Chapter III (1)

But there is still another class of chemical phenomena where the life without air of microscopic organisms is fully shown. Pasteur proved that in the special fermentation which bears the name of putrefaction the _primum movens_ of the putrefaction resides in microscopic vibrios of absolutely the same order as those which compose the butyric ferment. The fermentation of sugar, of mannite, of gums, of lactate of lime, by the butyric vibrio, so closely resembles the phenomena of putrefaction, that one might call these fermentations the putrefaction of sugar and of the other products.

If it has been thought right to call the fermentation of animal matters putrefaction, it is because at the moment of the decomposition of fibrine, of albumen, of blood, of gelatine, of the substance of the tendons, &c., the sulphur, and even the phosphorus, which enter into their composition give rise to putrid odours, due to the evil-smelling gases of sulphur and phosphorus.

The phenomena of putrefaction being then simply fermentations, differing only in regard to the chemical composition of the fermenting matters, Liebig naturally included them in his general theory of the decomposition of organic matters after death. At a period long antecedent to Pasteur's labours it had been established that there existed in putrefying matters fungi or microscopic animalculæ, and the idea had taken shape that these creatures might have an influence in the phenomena. The proofs were wanting, but the notion of a possible relation remained. We may read in his 'Lessons on Chemistry' with what disdain Liebig mentioned these hypothetical opinions.

'Those who pretend to explain the putrefaction of animal substances by the presence of animalculæ,' he wrote, 'reason very much like a child who would explain the rapidity of the Rhine by attributing it to the violent motions imparted to it in the direction of Bingen by the numerous wheels of the mills of Mayence. Is it possible to consider plants and animals as the causes of the destruction of other organisms when their own elements are condemned to undergo the same decompositions as the creatures which have preceded them? If the fungus is the cause of the destruction of the oak, if the microscopic animalcula is the cause of the putrefaction of the dead elephant, I would ask in my turn what is the cause which determines the putrefaction of the fungus or of the microscopic animalcula when life is withdrawn from these two organisms?'

Thirty-two years later, and after Pasteur had accumulated, during more than twenty years, proof upon proof that the theory of Liebig would not stand examination, a physician of Paris, M. Bouillaud, asked, with the insistent voice of a querulous octogenarian: 'Let M. Pasteur then tell us here, in presence of the Académie de Médecine, what are the ferments of the ferments.'

Before replying to this argument, which Liebig and M. Bouillaud believed to be irrefutable, Pasteur, wishing to mark all the phases of the phenomena, expounded in a short preamble the part played by atmospheric oxygen in the destruction of animal and vegetable matters after death. It is easy to understand, indeed, that fermentation and putrefaction only represent the first phase of the return to the atmosphere and to the soil of all that has lived. Fermentations and putrefactions give rise to substances which are still very complex, although they represent the products of decomposition of fermentable matters. When sugar ferments, a large proportion of it becomes gas; but alongside of the carbonic acid gas which is formed, and which is, indeed, a partial return of the sugar to the atmosphere, new substances, such as alcohol, succinic acid, glycerine, and materials of yeast, are produced. When the flesh of animals putrifies, certain products of decomposition, also very complex, are formed with the vapour of water and the other gases of putrefaction. Where, then, does nature find the agents of destruction of these secondary products?

The great fact of the destruction of animal and vegetable matters is accomplished by slow combustion, through the appropriation of atmospheric oxygen. Here, again, one must banish from science the preconceived views which assumed that the oxygen seized directly on the organic matter after death, and that this matter was consumed by purely chemical processes. It is life that presides over this work of death.

If fermentation and putrefaction are principally the work of microscopic anaérobies, living without free oxygen, the slow combustion is found very largely, if not exclusively, to depend upon a class of infinitely small aérobies. It is these last which have the property of consuming the oxygen of the air. It is these lower organisms which are the powerful agents in the return to the atmosphere of all which has lived. Mildew, mould, bacteria, which we have already noticed, monads, two thousand of which would go to make up a millimeter, all these microscopic organisms are charged with the great work of re-establishing the equilibrium of life by giving back to it all that it has formed.

To demonstrate the important part played everywhere by these microscopic organisms, Pasteur made two experiments. He first introduced into vessels air deprived of all dust. This process we shall have occasion to examine in all its details, in connection with the researches on spontaneous generation. In these vessels, containing pure air, were placed the water of yeast with sugar dissolved in it, milk, sawdust--all of which had been deprived by heat of the germs of the lower organisms. The vessels and their contents were then subjected to a temperature of twenty-five to thirty-five degrees Centigrade. In a series of parallel experiments, made under the same conditions and at the same temperature, Pasteur took no steps to prevent the germination of the little seeds of mould suspended in the air, or associated with the substances contained in the vessels, neither did he avoid other infinitely small germs of the class aérobies.

After some time the air of all the vessels of the two series was submitted to analysis, when, behold, a very interesting fact! In the vessels where life had been withdrawn from the organic matters--that is to say, where there were no germs--the air still contained a large proportion of oxygen. In the vessels, on the contrary, where the microscopic organisms had been allowed to develop, the oxygen was totally absent, having been replaced by carbonic acid gas. And, further, for this absorption and total consumption of the oxygen gas a few days had sufficed; while in the vessels without microscopic life there remained, after several years, a considerable quantity of oxygen in a free state, so weak is the proportion of oxygen that the organic matters consume directly and chemically when the infinitely small organisms are absent.

