carbonic acid, and, after the absorption, the remaining air contained:—
Oxygen . ... . ... . ... . ... . ... . ... . ... ... 19.7
Nitrogen . ... . ... . ... . ... . ... . ... . ... . 80.3
Taking into consideration the volume of this flask, this shows a minimum of 50 cc. (3.05 cub. in.) of oxygen to have been absorbed by the yeast. The liquid contained no more sugar, and the weight of the yeast, dried at a temperature of 100 degrees C (212 degrees F.), was 0.44 grammes. The ratio between the weights of yeast and sugar is 0.44/10=1/22.7 [Footnote: 200 cc. of liquid were used, which, as containing 3 per cent., had in solution 10 grammes of sugar.—D.C.R.]. On this occasion, where we had increased the quantity of oxygen held in solution, so as to yield itself for assimilation at the beginning and during the earlier developments of the yeast, we found instead of the previous ratio of 1/76 that of 1/23.
The next experiment was to increase the proportion of oxygen to a still greater extent, by rendering the diffusion of gas a more easy matter than in a flask, the air in which is in a state of perfect quiescence. Such a state of matters hinders the supply of oxygen, inasmuch as the carbonic acid, as soon as it is liberated, at once forms an immovable layer on the surface of the liquid, and so separates off the oxygen. To effect the purpose of our present experiment, we used flat basins having glass bottoms and low sides, also of glass, in which the depth of the liquid is not more than a few millimetres (less than 1/4 inch) (Fig. 5). The following is one of our experiments so conducted:—On April 16th, 1860, we sowed a trace of beer yeast ("high" yeast) in 200 cc. (7 fl. oz.) of a saccharine liquid containing 1.720 grammes (26.2 grains) of sugar-candy. From April 18th our yeast was in good condition and well developed. We collected it, after having added to the liquid a few drops of concentrated sulphuric acid, with the object of checking the fermentation to a great extent, and facilitating filtration. The sugar remaining in the filtered liquid, determined by Fehling's solution, showed that 1.04 grammes (16 grains) of sugar had disappeared. The weight of the yeast, dried at 100 degrees C. (212 degrees F.), was 0.127 gramme (2 grains), which gives us the ratio between the weight of the yeast and that of the fermented sugar 0.123/1.04=1/8.1, which is considerably higher than the preceding ones.
We may still further increase this ratio by making our estimation as soon as possible after the impregnation, or the addition of the ferment. It will be readily understood why yeast, which is composed of cells that bud and subsequently detach themselves from one another, soon forms a deposit at the bottom of the vessels. In consequence of this habit of growth, the cells constantly covering each other prevents the lower layers from having access to the oxygen held in solution in the liquid, which is absorbed by the upper ones. Hence, these which are covered and deprived of this gas act on the sugar without deriving any vital benefit from the oxygen—a circumstance which must tend to diminish the ratio of which we are speaking. Once more repeating the preceding experiment, but stopping it as soon as we think that the weight of yeast formed may be determined by the balance (we find that this may be done twenty-four hours after impregnation with an inappreciable quantity of yeast), in this case the ratio between the weights of yeast and sugar is gr/024 yeast/0 gr. 09 sugar=1/4. This is the highest ratio we have been able to obtain.
Under these conditions the fermentation of sugar is extremely languid: the ratio obtained is very nearly the same that ordinary fungoid growths would give. The carbonic acid evolved is principally formed by the decompositions which result from the assimilation of atmospheric oxygen. The yeast, therefore, lives and performs its functions after the manner of ordinary fungi: so far it is no longer a ferment, so to say; moreover, we might expect to find it to cease to be a ferment at all if we could only surround each cell separately with all the air that it required. This is what the preceding phenomena teach us; we shall have occasion to compare them later on with others which relate to the vital action exercised on yeast by the sugar of milk.
We may here be permitted to make a digression.
