My translation of page 81-100 of Kervegant.
In Jamaica, in the manufacture of full-bodied rums, the yield is only 55% of the Pasteur figure. Some losses, due to the formation of acids and aromatic esters, are also unavoidable. Floro, following the observations he has made, divides the various losses as follows: [I think Floro is M.B. Floro who wrote important articles about Jamaica rum in response to Arroyo regarding distillery losses for full flavored rum production.]
Losses that can be reduced
Alcohol in the residual sugars of the must . . . . . . . . . . . . . . 7.65%
lost through evaporation and entrainment . . . . . . . . . . . . . . 6.44
in the bottoms and overflows . . . . . . . . . . . . . . . . . . . . . . . . . 1.78
in the vinasses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0.041
in leaks, etc. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0.293
Losses due to your quality
Losses due to your quality . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17.20%
lost after the end of fermentation and before distillation. . 9.33
esterified during distillation . . . . . . . . . . . . . . . . . . . . . . . . . . . 1.80
Total . . . . . . . . . . . 44.543
In Martinique, yield in the winery usually varies, in the case of molasses rhummeries, between 65 and 80% of the Pasteur figure. It is higher in vesou distilleries, where it reaches 75 to 90%, and even exceptionally 93%. Distillation losses are also quite high. The proportion of alcohol in the vinasse usually varies between 0.5 and 2 p. 1000, ie 1 to 4% of the quantity produced (in well-developed equipment, however, the alcohol content of the vinasse may drop below 0.2%). It is necessary to add the losses in the bottom of the tank, which are thrown to the river. It is admitted that the average yield is, in vesou distillery, from 100 to 55 per ton of cane handled.
Under very favorable conditions, however, it would be possible to go beyond Pasteur’s performance in industrial practice. Thus Russy Grimaud’s beet distillery (1), using the process of yeast recovery, obtained per 100 Kg. of sucrose implemented 63.83 l. pure alcohol, a figure increased by the alcohol lost to evaporation and entrainment by carbon dioxide (1 to 2%) and that it would have been possible to recover by fermentation in closed vats or by washing the gases. Pérard (2), at the distillery of Baleycourt, operating in open vats by the same process of recovery of yeasts, observed the average yield, for the 1936-37 campaign, of 63.97 l. alcohol per 100 kg of sucrose used. In the cane molasses distillery at Puerto Rico, the application of the Arroyo process with thick musts would have been possible with certain vats to reach Pasteur’s yield (see Chapter V).
(1) Various processes for recovering alcohol lost in the vat room: Yeast recovery process. C. R 8. Cons. Int. des Ind. Agr., 1935.
(2) Bull. Ass. Chim. LV, 212, 1938.
Acetic fermentation.
Kutying, in 1837, for the first time attributed the production of acetic acid to the action of a microorganism. This theory, fought by Liebig and Berzelius, was definitively established by Pasteur in 1860. In 1906, Buchner isolated acetic bacteria diastase determining oxidation of alcohol, oxidase.
Mechanism of fermentation.
The reaction which gives rise to acetic acid was first considered to correspond to a simple oxidation, according to the formula:
CH3-CH2OH+02 ―> CH-CO2H+H20
The phenomenon is actually more complex. It is currently accepted that the first stage of the oxidation of ethyl alcohol would be acetaldehyde, which was isolated by Neuberg and North as a combination of sulfite:
CH2 – CH2OH +O2―> CH3-COH + H2O [Need to recheck these atom counts]
According to Neuberg and Windisch, a sort of reaction from Cannizaro would occur, giving rise to acetic acid and alcohol, which would enter the cycle to be oxidized again:
2CH3 – COH+H20 ―> CH3 – CO2H+CH3 – CH3OH
Kluyver and Donker, inspired by the theory of Wieland (1), considering the acetic fermentation as an oxidation-reduction phenomenon.
The two mechanisms seem to intervene one and the other according to the cases, and sometimes even simultaneously, in the organic oxidations (Keilin).
