Louis Pasteur (1822–95). Scientific Papers.
The
Harvard Classics. 1909–14. |
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| The Physiological
Theory of Fermentation |
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| II. Fermentation in
Saccharine Fruits Immersed in Carbonic Acid Gas |
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| THE THEORY which we
have, step by step, evolved, on the subject of the cause of the
chemical phenomena of fermentation, may claim a character of
simplicity and generality that is well worthy of attention.
Fermentation is no longer one of those isolated and mysterious
phenomena which do not admit of explanation. It is the consequence
of a peculiar vital process of nutrition which occurs under certain
conditions, differing from those which characterize the life of all
ordinary beings, animal or vegetable, but by which the latter may be
affected, more or less, in a way which brings them, to some extent
within the class of ferments, properly so called. We can even
conceive that the fermentative character may belong to every
organized form, to every animal or vegetable cell, on the sole
condition that the chemico-vital acts of assimilation and excretion
must be capable of taking place in that cell for a brief period,
longer or shorter it may be, without necessity for recourse to
supplies of atmospheric oxygen; in other words, the cell must be
able to derive its needful heat from the decomposition of some body
which yields a surplus of heat in the process. |
1 |
| As a consequence of these conclusions it should be
an easy matter to show, in the majority of living beings, the
manifestation of the phenomena of fermentation; for there are,
probably, none in which all chemical action entirely disappears,
upon the sudden cessation of life. One day, when we were expressing
these views in our laboratory, in the presence of M. Dumas, who
seemed inclined to admit their truth, we added: “We should like to
make a wager that if we were to plunge a bunch of grapes into
carbonic acid gas, there would be immediately produced alcohol and
carbonic acid gas, in consequence of a renewed action starting in
the interior cells of the grapes, in such a way that these cells
would assume the functions of yeast cells. We will make the
experiment, and when you come to-morrow”—it was our good fortune to
have M. Dumas working in our laboratory at that time—“we will give
you an account of the result.” Our predictions were realized. We
then endeavoured to find, in the presence of M. Dumas, who assisted
us in our endeavour, cells of yeast in the grapes; but it was quite
impossible to discover any. 1 |
2 |
| Encouraged by this result, we undertook fresh
experiments on grapes, on a melon, on oranges, on plums, and on
rhubarb leaves, gathered in the garden of the École Normale,
and, in every case, our substance, when immersed in carbonic acid
gas, gave rise to the production of alcohol and carbonic acid. We
obtained the following surprising results from some prunes de
Monsieur: 2—On July
21, 1872, we placed twenty-four of these plums under a glass bell,
which we immediately filled with carbonic acid gas. The plums had
been gathered on the previous day. By the side of the bell we placed
other twenty-four plums, which were left there uncovered. Eight days
afterwards, in the course of which time there had been a
considerable evolution of carbonic acid from the bell, we withdrew
the plums and compared them with those which had been left exposed
to the air. The difference was striking, almost incredible. Whilst
the plums which had been surrounded with air (the experiments of
Bérard have long since taught us that, under this latter condition,
fruits absorb oxygen from the air and emit carbonic acid gas in
almost equal volume) had become very soft and watery and sweet, the
plums taken from under the jar had remained very firm and hard, the
flesh was by no means watery, but they had lost much sugar. Lastly,
when submitted to distillation, after crushing, they yielded 6.5
grammes (99.7 grains) of alcohol, more than 1 per cent. of the total
weight of the plums. What better proof than these facts could we
have of the existence of a considerable chemical action in the
interior of fruit, an action which derives the heat necessary for
its manifestation from the decomposition of the sugar present in the
cells? Moreover, and this circumstance is especially worthy of our
attention, in all these experiments we found that there was a
liberation of heat, of which the fruits and other organs were the
seat, as soon as they were plunged in the carbonic acid gas. This
heat is so considerable that it may at times be detected by the
hand, if the two sides of the bell, one of which is in contact with
the objects, are touched alternately. It also makes itself evident
in the formation of little drops on those parts of the bell which
are less directly exposed to the influence of the heat resulting
from the decomposition of the sugar of the cells. 3 |
3 |
| In short, fermentation is a very general phenomenon.
