Memoir on alcoholic fermentation

1838 Charles Cagniard-Latour

Cagniard-Latour, C. 1838. Memoire sur la fermentation vineuse. Annales de chimie et de physique, Vol. 68, pages 206 222.

IT IS WELL KNOWN THAT WHEN FRESH beer yeast is mixed with a solution of sugar and added to a closed flask . . . and exposed to a temperature of 25¡ centigrade, very quickly, within a few minutes, the solution undergoes a fermentation which proceeds very rapidly if the amount of yeast added is considerable. However, when the solution does not contain the yeast and the sugar in the solution is pure, the same circumstances do not ensue, even after a very long time. It seemed important to examine under the microscope this material which had the property of causing the fermentation of sugar. This examination, which I have had the honour of reporting in a letter to the Academy, has led me to consider that the particles of which the yeast is composed are globular in form, and I have concluded that it is very probable that these particles are organised. (Footnote: Twenty-five years ago I first examined fresh yeast under the microscope. However, my instrument was very poor, and I concluded that the yeast was like a very fine sand composed of crystalloid particles. These observations were in error. The majority of the microscopic observations indicated in this memoir were performed on a microscope constructed by M. Georges Oberhauser. It enabled me to obtain enlargements of 300-400 times. For measuring the size of the globules, I used an ocular micrometer constructed by M. Charles Chevalier....)
The globules which I have observed are generally simple, transparent, spherical or slightly elongated, almost colourless. I have never seen them exhibit any movement which could be considered an indication of their independent action. On the other hand, the globules of yeast can appear in a liquid before the alcoholic fermentation has begun. But when the globular bodies, that is to say, the non-crystalline bodies, are beginning to produce themselves in a mucous liquid which, before the experiment has begun, does not reveal these globules, and when these bodies do not appear to have any power of locomotion, the microscopist will consider these bodies as simple plants. This is the way M. Turpin has viewed the protospheres which developed in a gelatinous medium, as I have reported to the Institute in a previous letter.
It is highly probable that the globules of yeast are organised, and they appear to belong to the vegetable kingdom. . .
But these plants, if one can give this name to such simple vesicles, are very small, and highly variable in size. The diameter of the largest globules does not exceed a hundredth of a millimeter. For the most part they are of such a size, that a cake of yeast of one cubic millimeter would contain at least a million globules.
On the assumption that the yeast globules had the faculty to reproduce themselves I performed a number of experiments to find out more about them....
My earlier observations (see the journal of the Institute, no. 185) had yielded the following principal results: ( l ) In order to liberate gas, the globules of yeast rise to the surface of the wort (of beer) and many of these globules are within the foam or scum which is produced during the fermentation, and can be readily observed with the microscope. ( 2 ) During their action on the wort, the globules diminish in size, and very probably produce seeds or reproductive bodies through this contraction, since one discovers very soon in the wort new globules, smaller in size. These globules, which are not observed at first, seem to have the ability to reproduce by budding or elongation, and because of this can form globules in chains.
which confirms my hypothesis that the yeast globules are organised and belong in the plant kingdom.
I examined samples removed hourly from the fermentation vessel. One hour after the addition of the yeast I already saw that the wort contained single globules each of which contained a small secondary globule attached to it. This small globule seemed to grow, since a little while later most of the pairs seemed to contain globules of the same size. The fourth sample revealed a very few paired globules. In order to assure myself that these pairs of globules were actually attached and not merely touching each other, I applied a force to the cover glass with a small needle while watching the globules under the microscope, and although this force produced a violent agitation of the globules, it did not affect the point of attachment of the pairs. It would appear that the individual globules become separated naturally as they grow to a certain age, since the commercial yeast generally consists of simple globules.... In the course of many fermentations which I have carried out with beer yeast, it has been possible to distinguish in certain (globules many granules, and sometimes a round or oval body, sometimes central and sometimes lateral, which one can presume to be a scar or umbilical mark which develops during the separation of the two globules....
The globules of ferment are able to develop very rapidly, because when one collects the total amount of yeast that is produced during the fermentation, one finds that this quantity weighs about seven times the initial quantity of yeast added, which agrees with the microscopic examination.
Considering the rapidity with which this increase in yeast is obtained, there is every reason to believe that this is the result of the reproduction of the globules of yeast which have been added. It appears that the globules reproduce in a liquid which contains the nutrients favourable for their growth. All brewers are aware that the beer wort ordinarily produces a larger amount of yeast than that added, but it has been supposed that this increase is due principally to a precipitation of the plant proteins which are present in the wort....
But while the wort is a medium in which reproduction of the ferment proceeds very readily, it is not the same as a simple solution of sugar, since an active yeast does not increase in weight in the latter, and actually loses its activity
M. Gay-Lussac has demonstrated in various experiments that oxygen has a great influence on the beginning of fermentation in certain liquids, especially in grape juice. But although oxygen is necessary for fermentation to begin, it is not necessary for fermentation to continue. Because of this fact and the knowledge that beer yeast is able to produce a fermentation in sugary materials without the presence of oxygen, M. Gay-Lussac has advanced the opinion that the power of fermentation rests in a large number of substances, but in a different state than in beer y east. The experiments to he presented below seem to indicate that this opinion is correct. I expressed the juice from a bunch of grapes while they were enclosed in a bell jar under an atmosphere of hydrogen and allowed this juice to remain under mercury for more than 15 hours. During this time I examined under the microscope a small amount of the deposit which separated from the juice and found it to be practically amorphous. But after I allowed a small amount of oxygen into the jar, which started the juice to ferment, I found a large deposit of yeast globules. One can surmise the  following from this: (1) The bodies of these small plants form a part of the material which deposited. (2) They had not germinated because they enclosed within the berries of the grapes. ( 3) This germination begins when they are exposed to oxygen and because of this beginning of development, they are able to act like the beer yeast. It has been reported to me that it is possible to cause the production of globules w hen egg white is diluted in water and heated, and that the globules of yeast arise from a nitrogenous material of animal or vegetable origin, which is present in the wort, and coagulates to form the globules. If this is so the yeast globules are no more living than those globules obtained during the coagulation of egg white by heat.
In order to clarify this point, I placed in a capsule in a bath heated to 90° centigrade, a mixture of 50 grams of water and one gram of egg white. When a portion of the albumin had been coagulated by the heat, I removed the capsule and after cooling, examined a small amount of the material which had formed at the surface of the liquid under the microscope. I found that this material contained a type of globule with a diameter averaging about a hundredth of a millimeter. But these globules resembled in general something crystalloid, and in none of them could be seen granules or an umbilical mark. It seems to me therefore, that the objection that the globules of ferment are analogous to those of coagulated egg white is not valid.
Furthermore, I have conducted a spontaneous fermentation of wort in a closed container, that is to say, a fermentation without the addition of yeast. This wort, even though it had been filtered, produced during its alcoholic fermentation a deposit of yeast. which upon examination under the microscope, proved to be composed of globules analogous to those in ordinary yeast. This fermentation took place much slower than those carried out in breweries, and one might expect, if the hypothesis that the globules are formed by a type of proteinaceous coagulation, that they would resemble those from coagulated egg white. But this is not the case. The globules in the deposit did not prove to be the same size as those in ordinary yeast, hut this does not argue against their
living nature, since the fermentation in this case had taken place over a very long time, and the globules may be quite different in age from those in ordinary yeast.
I carried out the same experiment in a flask which had been filled with carbon dioxide. Although the fermentation developed a little slower, the deposit obtained showed the same appearance under the microscope.
Since in these experiments the deposit formed showed the same appearance under the microscope as beer yeast, I have carried out a number of fermentations in closed vessels with various liquids, such as currant juice, grape juice, and prune juice, as well as in a solution of sugar with egg white. In all these cases the liquids were filtered before they were placed in their vessels. Examination of the deposits obtained has shown that in all cases they are composed to a large extent of globules which are analogous to those of beer yeast....
All who carry out fermentations on a large scale, such as brewers and distillers, know that in spite of the greatest care in their operations, their results are extremely variable. These irregularities are favourable to the hypothesis that the alcoholic fermentation is caused by bodies endowed with life, because who knows in how many different ways such similar bodies can be affected.... 