The first step would be dehydrogenation of the alcohol, giving rise to acetaldehyde, which after hydration would in turn undergo dehydrogenation:
Oxygen, necessary for the action of the acetic ferment, acts only as a hydrogen receptor:
2H2 + O2 ―> 2H2O
The conversion of ethyl alcohol to acetic acid could be achieved anaerobically by replacing the acetic bacteria with a hydrogen receptor (platinum black, palladium, methylene blue).
The theory of Kluyver and Donker helps explain the rapid progress of fermentation. The reaction indicated by Neuberg and Windisch, however, remains possible, but would only be a side reaction.
In addition to aldehyde and acetic acid, secondary products are still poorly known, such as fixed (lactic, succinic) or volatile acids (caproic, valerianic, etc.) during acetic fermentation. and esters (ethyl acetate, etc.). On the other hand, some bacteria completely oxidize the alcohol, giving carbon dioxide and water. [secondary products are where there may be a great benefit to rum.]
Acetic ferments can also attack alcohol other than ethyl alcohol, as well as sugars. The oxidizing power varies greatly depending on the species. Propyl and butyl alcohols are converted into propionic and butyric acids, glycerin into glycerol acid and dioxygenone. In sugars, the aldehyde function is transformed into an acid function: for example, glucose gives gluconic acid and, with certain bacteria, oxalic acid. Sucrose, levulose, and generally ketones are not attacked (Bertrand and Watermann).
Acetic ferments.
Acetic bacteria are formed by net-shaped cells, often joined in chains and generally immobile. They usually develop on the surface of alcoholic liquids in obligatory aerobiosis, forming at first thin veils, then becoming more or less thick, transparent or opaque, oily or dry, more or less wrinkled. In the young state, the veil is difficult to wet; once old, it breaks easily and can immerse itself in the liquid, forming a mucilaginous mass (mother of vinegar). They do not form endospores and do not liquefy gelatin.
There are many species of acetic bacteria, differentiated by the size and shape of cells, the nature of the veil formed, the appearance of colonies on solid media, physiological properties (acid and alcohol resistance, optimum temperature, oxidizing power, etc…).
The species found in distillery musts are generally weak producers of acids, they do not tolerate alcohol well, but are able to oxidize many sugars. They most often give thick veils.
Sugar cane juices contain several species of acetic bacteria. Tanaka, at Formosa, was able to isolate the following, each represented by several varieties: Acetobacter (Bacterium) xylinum Brown, A. acetosum Henneberg, A. Lindneri Henneberg; Bacterium aceti Brown, B. curvum Henneerg; Gluconoacetobacter liquefaciens, G. Asai.
Ashby has encountered two distinct species in Jamaican distilleries:
a) A bacterium that appears quickly in fermented musts with low or high acidity. It forms a delicate blue veil, which becomes white afterwards, but always remains fragile. In a glass container, the veil rises along the walls, above the surface of the liquid. The bacterium comes under the aspect of short, thick sticks, united in chains of short length and appearing yellow or yellowish brown when dyed by iodine. It resembles Bacterium Kutzingianum Hansen, except that it does not stain blue with iodine.
b) A bacterium which produces a veil on the surface of liquids of cartilaginous consistency, very resistant. It is in the form of long, narrow sticks, staining blue with iodine and sulfuric acid, and can be identified with Bacterium xylinum Brown.
The first of these ferments multiplies vigorously when the alcohol content is between 7 and 9%. It gives about 4% of acetic acid, after a fortnight (7.5% maximum). B. xylinum, on the contrary, can not develop when the alcoholic richness reaches 7°; it produces about 3% acetic acid in liquids containing 4.5% alcohol. It is therefore found mainly in fermented cane juice, while the previous species is found in molasses musts containing 6% alcohol or more.
According to Watts and Tempany, the acidification which affects the freshly expressed cane juice, before the alcoholic fermentation has developed (surissement du jus), is due to the action of an anaerobic ferment, directly attacking the sugar. The acidity formed accounts for about 2/3 of the fixed acids and 1/3 of the volatile acids, of which acetic acid predominates.
Importance in the distillery.
Acetification is a frequent condition in distiller’s musts. In the old days, when the conditions of its development were poorly known, it caused considerable losses, lowering the alcohol yield and spoiling the quality of the product obtained (excessive content of spirits in acetic acid).