In his work on fermentations, which M. Schutzenberger has recently published, the author criticises the deductions that we have drawn from the preceding experiments, and combats the explanation which we have given of the phenomena of fermentation. [Footnote: International Science Series, vol. xx, pp. 179-182. London, 1876.—D. C. R.] It is an easy matter to show the weak point of M. Schutzenberger's reasoning. We determined the power of the ferment by the relation of the weight of sugar decomposed to the weight of the yeast produced. M. Schutzenberger asserts that in doing this we lay down a doubtful hypothesis, and he thinks that this power, which he terms FERMENTATIVE ENERGY, may be estimated more correctly by the quantity of sugar decomposed by the unit-weight of yeast in unit-time; moreover, since our experiments show that yeast is very vigorous when it has a sufficient supply of oxygen, and that, in such a case, it can decompose much sugar in a little time, M. Schutzenberger concludes that it must then have great power as a ferment, even greater than when it performs its functions without the aid of air, since under this condition it decomposes sugar very slowly. In short, he is disposed to draw from our observations the very opposite conclusion to that which we arrived at.
M, Schutzenberger has failed to notice that the power of a ferment is independent of the time during which it performs its functions. We placed a trace of yeast in one litre of saccharine wort; it propagated, and all the sugar was decomposed. Now, whether the chemical action involved in this decomposition of sugar had required for its completion one day, or one month, or one year, such a factor was of no more importance in this matter than the mechanical labour required to raise a ton of materials from the ground to the top of a house would be affected by the fact that it had taken twelve hours instead of one. The notion of time has nothing to do with the definition of work. M. Schutzenberger has not perceived that in introducing the consideration of time into the definition of the power of a ferment, he must introduce at the same time, that of the vital activity of the cells which is independent of their character as a ferment. Apart from the consideration of the relation existing between the weight of fermentable substance decomposed and that of ferment produced, there is no occasion to speak of fermentations or of ferments. The phenomena of fermentation and of ferments have been placed apart from others, precisely because, in certain chemical actions, that ratio has been out of proportion; but the time that these phenomena require for their accomplishment has nothing to do with either their existence proper, or with their power. The cells of a ferment may, under some circumstances, require eight days for revival and propagation, whilst, under other conditions, only a few hours are necessary; so that, if we introduce the notion of time into our estimate of their power of decomposition, we may be led to conclude that in the first case that power was entirely wanting, and that in the second case it was considerable, although all the time we are dealing with the same organism—the identical ferment.
M. Schutzenberger is astonished that fermentation can take place in the presence of free oxygen, if, as we suppose, the decomposition of the sugar is the consequence of the nutrition of the yeast, at the expense of the combined oxygen, which yields itself to the ferment. At all events, he argues, fermentation ought to be slower in the presence of free oxygen. But why should it be slower? We have proved that in the presence of oxygen the vital activity of the cells increases, so that, as far as rapidity of action is concerned, its power cannot be diminished. It might, nevertheless, be weakened as a ferment, and this is precisely what happens. Free oxygen imparts to the yeast a vital activity, but at the same time impairs its power as yeast—qua yeast, inasmuch as under this condition it approaches the state in which it can carry on its vital processes after the manner of an ordinary fungus; the mode of life, that is, in which the ratio between the weight of sugar decomposed and the weight of the new cells produced will be the same as holds generally among organisms which are not ferments. In short, varying our form of expression a little, we may conclude with perfect truth, from the sum total of observed facts, that the yeast which lives in the presence of oxygen and can assimilate as much of that gas as is necessary to its perfect nutrition, ceases absolutely to be a ferment at all. Nevertheless, yeast formed under these conditions and subsequently brought into the presence of sugar, OUT OF THE INFLUENCE OF AIR, would decompose more IN A GIVEN TIME than in any other of its states. The reason is that yeast which has formed in contact with air, having the maximum of free oxygen that it can assimilate is fresher and possessed of greater vital activity than that which has been formed without air or with an insufficiency of air. M. Schutzenberger would associate this activity with the notion of time in estimating the power of the ferment; but he forgets to notice that yeast can only manifest this maximum of energy under a radical change of its life conditions; by having no more air at its disposal and breathing no more free oxygen. In other words, when its respiratory power becomes null, its fermentative power is at its greatest. M. Schutzenberger asserts exactly the opposite (p. 151 of his work— Paris, 1875) [Footnote: Page 182, English edition], and so gratuitously places himself in opposition to facts.