But can these microscopic organisms, after having decomposed or burnt up all these secondary products, be in their turn decomposed?

How, cried M. Bouillaud, repeating his question, can they be destroyed or decomposed? How can their materials, which are of the same order as those of all the living creatures of the earth, be gasified and caused to return to the atmosphere? After having been charged with the transformation of others, whose business will it be to transform them?

A ferment which has finished its work, replied Pasteur, and which for want of aliment cannot continue it, becomes in its turn an accumulation, so to speak, of dead organic matters. Such, for example, would be an accumulation of yeast exposed to the air. Leave this mass to itself in summer temperature, and you will see appear in the interior of the mass anaérobic vibrios and the putrefactions associated with their life when protected from contact with the air. At the same time, on the surface of the entire mass--that is to say, that which finds itself in immediate contact with the oxygen of the air--the germs of bacteria, the seeds of mould will grow, and, by fixing the oxygen, determine the slow combustions which gasify the mass. The ferments of ferments are simply ferments. As long as the aérobic ferments of the surface have at their disposal free oxygen, they will multiply and continue their work of destruction. The anaérobic vibrios perish for want of new matter to decompose, and they form, in their turn, a mass of organic matter which, by and by, becomes the prey of aérobies. The portion of the aérobies which has lived becomes the prey either of new aérobies of different species, or of individuals of their own species, so that from putrefaction to putrefaction, and from combustion to combustion, the organic mass with which we started finds itself reduced to an assemblage of anaérobic and aérobic germs--of those same germs which were mixed up in the original primitive organic substances.

Though a collection of germs becomes again in its turn a collection of organic matter, subject to the double action of the phenomena of putrefaction and of combustion, there need be no anxiety as to their ultimate destruction; in the final analysis they represent life under its eternal form, for life is the germ, and the germ is life.

* * * * *

Thus in the destruction of that which has lived, all reduces itself to the simultaneous action of these three great natural phenomena--fermentation, putrefaction, and slow combustion. A living organism dies--animal, or plant, or the remains of one or the other. It is exposed to the contact of the air. To the life which has quitted it succeeds life under other forms. In the superficial parts, which the air can reach, the germs of the infinitely small aérobies hatch and multiply themselves. The carbon, the hydrogen, and the nitrogen of the organic matters are transformed by the oxygen of the air, and under the influence of the life of these aérobies, into carbonic acid, vapour of water, and ammonia gas. As long as organic matter and air are present, these combustions will continue. While these superficial combustions are going on, fermentation and putrefaction are doing their work in the interior of the mass by the developed germs of the anaérobies, which not only do not require oxygen for their life, but which oxygen actually kills. Little by little, at length, by this work of fermentation and slow combustion, the phenomenon is accomplished. Whether in the free atmosphere, or under the earth, which is always more or less impregnated with air, all animal and vegetable matters end by disappearing. To arrest these phenomena an extremely low temperature is required. It is thus that in the ice of the Polar regions antediluvian elephants have been found perfectly intact. The microscopic organisms could not live in so cold a temperature. These facts still further strengthen all the new ideas as to the important part performed by these infinitely small organisms, which are, in fact, the masters of the world. If we could suppress their work, which is always going on, the surface of the globe, encumbered with organic matters, would soon become uninhabitable.

FOOTNOTE:

[8] [A millimeter is 1/25th of an inch.]

_ACETIC FERMENTATION._

THE MANUFACTURE OF VINEGAR.

Soon afterwards Pasteur came upon a most curious illustration of the 'fixation' of atmospheric oxygen by a microscopic organism--the transformation of wine into vinegar. As its name indicates, vinegar is nothing else than wine turned sour. Everybody has remarked that wine, left to itself, in circumstances which occur daily, is frequently transformed into vinegar. This is noticed more particularly when bottles, having been uncorked, are left in a half-empty condition. Sometimes, however, wine turns sour even in corked bottles. In this case we may be sure that the bottles have been standing upright, and that corks more or less defective have permitted the air to penetrate into the wine. The presence of air, in fact, is indispensable to the chemical act of transforming wine into vinegar. How does this air intervene? And what is the little microscopic creature which, in conjunction with the air, becomes the agent of this fermentation?

In a celebrated lecture given at Orleans at the request of the manufacturers of vinegar in that town, Pasteur, after having stated the two foregoing scientific questions, proceeded to examine the difference between wine and vinegar. What takes place in the fermentation of the juice of the grape which yields the wine? The sugar of this juice disappears, giving place to carbonic acid gas, which is exhaled during fermentation, and to alcohol, which remains in the fermented liquid. Formerly, chemists gave the name of 'spirit' to all volatile matters which could be collected from distillation. Now, when we distil wine and condense the vapour in a worm surrounded by cold water, we collect the spirit of wine at the extremity of the worm--this, when the water with which it is mixed during distillation is withdrawn from it, we designate by the name of alcohol. Vinegar contains no alcohol. When distilled it yields water and a spirit. But this spirit is acid, with a very pungent odour, and not inflammable like spirit of wine. Separated from the water which had accompanied it during the distillation, this spirit takes the name of acetic acid. This is the form in which it is used in smelling bottles--in those bottles of English salts the vapour of which is so penetrating.