The main factors that promote acetic fermentation are:
a) high fermentation temperatures, which reduce the vigor of alcoholic yeasts and are instead favorable to acetic bacteria. Optimum temperatures of propagation generally vary between 30° (Bacterium xylinum) and 36° (B. aceti);
b) insufficient initial acidity (especially in the case of vesou musts), which is detrimental to the activity of the yeast;
e) the low alcohol content of fermented musts;
d) the use of contaminated yeast or the lack of cleanliness of the vat room.
The acetic ferments, when they do not exert an action too predominant, can intervene usefully in the production of the bouquet of the spirits, especially those with developed aroma. Some practices used in rhummeries are intended to promote their development. This is the case of the preliminary treatment of defecation foam and cane juice in the presence of bagasse, in the manufacture of grand arôme rums in Jamaica. It should also be noted that in the past, producers of agricultural rum in Martinique sometimes abandoned the cane for several days after cutting before handling it, in order to obtain a more robust eau-de-vie.
Butyric and acetonobutyl fermentation.
The first observations on butyric fermentation were made by Pasteur (1861). The production of butyl alcohol by fermentation, reported by Fitz in 1876, was studied by Grimbert (1893) and Beijerinck. Duclaux showed that butyric and butyl fermentations were caused by the same organisms: when the artificially neutral medium is maintained by the addition of chalk, butyric acid is produced, whereas if the culture is allowed to acidify freely, normal butyl alcohol is formed. Finally, in 1912, Fernbach and Schoen found that besides butyl alcohol, acetone appeared in a definite proportion. [this may offer a clue to the usefulness of buffers in rum ferments.]
Mechanism of fermentation.
The mechanism of butyric fermentation is very complex and still poorly understood.
It is generally accepted that the degradation of the sugar molecule follows the same course as in the alcoholic fermentation, up to the acetaldehydestage. Then, the hydrogen, instead of being fixed on the aldehyde formed, stabilizes in the molecular state and emerges from the medium at the same time as the carbon dioxide. The acetaldehyde undergoes a condensation which transforms it into aldol, or β-oxybutyric aldehyde:
In a neutral medium, the hydrated aldol undergoes an intramolecular oxidation-reduction, which results in butyric acid:
In acidic medium, according to Schoen, there would be, oxidation-reduction of 2 molecules of aldol to give alcohol β-oxybutyl and β-oxybutyric acid, by the reaction of Cannizaro.
Oxybutyric acid would be oxidatively converted to acetoacetic acid, and oxybutyl alcohol, by concomitant reduction, to butyl alcohol. Finally, by decarboxylation, acetoacetic acid gives acetone and carbon dioxide:
The general formulas of the reactions would be the following:
a) Butyric fermentation:
(b) acetonobutyl fermentation:
In fact, these formulas are not exclusive. In addition to the main phenomenon, there are deviations from fermentation, which give rise to various secondary products: ethyl and isopropyl alcohols, acetic, formic and propionic acids, etc.. The substances formed vary in nature and quantity according to the ferment considered, the fermentable material, the reaction of the medium, the stage of the fermentation (at the beginning of the acetonobutyl fermentation, a little butyric acid always occurs).
By way of example, the following composition of the products obtained by Buchner and Mesenheimer (1) can be obtained by fermenting, by Bacillus butylicus Fitz, 100 grams of glucose in the presence of chalk and salts:
(1) Ber, Deut. Chem. Ges. XLI 1410, 1910.
Butyric ferments.
Butyric ferments take the form of large batons, often polymorphous (one finds forms in club, spindle, etc.), and are movable. They sporulate easily keeping for a long time easily as spores (even at temperatures of 100-105°). These ferments are generally anaerobic, but in addition to obligatory anaerobes, facultative anaerobes (Aero-bacillus genus) have been found, occuring optionally in response to circumstance. They prefer neutral or alkaline media and high temperatures (35°-40°).
True butyric ferments use ammonia as a nitrogen source. This differentiates them from putrefying bacteria, which attack proteic materials. There are, however, between these two groups transitional forms.