It is life without air, or life without free oxygen, or, more
generally still, it is the result of a chemical process accomplished
on a fermentable substance capable of producing heat by its
decomposition, in which process the entire heat used up is derived
from a part of the heat that the decomposition of the fermentable
substance sets free. The class of fermentations properly so called,
is, however, restricted by the small number of substances capable of
decomposing with the production of heat, and at the same time of
serving for the nourishment of lower forms of life, when deprived of
the presence and action of air. This, again, is a consequence of our
theory, which is well worthy of notice. |
4 |
| The facts that we have just mentioned in reference
to the formation of alcohol and carbonic acid in the substance of
ripe fruits, under special conditions, and apart from the action of
ferment, are already known to science. They were discovered in 1869
by M. Lechartier, formerly a pupil in the École Normale
Supérieure, and his coadjutor, M. Bellamy. 4 In 1821,
in a very remarkable work, especially when we consider the period
when it appeared, Bérard demonstrated several important propositions
in connection with the maturation of fruits: |
5 |
| I. All fruits, even those that are still green, and
likewise even those that are exposed to the sun, absorb oxygen and
set free an almost equal volume of carbonic acid gas. This is a
condition of their proper ripening. |
6 |
| II. Ripe fruits placed in a limited atmosphere,
after having absorbed all the oxygen and set free an almost equal
volume of carbonic acid, continue to emit that gas in notable
quantity, even when no bruise is to be seen—“as though by a kind of
fermentation,” as Bérard actually observes—and lose their saccharine
particles, a circumstance which causes the fruits to appear more
acid, although the actual weight of their acid may undergo no
augmentation whatever. |
7 |
| In this beautiful work, and in all subsequent ones
of which the ripening of fruits has been the subject, two facts of
great theoretical value have escaped the notice of the authors;
these are the two facts which Messrs. Lechartier and Bellamy pointed
out for the first time, namely, the production of alcohol and the
absence of cells of ferments. It is worthy of remark that these two
facts, as we have shown above, were actually fore-shadowed in the
theory of fermentation that we advocated as far back as 1861, and we
are happy to add that Messrs. Lechartier and Bellamy, who at first
had prudently drawn no theoretical conclusions from their work, now
entirely agree with the theory we have advanced. 5 Their
mode of reasoning is very different from that of the savants with
whom we discussed the subject before the Academy, on the occasion
when the communication which we addressed to the Academy in October,
1872, attracted attention once more to the remarkable observations
of Messrs. Lechartier and Bellamy. 6 M. Fremy,
in particular, was desirous of finding in these observations a
confirmation of his views on the subject of hemi-organism,
and a condemnation of ours, notwithstanding the fact that the
preceding explanations, and, more particularly our Note of 1861,
quoted word for word in the preceding section, furnish the most
conclusive evidence in favor of those ideas which we advocate.
Indeed, as far back as 1861 we pointed out very clearly that if we
could find plants able to live when deprived of air, in the presence
of sugar, they would bring about a fermentation of that substance,
in the same manner that yeast does. Such is the case with the fungi
already studied; such, too, is the case with the fruits employed in
the experiments of Messrs. Lechartier and Bellamy, and in our own
experiments, the results of which not only confirm those obtained by
these gentlemen, but even extend them, in so far as we have shown
that fruits, when surrounded with carbonic acid gas immediately
produce alcohol. When surrounded with air, they live in their
aërobian state and we have no fermentation; immersed immediately
afterwards in carbonic acid gas, they now assume their anaërobian
state, and at once begin to act upon the sugar in the manner of
ferments, and emit heat. As for seeing in these facts anything like
a confirmation of the theory of hemi-organism, imagined by M. Fremy,
the idea of such a thing is absurd. The following, for instance, is
the theory of the fermentation of the vintage, according to M.