In the formation of vinegar in contact with air the alcohol disappears, and is replaced by acetic acid. The air has thus given up something to the wine. Atmospheric air every one knows to be a mixture of nitrogen and oxygen, the nitrogen in the proportion of four-fifths of the total volume, and the oxygen of one-fifth. Well, in the transformation of wine into vinegar the nitrogen remains inactive. It is the oxygen alone which enters into combination with the alcohol. You ask for the proof of this? Take a bottle of wine turned sour, a bottle which at the same time is stopped hermetically; if the oxygen of the air contained in the bottle has combined with the alcohol, then, instead of air, there will be nothing in the bottle but nitrogen gas. Turn the bottle upside down and open it in a basin of water. The water of the basin will rush into the bottle to fill the partial vacuum created by the disappearance of the oxygen. The volume of water which enters the bottle is precisely equal to a fifth part of the total original volume of the air which the bottle contained at the time when it was closed. Moreover, it is easy to show that the gas which remains in the bottle has the properties of nitrogen gas. A lighted match is extinguished in it as if plunged into water, and a bird dies immediately in it of asphyxia.

If we confine our knowledge to what has gone before, it would seem that alcohol diluted with water and exposed to the air ought to furnish acetic acid. It is not so, however. Pure water alcoholised to the degree of ordinary wines may remain for whole years in contact with the air, without the least acetification. In this difference between natural wine and pure water alcoholised, and exposed to contact with air, we touch upon a vital point in the phenomena of fermentation. The celebrated theory of Liebig, which Pasteur was destined to overthrow, might be thus summed up:--If pure alcoholised water cannot become sour in contact with air, as is the case with wine, it is because the pure alcoholised water lacks the albuminoid substance which exists in the wine in a state of chemical alteration, and which is a ferment capable of causing the oxygen of the air to combine with the alcohol. And the proof, according to Liebig, that things act rigorously thus is, that if you add to the mixture of water and alcohol a little flour, or a little meat-juice, or even a minute quantity of any vegetable juice, the acetic fermentation arises, as if by compulsion. In other words, by the addition of a small quantity of any nitrogenised substance in process of alteration, you cause the union of the oxygen of the air with the alcohol.

There is doubtless always in the wine, when it turns sour, a necessary intermediary, producing the fixation of the oxygen of the air; since in no circumstances can pure alcohol, diluted to any degree whatever with pure water, transform itself into vinegar. But this necessary intermediary is not, as the German theory would have it, a dead albuminoid substance; it is a plant, and of all plants one of the simplest and most minute, which has been known from time immemorial under the name of flower of vinegar. This little fungus is invariably present on the surface of a wine which is being transformed into vinegar. Liebig was not ignorant of this, but he regarded it as a simple coincidence. Do we not know, said he, that whenever an infusion of organic matter is exposed to the air it becomes covered with a cryptogamic vegetation, or is invaded by a crowd of animalculæ? Is not vinegar a vegetable infusion? Vinegar affords a refuge to the flower of vinegar, just as it gives refuge to what are called the little eels of vinegar.

We can appreciate here the uncertainties of pure observation. The great art--and no one practised it better than Pasteur--consists in instituting decisive experiments which leave no room for an inexact interpretation of facts. These decisive proofs of the true part played by the little microscopic fungus, by this flower of vinegar, this mycoderma aceti, are thus formulated by Pasteur. It is but another example of the method which he used in alcoholic, lactic, and tartaric fermentations. The theories of Berzelius, of Mitscherlich, and of Liebig were destined again to receive the rudest shocks by the demonstration of these rigorous facts.

Let us place a little wine in a bottle, then hermetically seal it, and leave it to itself. In these conditions the wine becomes sour. But if we take the precaution of putting the bottle into hot water, so that the wine and the air in the bottle may be heated for some instants to a temperature of 60° Centigrade, and if, after cooling, we leave the bottle to itself, the wine in these conditions will never become transformed into vinegar. The heating, however, must have left intact the albuminoid or nitrogenous substances contained in the wine. These, then, cannot constitute the ferment of the vinegar. Can it be maintained that by heating the wine to 60° we have altered the albuminoid matter, which is, on this account, no longer able to act as a ferment, or, in other words, no longer able to determine the union of the oxygen of the air with the alcohol? This hypothesis falls to pieces before the following experiment. Open the bottle, blow into it with bellows, so that the once heated wine shall come into contact with ordinary air, and the acetification of the wine will take place.

But the master experiment is the following. We have seen that pure alcoholised water never turns sour unless some albuminoid matter is introduced into it. Pasteur saw that this albuminoid matter might be completely suppressed and replaced by saline crystallisable substances, alkaline and earthy phosphates, to which has been added a little phosphate of ammonia. In these conditions, especially if the alcoholised water be acidulated by small quantities of pure acetic acid, one actually sees the mycoderm developing, and the alcohol transforming itself into acetic acid. It is not possible to demonstrate in a more convincing manner that the albuminoid matters of the wine are not in this case the acetic ferment. These albuminoid matters, however, contribute to the acetic fermentation, but only as being an aliment to the mycoderma aceti, and notably a nitrogenous aliment. The true and only ferment of vinegar is the little fungus; it is the great agent of the phenomenon; it, indeed, accomplishes all.