I am acquainted with the principal literature concerning alcoholic fermentations but I have seen no work in which the microscope was used to study the phenomena on which it depends. (Footnote: Leeuwenhoek, in 1680, examined beer yeast under the microscope and stated that it is composed of globules, but he thouht they were a part of the grain which was used to make the wort. But this observation did not suggest to this author the most important point, that these globules are capable of germinating and growing in the wort during the fermentation. )
The principal results of the present work are: ( I ) The beer yeast is a mass of small globules which are able to reproduce, and consequently are organised, and are not a simple organic or chemical substance, as has been supposed. (2) These bodies appear to belong to the plant kingdom. (3) They seem to cause a decomposition of sugar only when they are alive, and one can conclude that it is very probable that the production of carbon dioxide and the decomposition of sugar and its conversion to alcohol are effects of their growth.
I would also like to remark that the yeast considered as an organised body merits the attention of physiologists for the following reasons: ( I ) It can grow and develop under certain conditions very rapidly, even under an atmosphere of carbon dioxide. (2) Its manner of growth presents details of a type which has not been observed in other microscopic objects composed of isolated globules. (3) It does not die w hen frozen or in the absence of water.
In conclusion I would like to add that the old question of the cause of alcoholic fermentation, which was proposed by the Institute, appears to be solved from the present results and those I have reported in 18 3 5 and 1836. These results indicate the conclusion that in general ferments, or at least those which cause alcoholic fermentation in the manner of yeast, are composed of organised microscopic bodies which are very simple, and the substances which they induce to ferment are purely chemical substances, since they are sugar and compounds related to sugar.

Comment by Thomas Brock
This work was performed independently of Schwann's, and confirms his for the most part. The student should note that the real reason these two workers came up with the same observations at the same time was probably due to the production of better microscopes, as Cagniard-Latour mentions. Since the yeast cell is quite small in comparison to plant and animal cells, it would take at least 400 magnifications to be able to discern cellular details. It was this greater magnification which enabled Cagniard-Latour to differentiate yeast cells from globules of coagulated egg albumin.
The author's observations on some of the physiological properties of the yeast are interesting. The yeast cell's ability to grow anaeobically and to withstand freezing and desiccation arc well confirmed today. Many other microorganisms, including Some pathogenic ones, also possess these properties.
This paper and the preceding one by Schwann gave Henle support for his early germ theory of disease.