4. Lastly, one characteristic of “high” yeast, which it shares in common

with some other ferments, although not with all, and which, from a

practical point of view, deserves special mention, is that as

fermentation proceeds the yeast rises to the surface of the liquid.

Whilst the process of the manufacture of beer by this ferment is going

on, the yeast is seen to work out of the bung-holes, flowing over in

considerable quantity. The ferment named after the author, as well as

“low” yeast, does not possess this property: it remains at the bottom of

the vessels. When “high” fermentation takes place in vessels that are

not filled, the ferment forms a thick layer, a kind of cap on the

surface of the beer. This characteristic may be witnessed even in the

fermentation of very small quantities of liquid. In our flasks, in which

the volume of fermenting wort does not exceed 100 c.c. or 150 c.c.

(about 4 or 5 fluid ounces), we may perceive, as the violence of

fermentation subsides, and the head falls, the sides of the vessel

covered to a height of from 1 cm. to 2 cm. (about 3/4-in.) above the

surface of the liquid, with particles of yeasty matter, in little

masses, or in a thin film, raised to that height by the head, and left

behind when that fell.

“Low” Ferment.—Whilst high yeast performs its functions in the breweries

in which it is used at somewhat high temperatures—namely, between 16° C.

and 20° C. (60° F. to 68° F.)—“low” yeast is never employed at a higher

temperature than 10° C. (50° F.), and it is even thought preferable that

it should not be subjected to more than 6° C., 7° C., or 8° C. (43° F.

to 46° F.). At these comparatively low temperatures “high” yeast would

have no perceptible action, whereas it is at such temperatures that

“low” yeast best performs its functions.

In our Memoir on alcoholic fermentation, published in 1860, in the

_Annales de Chimie et de Physique_, the idea of the identity of the two

yeasts was accepted; but we had at that time made no special

observations of our own on the subject.

Upon closer investigation we are inclined to believe that the two yeasts

are quite distinct. We might keep our “high” yeast at the lowest

temperatures that it can bear, and repeat our growths under these

conditions; or, on the other hand, we might subject the “low” yeast to

temperatures higher than those at which it ordinarily grows, without

ever succeeding in changing the first into the second or the second into

the first, supposing, of course, that each of our yeasts was pure to

begin with. If they were intermixed the change in the conditions of

development would cause one or the other to preponderate, and incline us

to believe that a transformation had really occurred.

[Illustration: Fig. 41.]

It is true that brewers generally are of a different opinion. Most of

them assert that “low” yeast cultivated at a high temperature becomes

“high” yeast; and conversely, that “high” yeast becomes “low” by

repeated growths at a low temperature. Many have told us that they have

proved this. Nevertheless it is our belief that the success of such

transformation has been but apparent, attributable in each case to the

fact, as we have just stated, of their having operated on a mixture of

the two yeasts.

Mitscherlich, and various authors after him, have asserted that “high”

yeast propagates by budding, and “low” yeast, on the contrary, by

spores, formed by the endogenous division of the protoplasm of the

cells, and set free by the rupture of the cell-wall, which then,

increasing in size, assume the character of ordinary cells. But we have

never been able to confirm this.

Fig. 41 represents a field of low yeast, taken from the deposit in a vat

after the fermentation of the beer was finished. The granular matter

mixed with the cells is altogether amorphous although in many cases

perfectly spherical. It is a product in no way related to this yeast

(see Plate I., No. 7).[100] “High” yeast and all the ferments of beer

have this kind of deposit associated with them. There is no doubt that

confused observations as regards these minute bodies have been the cause

of the error which we had to deal with in connection with a particular

mode of reproduction of low yeast, as to which we have already fully

expressed our views (Chap. V. § 1, p. 146).

Comparing Fig. 41 with Fig. 40 (p. 189), it may be seen that the general

aspect of low yeast is distinguished, in its early stages, although in

no very decided manner, from that of “high” yeast, by being slightly

smaller and less round or spherical in its cells than the latter.[101]

These differences, however, would escape an unpractised eye.

As to the case of “high” yeast, the deposits of “low” yeast after

fermentation appear as scattered, isolated cells; we do not find more

than two or three per cent. of united cells. Nevertheless the two yeasts

present, as we shall see, quite marked differences in the character of

their budding and multiplication.

[Illustration: Fig. 42.]

On May 28th, 1875, we put a trace of pure, unicellular, “low” yeast,

taken at the end of a fermentation, into a flask of wort. On May 29th,

sixteen hours after impregnation, the temperature during the night

having been 15° C. (59° F.), we made a sketch of the yeast before its

development had become apparent to the naked eye. No perceptible

development, that is to say, no visible deposit at the bottom of the

liquid and formation of patches of froth on the surface, took place

before May 30th. A mere glance at Fig. 42 will be sufficient to enable

us to detect a considerable difference between it and Fig. 40, which

represents the multiplication of the cells of “high” yeast. The cells of

the “low” yeast are slightly smaller and rather more oval, as we have

already had occasion to notice, and the budding processes are

considerably less ramified, in consequence of which there is a

comparative absence of globular clusters which are so striking a feature

in the development of “high” yeast, when examined early enough.

Moreover, if we cause our “low” yeast to age, by leaving it for a longer

or shorter time in the beer which it has formed, or if we exhaust it in

sweetened water by leaving it for whole months in a volume of sweetened

water considerably larger than what it is capable of fermenting, and

then proceed to revive it and cause it to propagate in an aerated

saccharine wort capable of nourishing it, this yeast will resume its

original aspect, as sketched and described. At most we shall observe

certain minute differences in the size of the cells in successive

growths. A very remarkable industrial characteristic of this yeast is

the fact that it never rises to the surface, no matter at what

temperature it may be working, whether between 6° C. and 8° C. or 15° C.

and 20° C.;[102] in other words, it is not buoyed up by the carbonic

acid gas when the fermentation is at its height. At the end of the

fermentation, the surface of the liquid and the sides of the vessel

above the level of the liquid are clean and not covered with the yeast,

which remains altogether at the bottom of the fermented liquid.

Moreover, the weight of new yeast which it yields is always less than

that yielded by “high” yeast, for the same quantity of fermentable

liquid, although greater than that which _saccharomyces pastorianus_

would give. Lastly, the beer possesses a flavour and delicacy which

cause it to be held in higher esteem by consumers than beers produced by

means of other ferments.[103]

§ IV.—On the Existence and Production of Other Species of Ferment.

Our present knowledge of the alcoholic ferments embraces the following,

without taking into account the ferment-form of _mucor_:—

The ferment named after the author, which is found associated with the

ferments of the grape and other domestic fruits, and with spontaneous

ferments in general.

The ferment of “high” beer.

The ferment of “low” beer.

To these must be added the ordinary ferment of wine, and that called

_apiculatus_, although, indeed, these last are of little practical

importance, since, in general, they soon become lost amongst others of

greater vitality, in the spontaneous fermentation of fruits. These are

not the only alcoholic ferments; a study of the germ-cells diffused over

the surface of fruits, grains, and stalks of all vegetables in different

countries, would doubtless lead to the discovery of many new ones. We

are even inclined to believe that one ferment might give rise to a

multitude of others. The investigations which we have undertaken in this

direction are as yet not far advanced; we may, however, be allowed

merely to state the principle which governs them. A ferment is a

combination of cells, the individuals of which must differ more or less

from each other. Each of these cells has certain generic and specific

peculiarities which it shares with the neighbouring cells; but over and

above this, certain peculiar characteristics which distinguish it, and

which it is capable of transmitting to succeeding generations. If,

therefore, we could manage with some species of ferment to isolate the

different cells that compose it, and could cultivate each of these

separately, we should obtain as many specimens of ferments, which would,

probably, be distinct from one another, inasmuch as each of them would

inherit the individual peculiarities of the cell from which it

originated. Our endeavours are directed to realizing this result

practically, by first thoroughly drying a ferment and reducing it to

fine powder. We have seen (Chap. III. § 6) that this mode of experiment

is practicable, that in a powder composed of yeast and plaster the

ferment preserves its faculty of reproduction for a very long time. If

we now drop a small quantity of this powder from a sufficient height,

and then, at a certain distance below the cloud of dust so formed, open

several flasks previously deprived of air and containing a fermentable

liquid that has been boiled, immediately closing them all up again,

under such circumstances it is conceivable that some of the cells of

yeast diffused in the cloud of dust, and separated widely in the act of

falling, will enter some of our flasks singly, and there develop an

appreciable weight of ferment, all the cells of which will have sprung

from the same mother-cell. We have proved that flasks may be easily

impregnated under these conditions, and our preliminary observations,

although incomplete, seem to favour the idea that numerous varieties of

ferment are to be obtained by these means.

Spontaneous ferments, properly so called, of which we have already

spoken, are, after all, the result of sowings of this kind. Originating

in liquids which have been boiled, and then left to themselves in

contact with the air in a place where cells or germs must have existed,

these ferments must necessarily often spring from single germs or from a

limited few, and this also would probably be a means of developing

distinct varieties of ferments.

Without dwelling longer on the practical consequences likely to result

from the ideas which we have just expressed, we shall proceed to

describe two new alcoholic ferments, which differ widely from those

already mentioned.

_New “High” Ferment._—We met with this ferment accidentally, under the

following circumstances:—On February 12th, 1873, we had brewed in the

laboratory about 2-½ hectolitres (rather over 50 gallons) of wort, 10

litres (about two gallons) of which were set aside to cool in a

white-iron trough, and left during the night exposed to free contact

with air in the underground part of the laboratory, where we have a

small experimental brewery. Next day we put some of this latter into a

bottle; the wort soon began to show evidence of change, various

productions made their appearance on the surface of the liquid, and a

deposit of yeast settled at the bottom. On May 23rd, perceiving bubbles

of gas and a steady fermentation set up in the wort, which remained all

the time corked up, and fearing that the bottle might burst by the

increasing internal pressure, we drew the cork. A considerable

liberation of gas at once took place, accompanied by a voluminous foam

which half emptied the bottle. A microscopical examination of the

deposit from the disturbed liquid led to the discovery of a very

homogeneous yeast, associated with various other organisms; it was

clearly a yeast which we had not hitherto met with amongst the

spontaneous ferments which we had had occasion to study. Thinking that

this might be a new species of ferment which would probably produce a

beer that was also unknown, we set to work to purify it by cultivation

in flasks of pure wort, during the months of May, June, and July. Our

last growths, of August 4th, 1873, were preserved, in order that we

might assure ourselves of the purity of the beer, and, consequently, of

the ferment. On November 15th its purity was established. On that date

we made some beer with this ferment, which had now been left to itself

for several months in contact with pure air. The beer which we obtained

resembled no known variety; consequently the ferment must itself have

been a distinct one, differing from others, especially those which we

have been considering in this chapter.

[Illustration: Fig. 43.]

Fig. 43 represents the rejuvenescence of this ferment. Comparing this

figure with Fig. 42, we see that this ferment presents a considerable

resemblance to “low” yeast in dimensions, method of budding, and oval

shape; but the feature which distinguishes it essentially from “low”

yeast is that it rises to the surface, like “high” yeast. Buoyed up by

the gas during fermentation, it forms a layer of yeast on the surface of

the fermenting liquid, where it remains after the head has fallen. Some

of this head of yeast likewise adheres to the sides of the vessel above

the level of the liquid.

In short, by the greater regularity of its forms and the uniform

dimensions of its cells, this ferment is to be easily distinguished from

_saccharomyces pastorianus_; its aspect, which is oval instead of

spherical, and the ramified form of its chains of cells, which is less

marked than in the case of “high” yeast, also prevent our confounding it

with the latter ferment; in its rising character it differs absolutely

from “low” yeast; lastly, it may be distinguished from all other

ferments by the flavour of the beer that it produces.

The ferment which we discovered in this accidental way may be utilized.

Indeed, we may ask, is it not to be found already in our beer? We are

inclined to believe that it is. After the war of 1870, some Viennese

traders established at Maisons-Alfort, near Paris, a manufactory of

yeast for bakers. They saccharified by means of malt a mixture of the

meals of rye, maize, and barley, which they then caused to ferment. One

day we had occasion to study the yeast produced in this establishment,

and although we did not submit it to a sufficient number of consecutive

experiments to enable us to speak positively, we are under the

impression that the yeast produced at Maisons-Alfort is a “high” one,

differing from what may be properly termed the “high” yeast of breweries

in which “high” fermentation is practised, but presenting a great

resemblance to the “high” yeast of which we have been speaking. It would

be interesting to confirm the opinion of their possible identity by

fresh studies, and the best way of doing this would be to compare the

qualities of beer which the two yeasts could produce.

_Caseous Ferment._—We give the title caseous for a reason that will

presently appear, to a ferment which we came across also accidentally.

We were trying different methods of purifying yeasts, and for this

purpose had composed a liquid formed of:

Ordinary wort 150 c.c.[104]

Water saturated with bi-tartrate of potash 50 c.c.

Alcohol of 90° 25 c.c.

Quantities of this liquid were placed in several of our double-necked

flasks, submitted to boiling, then, after cooling, impregnated with

different ferments, and kept in a water-bath at 50° C. (122° F.) for one

hour.