In presence of abundant air supply, yeast vegetates with extraordinary activity. We see this in the weight of new yeast, comparatively large, that may be formed in the course of a few hours. The microscope still more clearly shows this activity in the rapidity of budding, and the fresh and active appearance of all the cells. Fig. 6 represents the yeast of our last experiment at the moment when we stopped the fermentation. Nothing has been taken from imagination, all the groups have been faithfully sketched as they were. [Footnote: This figure is on a scale of 300 diameters, most of the figures in this work being of 400 diameters].
In passing it is of interest to note how promptly the preceding results were turned to good account practically. In well-managed distilleries, the custom of aerating the wort and the juices to render them more adapted to fermentation, has been introduced. The molasses mixed with water, is permitted to run in thin threads through the air at the moment when the yeast is added. Manufactories have been erected in which the manufacture of yeast is almost exclusively carried on. The saccharine worts, after the addition of yeast, are left to themselves, in contact with air, in shallow vats of large superficial area, realizing thus on an immense scale the conditions of the experiments which we undertook in 1861, and which we have already described in determining the rapid and easy multiplication of yeast in contact with air.
The next experiment was to determine the volume of oxygen absorbed by a known quantity of yeast, the yeast living in contact with air, and under such conditions that the absorption of air was comparatively easy and abundant.
With this object we repeated the experiment that we performed with the large-bottomed flask (Fig. 4), employing a vessel shaped like Fig. B (Fig. 7), which is, in point of fact, the flask A with its neck drawn out and closed in a flame, after the introduction of a thin layer of some saccharine juice impregnated with a trace of pure yeast. The following are the data and results of an experiment of this kind.
We employed 60 cc. (about 2 fluid ounces) of yeast-water, sweetened with two percent. of sugar and impregnated with a trace of yeast. After having subjected our vessel to a temperature of 25 degrees C. (77 degrees F.) in an oven for fifteen hours, the drawn-out point was brought under an inverted jar filled with mercury and the point broken off. A portion of the gas escaped and was collected in the jar. For 25 cc. of this gas we found, after absorption by potash 20.6, and after absorption by pyrogallic acid, 17.3. Taking into account the volume which remained free in the flask, which held 315 cc., there was a total absorption of 14.5 cc. (0.83 cub. in.) of oxygen. [Footnote: It may be useful for the non-scientific reader to put it thus: that the 25 cc. which escaped, being a fair sample of the whole gas in the flask, and containing (1) 25-20.6=4.4 cc., absorbed by potash and therefore due to carbonic acid, and (2) 20.6-17.3=3.3 cc., absorbed by pyrogallate, and therefore due to oxygen, and the remaining 17.3 cc. being nitrogen, the whole gas in the flask, which has a capacity of 312 cc., will contain oxygen in the above portion and therefore its amount may be determined provided we know the total gas in the flask before opening. On the other hand we know that air normally contains approximately, 1-5 its volume of oxygen, the rest being nitrogen, so that, by ascertaining the diminution of the proportion in the flask, we can find how many cubic centimeters have been absorbed by the yeast. The author, however, has not given all the data necessary for accurate calculation.—D.C.R.] The weight of the yeast, in a state of dryness, was 0.035 gramme.