Butyric bacteria can attack most carbohydrates, including starch. Cellulose, however, is not modified except by quite special species. The sugars are degraded either by oxidation or much more frequently by fermentation.
Butyric ferments are widespread in nature. They are encountered in milk, manure, soil, beet and cane juice, candy bars, and so on. There are many species, differentiated by their action on various sugars and proteic materials, by nature, and the amount of by-products they form, etc. Among those best studied are the following species:
—Bacillus butyricus Pasteur.— It determines the fermentation of lactate of lime, the milk having previously undergone the action of the lactic ferment, of glycerine, of certain sugars, giving rise, as principal products, to butyric acid, hydrogen and carbon dioxide in varying proportions. [I believe he is talking about lime water used in nixtamalization for making tortilla chips and stuff like that.]
—Clostridium butyricum Prazmowski.— Anaerobic and giving mostly butyric acid; it is the species that is most commonly found in distillery musts.
—Bacillus butylicus Fitz.— It ferments sucrose, glucose, mannite and glycerin, giving hydrogen, carbohydrates, lactic acid, succinic, butyric acid, butyl alcohol and a little ethyl alcohol.
—Bacillus orthobutylicus Grimbert, who ferment sugars, inulin, dextrin and starch, giving hydrogen, carbon dioxide, these butyric and acetic acids and normal butyl alcohol.
—Bacillus tetryl Arroyo.— Discovered by Arroyo on the roots of the Kassaer cane in Puerto Rico. It has been used industrially for the manufacture of acetone and butanol from cane molasses. [This likely relates to Arroyo’s patented work before joining the experiment station.]
Clostridium saccharolyticum Bergery, whose presence has been reported by Hall, James and Nelson in Barbados cane syrup.
Clostridium saccharo–butyricum Arroyo.— Found in Puerto Rico on annatto seeds and used for the production of butyric acid from cane molasses. It gives, according to Arroyo, in addition to hydrogen and carbon dioxide, about 93% of normal butyric acid, 4.1% of acetic acid, 1.9% of propionic acid and 1% of higher fatty acids (caproic acid, heptoic acid, etc.). It does not produce appreciable quantities of alcohols, aldehydes or ketones. The activity of the bacterium is stopped when the concentration of sugars in the medium exceeds 6 gr per 100 cc. or when the alcohol level reaches 8% by volume. It is the same when the pH goes down to around 4.0. [Very cool, I don’t think I know where this information comes from but hopefully it gives clues that will produce a paper.]
Importance in the distillery.
The distillery raw materials normally contain butyric ferments, in the state of bacteria or, more often, spores (molasses). The activity of these microorganisms is annihilated, when the fermentation is carried out with pure yeasts or in the presence of antiseptics. On the other hand, they can play an important role in spontaneous fermentations, especially when they have a long duration. Their intervention results in a decrease in alcohol yield, and also in the production of higher alcohols (butyl, propyl, etc.) and volatile acids (butyric, propionic, formic, etc.), which contribute to the formation of the bouquet of the eaux-de-vie.
Allan was able to isolate in Jamaica’s rhummeries, and more particularly liquids from the “muck hole”, various butyric bacteria. He found that if they do not get along well with cane juice alone, they develop vigorously in the addition of sugar solutions made of albuminoidal substances, as well as in vinasse with the addition of yeast extract. This last medium corresponds substantially, from the point of view of composition, to the must obtained by mixing vesou or molasses with vinasse. The bacteria grow well from 26° C, but have the optimum temperature 35°. The quantities of acids formed however remain rather low (0.3 – 0.4%), if the must is not neutralized, by means of lime for example. [always be buffering]
According to the same author, the bacteria predominate in the tanks towards the end of the long-term fermentation in distilleries producing grand arôme rum, to the point where the yeasts of the fermented liquid disappear completely. The intervention of butyric bacteria explains that the fusel-oil of Jamaican rum is formed essentially by normal butyl alcohol.