In operating under these conditions with brewers’ “high” yeast, say, for

instance, with what is called Dutch yeast, a kind well known in

distilleries, fermentation shows itself in the course of a few days, in

spite of the increased temperature to which our liquid, which is hopped

and slightly acid and alcoholic, has been subjected. The time required

for the resumption of fermentation depends both upon the degree of

temperature to which the yeast has been exposed and upon the duration of

its exposure. These, however, are not the points upon which we now wish

to dwell. It is of greater importance to notice that the new yeast has

none of the characteristics of “high” ferment, of which Dutch yeast

seems to be exclusively composed, if we do not take into account

impurities which cannot be avoided in a commercial product of this

nature. Other specimens of Dutch yeasts would give the same results.

[Illustration: Fig. 44.]

[Illustration: Fig. 45.]

Figs. 44 and 45 represent this new ferment magnified to the same extent

as the other ferments have generally been, that is 400/1; it will

readily be seen how different its form is from that of “high” yeast, how

far it is from having the spherical aspect and mode of budding

characteristic of that ferment. In Fig. 45 the ferment is represented in

a mass; in Fig. 44 we see the ramified groups, the cells and segments of

which form, after separation, the yeast of the deposit. It thus appears

to be composed of jointed branches of greater or less length, which, at

the junctions of the segments, put forth similar cells or segments of a

round, oval, pyriform, cylindrical, or other shape; in all its

characters recalling the description of _dematium_. Moreover, the cells

and segments exhibit a greater sharpness of outline, as well as a more

marked transparency and refractive power than are found in the majority

of ferments; but the most curious physical characteristic of this

ferment is its plasticity and elasticity, if we may use those terms. It

can only be made to diffuse through water with great difficulty; when

shaken up in it, it sinks to the bottom quickly as a clotted sediment,

and the supernatant liquid appears scarcely at all charged with globules

in suspension. Again, when placed on a microscope slide and compressed

by the cover-glass, it returns to its original form on removal of the

pressure. It is from these considerations that we have given to it the

name of _caseous ferment_.[105] Lastly, this ferment produces a beer of

a peculiar kind, which cannot be confounded with other kinds of beer

known in the present day. We should add that it preserves its

characteristics in repeated growths, and that we have never found it

reproduce ordinary “high” yeast.

When caseous ferment is sown in a saccharine medium charged with mineral

salts, its aspect, form, and mode of budding differ completely from what

they are when the ferment exists in a natural medium, such as wort or

other liquid adapted to the nutrition and life of ferments.

[Illustration: Fig. 46.]

Fig. 46 represents this ferment in course of development, forty-eight

hours after it had been sown in a saline medium (we employed Raulin’s

fluid, substituting bi-tartrate for the nitrate of ammonia). It will be

seen how different its aspect is from that of the preceding figures; it

is still capable, however, of resuming the forms of the latter if

cultivated afresh in natural saccharine worts.

“High” yeast from a “high” fermentation brewery in the Ardennes, after

having been exposed to heat under the conditions given above, likewise

produced caseous ferment, without a trace of “high” ferment, just as

happened in the case of the Dutch yeast. All the “high” yeasts used in

brewing seem to behave in the same manner.

What conclusion are we to draw from these facts? Apparently that “high”

yeast is modified by heat in an acid and alcoholic medium, giving rise

to caseous ferment. On the other hand, it might be conceived that the

“high” yeasts on which we experimented were not pure, but contained, in

a state of intermixture, some caseous ferment, and that by the

application of a temperature of 50° C. (122° F.) to our alcoholic

medium, the high ferment was all killed and the caseous ferment alone

survived. It is a remarkable fact that this latter hypothesis,

improbable as it seems, inasmuch as the microscope revealed no

intermixture of ferments, seems, nevertheless, to be a true one. As a

matter of fact, if we subject to a temperature of 50° C. for one hour in

the medium in which it acts, not the “high” yeast of commerce but “high”

yeast that is _absolutely pure_, this will perish utterly, and the wort

after cooling may remain for years in an oven without either undergoing

fermentation or developing any growth whatever of “high” ferment or

“caseous” ferment.

On the other hand, if we impregnate this same alcoholic liquid with some

of the caseous ferment and then heat the vessel to 50° C. for one hour,

the caseous ferment will go on reproducing itself after the liquid has

cooled down.[106]

It seems, therefore, impossible to admit that caseous ferment results

from a modification of “high” ferment, and we are led to believe that in

the preceding experiments it must have been the progeny of cells of

caseous ferment present in the “high” yeasts of commerce, which cells,

probably in consequence of their scarcity, the microscope was unable to

reveal, but which, nevertheless, did exist, and went on reproducing

themselves alone after the heating.

[Illustration: Fig. 47.]

This conclusion is supported by the following fact, which also tends to

prove that in the case of the “high” English _pale ales_, caseous

ferment plays a most important part. In the medium already described, we

sowed the deposit from a bottle of good English pale ale. After having

been heated the yeast went on growing, and we obtained the very

beautiful specimen of caseous ferment represented in Fig. 47. The two

dark globules are dead cells which had been killed. Two minute segments

of lactic ferment are also visible in the sketch—the yeast which we

sowed was, of course, impure—and their presence proves, we may observe,

by the way, that lactic ferment also can withstand a temperature of 50°

C. (122° F.) in the medium which we here employed. The yeast as sowed is

represented in Fig. 48; it reminds us forcibly of certain forms of the

caseous ferment. Amongst the globules, which for the most part were

transparent and very young, there were some which appeared aged and of a

yellowish colour and granular. These latter probably belonged to the

yeast of manufacture. Their shape distinguishes them from “high” yeast,

properly so called, as on the other hand it causes them to appear more

like cells of a recent growth to which, there is no doubt, beer, after

it is put in bottle, owes its effervescence and head. These various

circumstances incline us to believe that the caseous ferment forms part

of certain commercial yeasts, especially those used in the celebrated

breweries of Bass and Allsopp, at Burton-on-Trent, in the manufacture of

pale ale. Caseous yeast is, moreover, a “high” ferment, that is to say,

it rises to the surface.

[Illustration: Fig. 48.]

§ V.—On a New Race of Alcoholic Ferments: Aërobian Ferments.

Mention has already been made of certain researches which we undertook

with the object of ascertaining whether _mycoderma vini_, or

efflorescence of wine, and _mycoderma cerevisiæ_, or efflorescence of

beer, which grow equally well in all fermented liquids, have the power

of becoming transformed into actual alcoholic ferment. The result of

those researches was stated to be that these mycodermata do not become

transformed into ferment, properly so called, and that whenever any such

transformation has been supposed to have taken place, the ferment

produced was derived from germs introduced by the air or by the utensils

employed. What we did ascertain of the ferment-producing power of

_mycoderma vini_, was merely that this plant, when submerged, is capable

of causing sugar to ferment, in consequence of a certain continuous life

possible to its cells, apart from the oxidations resulting from the

presence of free oxygen, but without any generation of new cells taking

place.

Whilst engaged in these researches, we were pursuing others in relation

to the converse of the proposition just discussed, that is to say,

respecting the possibility of ferment becoming transformed into

_mycoderma vini_ or _mycoderma cerevisiae_. Our experiments in

connection with this subject chiefly consisted in various endeavours by

way of exhausting the yeast and subsequent revival of its growth. This

exhaustion was effected by growing the yeast in excess of sweetened

water, and at other times in unsweetened yeast-water, our efforts being

directed to deprive it of all power of fermenting. We afterwards caused

it to develop afresh in highly aerated, nutritious liquids, in order

that we might see how it reproduced itself, and if its new form were

that of a mycoderma. The yeast after having lost its power as a ferment,

and being no longer able to act in pure sweetened water, nevertheless

reproduced itself when placed in fermentable media, holding in solution

materials adapted to its nutrition; yet we never succeeded in obtaining

any organism besides the ferment, and, indeed, the identical variety of

ferment on which we had operated. In no case was _mycoderma vini_ or

_cerevisiæ_ produced, and we concluded that we were justified in stating

that whenever the _mycoderma vini_ appeared on the surface of a

fermented or fermentable liquid, its germ must have been introduced by

the surrounding air, or have previously existed in the liquid, and that

the reason why this germ multiplied so abundantly was because the liquid

in question had been peculiarly adapted to the vitality of the plant.

In a laboratory where alcoholic fermentations are studied, these germs

of _mycoderma vini_ exist in great abundance on the surfaces of

different objects. This fact admits of easy proof; we have merely to

open in such a laboratory some flasks containing yeast-water deprived of

air, or yeast-water sweetened, or any natural saccharine medium, or any

fermented liquid, which till the moment when our flasks were closed had

been kept boiling (Chap. IV.); it would be a very rare thing, indeed, if

_mycoderma vini_ did not develop in most of these flasks after the air

was readmitted, especially if, shortly before this operation, the dust

lying on the surface of the tables or floor of the laboratory had been

stirred up by dusting or sweeping.

This series of experiments, the salient points of which we have just

given, conducted with a view to ascertain whether yeast could be

transformed into mycoderma, has led the way to certain results of

special interest, results which concern all alcoholic ferments, and

which in all probability will be found in the long run to apply to all

aërobian ferments.

It being necessary for the conduct of our experiments to preserve our

yeast in a state of purity for an indefinite period, often for a great

length of time, in contact with pure air, we discovered that yeast was

possessed of extraordinary vitality, and that it rarely perished

completely throughout, inasmuch as we could almost invariably cause it

to revive by bringing it into contact with fresh, fermentable liquid.

This revival of the yeast—and it is to this point that we are most

anxious to direct the attention of our readers—is effected from two

distinct sources:—

1. By those cells of yeast which have not perished.

2. By cells of new formation.

We may give an example to explain this more clearly. In one of our

two-necked flasks we cause some pure wort to ferment by employing yeast

also in a state of purity. Fermentation completed, we leave the liquid

to itself, not touching the flask again. The fermented liquor covers a

deposit of yeast, apparently inert, and no trace of _mycoderma vini_

makes its appearance on the surface of the liquid. Let us suppose that

we go on daily for a considerable time introducing a little of the yeast

from this flask to a different flask of wort: the fresh flasks will

begin to ferment. The only appreciable difference which these successive

flasks will present, their impregnation having been effected at

intervals of twenty-four hours, will be that, _ceteris paribus_,

fermentation in them will be more and more slow in making its

appearance. This difference, as we have already explained, will be due

to the fact that the yeast in the first flask will, in the course of

time, undergo, in each of its cells, a process which we cannot better

describe than as a progressive _senescence_. The cells gradually become

filled with amorphous granulations, their interior becomes yellow, and

the protoplasm collects, either at the centre or near the borders; in

short, the vitality of the yeast becomes feeble. When, however, it is

taken out of the liquid in which it has fermented and introduced into a

fresh saccharine wort, it gradually resumes its transparency, and then

begins to germinate. These effects are the less rapidly brought about

the longer the cells remain exhausting themselves in the first fermented

liquid. They might be left in that liquid for such a length of time that

they would eventually perish, a fact which would manifest itself in

their absolute sterility and quiescence when sown in a fresh medium. In

general, however, matters are not carried far enough for this to take

place, and the yeast, preserved in a state of purity in its fermented

liquid, retains the capacity of revival, which may then go on

indefinitely. As a matter of observation, the cells of yeast, after

causing the liquid to ferment, instead of remaining inactive, and so by

living at their own expense gradually passing into a state of

exhaustion, begin to bud again; at least this is true of many of them.

Multiplying afresh in the fermented medium, under the influence of the

air, they form a kind of mycodermic film on the liquid surface, or a

ring round the sides of the flask, on a level with the liquid. This

development might often be mistaken for _mycoderma vini_ or _cerevisiæ_;

in reality there is not a single cell of _mycoderma_ formed. If we sow a

trace of the new growth in a saccharine medium it will behave exactly as

yeast would, budding and multiplying, and setting up fermentation in the

liquid. And thus, in spite of its mycodermic aspect, this growth is

nothing but yeast, since it gives rise to true alcoholic fermentation;

but it is a kind of yeast which, under the foregoing conditions, lives

after the manner of fungoid growths, absorbing the oxygen of the air and

emitting carbonic acid gas. It appears on the surface of all fermented

liquids, especially those which, like beer, contain carbohydrates, and

its quantity is the greater, and its action the more rapid, in

proportion as it has more perfect access to the air. We have termed this

yeast _aërobian ferment_ or _fungoid ferment_.

It may easily be understood how this kind of production has escaped

notice up to the present time. The conditions of our experiment were, in

many respects, novel; a saccharine liquid had never before been caused

to ferment by means of pure yeast, absolutely free from foreign germs; a

fermented liquid had not previously been exposed to contact with pure

air for an indefinite time. On the other hand, all ordinary fermented

liquids, when left to themselves in contact with air, are a ready prey

to _mycoderma vini_ or _aceti_ at their surface, and then give rise to

true fungoid growths. The appearance of these organisms, which always

takes place soon, has thus constantly concealed or prevented the

development of the true aërobian ferments. In repeating the experiment

described any alcoholic ferment may be used, and each one will be found

to produce its own peculiar fungoid form of ferment. Another point

worthy of notice is that these aërobian ferments, when they put forth

buds in the act of fermentation, reproduce the forms of the original

ferment, at least apparently so. In this respect they cannot be

distinguished, notwithstanding the fact, surprising as it seems, that

the two kinds of ferments are not identical. If we operate on a “low”

yeast its aërobian ferment will differ physiologically from the ferment

from which it sprung, presenting various special peculiarities which are

not to be found in the original “low” yeast. In most of our experiments

we have found the new aërobian ferment to be similar in its action to

“high” yeast, rising to the surface, and producing a beer which

possesses a greater fragrance than beer brewed with the identical “low”

yeast from which it was derived. Lastly, the properties of an aërobious

ferment are not peculiar to first growth, but are hereditary; by

repeating the growth of the first aërobian ferment we do not cause them

to disappear, we find them again in succeeding generations.