It follows that in the production of 35 milligrammes (0.524 grain) of yeast there was an absorption of 14 or 15 cc. (about 7/8 cub. in.) of oxygen, even supposing that the yeast was formed entirely under the influence of that gas: this is equivalent to not less than 414 cc. for 1 gramme of yeast (or about 33 cubic inches for every 20 grains). [Footnote: This number is probably too small; it is scarcely possible that the increase of weight in the yeast, even under the exceptional conditions of the experiment described, was not to some extent at least due to oxidation apart from free oxygen, inasmuch as some of the cells were covered by others. The increased weight of the yeast is always due to the action of two distant modes of vital energy— activity, namely, in presence and activity in absence of air. We might endeavor to shorten the duration of the experiment still further, in which case we would still more assimilate the life of the yeast to that of ordinary moulds.]
Such is the large volume of oxygen necessary for the development of one gramme of yeast when the plant can assimilate this gas after the manner of an ordinary fungus.
Let us now return to the first experiment described in the paragraph on page 292 in which a flask of three litres capacity was filled with fermentable liquid, which, when caused to ferment, yielded 2.25 grammes of yeast, under circumstances where it could not obtain a greater supply of free oxygen than 16.5 cc. (about one cubic inch). According to what we have just stated, if this 2.25 grammes (34 grains) of yeast had not been able to live without oxygen, in other words, if the original cells had been unable to multiply otherwise than by absorbing free oxygen, the amount of that gas required could not have been less than 2.25 X 4l4 cc., that is, 931.5 cc. (56.85 cubic inches). The greater part of the 2.25 grammes, therefore, had evidently been produced as the growth of an anaerobian plant.
Ordinary fungi likewise require large quantities of oxygen for their development, as we may readily prove by cultivating any mould in a closed vessel full of air, and then taking the weight of plant formed and measuring the volume of oxygen absorbed. To do this, we take a flask of the shape shown in Fig. 8, capable of holding about 300 cc. (10 1/2 fluid ounces), and containing a liquid adapted to the life of moulds. We boil this liquid, and seal the drawn-out point after the steam has expelled the air wholly or in part; we then open the flask in a garden or in a room. Should a fungus-spore enter the flask, as will invariably be the case in a certain number of flasks out of several used in the experiment, except under special circumstances, it will develop there and gradually absorb all the oxygen contained in the air of the flask. Measuring the volume of this air, and weighing, after drying, the amount of plant formed, we find that for a certain quantity of oxygen absorbed we have a certain weight of mycelium, or of mycelium together with its organs of fructification. In an experiment of this kind, in which the plant was weighed a year after its development, we found for 0.008 gramme (0.123 gram) of MYCELIUM, dried at 100 degrees C. (212 degrees F.), an absorption that amounted to not less than 43 cc. (2.5 cubic inches) of oxygen at 25 degrees. These numbers, however, must vary sensibly with the nature of the mould employed, and also with the greater or less activity of its development, because the phenomena is complicated by the presence of accessory oxidations, such as we find in the case of mycoderma vini and aceti, to which cause the large absorption of oxygen in our last experiment may doubtless be attributed. [Footnote: In these experiments, in which the moulds remain for a long time in contact with a saccharine wort out of contact with oxygen—the oxygen being promptly absorbed by the vital action of the plant (see our Memoire sur les Generations dites Spontanees, p. 54. note)—there is no doubt that an appreciable quantity of alcohol is formed because the plant does not immediately lose vital activity after the absorption of oxygen.
A 300 cc. (10-oz.) flask, containing 100 cc. of must, after the air in it had been expelled by boiling, was open and immediately re-closed on August 15th, 1873. A fungoid growth—a unique one, of greenish-grey colour—developed from spontaneous impregnation, and decolourized the liquid, which originally was of a yellowish- brown. Some large crystals, sparkling like diamonds, of neutral tartrate of lime, were precipitated, about a year afterwards, long after the death of the plant, we examined this liquid. It contained 0.3 gramme (4.6 grains) of alcohol, and 0.053 gramme (0.8 grain) of vegetable matter, dried at 100 degrees C. (212 degrees F.). We ascertained that the spores of the fungus were dead at the moment when the flask was opened. When sown, they did not develop in the least degree.]