Fremy. 7 |
8 |
| “To speak here of alcoholic fermentation alone,” 8 our
author says, “I hold that in the production of wine it is the juice
of the fruit itself that, in contact with air, produces grains of
ferment, by the transformation of the albuminous matter; Pasteur, on
the other hand, maintains that the fermentation is produced by germs
existing outside of the grapes.” |
9 |
| Now what bearing on this purely imaginary theory can
the fact have, that a whole fruit, immersed in carbonic acid gas,
immediately produces alcohol and carbonic acid? In the preceding
passage which we have borrowed from M. Fremy, an indispensable
condition of the transformation of the albuminous matter is the
contact with air and the crushing of the grapes. Here, however, we
are dealing with uninjured fruits in contact with carbonic acid
gas. Our theory, on the other hand, which, we may repeat, we
have advocated since 1861, maintains that all cells become
fermentative when their vital action is protracted in the absence of
air, which are precisely the conditions that hold in the experiments
on fruits immersed in carbonic acid gas. The vital energy is not
immediately suspended in their cells, and the latter are deprived of
air. Consequently, fermentation must result. Moreover, we may add,
if we destroy the fruit, or crush it before immersing it in the gas,
it no longer produces alcohol or fermentation of any kind, a
circumstance that may be attributed to the fact of the destruction
of vital action in the crushed fruit. On the other hand, in what way
ought this crushing to affect the hypothesis of hemi-organism? The
crushed fruit ought to act quite as well, or even better than that
which is uncrushed. In short, nothing can be more directly opposed
to the theory of the mode of manifestation of that hidden force to
which the name of hemi-organism has been given, than the discovery
of the production of these phenomena of fermentation in fruits
surrounded with carbonic acid gas; whilst the theory, which sees in
fermentation a consequence of vital energy in absence of air, finds
in these facts the strictest confirmation of an express prediction,
which from the first formed an integral part of its statement. |
10 |
| We should not be justified in devoting further time
to opinions which are not supported by any serious experiment.
Abroad, as well as in France, the theory of the transformation of
albuminous substances into organized ferments had been advocated
long before it had been taken up by M. Fremy. It no longer commands
the slightest credit, nor do any observers of note any longer give
it the least attention; it might even be said that it has become a
subject of ridicule. |
11 |
| An attempt has also been made to prove that we have
contradicted ourselves, inasmuch as in 1860 we published our opinion
that alcoholic fermentation can never occur without a simultaneous
occurrence of organization, development, and multiplication of
globules; or continued life, carried on from globules already
formed. 9 Nothing,
however, can be truer than that opinion, and at the present moment,
after fifteen years of study devoted to the subject since the
publication to which we have referred, we need no longer say, “we
think,” but instead, “we affirm,” that it is correct. It is, as a
matter of fact, to alcoholic fermentation, properly so called, that
the charge to which we have referred relates—to that fermentation
which yields, besides alcohol, carbonic acid, succinic acid,
glycerine, volatile acids, and other products. This fermentation
undoubtedly requires the presence of yeast-cells under the
conditions that we have named. Those who have contradicted us have
fallen into the error of supposing that the fermentation of fruits
is an ordinary alcoholic fermentation, identical with that produced
by beer yeast, and that, consequently, the cells of that yeast must,
according to our own theory, be always present. There is not the
least authority for such a supposition. When we come to exact
quantitative estimations—and these are to be found in the figures
supplied by Messrs. Lechartier and Bellamy—it will be seen that the
proportions of alcohol and carbonic acid gas produced in the
fermentation of fruits differ widely from those that we find in
alcoholic fermentations properly so called, as must necessarily be
the case since in the former the fermentation is effected by the
cells of a fruit, but in the latter by cells of ordinary alcoholic
ferment. Indeed we have a strong conviction that each fruit would be
found to give rise to special action, the chemical equation of which
would be different from that in the case of other fruits. As for the
circumstance that the cells of these fruits cause fermentation
without multiplying, this comes under the kind of activity which we
have already distinguished by the expression continuous life in
cells already formed. |
12 |
| We will conclude this section with a few remarks on
the subject of equations of fermentations, which have been suggested
to us principally in attempts to explain the results derived from
the fermentation of fruits immersed in carbonic acid gas. |
13 |
| Originally, when fermentations were put amongst the
class of decompositions by contact-action, it seemed probable, and,
in fact, was believed, that every fermentation has its own