Notwithstanding these facts, it would be difficult to discover any very

appreciable differences between the forms of the cells of any particular

yeast and those of its aërobian ferment in course of development. So

true is this, that the aërobian ferment of _saccharomyces pastorianus_

might even be caused to put on the forms of _dematium pullulans_, which

we have had occasion to observe specially characterize this ferment

after the cells have been subjected to a prolonged process of

senescence.[107] This is evident from the following example, which will

once again show the remarkable extent to which the forms of a particular

organism may be varied by changes in composition of the nutritive

medium:—

On August 6th, 1873, we took some of the ferment _saccharomyces

pastorianus_ from a flask of wort that had undergone fermentation, and

sowed a scarcely perceptible quantity of it in another flask containing

a saline medium, composed as follows:—

Water containing about 10 per cent. of 150 c.c. (5-1/4 fl.

sugar-candy oz.)

Ash of yeast 0·5 gramme (8 grs.)

Ammonic bitartrate 0·2 “ (3 grs.)

Ammonic sulphate 0·2 ” (3 grs.)

In the course of the following days the ferment began to develop,

although with difficulty, the fermentation revealing itself by

collections of bubbles appearing here and there on the surface of the

liquid. We left the flask undisturbed till the 25th of November

following. On that day we found a very white deposit of ferment covering

the yeast-ash that had not been taken into solution, and a ring of

aërobian ferment on a level with the surface of liquid; all the sugar

had disappeared; the liquid contained 5·2 per cent. of alcohol, by

volume, at a temperature of 15° C. (59° F.); and, lastly, in consequence

of the purity of the materials employed, there was no trace of the

formation of fungoid growths, whether of _mycoderma vini_ or of

_mycoderma cerevisiæ_, on the surface of the liquid, or of vibrios or

lactic-ferment below the surface.

Thus then we see—and several other examples throughout this work confirm

the fact—that saccharine liquids holding mineral salts in solution are

as capable of complete fermentation as any media of natural composition.

It is true that ferment develops slowly and with difficulty in them, and

at times takes on rather curious forms, but, nevertheless, it does

develop in the media and carry on a fermentation in which not the

minutest particle of sugar is left undecomposed. This is true, at least,

in the case of _saccharomyces pastorianus_, but there are other ferments

which in such media are checked in their multiplication and in their

continued action on sugar. One condition indispensable to the

accomplishment of fermentation in such a sweetened mineral medium, by

means of _saccharomyces pastorianus_, is the absolute purity of the

materials and of the ferment. It is necessary that the life and

physiological action of the latter should be in no way interfered with

by the presence of other microscopic organisms. We shall have occasion

to revert to this important detail in connection with our growths.

[Illustration: Fig. 49.]

Fig. 49 represents the ferment as it appeared when examined on August

11th, 1873. We can no longer recognize in it any _saccharomyces

pastorianus_. The general appearance is spherical, and there are a

number of clusters of budding cells which remind one at first sight of

the mode of germination of brewers’ “high” yeast. At _a_, _a_, _a_, we

see globules from which irregular abortive filaments have sprung, a

proof of difficult germination. No such monstrosities could ever have

occurred if we had used beer-wort or must as our nutritive medium.

On November 25th we made another examination and sketch of the ferment,

the appearance of which did not differ materially from that given above.

The general appearance was the same, consisting mostly of globules

joined together in clusters of two or three or more. No separation, such

as occurs in the case of ferment formed in natural worts, had taken

place. The ferment, moreover, was very irregular, and comprised cells of

all sizes. We sowed some of it in a flask of pure wort. On November 26th

there was no apparent development: on November 27th, however, not more

than forty-eight hours after impregnation, there was a considerable

deposit of white ferment at the bottom of the liquid, and fermentation

was so active that the surface of the liquid was covered with an

abundant froth. This shows us the wonderful vitality and recuperative

power possessed by germs which, left to themselves for about four

months, revived so readily. It proves too that the reviving influences

took effect on some aërobian ferment. From the mode of life of this

latter being similar to that of a surface fungoid growth, it does not

become exhausted as the cells of ordinary ferment do. Now the cells

which, sown on August 6th, had become exhausted by prolonged stay in the

mineral liquid, and were almost inert, would have required several days

for their revival; but in the experiment described the revival was

rapid, and this rapidity proves, as we have said, that the revival must

have taken place in cells of aërobian ferment.

[Illustration: Fig. 50.]

Taking some fresh yeast from the bottom of the liquid we examined and

made a sketch of it (Fig. 50). The field was filled with round and oval

cells, jointed and ramified filaments, budding and multiplying in the

most remarkable manner, reminding us of the germination of the cells of

yeast exhausted in sweetened water, and also of the germination in the

form of _dematium pullulans_ of certain germ-cells which are spread over

the surface of sweet, domestic fruits. We could never grow tired, as we

wrote it in our original notes, of sketching this beautiful plant, which

establishes very clearly a transition between one of the best defined

cellular ferments, viz., _saccharomyces pastorianus_, and certain forms

of very common fungoid growths, those of _dematium_, and even of the

most common mould, _mucor mucedo_ or _racemosus_, when it vegetates

beneath the surface of a liquid and acts as a ferment.[108] We have

here, as in these cases, filamentous chains branching into other similar

chains, composed of more or less elongated cells, which at length fall

off and germinate exactly as the conidia-bearing _hyphae_ of _mucor_ do.

The aërobian ferment of “high” yeast, in whatever medium we cultivated

it, presented no peculiarity, as far as its forms were concerned. It was

composed of cells of spherical shape, like ordinary “high” yeast, and

germinated in the same way as the latter.

[Illustration: Fig. 51.]

Fig. 51 represents the revival of this aërobian ferment. We recognize

here the branched mode of budding and spherical contour characteristic

of “high” yeast proper. Nor does the aërobian ferment of “low” yeast

present any special peculiarities, in forms, dimensions, and mode of

growth closely resembling the “low” yeast from which it is derived. At

the commencement of its restoration, however, if this is performed in

sweetened water, the cells in the groups are larger than those which are

subsequently developed.

[Illustration: Fig. 52.]

Fig. 52 represents the aërobian ferment of yeast used in

“low”-fermentation breweries, examined forty-eight hours after pitching.

We find that groups resembling that at a are of very rare occurrence.

They are to be seen only at the very beginning, generally only for the

first few hours of the renewed activity. Very soon, however, they

develop cells which are of the size of the oval cells budding at _b_.

[Illustration: Fig. 53.]

Fig. 53 represents the aërobian caseous yeast which forms rather

rapidly, in thick, greasy-looking pellicles, on the surface of liquids

which have been fermented by means of caseous ferment. The larger form

of cells, _a_ and _b_, is not often met with.

[Illustration: Fig. 54.]

On May 27th, 1875, we sowed, in a flask of wort, a trace of a pellicle

of this kind, which had formed on the surface of a flask in which

fermentation had been set up by means of caseous yeast in May of the

preceding year. On May 30th fermentation began to reveal its presence by

a voluminous froth, and the newly-formed yeast had reached the bottom of

the flask. A small quantity was taken out by a capillary glass tube, and

a sketch of the ferment made; this is given in Fig. 54. Amongst the

cells which occupy the field there are groups of some of larger size.

These are not distinct forms mixed with the others, but simply another

illustration of the fact that old cells in course of revival, especially

when they have been exhausted in sweetened water, as we have just

observed of the aërobian ferment of “low” yeast, commence with forms of

larger diameter or more elongated than the ordinary forms peculiar to

the ferment which at a later stage are developed from them. We have seen

how marked and exaggerated this feature was in the case of

_saccharomyces pastorianus_.

Let us again call attention to the forms of aërobian ferment furnished

by the yeast which we have already described under the name of _new

“high” yeast_. Fig. 55 represents this aërobian ferment, as taken on

November 27th, 1873, from a pellicle of rather greasy and moist

appearance, on the surface of a flask of fermented beer-wort which had

been impregnated on July 21st, 1872. It might readily be mistaken for

ordinary “high” yeast, yet no two ferments can be more distinct.

[Illustration: Fig. 55.]

[Illustration: Fig. 56.]

On November 27th, 1873, we sowed a trace of this ferment in a flask of

wort. From the 29th, with a continuous temperature of 25° C. (77° F.), a

considerable deposit of yeast began to form, and the froth of

fermentation covered the whole surface of the liquid. We took a little

of this deposit for examination; it is represented in Fig. 56. The field

is occupied with oval cells of great uniformity. We recognize the aspect

of the original yeast (Fig. 43). Here and there, indeed, we come across

some cells of larger size, such as those at _a_ and _b_, which is

another illustration of the remark that we have just made respecting the

forms which revived exhausted cells take on at the commencement of a new

germination.

The physical aspect of the several aërobian ferments is in general so

characteristic that we are often able by simple inspection to

distinguish between them as they occur on the surface of liquids.

_Saccharomyces pastorianus_ in its aërobian state forms a crown of cells

round the sides of the vessel at the surface of the liquid, which crown

is broken up by the least agitation of the liquid; its vitality

continues for years.

The aërobian ferment of “high” yeast appears in the form of small

isolated teats on the surface of the fermented liquid. It develops

rather sluggishly, and has no great vitality.

The aërobian form of “low” yeast develops as a somewhat fragile layer,

the least agitation precipitating it to the bottom of the vessel in a

cloud of very small irregular flocks, that do not diffuse through the

liquid as they fall. With free access to air it retains life for a long

time.

The aërobian ferment of caseous yeast forms a continuous greasy-looking

pellicle, gradually thickening, which breaks up into fragments when

shaken. With a supply of air it lives very long, and the pellicle

gradually increases in thickness.

In reviewing these ferments we may naturally ask ourselves the question

whether the “high” ferments of which we have spoken—the industrial one

concerned in the “high” fermentation of breweries, and the other which

we have termed _new “high” ferment_—are not aërobian ferments of “low”

yeasts. We are inclined to think that the ferment which in the preceding

paragraph we termed _new “high” ferment_, may, perhaps, be the aërobian

form of the “low” yeast employed by Alsatian and German brewers. We have

studied this new “high” ferment side by side with the aërobian ferment

of “low” yeast, and the result we have arrived at is, that in appearance

and mode of germination, as well as in the flavour and quality of the

beers which they produce, they greatly resemble one another. In the last

respect, however, we cannot say that the identity is quite absolute, and

hence it is with some doubt that we suggest the possible identity of the

two ferments. As regards the ordinary “high” yeast of breweries, it may

well be supposed, both from its power of rising to the surface during

fermentation and from the peculiar smell and flavour of its beer, that

we have in it the aërobian ferment of some “low” yeast, as to the

identity of which, however, we can say nothing, having no knowledge as

to where it is to be found; or, indeed, any certainty that such a yeast

actually exists.

In writing these lines an idea suggests itself which might be profitably

made the subject of serious experimental study. What would be the

peculiar properties of the aërobious ferment-form of an aërobian yeast?

Certain facts incline us to believe that these forms differ from each

other just as a “low” yeast differs from its aërobian ferment. If this

were actually the case it would be very interesting to compare the

peculiar properties of an indefinite series of aërobian ferments, all

derived from a common origin. We find recorded in our laboratory notes

that a certain aërobian ferment of the second generation produced a beer

different from that produced by the same ferment of the first

generation, being possessed of a fragrance so marked that, on entering

our laboratory, in which only a few litres of this beer were fermenting,

we were at once struck by the powerful odour which it emitted.

§ VI.—The Purification of Commercial Yeasts.

We have already stated that the researches detailed in the preceding

chapter require for their successful prosecution that the ferments on

which we experiment should be absolutely free from germs of other

organisms, and we have shown how impossible it would be, if this

condition were not complied with, to follow for weeks or months,

sometimes even years, the changes which occur in a yeast maintained in

contact with air, either in sweetened water or in a liquid which has

fermented under its influence. Equally necessary is it that the

saccharine worts employed should also be exempt from these impurities,

as well as the air, which is being constantly renewed at the surface of

the liquids. These last conditions may be realized by the adoption of

our double-necked flasks, with which a laboratory for research of this

kind should be furnished, always ready for use, filled with the

different kinds of liquids that may be required.