Is there not a great charm in seeing an obscure subject clearly illuminated by facts well understood and well interpreted? If in a bottle containing wine and air and raised to a temperature of 50° or 60° the wine never turns sour, it is because the germs of the mycoderma aceti, which the wine and the air hold in suspension, are deprived of all vitality by the heat. Placed, however, in contact with ordinary air, this once-heated wine can turn sour; because, though the germs of the mycoderma aceti contained at first in the wine are killed, this is not the case with those derived from the surrounding air. Pure alcoholised water never turns sour, even in contact with ordinary air, and with whatever germs this air may carry, or that may be found in the dust of the vessels which receive it. The reason is that these germs cannot become fertile because of the absence of their indispensable food. Wine in bottles well filled and laid flat do not acetify; this is because the mycoderm cannot multiply for lack of oxygen. Without doubt the air constantly penetrates through the pores of the cork, but always in such feeble quantities that the colouring matters of the wine, and other more or less oxydisable constituents, take possession of it without leaving the smallest quantity for the germs of the mycoderm which are generally suspended in the wine. When the bottle is upright the conditions are quite altered. The desiccation of the cork renders it much more permeable to the air, and the germs of the mycoderm on the surface of the liquid, if any exist there, are enveloped by air.

Thus, to recapitulate in a few words the principles which have just been established; it is easy to see that the formation of vinegar is always preceded by the development, on the surface of the wine, of a little plant formed of strangulated particles, of an extreme tenuity, and the accumulation of which sometimes takes the form of a hardly visible veil, sometimes of a wrinkled film of very slight thickness, and greasy to the touch, because of the various fatty matters which the plant contains.

This cryptogam has the singular property of condensing considerable quantities of oxygen and of provoking the _fixation_ of this gas upon the alcohol, which is thereby transformed into acetic acid. The little mycoderm is not less exacting than larger vegetables. It must have its appropriate aliments. Wine offers them in abundance: nitrogenous matters, the phosphates of magnesia and of potash. The mycoderm thrives, moreover, in warm climates. To cultivate it in temperate regions like ours it is well to warm artificially the places where it is cultivated. But if wine contains within itself all the elements necessary to the life of the little mycoderm, this life is further promoted by rendering the wine more acid through the addition of acetic acid.

What, then, can be more simple than to produce vinegar from wine--a manufacture which justly makes the reputation of the town of Orleans? Take some wine, and after having mixed with it one-fourth or one-third of its volume of vinegar already formed, sow on its surface the little plant which does the work of acetification. It is only necessary to skim off, by means of a wooden spatula, a little of the mycodermic film from a liquid covered with it, and to transfer it to the liquid to be acetified. The fatty matters which it contains render the wetting of it difficult. Thus, when we plunge into the liquid the spatula covered with the film, the latter detaches itself and spreads out over the surface instead of falling to the bottom. When we operate in summer, or in a room heated to 15° or 25° Centigrade in winter, in twenty-four or forty-eight hours at most, the mycoderm covers the whole liquid, so easy and rapid is its development. After some days all the wine has become vinegar.

On one occasion, in a discussion which he was holding at the Academy of Sciences, Pasteur, wishing to affirm the prodigious activity of the life and multiplication of this little organism, expressed himself thus:--

'I would undertake in the space of twenty-four hours to cover with mycoderma aceti a surface of vinous liquid as large as the hall in which we are here assembled. I should only have to sow in it the day before almost invisible particles of newly-formed mycoderma aceti.'

Let the reader try to imagine the millions upon millions of little mycoderma particles which would come to life in that one day.

But how is the mycoderm seed to be obtained in the first instance? Nothing more simple. The mycoderma aceti is one of those little so-called 'spontaneous' productions which are sure to appear of themselves on the surface of liquids or infusions suitable to their development. In wine, in vinegar, or suspended in air, everywhere around us, in our towns, in our houses, there exist germs of this little plant. If we wish to procure some fresh mycoderm it is only necessary to put a mixture of wine and vinegar into a warm place. In a few days, generally, if not always, there appear here and there little greyish patches scattering the light instead of regularly reflecting it, as does the surrounding liquid. These specks go on increasing progressively and rapidly. This is the mycoderma aceti raised from the seeds which the wine or the added vinegar contained, or which the air deposited; just as we see a field covered with divers weeds by seeds naturally distributed in the earth, or which have been brought to it by the wind or by animals. Even in this last circumstance the comparison holds good, for after you have put wine or vinegar in a warm place there soon appear, whence we know not, little reddish flies, so commonly seen in vinegar manufactories, and in all places where vegetable matter is turning sour. With their feet, or with their probosces, these flies transport the seed.

* * * * *

At Orleans the process for the manufacture of vinegar is very simple. Barrels ranged over each other have on each of their vertically-placed bottoms a circular opening some centimeters in diameter, and a smaller hole adjacent, called _fausset_, for the air to pass in and out when the large opening is closed, either by the funnel, through which the wine is introduced, or by the syphon, which is used for drawing off the vinegar. These barrels, of which the capacity is 230 litres, are half filled. The manual labour consists in keeping up a suitable temperature in the vessel, and in drawing from it every eight days about eight or ten litres of vinegar, which are replaced by eight or ten litres of wine.