well-defined equation which never varied. In the present day, on the
contrary, it must be borne in mind that the equation of a
fermentation varies essentially with the conditions under which that
fermentation is accomplished, and that a statement of this equation
is a problem no less complicated than that in the case of the
nutrition of a living being. To every fermentation may be assigned
an equation in a general sort of way, an equation, however, which,
in numerous points of detail, is liable to the thousand variations
connected with the phenomena of life. Moreover, there will be as
many distinct fermentations brought about by one ferment as there
are fermentable substances capable of supplying the carbon element
of the food of that same ferment, in the same way that the equation
of the nutrition of an animal will vary with the nature of the food
which it consumes. As regards fermentation producing alcohol, which
may be effected by several different ferments, there will be as in
the case of a given sugar, as many general equations as there are
ferments, whether they be ferment-cells properly so called, or cells
of the organs of living beings functioning as ferments. In the same
way the equation of nutrition varies in the case of different
animals nourished on the same food. And it is from the same reason
that ordinary wort produces such a variety of beers when treated
with the numerous alcoholic ferments which we have described. These
remarks are applicable to all ferments alike; for instance, butyric
ferment is capable of producing a host of distinct fermentations, in
consequence of its ability to derive the carbonaceous part of its
food from very different substances, from sugar, or lactic acid, or
glycerine, or mannite, and many others. |
14 |
| When we say that every fermentation has its own
peculiar ferment, it must be understood that we are speaking of the
fermentation considered as a whole, including all the accessory
products. We do not mean to imply that the ferment in question is
not capable of acting on some other fermentable substance and giving
rise to fermentation of a very different kind. Moreover, it is quite
erroneous to suppose that the presence of a single one of the
products of a fermentation implies the co-existence of a particular
ferment. If, for example, we find alcohol among the products of a
fermentation, or even alcohol and carbonic acid gas together, this
does not prove that the ferment must be an alcoholic ferment,
belonging to alcoholic fermentations, in the strict sense of the
term. Nor, again, does the mere presence of lactic acid necessarily
imply the presence of lactic ferment. As a matter of fact, different
fermentations may give rise to one or even several identical
products. We could not say with certainty, from a purely chemical
point of view, that we were dealing, for example, with an alcoholic
fermentation properly so called, and that the yeast of beer must be
present in it, if we had not first determined the presence of all
the numerous products of that particular fermentation under
conditions similar to those under which the fermentation in question
had occurred. In works on fermentation the reader will often find
those confusions against which we are now attempting to guard him.
It is precisely in consequence of not having had their attention
drawn to such observations that some have imagined that the
fermentation in fruits immersed in carbonic acid gas is in
contradiction to the assertion which we originally made in our
Memoir on alcoholic fermentation published in 1860, the exact words
of which we may here repeat:—“The chemical phenomena of fermentation
are related essentially to a vital activity, beginning and ending
with the latter; we believe that alcoholic fermentation never
occurs”—we were discussing the question of ordinary alcoholic
fermentation produced by the yeast of beer—“without the simultaneous
occurrence of organization, development, and multiplication of
globules, or continued life, carried on by means of the globules
already formed. The general results of the present Memoir seem to us
to be in direct opposition to the opinions of MM. Liebig and
Berzelius.” These conclusions, we repeat, are as true now as they
ever were, and are as applicable to the fermentation of fruits, of
which nothing was known in 1860, as they are to the fermentation
produced by the means of yeast. Only, in the case of fruits, it is
the cells of the parenchyma that function as ferment, by a
continuation of their activity in carbonic acid gas whilst in
the other case the ferment consists of cells of yeast. |
15 |
| There should be nothing very surprising in the fact
that fermentation can originate in fruits and form alcohol without
the presence of yeast, if the fermentation of fruits were not
confounded completely with alcoholic fermentation yielding the same
products and in the same proportions. It is through the misuse of
words that the fermentation of fruits has been termed alcoholic, in
a way which has misled many persons. 10 In this
fermentation, neither alcohol nor carbonic acid gas exists in those
proportions in which they are found in fermentation produced by
yeast; and, although we may determine in it the presence of succinic
acid, glycerine, and a small quantity of volatile acids 11 the
relative proportions of these substances will be different from what