In general, the inconveniences resulting from the impurity of a yeast

employed do not immediately manifest themselves, in consequence of the

enormous preponderance of the true yeast, which, in comparison with the

foreign germs that contaminate it, may be so great that microscopical

examination fails to reveal even the presence of these latter. Again, it

is a well-known fact that the abundance of one growth in a limited

medium operates to the prejudice of a less abundant one, inasmuch as the

first consumes the nutritive materials at the expense of the second, and

more particularly the needful amount of oxygen. It follows, that when a

saccharine liquid is impregnated with commercial yeast, nothing but

yeast may be detected for a time, and one is led to believe in the

purity of the subsequent growth. This, however, supposes that the

external conditions, as well as those of the medium of growth, are

equally adapted to the life of the yeast and that of those organisms

present as impurities; for if these conditions rather favoured the

nutrition of the latter, we should be sure to find their proper

developments appearing at an early stage. For example, when the growth

of yeast becomes sluggish, we have invariably the development of such

after-growths. The principal germs, having exhausted the saccharine

liquid which has fermented under their influence and is no longer

adapted for their growth, cease to develop, and have their place taken

by ferments of disease, spores of moulds, mycodermata, &c., the growth

of which proceeds more or less rapidly, in proportion as the character

of the liquid and the surrounding temperature are more or less suited to

their growth.

Here, too, we have an explanation of the rapid change that occurs in

brewers’ yeast when left to itself after fermentation. In such a mass of

cells, kept apart from any food-supply, and only with difficulty able to

keep themselves in life by consuming their own soluble contents, we have

an excellent field for the development of foreign germs. In this way we

may have a rapid putrefaction in yeast, to which there will be a

correspondingly rapid growth of organisms in the liquid, where they

find, as well as in the yeast-cells, appropriate nourishment. Nothing

could better confirm this view of the matter than the array of facts, by

way of antithesis, already described, in which we have seen a pure yeast

remain for an indefinite time in contact with pure air, without

undergoing any putrefaction, or manifesting other changes than those

which result from the combustions peculiar to living cells when left to

support themselves, in a moist state, in contact with oxygen.

In the process of brewing, as soon as fermentation is finished, or

rather, as soon as certain physical effects are produced, for instance,

when the beer falls bright, or, as the French say technically, when the

yeast breaks up,[109] the beer is racked; subsequently the yeast, which

is left in a plastic layer at the bottom of the vessels, is collected,

washed, and kept under water in a cool place, to be used again in the

course of twenty-four or forty-eight hours. Brewers never care to keep

their yeast for a longer time before using it, especially in summer. We

can understand how this practice prevents the foreign germs which are

mixed with the yeast from living and reproducing; but although the

conditions of brewing, as far as the treatment of the yeast is

concerned, may, in a certain measure, prevent the development of these

germs of disease, nevertheless they are there, and from their extreme

minuteness, pass into the beer in greater or less number, however bright

it may have been rendered by racking. There they only await conditions

favourable to their existence to enable them to develop, and to affect

more or less injuriously the qualities of that delicate beverage.

On December 15th, 1872, we bought nine samples of beer in different

large cafés in Paris, which had all come from the best breweries of

Strasburg, Nancy, Vienna, and Burton. After leaving them for twenty-four

hours, we decanted all our samples, and then sowed a drop of the deposit

of each in flasks of pure wort. On January 2nd, 1873, we examined the

ferments formed in these worts, which had been kept in an oven at a

temperature of 20° C. (68° F.), and also tasted the beers produced; they

all had an abominable taste, and each contained diseased ferments.

At the same time, by way of comparison, we impregnated other flasks of

wort with pure ferments. None of the beers of this series acquired a bad

taste or produced foreign ferments; they only became flat.

When we review the operations of the brewer’s art, we are surprised by

the comparative perfection to which that art has been brought by the

laborious experience of years, and the more so when we consider that, as

regards the question of the diseases of beer, the brewer has never been

guided by any such rigorous principles as those which we have explained

in this work. We have already given proofs of this in our first chapter.

The beer is racked and separated from its yeast before fermentation has

entirely ceased. The principal reason for this is that it is necessary

that the beer, after being run into cask, should work again and undergo

a secondary fermentation, in order that it may not be invaded by the

parasites, of which we have already spoken, as would not fail to be the

case if the beer were suffered to remain in a state of perfect

quiescence. Not only is the beer racked before it has attained its limit

of attenuation, but in addition to this, and also with the view of

checking the development of parasites, it is placed in cellars sensibly

cooler than the temperature of fermentation, low as that is in the case

of “low” beers: the temperature of the cellars being not higher than 2°

or 3° C. (36° F.).

Unfortunately, the requirements of trade prevent our complying with

these exigencies to the end. When the beer is sold it is conveyed away,

no matter what the season may be, and deposited in the retailer’s

cellar, for a longer or shorter time, according to the variations of

consumption. On a warm day beer will be in great demand; the next day,

if rain or cold have come on, the demand will be very limited, since

beer is, in our climate at least, a drink for hot weather. From causes

of this nature, the beer may have to remain a long time in the cellars

of the retailers or consumers. By way of precaution, indeed, it is put

into very small casks, which permit of a frequent renewal of the supply,

and is conveyed to distances by express trains, and during the night; it

is even sent away in wagons provided with a kind of double case, the

outer jacket being filled with ice, which keeps the air surrounding the

casks constantly cold. Such are some of the troublesome measures taken

to obviate the danger that we have pointed out. They operate very

injuriously in restricting the trade and raising the price of beer. It

is a matter of extreme importance, then, that our produce should be

better removed from the action of those microscopic enemies which beer

contains; in other words, that this beverage should have less cause to

fear circumstances favourable to the development of the germs of

impurity with which it is always contaminated, as a natural consequence

of the methods of manufacture at present adopted. The question of

alteration in the flavour of beer should be regarded from another point

of view which merits equal attention. We have seen that there are

different kinds of beer, each of which corresponds to a special ferment

from which it derives its flavour and aroma, and, in a word, everything

which gives it a value in the eyes of the consumer. It very often

happens, especially in badly-managed breweries, and more particularly in

those in which several beers are manufactured, that the yeast is a

mixture of different ferments. The evil effects of such a mixture are

experienced in the course of manufacture, and still more so in the beer

after manufacture. Brewers in good “low” fermentation breweries, who

brew what is called _stock_ beer, during the winter months, for

consumption in summer, up to August and September, are very anxious to

prevent the development of a _vinous_ flavour in beers of this kind.

According to our observations, this vinous flavour seems to be

principally due to an intermixture with the pitching yeast of

_saccharomyces pastorianus_ or its varieties, one of the peculiarities

of which ferment is that in the course of time it imparts a decided

vinous flavour to beer. If this ferment were not present amongst the

yeast-cells—and here we are speaking of an absolute, so to say,

mathematically absolute absence—the beer produced would gradually grow

old in the store cellars, without ever acquiring any vinous flavour,

properly so called.

This vinous flavour develops more especially in English beers when these

are kept. It is an easy matter to show that in English beers, after

their manufacture, _saccharomyces pastorianus_ and the ferment which we

have termed _caseous_, which also imparts a peculiar flavour, form

almost exclusively, notwithstanding the fact that the yeast used in the

manufacture of English beer is a ferment essentially distinct from

_saccharomyces pastorianus_.

The secondary fermentation which takes place in “high” and “low” beers

stored in cask after manufacture, is very often due to this same

ferment, which may be recognized by elongated jointed cells, at times

more or less ramified, as well as by the influence which it exercises

upon the flavour of the beer.

We may add that the general result of our researches has convinced us

that “high” yeast cannot transform itself, any more than “low” yeast

can, into the ferment of which we are speaking, and that whenever a beer

produced by means of “high” or “low” yeast develops a foreign ferment,

this ferment must have existed in the original yeast in the form of

germs, which, from their extreme scarcity, often fail to be detected by

means of the microscope. The best proof that we can give of this is the

fact that a beer produced by means of “high” or “low” yeast, if left to

itself for months or years, will never contain in its deposit anything

besides the yeast that was used in its manufacture, provided that that

was pure to begin with. Now this can never be the case in dealing with

actual commercial beers, no matter what they may be or in what brewery

they may be produced. In all beers, in the course of time, in addition

to diseased ferments, ferments essentially different from those used in

their manufacture will appear, and notably _saccharomyces pastorianus_;

this result must be attributed to the general impurity of commercial

yeasts.

In certain cases the intermixture of ferments is to be feared almost as

much as the presence of diseased ferments, when these latter have not

developed to any great extent. We have often seen our fermentations

invaded by ferments differing absolutely from those which we originally

employed. The repetition of growths, and more particularly changes in

the composition of our fermentable media, purposely made with the view

of attaining certain results, often produce complications of this kind.

For a long time we were unable to realize the true significance of the

results of some of our experiments, in consequence of the facts which we

have just explained, as well as those detailed in the preceding

paragraph, having escaped our notice; indeed, our ignorance of those

facts added greatly to the difficulty and length of our researches. Our

labours from the commencement of this work to the date of its

publication have extended over not less than five years, and no one can

know better than ourselves with what advantage we might devote a still

longer time to it; but, as Lavoisier says, one would never give anything

to the world if he delayed doing so until he fully attained unto his

ideal aims, which always seem more distant the more one increases one’s

efforts in the attempt.

Our preceding observations show how extremely important it is to employ

pure yeasts to obtain, on the one hand, well flavoured beers, whilst

adhering to the processes at present existing in breweries, and on the

other, beers of good keeping qualities, less liable to injury, less

dependent on actual commercial requirements, capable, that is, of

withstanding conditions favourable to the development of ferments

prejudicial to the soundness of the produce, what we have named ferments

of disease.

In the case of intermixture of alcoholic ferments, we may sometimes

manage to effect their separation by taking advantage of their unequal

vitalities in different media of cultivation. On December 17th, 1872, we

made a powder of commercial Dutch yeast and plaster, as described in

Chapter III. § 6. The Dutch yeast was a “high” ferment.

On July 25th, 1873, we sowed a portion of this dried mixture in a flask

of pure wort. From July 27th patches of bubbles from fermentation were

visible on the surface.

On August 2nd the fermentation was completed. The yeast, examined under

the microscope, was _apparently_ pure, formed of spherical cells of a

fine “high” ferment. We poured away the fermented liquid, observing

every necessary precaution, and left in the flask almost all the deposit

of yeast, and not more than one or two cubic centimetres (about half a

tea-spoonful) of beer.

On November 15th following the yeast, examined afresh, still seemed pure

and still exhibited the form of round cells of “high” yeast, only that

they had taken on a very aged aspect, showing a double contour, and

filled with granulations collected irregularly about the centre. Such

are the precise characteristics of dead cells; nevertheless it was still

possible that some living cells yet remained. To assure ourselves of

this we took some of the yeast and placed it in a flask of pure wort. On

the 19th a little froth from fermentative action appeared on the

surface. We then examined the yeast and discovered that it was no longer

“high” yeast, but a small ferment of rather irregular appearance, in

which the jointed cells of _saccharomyces pastorianus_, as it usually

appears after a succession of growths, predominated. It must not be

imagined here that what we saw was a transformation of one yeast into

another. The phenomena are to be explained much more simply. The Dutch

yeast employed being very impure must have contained traces of foreign

ferments, especially of _saccharomyces pastorianus_. Reduced to a dry

powder on December 17th, 1872, the two or more varieties of cells

comprising it had preserved their vitality in consequence of the

plaster, and this vitality had continued at all events until July 25th,

1873. Subsequently, when cultivated in wort, they had multiplied in that

medium. The _saccharomyces_ had revived like the rest, but its quantity,

compared with the high Dutch yeast, was so small that the microscopical

observations made on August 2nd, when the flask was decanted, failed to

discover its presence. Between August 2nd and November 15th the high

yeast must have perished entirely: the cells of _saccharomyces_, on the

contrary, still maintained their vitality, and these alone multiplied in

the flask of wort impregnated on November 15th. Here we have an example

of the separation of alcoholic ferments, through the unequal resistance

they sometimes offer to adverse conditions to which they may be

subjected. We may also conclude that if we had prepared a quantity of

beer with the “high” yeast, which in our experiment of August 2nd, 1873,

seemed to have developed in a state of entire purity, this beer when

made and stored in cask or bottle could not have failed to undergo a

secondary fermentation, in consequence of a development of

_saccharomyces pastorianus_.

Let us take, as another example of purification of the same kind, the

case of the different ferments of the vintage. When must begins to

ferment the apiculated ferment invariably appears, and becomes

afterwards associated, more or less, with the _saccharomyces

pastorianus_, in the presence of which the multiplication of the

apiculated ferment soon ceases. _Saccharomyces pastorianus_, in its

turn, is gradually displaced by the ferment which we have termed the

ordinary ferment of wine, and which Dr. Rees has named _saccharomyces

ellipsoïdeus_. On the subject of these changes in the proportion of the

ferments of wine, the Note which we published in 1862 in the _Bulletin

de la Société chimique_ may be consulted. Now, these various ferments

mutually interfere with each other: whereas if _saccharomyces

apiculatus_ were there alone it would multiply to a greater extent, and

with greater advantage to the fermentation of the must. This result is

obtained by filtering the must, as we have already observed.

It is evident from what we have just said that the principal part of the

deposits of yeast in the sediment of fermented grapes, at the time when

the wine is first racked, which in the Jura, is called _l’entonnaison_,

is composed of the ordinary ferment of wine, the _saccharomyces

ellipsoïdeus_, and that the cells of apiculated ferment are scarcely

discoverable with the microscope, being scattered amongst an infinite

multitude of other ferments.[110]

[Illustration: Fig. 57.]