Finally, Arroyo, recently in Puerto Rico, by fermenting musts of cane molasses with Pombé yeast in symbiosis with Clostridium saccharobutyricum, was able to obtain a rum having all the characteristics of Jamaican rums, as regards the bouquet and the chemical composition. This author has observed that by cultivating the bacterium in symbiosis with yeast, its multiplication and the formation of alcohol were considerably accelerated. The duration of the fermentation of the yeast, from 70 to 96 hours in the case of yeast alone, was reduced to 28-48 hours. The author attributes this fact to the action of radiation emitted by the bacterium and comparable to the mitogenic rays of Gurwitsch (1).
(1) Mitogenetic radiation, discovered by Gurwitsch (Arch, Mikrosk Anat. und Entro Mech. C. 11, 1923), are issued by certain living organisms at certain stages of development. They pass through quartz, but not glass, and when they meet with other growing tissues, they can act upon them, accelerating their development or reproduction. Arroyo has found that various bacteria can emit similar rays, acting on the speed of multiplication and the zymogenic power of the yeasts, even when the bacterium is separated from the yeast by a wall of quartz.
Lactic fermentation.
It was Pasteur who isolated, in 1857, the first lactic ferments. Today we can distinguish between true lactic ferments, which produce lactic acid almost alone, and lactic pseudo-ferments, which at the same time give rise to significant quantities of various by-products (acetic acid, ethyl alcohol, CO2, H, etc.).
Mechanism of fermentation.
It was once thought that the production of lactic acid from hexoses was by simple splitting, according to the formula:
C6H12O6 —> 2C3H6O3
But, as during the lactic fermentation, several of the bodies isolated in the alcoholic fermentation (hexose-phosphate, acetaldehyde, methylglyoxat) have been found, the mechanism of these two fermentations is now considered to be analogous.
According to Neuberg, sugar would give rise, through hexose phosphates, methylglyoxal hydrate. This would then be transformed into lactic acid, by intramolecular oxidation-reduction (caused by glyoxalase):
According to Meyerhoff, lactic fermentation could be compared to the production of lactic acid in the muscle, under the action of lactic zymase (isolated from the muscle in 1927). The evolution of sugars would be the same as in alcoholic fermentation, up to the pyruvic acid stage. It would then undergo an oxidation-reduction, in the presence of phosphoglycerol, to give lactic acid and phosphoglyceral:
The phosphoglyceral thus formed would react on a second molecule of pyruvic acid, to give still lactic acid and phosphoric acid, which would enter the reaction to regenerate the pyruvic acid.
This mechanism is that of true lactic fermentation. The transformations are much more complex and poorly known, in the case of pseudo-lactic ferments, which produce at the expense of sugars large quantities of gas (CO2 and H) and give rise to various other acids (acetic, succinic, formic) and ethyl alcohol.
Lactic ferments.
These ferments, very numerous, are in the form of batons of variable size or cocci, isolated, united two by two diplobacilli, diplococci), or arranged in chains more or less long (streptobacilli, streptococci); they are immobile and do not form spores.
Some species are anaerobic, others are aerobic or anaerobic, others are indifferent to oxygen. They are sensitive to acidity: they produce only 2% maximum lactic acid and generally much less. They prefer neutral environments. Not sporulating, they are usually destroyed by heating at 65-70° for 5 minutes. The optimum growth temperature may be relatively high for some species (40-50° for bacteria of the genus Thermobacterium), lower for others (30°). True lactic ferments require peptones for their nitrogen nutrition; the lactic pseudo-ferments are content with amino acids or with ammoniacal salts. Very common in nature, they are found in milk, distiller’s musts, manure, etc.
The lactic ferments found in the distillery belong especially to the group of lactobacilli, anaerobic rod-shaped bacteria, often joined by 2 or in chains, able to withstand high doses of lactic acidity and giving only traces of products other than lactic acid.
Lactabocillus Delbruckii Leichman and L. Lindneri were isolated from cane molasses.
The first of these organisms is in the form of rods of 2.7 mus to 8 mus long and 0.4-0.7 mu wide, either isolated, or combined 2 by 2 or sometimes very long chains. It does not grow in milk and thrives especially well in non-hopped wort and distillery musts. Optimum temperature 45°. The amount of acid formed is up to 1.6%. Acidification is slowed by 4% and stopped by 10% alcohol.