they are in the case of alcoholic fermentation. |
16 |
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| Note 1. To determine the absence of
cells of ferment in fruits that have been immersed in carbonic acid
gas, we must first of all carefully raise the pellicle of the fruit,
taking care that the subjacent parenchyma does not touch the surface
of the pellicle, since the organized corpuscles existing on the
exterior of the fruit might introduce an error into our
microscopical observations. Experiments on grapes have given us an
explanation of a fact generally known, the cause of which, however,
had hitherto escaped our knowledge. We all know that the taste and
aroma of the vintage, that is, of the grapes stripped from the
bunches and thrown into tubs, where they get soaked in the juice
that issues from the wounded specimens, are very different from the
taste and aroma of an uninjured bunch. Now grapes that have been
immersed in an atmosphere of carbonic acid gas have exactly the
flavour and smell of the vintage; the reason is that, in the vintage
tub, the grapes are immediately surrounded by an atmosphere of
carbonic acid gas, and undergo, in consequence, the fermentation
peculiar to grapes that have been plunged in this gas. These facts
deserve to be studied from a practical point of view. It would be
interesting, for example, to learn what difference there would be in
the quality of two wines, the grapes of which, in the one case, had
been perfectly crushed, so as to cause as great a separation of the
cells of the parenchyma as possible; in the other case, left, for
the most part, whole, as in the case in the ordinary vintage. The
first wine would be deprived of those fixed and fragrant principles
produced by the fermentation of which we have just spoken, when the
grapes are immersed in carbonic acid gas. By such a comparison as
that which we suggest we should be able to form a priori
judgment on the merits of the new system, which has not been
carefully studied, although already widely adopted, of milled,
cylindrical crushers, for pressing the vintage. [back] |
| Note 2. We have sometimes found
small quantities of alcohol in fruits and other vegetable organs,
surrounded with ordinary air, but always in small proportion, and in
a manner which suggested its accidental character. It is easy to
understand how, in the thickness of certain fruits, certain parts of
those fruits might be deprived of air, under which circumstances
they would have been acting under conditions similar to those under
which fruits act when wholly immersed in carbonic acid gas.
Moreover, it would be useful to determine whether alcohol is not a
normal product of vegetation. [back] |
| Note 3. In these studies of plants
living immersed in carbonic acid gas, we have come across a fact
which corroborates those which we have already given in reference to
the facility with which lactic and viscous ferments, and, generally
speaking, those which we have termed the disease ferments of beer,
develop when deprived of air, and which shows, consequently, how
very marked their aërobian character is. If we immerse beet-roots or
turnips in carbonic acid gas, we produce well-defined fermentations
in those roots. Their whole surface readily permits the escape of
the highly acid liquids, and they become filled with lactic,
viscous, and other ferments. This shows us the great danger which
may result from the use of pits, in which the beet-roots are
preserved, when the air is not renewed, and that the original oxygen
is expelled by the vital processes of fungi or other deoxidizing
chemical actions. We have directed the attention of the
manufacturers of beet-root sugar to this point. [back] |
| Note 4. Lechartier and Bellamy,
Comptes rendus de l’Académie des Sciences, vol., lxix., pp.,
366 and 466, 1869. [back] |
| Note 5. Those gentlemen express
themselves thus: “In a note presented to the Academy in November,
1872, we published certain experiments which showed that carbonic
acid and alcohol may be produced in fruits kept in a closed vessel,
out of contact with atmospheric oxygen, without our being able to
discover alcoholic ferment in the interior of those fruits. “M.
Pasteur, as a logical deduction from the principle which he has
established in connection with the theory of fermentation, considers
that the formation of alcohol may be attributed to the fact that
the physical and chemical processes of life in the cells of fruit
continue under new conditions, in a manner similar to those of the
cells of ferment. Experiments, continued during 1872, 1873, and
1874, on different fruits have furnished results all of which seem
to us to harmonize with this proposition, and to establish it on a
firm basis of proof.”—Comptes rendus, vol. lxxix p. 949,
1874. [back] |
| Note 6. PASTEUR, Faites nouveaux pour servir à la
connaissance de la théorie des fermentations proprement dites.
(Comptes rendus de l’Académie des Sciences, vol. lxxv., p.
784.) See in the same volume the discussion that followed; also,
PASTEUR, Note sur la production de l’alcool
par les fruits, same volume, p. 1054, in which we recount the
observations anterior to our own, made by Messrs. Lechartier and
Bellamy in 1869. [back] |
| Note 7. Comptes rendus,
meeting of January 15th, 1872. [back] |
| Note 8. As a matter of fact, M.