We procured from Arbois, on January 20th, 1875, some wine yeast taken

from a large barrel of the preceding vintage, racked on January 18th.

The ferment was very irregular. Some of its cells were very old, of a

yellowish colour, and full of granulations—amongst these a certain

number formed jointed segments, rather elongated, and probably belonging

to _saccharomyces pastorianus_. The other cells were transparent, and

apparently still young. This mixture of the two ferments is represented

in Fig. 57. No doubt if we had searched carefully we should also have

found some cells of _saccharomyces apiculatus_. On January 21st we sowed

a small quantity of this raw yeast in a flask of sweetened water. On the

24th we poured off the liquid, and supplied the deposit with fresh

sweetened water. The exterior temperature was 12° C. (54° F.). On the

27th we took some of the deposit and put it into a flask of wort. The

following days there was a development of yeast, accompanied by

fermentation. We obtained, however, neither the large forms of the

ferments of fruits, nor those of the more minute ferments represented in

Plate XI. The _saccharomyces pastorianus_, represented in the yeast

which we sowed by aged, granular, elongated cells, had, therefore, not

revived. Fearing that this result might have been attributable to

insufficiency of the exhaustion, which had only lasted for a few days,

we raised the temperature of the flask of sweetened water to 25° C. (77°

F.), at which we kept it until February 20th. On that day we sowed some

of this yeast in wort. There was a very perceptible revival the next

day, but it was still impossible to detect with the microscope the forms

we have just mentioned, nor did _saccharomyces pastorianus_ appear in

fresh, succeeding growths.

[Illustration: Fig. 58.]

Fig. 58 represents the yeast formed, which evidently had sprung from the

transparent cells seen in Fig. 57, and doubtless belonging to the

ordinary ferment of wine, _saccharomyces ellipsoïdeus_. Here we have

another example of the natural separation of ferments brought about by

the death of one or two of them, or by extreme differences in the time

of their revival.

We cultivated this yeast (Fig. 58), to some considerable extent, in

beer-wort. It produced a peculiar beer, of vinous character, in fact a

true _barley wine_. This proves, we may here remark, that ordinary wine,

in its flavour and quality, depends to a great extent on the specific

nature of the ferments which develop during the fermentation of the

vintage; and we may fairly assume that if we were to subject the same

must to the action of different ferments we should obtain wines of

different characters. With a view to the practical application of this

idea, it would be well to undertake new studies in this direction; and

the methods of cultivating and managing ferments, explained in this

work, would be of great value in such researches.

The purification of ferments may be accomplished by various methods,

according as we have to deal with an intermixture of ferments, or to

regard as our principal object the expulsion of ferments of disease,

such as vibrio germs, lactic ferment, the filamentous ferment of turned

beer, _mycoderma aceti_ or _mycoderma vini_.

One method of easy application consists in sowing the yeast in water

sweetened with 10 per cent. of sugar. This liquid should be first

boiled, and preserved in the two-necked flasks which we have so often

described. Sweetened water is a very exhaustive medium for ferments, and

the organisms mixed with them. A great many cells perish in it, and the

chances are that the foreign germs, which are always scarce in

comparison with the great number of cells of ferment, may be amongst

those which die, or those which become so exhausted that when the yeast,

after this treatment, is sown in wort, they disappear, and allow those

cells which have remained vigorous enough to develop alone. The addition

of a little tartaric acid to the saccharine solution—say, from 1/1000 to

2/1000 part by weight—often facilitates the destruction of certain germs

of impurity. _Mycoderma aceti_ and _mycoderma vini_ do not find suitable

life-conditions in the sweetened water; they soon disappear if

cultivated alternately in sweetened water and wort.

In the place of flasks we may make use simply of shallow basins, covered

with sheets of glass, such as we have already had occasion to describe,

for cultivating yeast in wort after it has been for a longer or shorter

time in the sweetened water. The success of these methods of

purification is mainly due to the fact that wort is highly aerated, and

experience shows that the principal disease-ferments of beer are as much

checked in their development by the presence of air as they are favoured

by its absence, the inverse of which holds good in the case of alcoholic

ferments. So true is this that, working with commercial yeast, which is

invariably impure, it would be impossible in our opinion to make beer in

closed vessels; and, indeed, as a matter of fact, one has never

succeeded in doing this, although the attempt has often been made. To do

so requires, much more than in methods actually in use, the employment

of pure yeast.

There is, therefore, this advantage in cultivating yeasts in shallow

basins, that the multiplication of the alcoholic ferments is promoted,

and that of most of the disease-ferments is checked. There is an

exception, indeed, in the case of mycodermata; but of all

disease-ferments these are the most easily got rid of, by repeating our

growths before they make their appearance. Notwithstanding this, our

two-necked flasks, which also contain much air at first, are to be

preferred to the shallow basins, inasmuch as they are a perfect

safeguard against the germs floating in the surrounding air, as well as

those of the ferment _saccharomyces pastorianus_.

Another method is suggested to us by the curious results of which we

have already spoken, obtained by sowing yeasts in a wort rendered acid

and alcoholic by the addition of bi-tartrate of potash and alcohol.

Experience proves that many disease-ferments find great difficulty in

withstanding a succession of growths in wort to which 1-½ per cent. of

tartaric acid and from 2 to 3 per cent. of alcohol have been added. Such

a mixture, however, is equally well adapted to the requirements of

_saccharomyces pastorianus_, and we must always assure ourselves that

this organism has not taken the place of the yeast we are endeavouring

to purify. Growths at a very low temperature are of great help in

enabling us to get rid of all ferments that are foreign to “low” yeast,

and should be resorted to in all cases where this yeast is to be

purified.

Another method of purification, which is perhaps quicker, although

inferior in other respects, consists in the employment of carbolic

acid—that is to say, in purifying our yeast by successive growths, we

may add to every 100 c.c. (3-½ fluid ounces) of wort that we employ from

ten to twelve drops of phenol water, containing 10 per cent. of the

acid. The action of the phenol, which at first is invariably combined

with that of the oxygen of the air, tends to destroy the vitality of

many of the cells sown, involving to some extent also the yeast which we

are interested in preserving. But amongst the number of cells that are

affected those which are less abundant, that is to say, those which are

present as impurities, are paralyzed relatively in much greater

proportion. If the acid does not destroy them it greatly checks their

development, and the cells of yeast, which multiply continuously in vast

numbers (for the fermentation goes on in spite of the phenol, if this is

added in small quantity), gradually choke the foreign germs in a

succession of growths.

By these different means, which are employed separately or combined with

one another, we generally manage to obtain the yeast which we wish to

purify in a very pure state. We need scarcely add that it is always

well, in the case of our purifications, to begin with specimens which

are already as pure as it is possible to obtain them. In making our

choice the microscope is our best guide, but it is not a sufficient one.

We should be strangely deceived if we believed in the purity of a yeast

for the sole reason that when examined under the microscope it appeared

to contain nothing of a foreign nature. The best means of assuring

ourselves of the purity of a yeast consists in making some beer in one

of our two-necked flasks, and leaving this flask, after fermentation, in

an oven at a temperature of 20° or 25° C. (68° to 77° F.). If the beer,

in the course of a few weeks, does not thicken, or become covered with

efflorescence, if its deposit is microscopically pure, if, in short, it

only tastes flat, we may have every confidence in the purity of the

yeast which produced it. After we have purified a yeast we are,

unfortunately, never sure that it has not undergone some change in the

course of the manipulations to which it has been subjected in

purification. It is indispensable, therefore, that we should test it,

and see if the flavour of the beer produced by it is really the one that

we want—viz., that of the beer from which we took the yeast that we

submitted to purification.

In the course of a series of practical experiments that we were carrying

out in the large brewery of Tourtel, at Tantonville, in 1875, in

connection with the new process of brewing, which will be explained in

Chap. VII., the following circumstance occurred. We had purified some of

the yeast of the brewery, by means of a succession of growths and adding

a few drops of phenol, and had obtained a yeast of irreproachable

purity. It happened that this yeast, which was repeatedly cultivated in

the brewery during the summer of 1875, from six to ten hectolitres (130

to 220 gallons) of wort being used on each occasion, always produced a

beer that had a yeast-bitten flavour and defective clarifying powers,

notwithstanding that it possessed remarkable keeping properties, which

it owed to the pureness of the ferment employed. As a matter of fact,

the beer suffered no injury from journeys of more than 300 miles, by

slow trains, in ordinary casks, containing from 50 to 100 litres (10 to

20 gallons), during the great heats of June and July, or from being

subsequently stored for two months in a cellar, the temperature of which

rose during that time from 12° to 18° C. (54° to 65° F.). The

temperature of fermentation had been 13° C. (55° F.). Beer from the same

brewery, made with the same wort by the ordinary process, did not remain

sound for three weeks in this same cellar.

To what may we attribute the peculiarity of the beer as just described?

It is probable that during our processes of purification some ferment

had taken the place of the principal yeast. Commercial yeasts, even

those with which the brewer is thoroughly satisfied, generally contain

various ferments, which are maintained in their relative proportions, or

very nearly so, by the uniform conditions under which work is carried on

in a brewery; but these proportions, it is obvious, might be very

seriously affected by any radical change in the conditions of growth.

Footnote 82:

Extract from a Note which I inserted in 1862 in the _Bulletin de la

Société chimique_ of Paris.

Footnote 83:

Schützenberger, in his work on “Fermentation,” following Dr. de

Vauréal. Paris, 1875, p. 278. [See pp. 61, 62 English version in

International Scientific Series (H. S. King & Co., London, 1876). This

appears to be the only reference to this subject in the English

copy.—D. C. R.]

Footnote 84:

The principal result of Dr. Rees’ labours consists in the discovery of

a sporulation peculiar to yeast cells, that is to say, to a formation

in the interior of these cells, and under particular conditions—such

as when the growth occurs on slices of cooked potatoes, carrots,

&c.—of two, three, or four smaller cells, which, when placed in

fermentable liquids, act like the germinating spores of ferments. The

mother-cell may be regarded as an _ascus_, and the daughter-cells as

_ascospores_, and so the genus _saccharomyces_ may be classified among

the group of fungi termed _ascomycetes_. These facts have been

frequently confirmed, notably by Dr. Engel, professor of the Faculty

of Medicine, at Nancy. Previously to Dr. Rees’ discovery, M. de Seynes

(_Comptes rendus_, t. lxvii., 1868) had described an endogenous

formation of spores in _mycoderma vini_, particularly in the elongated

cells, followed by the rupture of the mother-cell, and subsequent

absorption of cell-walls and other contents after the issue of the

endospores, which we have just termed _ascospores_. We ourselves had

also previously called attention to those refractive corpuscles which

appear amongst vibrios as probably being reproductive corpuscles, and

we had likewise witnessed the reabsorption of the parts surrounding

them. The plate on page 228 of our “Studies on the Silkworm Disease”

represents the phenomena in question.

Footnote 85:

See _Comptes rendus de l’Académie des Sciences_, vol. lviii. p. 144.

Footnote 86:

The plates referred to in this paragraph were exhibited at a meeting

of the Academy of Sciences, November 18, 1872, and commented upon by

the perpetual secretary, M. Dumas.

Footnote 87:

For these observations, we employed small glass cells, which we made

out of some St. Gobain glass by punching holes through it, and then

cementing on one side one of the little glasses used for covering

objects in microscopical examinations. In this manner we made small

troughs, in which we placed some wort that had been boiled, and a drop

of the water in which grapes had been washed. To prevent evaporation

we covered the cells with a sheet of glass. We examined the liquid in

these cells by inclining our microscope to the angle required.[88]

[Illustration: Fig. 29.]

We also made use of cells similar to those employed by MM. Van Tieghem

and Lemonnier[89] in their researches on _mucorines_ (Fig. 30).

[Illustration: Fig. 30.]

An apparatus similar to that employed by M. Duclaux in 1853[90] would

do equally well. We should be able to work with even greater facility

if we employed bulbs like some which we ordered in Germany, some

twelve years ago, of the well-known glass-blower, Geissler. We have

heard that these bulbs now sold by that maker are much used by German

microscopists. They consist of a tube blown out into a flat bulb, the

sides of which, in the centre, come sufficiently close together to

enclose but a very thin layer of liquid, and to admit of microscopical

examination. We may fill these tube-bulbs completely with liquid, to

the exclusion of air or we may surround the central drop with air.

[Illustration: Fig. 31.]

Footnote 88:

In our essay on acetic fermentation, published in 1864, we have

already described this apparatus, which we employed to follow the

multiplication of the jointed filaments of _mycoderma aceti_. See

PASTEUR, _Etudes sur le vinaigre_, p. 64, Paris, 1868.

Footnote 89:

VAN TIEGHEM and LEMONNIER, _Annales des Sciences naturelles_, 5th

series, _Botanique_, t. xvii. 1873.

Footnote 90:

DUCLAUX, _Comptes rendus des séances de l’Académie des Sciences_, t.

lvi. p. 1225.