Lactic acid bacteria meet quite frequently in rhummerie musts (Ficker and Szügs). They would predominate in the fermentation of defecation scums (Ashby). A representative of the group of pseudo-lactic ferments, Leuconostoc mesenteroides, is easily developed in musts of vesou or molasses with neutral or alkaline reaction.
Lactic acid bacteria must generally be considered as a fermentation disease, the development of which is favored by high fermentation temperatures, insufficient acidification of musts, lack of cleanliness. However, secondary products (acetic, formic, etc.) to which various species give rise can play a useful role in the production of the aroma.
In the distillery of starchy material, lactic acid bacteria were used to obtain the acidification of musts. These are left to spontaneous lactic fermentation, at high temperatures (50-55 °), or sometimes seeded with a pure culture of Lactobacillus Delbruckii. [Sour mash whiskey process]
Mycodermic fermentation
In this fermentation, which occurs easily if the vats are left to their own after the end of the alcoholic fermentation, the alcohol is completely oxidized, with production of water and carbon dioxide.
The agents of this fermentation are asporogenous yeasts, of the genus Mycoderma. These are most often in the form of elongate cells, cylindrical, transparent protoplasm and vacuolized, showing a tendency to remain united in thin chains. They are aerobic and form on the surface of liquids, from the beginning of the fermentation, a folded veil, filled with air bubbles (levures à voile).
Mycoderms are very common in the air and live mostly in liquids containing alcohol. Many species have been described. Of particular note is Mycoderma cerevsiae Desm, which is found in breweries and alters beer as well as Mycoderma vini Desm, which causes the flower of wine.
While some species do not tolerate more than 1%, others support 15% alcohol. Mycoderms prefer mediums rich in organic matter. They poorly support acidity, their growth usually being stopped by 2% acetic acid. However, they can attack organic acids (acetic acid, malic acid, tartaric acid, etc.) if the concentration of these acids is low. Rarely, do they ferment some sugars, giving a little alcohol.
The main products of fermentation are water and carbon dioxide. Secondary products include small amounts of organic acids (formic, succinic, maline, acetic, etc.) and esters (ethyl acetate).
The mycoderms, if they play a harmful part in burning the alcohol formed, may in certain cases intervene happily, producing aromatic principles. It is to these ferments that the flavor of certain sherry wines is due, which age several years in barrels unfilled and covered with Mycoderma vini. Note also that in rhummerie, to obtain a more “bouquetéd” brandy, the vats are sometimes allowed to “blanch”, that is to say, to cover themselves with a mycodermic veil, before proceeding to their distillation. In the manufacture of certain grand arôme rums, it is the rule to abandon the vats for 3 or 4 days after the end of the alcoholic fermentation.
Putrid fermentation.
The putrescent ferments are distinguished from the ferments previously studied by the fact that they cause an alkalinization of the medium: the nitrogenous matters are decomposed first into amino acids, then into various volatile acids (carbonic, sulphohydric, acetic, propionic, butyric, valeric, etc.), which combine with the produced ammonia. There is also release of aromatic or heterocyclic products. Aerobic bacteria push further degradation than anaerobic species. The nature of the attacked amino acid, the conditions of the fermentation intervene, at the same time as the microbial species, to modify the nature of the products formed.
Many putrid ferments also attack sugars and some of them, which are transitional species with lactic and butyric ferments, give rise to free fatty acids. The action of these organizations on sugars is, however, very limited.
We usually divide fermentations of putrefaction in anaerobic bacteria, which develop in depth, and in aerobic bacteria that form veils on the surface of liquids. The former are close to butyric pseudo-fermentations, of which they are distinguished by their property of attacking albuminoid substances. The latter are mainly represented by the groups Bacillus fluorescens, B. proteus, and B. subtilis.
Fluorescent bacteria, which are very common in water, air and the upper layers of the soil, are mobile, non-sporulating, usually gram-negative, indol-producing.