Fremy applies his theory of hemi-organism, not only to the alcoholic
fermentation of grape juice, but to all other fermentations. The
following passage occurs in one of his notes (Comptes rendus de
l’Académie, vol. lxxv., p. 979, October 28th, 1872):
“Experiments on Germinated Barley.—The object of these was to show
that when barley, left to itself in sweetened water, produces in
succession alcoholic, lactic, butyric, and acetic fermentations,
these modifications are brought about by ferments which are produced
inside the grains themselves, and not by atmospheric germs. More
than forty different experiments were devoted to this part of my
work.” Need we add that this assertion is based on no substantial
foundation? The cells belonging to the grains of barley, or their
albuminous contents, never do produce cells of alcoholic ferment, or
of lactic ferment, or butyric vibrios. Whenever those ferments
appear, they may be traced to germs of those organisms, diffused
throughout the interior of the grains, or adhering to the exterior
surface, or existing in the water employed, or on the side of the
vessels used. There are many ways of demonstrating this, of which
the following is one: Since the results of our experiments have
shown that sweetened water, phosphates, and chalk very readily give
rise to lactic and butyric fermentations, what reason is there for
supposing that if we substitute grains of barley for chalk, the
lactic and butyric ferments will spring from those grains, in
consequence of a transformation of their cells and albuminous
substances? Surely there is no ground for maintaining that they are
produced by hemi-organism, since a medium composed of sugar, or
chalk, or phosphates of ammonia, potash, or magnesia contains no
albuminous substances. This is an indirect but irresistible argument
against the hemi-organism theory. [back] |
| Note 9. PASTEUR, Mémoire sur la fermentation
alcoolique, 1860; Annales de Chimie et de Physique. The
word globules is here used for cells. In our researches we have
always endeavoured to prevent any confusion of ideas. We stated at
the beginning of our Memoir of 1860 that: “We apply the term
alcoholic to that fermentation which sugar undergoes under the
influence of the ferment known as beer yeast.” This is, the
fermentation which produces wine and all alcoholic beverages. This,
too, is regarded as the type for a host of similar phenomena
designated, by general usage, under the generic name of
fermentation, and qualified by the name of one of the
essential products of the special phenomenon under observation.
Bearing in mind this fact in reference to the nomenclature that we
have adopted, it will be seen that the expression alcoholic
fermentation cannot be applied to every phenomenon of
fermentation in which alcohol is produced, inasmuch as there may be
a number of phenomena having this character in common. If we had not
at starting defined that particular one amongst the number of very
distinct phenomena, which, to the exclusion of the others, should
bear the name of alcoholic fermentation, we should inevitably have
given rise to a confusion of language that would soon pass from
words to ideas, and tend to introduce unnecessary complexity into
researches which are already, in themselves, sufficiently complex to
necessitate the adoption of scrupulous care to prevent their
becoming still more involved. It seems to us that any further doubt
as to the meaning of the words alcoholic fermentation, and
the sense in which they are employed, is impossible, inasmuch as
Lavoisier, Gay-Lussac, and Thénard have applied this term to the
fermentation of sugar by means of beer yeast. It would be both
dangerous and unprofitable to discard the example set by these
illustrious masters, to whom we are indebted for our earliest
knowledge of this subject. [back] |
| Note 10. See, for example, the
communications of MM. Colin and Poggiale, and the discussion on
them, in the Bulletin de l’Académie de Médecine, March 2d,
9th, and 30th, and February 16th and 23rd, 1875. [back] |
| Note 11. We have elsewhere
determined the formation of minute quantities of volatile acids in
alcoholic fermentation. M. Béchamp, who studied these, recognized
several belonging to the series of fatty acids, acetic acid, butyric
acid, &c. “The presence of succinic acid is not accidental, but
constant; if we put aside volatile acids that form in quantities
which we may call infinitely small, we may say that succinic acid is
the only normal acid of alcoholic fermentation.”—PASTEUR, Comptes rendus de l’Académie, vol.
xlvii., p. 224, 1858. [back] |
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