Footnote 91:

In experiments of this kind there is always a slight increase in the

volume of air in the jar. This increase may be very perceptible even

when the experiment made with fresh grapes, in August, for instance,

causes no fermentation due to the action of yeast. After the oxygen of

the air has been absorbed and replaced by carbonic acid gas, either by

direct oxidation or by the action of moulds, the grapes, although

crushed, act like fruits plunged into carbonic acid gas[92], and this

effect is even more marked in the case of imperfectly crushed grapes.

The reason is, that the crushing is never so perfect as to injure all

the cells of the parenchyma. We may easily convince ourselves that the

experiment on the liberation of carbonic acid gas and the formation of

alcohol by grapes and fruits in general when plunged into carbonic

acid succeeds very well in the case of fragments of fruits or grapes,

and succeeds better the less the parts are crushed.

Footnote 92:

See paragraph: Fermentation in saccharine fruits immersed in carbonic

acid gas, Chap. vi, § 2, p. 266.

Footnote 93:

Dr. Rees has given the name _saccharomyces ellipsoideus_ to the

ferment of wine represented in Plates VIII. to XI. of our “Studies on

Wine,” which we have termed the ordinary ferment of wine, from its

being the most abundant of the ferments found at the end of the

fermentation that produces the wine.

Footnote 94:

The alcoholic ferments in general, subjected to these weakening

influences, have not all the same power of resistance. That one which

seems to possess this power in the highest degree is the

_saccharomyces pastorianus_, which ferment we had in view in writing

the above.

Footnote 95:

The term _exhaustion_ (_épuisement_), which we have just used, was,

perhaps, not altogether felicitously chosen. No doubt we exhaust the

cells of yeast when we sow an imponderable weight of them in a large

quantity of sweetened water; it might, however, be better to say that

in such a case we adopt a particular method of preserving the vitality

of the cells, without suffering them to die of exhaustion, or to

multiply by budding. We may remark that the yeast, in this case,

exists in a state of latent life, which resembles that of cells on the

surface of fruit. The cells on the surface of fruits, bunches, or

barks, can no more find around them sufficient aliment for their

propagation than can our yeast-cells in a great excess of sweetened

water. We would not, however, say of the spores on the surface of

fruits, or their woods, that they are in a state of exhaustion; the

term would be misapplied.

Footnote 96:

See foot note p. 79.—D. C. R.

Footnote 97:

M. Béchamp (_Comptes rendus_, November 18th, 1872) asserts that _the

air has no direct influence on the production of ferment or on the

process of alcoholic fermentation_. That experienced chemist deduces

this erroneous assertion from experiments on sweetened water, to which

bunches of grapes, petals of corn-poppies and petals of _robinia

pseudo-acacia_ had been added. As may be seen in our “Studies on Wine”

(p. 7, 1st edition, 1866), these experiments conducted by M. Béchamp

in 1872 were merely a reproduction of those made long before with vine

leaves, petals of elder-flowers, leaves of sorrel, &c., by the Marquis

de Bullion, Fabroni, and other experimentalists. M. Béchamp has

modified his later experiments by not adding the bunches of grapes,

leaves, &c., to the sweetened water before having introduced carbonic

acid gas into the liquid. Fermentation having still taken place in

spite of this change, M. Béchamp wrongly concluded that _air has no

direct influence on the production of yeast on an alcoholic

fermentation_. The introduction of the carbonic acid gas could not

remove all the air imparted to the sweetened water by the objects

placed in it, and it was this air which remained adhering to these

objects that permitted the production of fermentation. We may avail

ourselves of the opportunity here presented to add that, in this same

Note of November, 1872, M. Béchamp commences by making various

assertions concerning the forms assumed by cells of the alcoholic

ferment of the grape when in process of fermentation. This question

was discussed by us ten years before, and our conclusions supported by

sketches, in a Note which appeared in the _Bulletin de la Société

chimique de Paris_, for 1862.

Footnote 98:

The germs of ferments are less widely diffused than M. de Bary

supposes, as may be seen from our observations in Chap. III. See, too,

our Memoir of 1862, _Sur les Générations dites Spontanées_, p. 49. It

is only in a laboratory devoted to researches on fermentation, or

places such as vaults, cellars, and breweries, that the air holds

appreciably in suspension cells of ferments, ready to germinate in

saccharine media. If we except these particular circumstances, ferment

is not very largely diffused, save on the surface of fruits and the

wood of the trees which bear them, and perhaps, also, on some other

plants. The particles of dust held in suspension in any atmosphere

whatever rarely produce fermentation in pure must even when we take

all possible precautions, so that the action be not overlooked; for

true fermentation may be hidden by fungoid growths, when there is much

air and but a small quantity of saccharine liquid present.

Footnote 99:

In these experiments the apiculated ferment appeared sometimes, but

much less frequently than _saccharomyces pastorianus_. We also met

with the ellipsoidal ferment. We should probably have a greater

variety of ferments if our experiments could be conducted in the open

air, but insects and particles of dust of all kinds brought by the

wind render experiments under such conditions difficult and

untrustworthy. In a laboratory we have not these difficulties to

contend against, but, unfortunately, the operations ordinarily carried

on there cause the results of our experiments to be of a less general

character than they would be if obtained in free contact with country

air.

Footnote 100:

[A rather serious clerical error appears to have here crept into the

original, for on referring to Plate I. and the letterpress descriptive

of No. 7 (p. 5), we find it applies to a very formidable species of

diseased ferment, whereas the author is here speaking of an amorphous

deposit, harmless in character, and more or less associated with all

yeasts. Doubtless No. 7 should stand No. 6, see p. 6.—D. C. R.]

Footnote 101:

[We would here call the reader’s attention to the following extract

from Dr. Graham’s appreciative review of this work in “Nature,”

January 11th, 1877. He says: “M. Pasteur seems to be in error in

stating (p. 190, Fr. ed.) that the bottom yeast may be distinguished

by being less spherical than top yeast. It is true that in London and

Edinburgh yeast, the cells will be found usually round; hard water,

however, such as that at Burton, or artificially made so, yields yeast

in which the cells are distinctly ovoid in appearance, resembling very

closely Bavarian bottom yeast.”—D. C. R.]

Footnote 102:

[43° F. to 46° F. or 59° F. to 68° F.]

Footnote 103:

[On this point again Dr. Graham expresses some dissent (“Nature,” loc.

cit.): “Here surely M. Pasteur must be thinking rather of the inferior

products of the surface fermentation in France and Germany, than of

those of England and Scotland.”—D. C. R.]

Footnote 104:

[28·4 c.c. = 1 fl. oz. approximately.]

Footnote 105:

[M. Pasteur has evidently employed the word “caseous” to express the

curdy nature of the ferment he is describing, its plasticity and other

peculiarities of physical character; but we are, nevertheless, tempted

to suggest that he may have had in mind also the peculiar “cheesy”

odour given off by these very yeasts, which he refers to in the text

as containing a considerable intermixture of “caseous ferment.”—F. F.]

Footnote 106:

The caseous ferment, however, must not be exposed to heat, under the

afore-mentioned conditions, when it is too young. At the commencement

of its development, for instance, within a few days of having been

sown. In such case, it would be in danger of perishing, probably in

consequence of the tenderness of its tissues. At the end of a

fermentation, and even several months afterwards, it might be safely

heated to 50° C. (122° Fahr.) without any harm to it. “Low” yeast also

can withstand a temperature of 50° C. in the medium in question.

Footnote 107:

Although we believe that the aërobian ferment of a particular yeast

may be produced by a kind of transformation of the cells of the

latter, yet we admit that this question is open to some doubt. The

facts which we unexpectedly discovered in connection with the

_caseous_ ferment should make us extremely careful, and disposed to

inquire whether aërobian ferments do not originally, in a state of

intermixture, form part of the ferments from which they spring. One

reason which might incline us to believe this, is the fact that a

ferment sometimes perishes without the appearance of aërobian ferment

on the surface. There is nothing very natural indeed in the hypothesis

that we advance, which sets aside the supposed intermixture; but, on

the other hand, if the aërobian ferment is a particular ferment,

simply intermixed with some other variety and developed by change of

conditions, how are we to account for its great resemblance in

appearance and mode of budding to the ferment on the surface of which

it appears? This resemblance, however, might be accounted for very

naturally if the two ferments were originally related.

Footnote 108:

We insist on this fact, that Fig. 50 represents the forms on revival

of the aërobian ferment of _saccharomyces pastorianus, when this has

grown in a mineral medium_. When produced on the surface of fermented

wort, the aërobian ferment of which we are speaking presents no

peculiarity, nor is there any irregularity in its forms or in its

development, and when we proceed to cultivate it in a natural

saccharine medium, or in wort, it does not produce any forms of

_dematium_, as in the preceding case; but the reason of this is that,

in consequence of the nature of the first medium, which is better

adapted to its nutrition, it assumes at once, in the second medium,

the forms of deposit-yeast in the course of ordinary germination.

Footnote 109:

“La cassure de la levûre.”

Footnote 110:

We have reason to believe that the ratio of the proportions of these

ferments depends greatly on the climatic conditions preceding the

period of vintage, on the state of dryness or humidity, as well as the

temperature at the time of gathering the grapes, and also on the

nature of the vines.

CHAPTER VI.

THE PHYSIOLOGICAL THEORY OF FERMENTATION.

§ I.—On the Relations existing between Oxygen and Yeast.

The object of all science is a continuous reduction of the number of

unexplained phenomena. It is observed, for instance, that fleshy fruits

are not liable to fermentation so long as their epidermis remains

uninjured. On the other hand, they ferment very readily when they are

piled up in heaps, more or less open, and immersed in their saccharine

juice. The mass becomes heated and swells; carbonic acid gas is

disengaged, and the sugar disappears and is replaced by alcohol. Now, as

to the question of the origin of these spontaneous phenomena, so

remarkable in character as well as usefulness for man’s service, modern

knowledge has taught us that fermentation is the consequence of a

development of vegetable cells, the germs of which do not exist in the

saccharine juices within fruits; that many varieties of these cellular

plants exist, each giving rise to its own particular fermentation. The

principal products of these various fermentations, although resembling

each other in their nature, differ in their relative proportions and in

the accessory substances that accompany them, a fact which alone is

sufficient to account for wide differences in the quality and commercial

value of alcoholic beverages.

Now that the discovery of ferments and their living nature, and our

knowledge of their origin, may have solved the mystery of the

spontaneous appearance of fermentations in natural saccharine juices, we

may ask whether we must still regard the reactions that occur in these

fermentations as phenomena inexplicable by the ordinary laws of

chemistry. We can readily see that fermentations occupy a special place

in the series of chemical and biological phenomena. What gives to

fermentations certain exceptional characters, of which we are only now

beginning to suspect the causes, is the mode of life in the minute

plants designated under the generic name of _ferments_, a mode of life

which is essentially different from that in other vegetables, and from

which result phenomena equally exceptional throughout the whole range of

the chemistry of living beings.

The least reflection will suffice to convince us that the alcoholic

ferments must possess the faculty of vegetating and performing their

functions out of contact with air. Let us consider, for instance, the

method of vintage practised in the Jura. The bunches are laid at the

foot of the vine in a large tub, and the grapes there stripped from

them. When the grapes, some of which are uninjured, others bruised, and

all moistened by the juice issuing from the latter, fill the tub—where

they form what is commonly called the _vintage_—they are conveyed in

barrels to large vessels fixed in cellars of a considerable depth. These

vessels are not filled to more than three-quarters of their capacity.

Fermentation soon takes place in them, and the carbonic acid gas finds

escape through the bunghole, the diameter of which, in the case of the

largest vessels, is not more than ten or twelve centimetres (about four

inches). The wine is not drawn off before the end of two or three

months. In this way it seems highly probable that the yeast which

produces the wine under such conditions must have developed, to a great

extent at least, out of contact with oxygen. No doubt oxygen is not

entirely absent from the first; nay, its limited presence is even a

necessity to the manifestation of the phenomena which follow. The grapes

are stripped from the bunch in contact with air, and the must which

drops from the wounded fruit takes a little of this gas into solution.

This small quantity of air so introduced into the must, at the

commencement of operations, plays a most indispensable part, it being

from the presence of this that the spores of ferments which are spread

over the surface of the grapes and the woody part of the bunches derive

the power of starting their vital phenomena.[111] This air, however,

especially when the grapes have been stripped from the bunches, is in

such small proportion, and that which is in contact with the liquid mass

is so promptly expelled by the carbonic acid gas, which is evolved as

soon as a little yeast has formed, that it will readily be admitted that

most of the yeast is produced apart from the influence of oxygen,

whether free or in solution. We shall revert to this fact, which is of

great importance. At present we are only concerned in pointing out that,

from the mere knowledge of the practices of certain localities, we are

induced to believe that the cells of yeast, after they have developed

from their spores, continue to live and multiply without the

intervention of oxygen, and that the alcoholic ferments have a mode of

life which is probably quite exceptional, since it is not generally met

with in other species, vegetable or animal.