Those of the proteus group are generally mobile, very polymorphic and not sporulating. They attack energetically the proteic matters and ferment the sugars, with production of hydrogen, carbon dioxide, acetic and succinic acids. The presence of the ferments of this group has been reported in the manufacture of Jamaican rum, and in particular in the liquid of the “muck hole” (Allan).
Bacillus subtilis Ehrenberg is a mobile, rod, quite often polyphobic, gram-positive; it produces very heat-resistant spores and has an optimum development temperature of about 40°. The final molasses of Cuba, along with B. subtilis properly so called, have been found to contain Bacillus vulgatus Migula, B mensentericus Trevisan, B. mensentericus fuscus Flugge, B. liodermos Flugge, and B. levaniformans Greig Smith.
Putrid bacteria usually have a very minor importance in the distillery. Although they are commonly found in certain raw materials (molasses), their action on sugars and albuminoid substances remains very limited, because of their sensitivity to acids; it is rare to see putrefying bacteria withstand a pH below 4.
In some cases, however, they can intervene by destroying a certain amount of sugar and giving rise to aromatic products acting on the bouquet of the brandy, favorably (production of certain volatile fatty acids), or unfavorably (production of H2S, indol, etc.). It would play a role in the production of Jamaican grand arôme rum (Allan, Cousins).
Gummy fermentation.
In this fermentation, the sugar juices are transformed into a gelatinous and viscous mass, under the action of various microorganisms which produce, from the sugars, gummy substances known under the names of dextran and levulan. These substances have been assigned the same crude formula as cellulose, (C6H10O5)n, but they correspond, according to some authors, to hydrated products of variable composition.
Dextran is a colorless gum that swells in water and dissolves in alkaline liquors. It is dextrorotatory (αⁿ=+200°) according to Scheibler and gives by hydrolysis of glucose.
Levulan has the appearance of a gum which becomes brittle by dessication. It melts around 250°, dissolves in hot water and gives by cooling, a gelatinous mass. Its rotatory power is equal to 221°. It provides by hydrolysis of levulose. It does not reduce, no more than dextran, Fehling’s liquor.
Frai de grenouille. [frog eggs]
The most well-known microorganism producing gums is Leuconostoc mesenteroides Van Tieghem, which determines the affection for a long time known as gomme de sucrerie, or frai de grenouille. This disease, reported as early as 1822 by Vauquelin in bottles of cane juice shipped from Martinique in France, appears both in beet sugar (mainly when sugar beets are not ripe enough) and in cane sugar (Java, Hawaii, Cuba, Louisiana, etc.). In Louisiana, canes damaged by frost often undergo, at the arrival of warm weather, gummy fermentation.
The affected sugar juices are transformed into compact gelatinous masses composed of insoluble, colorless or pink lumps, impaled in a viscous liquor. These masses, of firm and elastic consistency, resemble frai de grenouille.
Leuconostoc mesenteroides is a streptococcus, in the form of spherical grains surrounded by a glairy sheath (dextran). In the young state, it affects the appearance of a string of small grains surrounded by gelatin, then long refractory puddings, which curl irregularly while keeping in their axis the rosaries of grains that gave birth to them. The different tubes, curling up on themselves, form mucous bodies, constantly enlarging, and whose contoured surface resembles that of the brain, then masses more and more voluminous. The fermentation terminated, the grains soften and the rosaries break up.
Leuconostoc, which has been found in water and soil by Berijerinck, prefers alkaline media; its development is slow in neutral or acidic juices. The optimum temperature is around 36°; however, it develops mainly at relatively low indoor temperatures at 20°, unfavorable to the action of yeasts (Owen).
Leuconostoc attacks sucrose and glucose, with production of dextran, mannite and fatty acids (lactic, etc.) with lactose and maltose, it gives rise to lactic acid, but without forming a sheath. It secretes sucrase, which inverts to sucrose.