A barrel in which this give-and-take of wine and vinegar goes on is technically called a 'mother.' The starting of a 'mother' is not a rapid process. We begin by introducing into the barrel 100 litres of very good and very limpid vinegar; then two litres only of wine are added. Eight days after, three litres of wine are added, a week later four or five, until the barrel contains about 180 to 200 litres. Then for the first time vinegar is drawn off in sufficient quantity to bring back the volume of the liquid to about 100 litres. At this moment the labours of the 'mother' begin. Henceforward ten litres of vinegar may be drawn off every eight days, to be replaced by ten litres of wine. This is the maximum that a cask can yield in a week. When the casks work badly, as is often the case, it is necessary to diminish their production.

This Orleans system has many drawbacks. It requires three or four months to prepare what is called a 'mother,' which must be nourished with wine _very_ regularly once a week under penalty of seeing it lose all its power. Then it is necessary to continue the manufacture at all times, whether the vinegar be required or not. To reconstitute a 'mother,' one must begin from the very beginning, a process which involves a loss of three or four months' time. Lastly--a condition which is at times very inconvenient--a 'mother' cannot be transported from one place to another, or even from one part of the same locality to another. The 'mother,' in fact, must rest immovable.

Pasteur advised the suppression of the 'mothers.' He recommended an apparatus, which is simply a vat, placed in a chamber the temperature of which can be raised to 20° or 25° Centigrade. In these vats vinegar already formed is mixed with wine. On the surface is sown the little plant which converts the wine into vinegar. The mode of sowing it has been already explained. The acetification begins with the development of the plant.

A great merchant of Orleans, who had from the first adopted Pasteur's process, and who had won the prize offered by the 'Society for the Encouragement of National Industry' for a manufactory perfected after these principles, has stated that at the end of nine or ten days, sometimes even in eight, all the acetified wine is converted into vinegar. From a hundred litres of wine he drew off ninety-five litres of vinegar. After the great rise of temperature observed at the moment of the formation of the vinegar, and which is caused by the chemical union of the alcohol and the oxygen of the air, the vinegar is allowed to cool. It may then be drawn from the vat, introduced into barrels, refined, and straightway delivered, fit for consumption. When the vat is quite emptied, and well cleaned, a new mixture is made of vinegar and wine, the little plant is sown as before, and the same facts are reproduced in the second as in the first operation.

* * * * *

In the vessels where vinegar is preserved, whether in the manufactories, in private houses, or in grocers' shops, it often happens that the liquid becomes turbid, and impoverished in an extraordinary manner; it even ends in putrefaction, if a remedy be not promptly applied. Pasteur has pointed out the cause of these phenomena. After the alcohol has become acetic acid by the combustive action of the mycoderm, the question remains, what becomes of the mycoderm? Most frequently it falls to the bottom of the vessel, having no more work to accomplish. This is a phase of the manufacture which must be watched with care. It is shown by the experiments of Pasteur that the mycoderma aceti can live on vinegar already formed, maintaining its power of fixing the oxygen on certain constituents of the liquid. In this case the acetic acid itself is the seat of the chemical action--in other words, the oxygen unites with the carbon of the acetic acid, and transforms it into carbonic acid, and as the acetic acid has a composition which can be represented by carbon and water, it follows that if the combustion is allowed to take its course, instead of vinegar we have eventually nothing but water mixed with a small proportion of nitrogenous and mineral matters, and the remains of the mycoderm. We have thus an ordinary organic infusion exempt from all acidity, and one which could not be better fitted to become the prey of the vibrios of putrefaction or of the aérobic mucors. By these mucors, moreover, which form a film on the surface of the liquid after the mycoderm has fallen, the anaérobic vibrios, protected from the action of the air, can come into active existence. Here we find ourselves in presence of one of those double phenomena, of putrefaction in the deeper parts of the liquid, and of combustion at the surface which is in contact with the air. Nothing is more prejudicial to the quality of the vinegar than the setting in of this combustion after the vinegar has been formed, and when it contains no more alcohol. The first materials of the vinegar upon which the oxygen transmitted by the mycoderm fixes are, in fact, the ethereal and aromatic constituents which give to vinegar its chief value.

Another cause of the deterioration of the quality of vinegar, which is sometimes very annoying to the manufacturer, consists in the frequent presence of little eel-like organisms, very curious when viewed with a strong magnifier. Their bodies are so transparent that their internal organs can be easily distinguished. These eel-like creatures multiply with extraordinary rapidity. Certainly there is not a single barrel of vinegar manufactured by the Orleans system which does not contain them in alarming numbers. Prior to Pasteur's investigations, the ignorance regarding these organisms was such that they were actually considered necessary to the production of the vinegar; whereas they are, on the contrary, most inimical to it, and must, if possible, be got rid of. This is, moreover, rendered desirable by the repugnance which is naturally felt to using a liquid defiled by the presence of such animalcules--a repugnance which becomes almost insurmountable to anyone who has once seen through a microscope the swarms contained in a drop of vinegar. The mischief wrought by these little beings in the manufacture of vinegar results from the fact that they require air to live. The effect can easily be perceived by filling to the brim a bottle of vinegar, corking it, and then comparing it with a similar bottle half filled with the same vinegar, and left uncorked in contact with the air. In the first bottle, the motions of the eel-like creatures become gradually slower, until after a few days they cease to multiply and fall lifeless to the bottom of the vessel. In the second bottle, on the contrary, they continue to swarm and move about. This need of oxygen is further demonstrated by the fact that, if the vinegar reaches a certain depth in the bottle, life is suspended in the lower parts, and the little eel-like organisms, in order to breathe more freely, form a crawling zone in the upper layers of the liquid.