Another equally exceptional characteristic of yeast and fermentation in

general consists in the small proportion which the yeast that forms

bears to the sugar that decomposes. In all other known beings the weight

of nutritive matter assimilated corresponds with the weight of food used

up, any difference that may exist being comparatively small. The life of

yeast is entirely different. For a certain weight of yeast formed, we

may have ten times, twenty times, a hundred times as much sugar, or even

more decomposed, as we shall experimentally prove by-and-bye; that is to

say, that whilst the proportion varies in a precise manner, according to

conditions which we shall have occasion to specify, it is also greatly

out of proportion to the weight of the yeast. We repeat, the life of no

other being, under its normal physiological conditions, can show

anything similar. The alcoholic ferments, therefore, present themselves

to us as plants which possess at least two singular properties: they can

live without air, that is, without oxygen, and they can cause

decomposition to an amount which, though variable, yet, as estimated by

weight of product formed, is out of all proportion to the weight of

their own substance. These are facts of so great importance, and so

intimately connected with the theory of fermentation, that it is

indispensable to endeavour to establish them experimentally, with all

the exactness of which they will admit.

The question before us is whether yeast is in reality an anaërobian

plant, and what quantities of sugar it may cause to ferment, under the

various conditions under which we cause it to act.

The following experiments were undertaken to solve this double

problem:—We took a double-necked flask, of three litres (five pints)

capacity, one of the tubes being curved and forming an escape for the

gas; the other one, on the right hand side (Fig. 59), being furnished

with a glass tap. We filled this flask with pure yeast-water, sweetened

with 5 per cent. of sugar candy, the flask being so full that there was

not the least trace of air remaining above the tap or in the escape

tube; this artificial wort had, however, been itself aerated. The curved

tube was plunged in a porcelain vessel full of mercury, resting on a

firm support. In the small cylindrical funnel above the tap, the

capacity of which was from 10 c.c. to 15 c.c. (about half a fluid ounce)

we caused to ferment, at a temperature of 20° or 25° C. (about 75° F.),

five or six cubic centimetres of the saccharine liquid, by means of a

trace of yeast, which multiplied rapidly, causing fermentation, and

forming a slight deposit of yeast at the bottom of the funnel above the

tap. We then opened the tap, and some of the liquid in the funnel

entered the flask, carrying with it the small deposit of yeast, which

was sufficient to impregnate the saccharine liquid contained in the

flask. In this manner it is possible to introduce as small a quantity of

yeast as we wish, a quantity the weight of which, we may say, is hardly

appreciable. The yeast sown multiplies rapidly and produces

fermentation, the carbonic acid gas from which is expelled into the

mercury. In less than twelve days all the sugar had disappeared, and the

fermentation had finished. There was a sensible deposit of yeast

adhering to the sides of the flask; collected and dried it weighed 2·25

grammes (34 grains). It is evident that in this experiment the total

amount of yeast formed, if it required oxygen to enable it to live,

could not have absorbed, at most, more than the volume which was

originally held in solution in the saccharine liquid, when that was

exposed to the air before being introduced into the flask.

[Illustration: Fig. 59.]

[Illustration: Fig. 60.]

Some exact experiments conducted by M. Raulin in our laboratory have

established the fact that saccharine worts, like water, soon become

saturated when shaken briskly with an excess of air, and also that they

always take into solution a little less air than saturated pure water

contains under the same conditions of temperature and pressure. At a

temperature of 25° C. (77° F.) therefore, if we adopt the coefficient of

the solubility of oxygen in water given in Bunsen’s tables, we find that

1 litre (1-3/4 pints) of water saturated with air contains 5·5 c.c. (0·3

cubic inch) of oxygen. The three litres of yeast-water in the flask,

supposing it to have been saturated, contained less than 16·5 c.c. (1

cubic inch) of oxygen, or, in weight, less than 23 milligrammes (0·35

grains). This was the maximum amount of oxygen, supposing the greatest

possible quantity to have been absorbed, that was required by the yeast

formed in the fermentation of 150 grammes (4·8 Troy ounces) of sugar. We

shall better understand the significance of this result later on. Let us

repeat the foregoing experiment, but under altered conditions. Let us

fill, as before, our flask with sweetened yeast-water, but let this be

first boiled, so as to expel all the air it contains. To effect this we

arrange our apparatus as represented in the accompanying sketch (Fig.

60). We place our flask, A, on a tripod above a gas flame, and in place

of the vessel of mercury substitute a porcelain dish, under which we can

put a gas flame, and which contains some fermentable, saccharine liquid,

similar to that with which the flask is filled. We boil the liquid in

the flask and that in the basin simultaneously, and then let them cool

down together, so that as the liquid in the flask cools some of the

liquid is sucked from the basin into the flask. From a trial experiment

which we conducted, determining the quantity of oxygen that remained in

solution in the liquid after cooling, according to M. Schützenberger’s

valuable method, by means of hydrosulphite of soda,[112] we found that

the three litres in the flask, treated as we have described, contained

less than one milligramme (0·015 grain) of oxygen. At the same time we

conducted another experiment, by way of comparison (Fig. 61). We took a

flask, B, of larger capacity than the former one, which we filled about

half with the same volume as before of a saccharine liquid of

identically the same composition. This liquid had been previously freed

from alterative germs by boiling. In the funnel surmounting A, we put a

few cubic centimetres of saccharine liquid in a state of fermentation,

and when this small quantity of liquid was in full fermentation, and the

yeast in it was young and vigorous, we opened the tap, closing it again

immediately, so that a little of the liquid and yeast still remained in

the funnel. By this means we caused the liquid in A to ferment. We also

impregnated the liquid in B with some yeast taken from the funnel of A.

We then replaced the porcelain dish in which the curved escape tube of A

had been plunged, by a vessel filled with mercury.

[Illustration: Fig. 61.]

The following is a description of two of these comparative fermentations

and the results they gave.

The fermentable liquid was composed of yeast-water sweetened with 5 per

cent. of sugar-candy; the ferment employed was _saccharomyces

pastorianus_.

The impregnation took place on January 20th. The flasks were placed in

an oven at 25° C. (77° F.).

_Flask A, without air._

January 21st.—Fermentation commenced; a little frothy liquid issued

from the escape-tube and covered the mercury.

The following days, fermentation was active. Examining the yeast

mixed with the froth that was expelled into the mercury by the

evolution of carbonic acid gas, we found that it was very fine,

young, and actively budding.

February 3rd.—Fermentation still continued, showing itself by a

number of little bubbles rising from the bottom of the liquid, which

had settled bright. The yeast was at the bottom in the form of a

deposit.

February 7th.—Fermentation still continued, but very languidly.

February 9th.—A very languid fermentation still went on, discernible

in little bubbles rising from the bottom of the flask.

_Flask B, with air._

January 21st.—A sensible development of yeast.

The following days, fermentation was active, and there was an

abundant froth on the surface of the liquid.

February 1st.—All symptoms of fermentation had ceased.

As the fermentation in A would have continued for a long time, being so

very languid, and as that in B had been finished for several days, we

brought to a close our two experiments on February 9th. To do this we

poured off the liquids in A and B, collecting the yeasts on tared

filters. Filtration was an easy matter, more especially in the case of

A. Examining the yeasts under the microscope, immediately after

decantation, we found that both of them remained very pure. The yeast in

A was in little clusters, the globules of which were collected together,

and appeared by their well defined borders to be ready for an easy

revival in contact with air.

As might have been expected, the liquid in the flask B did not contain

the least trace of sugar; that in the flask A still contained some, as

was evident from the non-completion of fermentation, but not more than

4·6 grammes (71 grains). Now, as each flask originally contained 3

litres of liquid, holding in solution 5 per cent. of sugar, it follows

that 150 grammes (2,310 grains) of sugar had fermented in the flask B,

and 145·4 grammes (2,239·2 grains) in the flask A. The weights of yeast

after drying at 100° C. (212° F.) were—

For the flask B, with air 1·970 grammes (30·4 grains).

For the flask A, without air 1·368 grammes.[113]

The proportions were 1 of yeast to 76 of fermented sugar in the first

case, and 1 of yeast to 89 of fermented sugar in the second.

From these facts the following consequences may be deduced:

1. The fermentable liquid (flask B), which since it had been in contact

with air, necessarily held air in solution, although not to the point of

saturation, inasmuch as it had been once boiled to free it from all

foreign germs, furnished a weight of yeast sensibly greater than that

yielded by the liquid which contained no air at all (flask A), or, at

least, which could only have contained an exceedingly minute quantity.

2. This same slightly aerated fermentable liquid fermented much more

rapidly than the other. In eight or ten days it contained no more sugar;

while the other, after twenty days, still contained an appreciable

quantity.

Is this last fact to be explained by the greater quantity of yeast

formed in B? By no means. At first, when the air has access to the

liquid, much yeast is formed and little sugar disappears, as we shall

prove immediately; nevertheless the yeast formed in contact with the air

is more active than the other. Fermentation is correlative, first to the

development of the globules, and then to the continued life of those

globules once formed. The more oxygen these last globules have at their

disposal during their formation, the more vigorous, transparent, and

turgescent, and, as a consequence of this last quality, the more active

they are in decomposing sugar. We shall revert hereafter to these facts.

3. In the airless flask the proportion of yeast to sugar was 1/89; it

was only 1/76 in the flask which had air at first.

The proportion that the weight of yeast formed bears to the weight of

the sugar is, therefore, variable, and this variation depends, to a

certain extent, upon the presence of air and the possibility of oxygen

being absorbed by the yeast. We shall presently show that yeast

possesses the power of absorbing that gas and emitting carbonic acid,

like ordinary fungi, that even oxygen may be reckoned amongst the number

of food-stuffs that may be assimilated by this plant, and that this

fixation of oxygen in yeast, as well as the oxidations resulting from

it, have the most marked effect on the life of yeast, on the

multiplication of its cells, and on their activity as ferments acting

upon sugar, whether immediately or afterwards, apart from supplies of

oxygen or air.

In the preceding experiment, conducted without the presence of air,

there is one circumstance particularly worthy of notice. This experiment

succeeds, that is to say, the yeast sown in the medium deprived of

oxygen develops, only when this yeast is in a state of great vigour. We

have already explained the meaning of this last expression. But we wish

now to call attention to a very evident fact in connection with this

point. We impregnate a fermentable liquid; yeast develops and

fermentation appears. This lasts for several days and then ceases. Let

us suppose that, from the day when fermentation first appears in the

production of a minute froth, which gradually increases till it whitens

the surface of the liquid, we take, every twenty-four hours, or at

longer intervals, a trace of the yeast deposited on the bottom of the

vessel and use it for starting fresh fermentations. Conducting these

fermentations all under precisely the same conditions of temperature,

character, and volume of liquid, let us continue this for a prolonged

time, even after the original fermentation is finished. We shall have no

difficulty in seeing that the first signs of action in each of our

series of second fermentations appear always later and later in

proportion to the length of time that has elapsed from the commencement

of the original fermentation. In other words, the time necessary for the

development of the germs and the production of that amount of yeast

sufficient to cause the first appearance of fermentation varies with the

state of the impregnating cells, and is longer in proportion as the

cells are further removed from the period of their formation. It is

essential, in experiments of this kind, that the quantities of yeast

successively taken should be as nearly as possible equal in weight or

volume, since, _ceteris paribus_, fermentations manifest themselves more

quickly the larger the quantity of yeast employed in impregnation.

If we compare under the microscope the appearance and character of the

successive quantities of yeast taken, we shall see plainly that the

structure of the cells undergoes a progressive change. The first sample

which we take, quite at the beginning of the original fermentation,

generally gives us cells rather larger than those later on, and

possessing a remarkable tenderness. Their walls are extremely thin, the

consistency and softness of their protoplasm is akin to fluidity, and

their granular contents appear in the form of scarcely visible spots.

The borders of the cells soon become more marked, a proof that their

walls undergo a thickening; their protoplasm also becomes denser, and

the granulations more distinct. Cells of the same organ, in the states

of infancy and old age, should not differ more than the cells of which

we are speaking, taken in their extreme states. The progressive changes

in the cells, after they have acquired their normal form and volume,

clearly demonstrate the existence of a chemical work of a remarkable

intensity, during which their weight increases, although in volume they

undergo no sensible change, a fact that we have often characterized as

“the continued life of cells already formed.” We may call this work a

process of maturation on the part of the cells, almost the same that we

see going on in the case of adult beings in general, which continue to

live for a long time, even after they have become incapable of

reproduction, and long after their volume has become permanently fixed.

This being so it is evident, we repeat, that, to multiply in a

fermentable medium, quite out of contact with oxygen, the cells of yeast

must be extremely young, full of life and health, and still under the

influence of the vital activity which they owe to the free oxygen which

has served to form them, and which they have perhaps stored up for a

time. When older, they reproduce themselves with much difficulty when

deprived of air, and gradually become more languid; and if they do

multiply, it is in strange and monstrous forms. A little older still,

they remain absolutely inert in a medium deprived of free oxygen. This

is not because they are dead; for in general they may be revived in a

marvellous manner in the same liquid if it has been first aerated before

they are sown. It would not surprise us to learn that at this point

certain preconceived ideas suggest themselves to the mind of an

attentive reader on the subject of the causes that may serve to account

for such strange phenomena in the life of these beings which our

ignorance hides under the expressions of _youth_ and _age_; this,

however, is a subject that we cannot pause to consider here.