Connecting these observations with the other fact that the vinegar is formed by the action of the mycodermic film on its surface, we can understand at once that the mycoderm and the little eels continually carry on a struggle for existence, since both of these living things--the one animal the other vegetable--imperiously demand the same aliment, oxygen. They live, moreover, in the same superficial layers, a circumstance which gives rise to very curious phenomena. When, for one reason or another, the film of mycoderm is not formed, or when there is any delay in its production, the little eels invade in such great numbers the upper layers of the liquid that they absorb all the oxygen. The little plant has in consequence great difficulty in developing itself or even in beginning its life. Reciprocally, when the work of acetification is active, and when the mycoderm has occupied the upper layers, it gradually drives away the eels, which take refuge, not deep down, where they would perish, but against the moist sides of the barrel or the vat. There they form a thick whitish scum all in motion. It is a very curious spectacle. Here their enemy, the mycoderm, can no longer injure them to the same extent, since they are surrounded with air; and here they wait with impatience for the moment when they can again take their place in the liquid, and, in their turn, fight against the mycoderm. In Pasteur's process, where the vats are very often cleansed, it is easy to keep them free from these little animalcules; they have not time to multiply to a hurtful extent. Indeed, if the operation be well conducted, they do not make their appearance at all.

* * * * *

Nearly all Pasteur's publications have had from the moment of their appearance to undergo the severest criticism. Their novelty caused them to clash with the prejudices and errors current in science. His researches on fermentation provoked lively opposition. Liebig did not accept without recrimination a series of researches which concurred in upsetting the theory he had enunciated and defended in all his works. After having kept silence for ten years, he published, at Munich, where he was professor, a long memoir entirely directed against Pasteur's results. In 1870, on the eve of the war, Pasteur, who was at that time returning from a scientific journey into Austria, determined to pass by Munich, with the view of attempting to convince his distinguished adversary. Liebig received him with great courtesy, but, hardly recovered from an illness, he alleged his convalescence as a reason for declining all discussion.

Then followed the Franco-German war. Hardly was it terminated when Pasteur brought before the Academy of Sciences at Paris a defence of what he had published, as a sort of challenge to his illustrious opponent. The memoir of Liebig was filled with the most skilful arguments.

'I pondered it for nearly ten years before producing it,' he wrote. Pasteur, putting aside all subtleties of argument, went straight to the two objections of the German chemist which lay at the root of the discussion.

It may be remembered that one of the most decisive proofs by which Pasteur overthrew Liebig's theory resulted from the experiments in which by the aid of mineral bodies and fermentable matter he produced a special living ferment for each definite fermentation. By removing all nitrogenous organic matter, which in Liebig's theory constitutes the ferment, Pasteur established, at one and the same time, the life of the ferment and the absence of all action of albuminoid matter in process of alteration. Liebig here formally contested the fact that Pasteur had been able to produce yeast and alcoholic fermentation in a sweetened mineral medium by sowing therein an infinitesimal quantity of yeast. It is certain that, ten years previously, when Pasteur announced the production of yeast life and alcoholic fermentation under such conditions, his experiment was one so difficult to perform that it sometimes happened to Pasteur himself to be unable to reproduce it. The cells of yeast sown in the sweetened mineral medium found themselves often associated with other microscopic organisms, which were singularly hurtful to the life of the yeast. Pasteur was at this period far from being familiarised with the delicacy which such experiments require, and he did not yet know all the precautions indicated later on, which were indispensable to success. Though in his original memoir of 1860 Pasteur had pointed out the difficulties of his experiment, these difficulties existed nevertheless. Liebig took hold of them with skill, exaggerated them; saw, so to speak, nothing but them; and declared that the results announced never could have been obtained. But in 1871 the fundamental experiment of Pasteur, on the life of yeast in a sweetened mineral medium, had become a trifle for him. He knew exactly how to form media deprived of all foreign germs, how to prepare pure yeast, and how to prevent the introduction of new germs, which could develop in the liquids and hinder the life of the yeast.

'Choose,' said he to Liebig, 'from the members of the Academy one or several, and ask them to decide between you and me. I am ready to prepare before you and before them, in a sweetened mineral medium, as much yeast as you can reasonably ask for, and with substances provided by yourself.'

Liebig's second objection had reference to acetic fermentation. The process of acetification known as that of 'beech shavings' is widely practised in Germany and even in France. It consists in causing alcohol diluted with water and with the addition of some _millièmes_ of acetic acid to trickle slowly into barrels or vats filled with shavings of beech, either massed together without order or disposed in layers after having been rolled up like the spring of a watch. Openings formed in the sides of the barrel, and in a double bottom upon which the shavings rest, permit the access of the air, which rises into the barrel as it would in a chimney, and yields all or part of its oxygen to the alcohol to convert it into acetic acid. All writers prior to Pasteur, and Liebig in particular, maintained that the shavings acted like porous bodies in the same manner as finely divided platinum. The acetic acid, they said, was formed by a direct oxidation, without any other influence than the porosity of the wood. This view of the subject was rendered plausible by the fact that in many manufactories the alcohol employed is that of distillation, which contains no albuminoid substances. Moreover, the duration of the shavings is in a sense indefinite.