At this point we must observe—for it is a matter of great

importance—that, in the operations of the brewer there is always a time

when the yeasts are in this state of vigorous youth of which we have

been speaking, acquired under the influence of free oxygen, since all

the worts and all the yeasts of commerce are necessarily manipulated in

contact with air, and so impregnated more or less with oxygen. The yeast

immediately seizes upon this gas and acquires a state of freshness and

activity, which permits it to live afterwards out of contact with air,

and to act as a ferment. Thus, in ordinary brewery practice, we find the

yeast already formed in abundance even before the earliest external

signs of fermentation have made their appearance. In this first phase of

its existence, yeast lives chiefly like an ordinary fungus.

From the same circumstances it is clear that the brewer’s fermentations

may, speaking quite strictly, last for an indefinite time, in

consequence of the unceasing supply of fresh wort, and from the fact,

moreover, that the exterior air is constantly being introduced during

the work, and that the air contained in the fresh worts keeps up the

vital activity of the yeast, as the act of breathing keeps up the vigour

and life of cells in all living beings. If the air could not renew

itself in any way, the vital activity which the cells originally

received, under its influence, would become more and more exhausted, and

the fermentation eventually come to an end.

We may recount one of the results obtained in other experiments similar

to the last, in which, however, we employed yeast which was still older

than that used for our experiment with flask A (Fig. 60), and moreover

took still greater precautions to prevent the presence of air. Instead

of leaving the flask, as well as the dish, to cool slowly, after having

expelled all air by boiling, we permitted the liquid in the dish to

continue boiling whilst the flask was being cooled by artificial means;

the end of the escape tube was then taken out of the still boiling dish

and plunged into the mercury trough. In impregnating the liquid, instead

of employing the contents of the small cylindrical funnel whilst still

in a state of fermentation, we waited until this was finished. Under

these conditions, fermentation was still going on in our flask, after a

lapse of three months. We stopped it and found that 0·255 gramme (3·9

grains) of yeast had been formed, and that 45 grammes (693 grains) of

sugar had fermented, the ratio between the weights of yeast and sugar

being thus 0·255/45 = 1/176. In this experiment the yeast developed with

much difficulty, by reason of the conditions to which it had been

subjected. In appearance the cells varied much, some were to be found

large, elongated, and of tubular aspect, some seemed very old and were

extremely granular, whilst others were more transparent. All of them

might be considered abnormal cells.

In such experiments we encounter another difficulty. If the yeast sown

in the non-aerated fermentable liquid is in the least degree impure,

especially if we use sweetened yeast-water, we may be sure that

alcoholic fermentation will soon cease, if, indeed, it ever commences,

and that accessory fermentations will go on. The vibrios of butyric

fermentation, for instance, will propagate with remarkable facility

under these circumstances. Clearly then, the purity of the yeast at the

moment of impregnation, and the purity of the liquid in the funnel, are

conditions indispensable to success.

To secure the latter of these conditions, we close the funnel, as shown

in Fig. 60, by means of a cork pierced with two holes, through one of

which a short tube passes, to which a short length of india-rubber

tubing provided with a glass stopper is attached; through the other hole

a thin curved tube is passed. Thus fitted, the funnel can answer the

same purposes as our double-necked flasks. A few cubic centimetres of

sweetened yeast-water are then put in it and boiled, so that the steam

may destroy any germs adhering to the sides. When cold the liquid is

impregnated by means of a trace of pure yeast, introduced through the

glass-stoppered tube. If these precautions are neglected it is scarcely

possible to secure a successful fermentation in our flasks, because the

yeast sown is immediately held in check by a development of anaërobian

vibrios. For greater security, we may add to the fermentable liquid, at

the moment when it is prepared, a very small quantity of tartaric acid,

which will prevent the development of butyric vibrios.

[Illustration: Fig. 62.]

The variation of the ratio between the weight of the yeast and that of

the sugar decomposed by it now claims special attention. Side by side

with the experiments which we have just described, we conducted a third

lot by means of the flask C (Fig. 62), holding 4·7 litres (8-1/4 pints),

and fitted up like the usual two-necked flasks, with the object of

freeing the fermentable liquid from foreign germs, by boiling it to

begin with, so that we might carry on our work under conditions of

purity. The volume of yeast-water (containing 5 per cent. of sugar) was

only 200 c.c. (7 fl. oz.), and consequently, taking into account the

capacity of the flask, it formed but a very thin layer at the bottom. On

the day after impregnation the deposit of yeast was already

considerable, and forty-eight hours afterwards the fermentation was

completed. On the third day we collected the yeast, after having

analyzed the gas contained in the flask. This analysis was easily

accomplished by placing the flask in a hot-water bath, whilst the end of

the curved tube was plunged under a cylinder of mercury. The gas

contained 41·4 per cent. of carbonic acid, and, after the absorption,

the remaining air contained—

Oxygen 19·7

Nitrogen 80·3

——-

100·0

Taking into consideration the volume of the flask, this shows a minimum

of 50 c.c. (3·05 cub. in.) of oxygen to have been absorbed by the yeast.

The liquid contained no more sugar, and the weight of the yeast, dried

at a temperature of 100° C. (212° F.), was 0·44 gramme (6·8 grains). The

ratio between the weight of the yeast and that of the sugar was,

therefore, 0·44/[114]10 = 1/22·7. On this occasion, where we had

increased the quantity of oxygen held in solution, so as to yield itself

for assimilation at the beginning and during the earlier developments of

the yeast, we found instead of the previous ratio of 1/76 that of 1/23.

[Illustration: Fig. 63.]

The next experiment was to increase the proportion of oxygen to a still

greater extent, by rendering the diffusion of gas a more easy matter

than it is in a flask, the air in which is in a state of perfect

quiescence. Such a state of matters hinders the supply of oxygen,

inasmuch as the carbonic acid, as soon as it is liberated, at once forms

an immovable layer on the surface of the liquid, and so separates off

the oxygen. To effect the purpose of our present experiment, we used

flat basins having glass bottoms and low sides, also of glass, in which

the depth of the liquid is not more than a few millimetres (less than

1/4-inch) (Fig. 63). The following is one of our experiments so

conducted:—On April 16th, 1860, we sowed a trace of beer yeast (“high”

yeast) in 200 c.c. (7 fl. oz.) of a saccharine liquid containing 1·720

grammes (26·2 grains) of sugar-candy. From April 18th our yeast was in

good condition and well developed. We collected it, after having added

to the liquid a few drops of concentrated sulphuric acid, with the

object of checking the fermentation to a great extent, and facilitating

filtration. The sugar remaining in the filtered liquid, determined by

Fehling’s solution, showed that 1·04 grammes (16 grains) of sugar had

disappeared. The weight of the yeast, dried at 100° C. (212° F.), was

0·127 gramme (2 grains), which gives us the ratio between the weight of

the yeast and that of the fermented sugar

[Illustration: Formula 1: 0·127/1·04 = 1/8·1.]

which is considerably higher than the preceding ones.

We may still further increase this ratio by making our estimation as

soon as possible after the impregnation, or the addition of the ferment.

It will be readily understood why yeast, which is composed of cells that

bud and subsequently detach themselves from one another, soon forms a

deposit at the bottom of the vessels.

In consequence of this habit of growth, the cells constantly covering

each other prevents the lower layers from having access to the oxygen

held in solution in the liquid, which is absorbed by the upper ones.

Hence, those which are covered and deprived of this gas act on the sugar

without deriving any vital benefit from the oxygen—a circumstance which

must tend to diminish the ratio of which we are speaking. Once more

repeating the preceding experiment, but stopping it as soon as we think

that the weight of yeast formed may be determined by the balance (we

find that this may be done twenty-four hours after impregnation with an

inappreciable quantity of yeast) in this case the ratio between the

weights of yeast and sugar is

[Illustration: Formula 2: (0^{gr}·024 yeast)/(0^{gr}·098 sugar) = 1/4.]

This is the highest ratio that we have been able to obtain.

Under these conditions the fermentation of sugar is extremely languid:

the ratio obtained is very nearly the same that ordinary fungoid growths

would give. The carbonic acid evolved is principally formed by the

decompositions which result from the assimilation of atmospheric oxygen.

The yeast, therefore, lives and performs its functions after the manner

of ordinary fungi: so far it is no longer a ferment, so to say;

moreover, we might expect to find it cease to be a ferment at all if we

could only surround each cell separately with all the air that it

required. This is what the preceding phenomena teach us; we shall have

occasion to compare them later on with others which relate to the vital

action exercised on yeast by the sugar of milk.

We may here be permitted to make a digression.

In his work on fermentations, which M. Schützenberger has recently

published, the author criticises the deductions that we have drawn from

the preceding experiments, and combats the explanation which we have

given of the phenomena of fermentation.[115] It is an easy matter to

show the weak point of M. Schützenberger’s reasoning. We determined the

power of the ferment by the relation of the weight of sugar decomposed

to the weight of yeast produced. M. Schützenberger asserts that in doing

this we lay down a doubtful hypothesis, and he thinks that this power,

which he terms _fermentative energy_, may be estimated more correctly by

the quantity of sugar decomposed by the unit-weight of yeast in

unit-time; moreover, since our experiments show that yeast is very

vigorous when it has a sufficient supply of oxygen, and that, in such a

case, it can decompose much sugar in a little time, M. Schützenberger

concludes that it must then have great power as a ferment, even greater

than it has when it performs its functions without the aid of air, since

under this condition it decomposes sugar very slowly. In short, he is

disposed to draw from our observations the very opposite conclusion to

that which we arrived at.

M. Schützenberger has failed to notice that the power of a ferment is

independent of the time during which it performs its functions. We

placed a trace of yeast in one litre of saccharine wort; it propagated,

and all the sugar was decomposed. Now, whether the chemical action

involved in this decomposition of sugar had required for its completion

one day, or one month, or one year, such a factor was of no more

importance in this matter than the mechanical labour required to raise a

ton of materials from the ground to the top of a house would be affected

by the fact that it had taken twelve hours instead of one. The notion of

time has nothing to do with the definition of work. M. Schützenberger

has not perceived that in introducing the consideration of time into the

definition of the power of a ferment, he must introduce, at the same

time, that of the vital activity of the cells, which is independent of

their character as a ferment. Apart from the consideration of the

relation existing between the weight of fermentable substance decomposed

and that of ferment produced, there is no occasion to speak of

fermentations or of ferments. The phenomena of fermentation and of

ferments have been placed apart from others, precisely because, in

certain chemical actions, that ratio has been out of proportion; but the

time that these phenomena require for their accomplishment has nothing

to do either with their existence proper, or with their power. The cells

of a ferment may, under some circumstances, require eight days for

revival and propagation, whilst, under other conditions, only a few

hours are necessary; so that, if we introduce the notion of time into

our estimate of their power of decomposition, we may be led to conclude

that in the first case that power was entirely wanting, and that in the

second case it was considerable, although all the time we are dealing

with the same organism—the identical ferment.

M. Schützenberger is astonished that fermentation can take place in the

presence of free oxygen, if, as we suppose, the decomposition of the

sugar is the consequence of the nutrition of the yeast, at the expense

of the combined oxygen, which yields itself to the ferment. At all

events, he argues, fermentation ought to be slower in the presence of

free oxygen. But why should it be slower? We have proved that in the

presence of oxygen the vital activity of the cells increases, so that,

as far as rapidity of action is concerned, its power cannot be

diminished. It might, nevertheless, be weakened as a ferment, and this

is precisely what happens. Free oxygen imparts to the yeast an increased

vital activity, but at the same time impairs rapidly its power as

yeast—_quâ_ yeast, inasmuch as under this condition it approaches the

state in which it can carry on its vital processes after the manner of

an ordinary fungus; the mode of life, that is, in which the ratio

between the weight of sugar decomposed and the weight of the new cells

produced will be the same as holds generally among organisms which are

not ferments. In short, varying our form of expression a little, we may

conclude with perfect truth, from the sum total of observed facts, that

the yeast which lives in the presence of oxygen and can assimilate as

much of that gas as is necessary to its perfect nutrition, ceases

absolutely to be a ferment at all. Nevertheless, yeast formed under

these conditions and subsequently brought into the presence of sugar,

_out of the influence of air_, would decompose more _in a given time_

than in any other of its states. The reason is that yeast which has

formed in contact with air, having the maximum of free oxygen that it

can assimilate, is fresher and possessed of greater vital activity than

that which has been formed without air or with an insufficiency of air.

M. Schützenberger would associate this activity with the notion of time

in estimating the power of the ferment; but he forgets to notice that

yeast can only manifest this maximum of energy under a radical change of

its life-conditions; by having no more air at its disposal and breathing

no more free oxygen. In other words, when its respiratory power becomes

null, its fermentative power is at its greatest. M. Schützenberger

asserts exactly the opposite (p. 151 of his work—Paris, 1875),[116] and

so gratuitously places himself in opposition to facts.

[Illustration: Fig. 64.]

In presence of abundant air-supply, yeast vegetates with extraordinary

activity. We see this in the weight of new yeast, comparatively large,

that may be formed in the course of a few hours. The microscope still

more clearly shows this activity in the rapidity of budding, and the

fresh and active appearance of all the cells. Fig. 64 represents the

yeast of our last experiment at the moment when we stopped the

fermentation. Nothing has been taken from imagination, all the groups

have been faithfully sketched as they were.[117]