According to Pasteur, the shavings perform only a passive part in the manufacture. They promote the division of the liquid and cause a considerable augmentation of the surface exposed to the air. They moreover serve as a support for the ferment, which is still, according to him, the mycoderma aceti, under the mucous form proper to it when submerged.

Certainly appearances were far from being favourable to this view. When the shavings of a barrel which has been in work for several months or even for several years are examined, they are found to be extraordinarily clean. It might be said that they had just been carefully washed. Pasteur has shown that this is but a deceptive appearance, and that in reality these shavings are partly or wholly covered with a mucous film of mycoderma aceti of excessive tenuity. It is necessary to scrape the surface of the wood with a scalpel and examine the scrapings with the microscope to be assured of the presence of this pellicle.

Liebig, who somewhere speaks, not without a certain contempt, of the microscope, denied formally the exactitude of these assertions.

'With diluted alcohol, which is used for the rapid manufacture of vinegar,' he wrote, 'the elements of nutrition of the mycoderm are excluded, and the vinegar is made without its intervention.' He asserted also in his memoir of 1869 that he had consulted the head of one of the principal manufactories of vinegar in Germany, that in this manufactory the diluted alcohol did not receive during the whole course of its transformation any foreign addition, and that beyond the air and the surfaces of wood and charcoal--for charcoal is sometimes associated with the beech shavings--nothing can act upon the alcohol. Liebig added that the director of the manufactory did not believe at all in the presence of the mycoderm, and that finally he, Liebig, in examining the shavings which had been used for twenty-five years in the manufactory, saw no trace of mycoderm on their surface.

The argument appeared conclusive. How, in fact, could we understand the production of a plant containing within itself nitrogen and mineral elements which was nevertheless to be nourished by water and alcohol.

'You do not take into account,' replied Pasteur, 'the nature of the water which serves to dilute your alcohol. This water, like all ordinary waters, even the purest, contains salts of ammonia and mineral matters which are capable of nourishing the plant. Finally, you have not rightly examined with the microscope the surface of the shavings, otherwise you would have seen the little particles of the mycoderma aceti united, in some cases, to a thin film which can even be lifted up. I propose to you, moreover, to send to the Academic Commission charged with the decision of the debate, some shavings that you have obtained yourself in the manufactory at Munich, and in the presence of its director. I will undertake to prove before the members of the commission the presence of the mycoderm on the surface of these shavings.'

Liebig did not accept this challenge. To-day the question is decided.

_THE QUESTION OF SPONTANEOUS GENERATION._

'All dry bodies,' said Aristotle, 'which become damp, and all damp bodies which are dried, engender animal life.' Bees, according to Virgil, are produced from the corrupted entrails of a young bull. At the time of Louis XIV. we were hardly more advanced. A celebrated alchemist doctor, Van Helmont, wrote: 'The smells which rise from the bottom of morasses produce frogs, slugs, leeches, grasses, and other things.' But most extraordinary of all was the true recipe given by Van Helmont for producing a pot of mice. It suffices to press a dirty shirt into the orifice of a vessel containing a little corn. After about twenty-one days, the ferment proceeding from the dirty shirt modified by the odour of the corn effects the transmutation of the wheat into mice. Van Helmont, who asserted that he had witnessed the fact, added with assurance:

'The mice are born full grown; there are both males and females. To reproduce the species it suffices to pair them.'

'Scoop out a hole,' said he again, 'in a brick, put into it some sweet basil, crushed, lay a second brick upon the first so that the hole may be perfectly covered. Expose the two bricks to the sun, and at the end of a few days the smell of the sweet basil, acting as a ferment, will change the herb into real scorpions. An Italian naturalist, Redi, was the first to subject this question of spontaneous generation to a more attentive examination. He showed that maggots in meat are not spontaneously generated, but that they are the larvæ of flies' eggs. To prevent the production of maggots, Redi showed that it was only necessary to surround the meat with fine gauze before exposing it to the air. As no flies could alight upon meat thus protected, there were no eggs deposited, and consequently neither larvæ nor maggots. But at the moment when the doctrine of spontaneous generation began to lose ground by the limitation of its domain, the discovery of the microscope brought to this doctrine new and formidable support. In presence of the world of animalculæ, the partisans of spontaneous generation raised a note of triumph. 'We may have been mistaken,' they said, 'as to the origin of mice and maggots, but is it possible to believe that microscopic organisms are not the outcome of spontaneous generation? How can we otherwise explain their presence and rapid multiplication in all dead animal or vegetable matter in process of decomposition?'

Buffon lent the authority of his name to the doctrine of spontaneous generation. He even devised a system to explain this hypothesis. In 1745 two ecclesiastics entered upon an eager controversy for and against this question. While the English Catholic priest Needham adopted the theory of spontaneous generation, the Italian priest Spallanzani energetically opposed it; but while in the eyes of the public the Italian remained master of the dispute, his success was more apparent than real, more in word than in deed.

The problem was again brought forward in a more emphatic manner in 1858. M. Pouchet, director of the Museum of Natural History at Rouen, in addressing the Academy of Sciences, declared that he had succeeded in demonstrating in a manner absolutely certain the existence of microscopic living organisms, which had come into the world without germs, and consequently without parents similar to themselves.

How came Pasteur to throw himself into this discussion, at first sight so far removed from his other occupations? The results of his researches on fermentation led him to it as a sort of duty. He was carried on by a series of logical deductions. Let us recall to mind, for example, the experiment in which Pasteur exposed to the heat of the sun water sweetened with sugar and mixed with phosphates of potash and magnesia, a little sulphate of ammonia, and some carbonate of lime. In these conditions the lactic fermentation was often seen to develop itself--that is to say, the sugar became lactic acid, which combined with the lime of the carbonate to form lactate of lime. This salt crystallises in long needles, and ends sometimes by filling the whole vase, while a little organised living thing is at the same time produced and multiplied. If the experiment is carried on further, another fermentation generally succeeds to this one. Moving vibrios make their appearance and multiply, the lactate of lime disappears, the fluidity returns to the mass, and the lactate finds itself replaced by butyrate of lime. What a succession of strange phenomena! How did life appear in this sweetened medium, composed originally of such simple elements, and apparently so far removed from all production of life? This lactic ferment, these butyric vibrios, whence do they come? Are they formed of themselves? or are they produced by germs? If the latter, whence do the germs come? The appearance of living organised ferments had become for Pasteur the all-important question, since in all fermentations he had observed a correlation between the chemical action set up and the presence of microscopic organisms. Prior to the establishment of the facts already mentioned, these difficulties did not exist. The theory of Liebig was universally accepted.

Thus the question as to the origin of microscopic organisms and the part played by them in fermentation was imposed as a necessity on Pasteur. He could not proceed further in his researches without having solved this question.

In the month of October, 1857, Pasteur was called to Paris. After having been made dean at an incredibly early age, he was now, at the age of thirty-five, entrusted with the scientific studies at the École Normale Supérieure. But if the position was flattering, it did not give to Pasteur what he most desired. As he had no Professor's chair, he had no laboratory. In those days science, and the higher education in science, were at a discount. It was the period when Claude Bernard lived in a small damp laboratory, when M. Berthelot, though known through his great labours, was still nothing more than an assistant in the Collège de France.

At the time here referred to, the Minister of Public Instruction said to Pasteur, 'There is no clause in the budget to grant you 1,500 francs a year to defray the expense of experiments.' Pasteur did not hesitate to establish a laboratory at his own expense in one of the garrets of the École Normale. We can readily imagine the modesty of such an establishment in such a place. Dividing his time between his professional duties and his laboratory experiments, Pasteur never went out but to talk over his daily researches with M. Biot, M. Dumas, M. de Senarmont, and M. Balard. M. Biot especially was his habitual confidant. The day when M. Biot learned that Pasteur proposed to study the obscure question of spontaneous generation, he strongly dissuaded him from entangling himself in this labyrinth. 'You will never escape from it,' said he, 'you will only lose your time;' and when Pasteur attempted some timid observations with the view of showing that in the order of his studies it was indispensable for him to attack this problem, M. Biot grew angry. Although endowed, as Sainte-Beuve has said, with all the qualities of curiosity, of subtlety, of penetration, of ingenious exactitude, of method, and of perspicuity, with all the qualities, in short, essential and secondary, M. Biot treated the project of Pasteur as a presumptuous adventure.

Bolder than M. Biot, but with a circumspection always alive, M. Dumas declared to Pasteur, without, however, further insisting upon the point, that he would not advise anyone to occupy himself too long with such a subject. M. de Senarmont alone took the part of Pasteur, and said to M. Biot:

'Let Pasteur alone. If there is nothing to be found in the path which he has entered upon, do not be alarmed, he will not continue in it. But,' added he, 'I should be surprised if he found nothing in it.'

M. Pouchet had previously stated the problem with precision:

'The opponents of spontaneous generation assert that the germs of microscopic organisms exist in the air, which transports them to a distance. What, then, will these opponents say if I succeed in inducing the generation of living organisms, while substituting artificial air for that of the atmosphere?'

Pouchet then devised this ingenious experiment. He filled a bottle with boiling water, hermetically sealed it with the greatest care, and plunged it upside down into a basin of mercury. When the water was quite cold he uncorked the bottle under the metal, and introduced into it half a litre of pure oxygen gas, which is as necessary to the life of the smallest microscopic organism as it is to that of the larger animals and vegetables. Up to this time there was nothing in the vessel but pure water and oxygen. Pouchet then introduced a minute bunch of hay which had been enclosed in a corked bottle, and exposed in a stove for a long time to a temperature of more than 100 degrees. At the end of eight days a mouldiness was developed in this infusion of hay. 'Where does this come from?' cried M. Pouchet triumphantly. Certainly not from the oxygen, which had been prepared from a chemical compound at the temperature of incandescence. The water had been equally deprived of germs, since at the boiling temperature all germs would have been destroyed. The hay also could not have contained them, for it had been taken from a stove heated to 100 degrees. As it was urged, however, that certain organisms could resist this temperature, M. Pouchet heated the hay from 200 to 300 degrees, or to any temperature that might be desired.

Pasteur came to disturb the triumph of M. Pouchet.