According to Sacchetti, the “gomme de sucrerie” is due to the symbiosis of two microbes, Leuconostoc mcgenteroides and Bacillus vulgatus Migula, giving birth first to dextran and the second to levulan, and a yeast, Saccharomyces cartilaginosus Lindner. This symbiosis was confirmed by Monoyer, who described the yeast associated under the name of Torulopsis Canbresieri Mon. n. sp. Leuconostoc is found in the central part of the gummy particles, the bacillus in the middle part, while the yeast forms an outer crust, which protects the bacteria against the action of dissolved oxygen in the liquid.
Various species of Coccaceae similar to Leuconostoc have been studied by Schöne, Zettnow, and Gonnerman. They are differentiated by their size, their behavior on solid or liquid media, the nature of the products formed, etc.
Other producers of gum.
Various other microorganisms that produce gums from sugar have been described. Let’s mention the following:
—Bacterium gelatinosum beta Glaser, which forms a mucous veil on the surface of beet juice, much like that produced by Leuconostoc. It liquefies gelatin and interchanges sucrose, Optimum temperature: 40°.
—Bacillus viscosus sacchari Kramer;
—Semiclostridium commune Maassen, which forms levulan at the expense of sucrose, but does not attack the invert sugar.
—Bacillus levaniformans Greig Smith. Found by Greig Smith (1) in 1901, in cane juice and raw sugars in Queensland, the microbe, like the previous one, attacks only sucrose, with levulan production. The same is true of Bacterium gummosum Ritsert.
(1) J. Chem. Ind. XXI, 1381, 1902.
Semiclostridium commune forms sheaths like Leuconostoc, but the others do not produce them.
Following Owen (2), the bacteria of the group mesentericus (bacilli of the potato): Bacillus mesentericus, B. vulgatus. B. liodermos, acquire the property, if grown for several generations in saccharide media, to transform sucrose into levulan. These microbes, which are normally found in the soil, can acquire this property in the neighborhood of sugar houses. Introduced in the juice by the bagasse and the particles of earth adhering to the canes, they are found in press cakes and final molasses, because they are likely to resist, in the state of spores, at the high temperatures of sugar manufacture. Press cakes are commonly used as fertilizers and the molasses bacteria also return to the soil when they are used for feeding animals, resulting in a gradual acclimatization of the enzymes to sugary solutions and a dissemination of these organisms in the soil of cane plantations.
(2) J. Bacteriology VIII, 421, 1923.
Owen considers that many gum-producing microorganisms, described by the authors as distinct species, are actually only forms derived from the bacteria of the mesentericus group. This would be the case with Bacillus levaniformans, which would become B. mesentericus levaniformans.
These ferments produce levulan directly from sucrose: the previous inversion of it by sucrase decreases and slows down the formation of gum. They prefer alkaline media, the optimum pH being between 6.7 and 7 (Owen).
Cellulose production.
Browne reported that in Louisiana, an aerobic fermentation giving rise to cellulan is quite common. Cane juice forms gelatinous masses, which can weigh several pounds and have a stratified appearance.
The membranes constituting the strata are of cellulosic nature. Under the microscope, they appear to be formed by chains of bacteria, intermingled with streptococci embedded in a cellulosic sheath. Bacteria are usually mixed with yeast cells.
The cellulan producing organism is very similar, if not identical, to Bacterium xylinum.
Fermentation by the mucédinées [molds].
The presence in cane molasses of many moldy sugars has been reported. Church and Thom, for example, found the following species: Aspergillus niger Van Tieghem, A. flavus Link, A. nidulans Winter, Aspergillus sp. (blue); Citromyces spp., Penicillium expansum, P. diverticatum, P. luteum, P. roseum, P. purpurogenum.
Molds develop on the other hand as parasites on sugars, sometimes even on vats and walls of poorly maintained distilleries.
These organisms attack a variety of foods. They can notably metabolize sugars, giving carbon dioxide and water. Between this final term and the ternary compounds, we find various intermediate bodies, corresponding to incomplete oxidations: saccharic, oxalic, citric gluconic acids, etc. These acids usually appear only fleetingly and are soon metabolized in their turn. But under certain conditions, they can remain in the liquid and contribute to the formation of the bouquet.
Among the most interesting products of this fermentation are the oxalic acid, which appears mainly in nitrogen and sugar-rich media (15-20%), and citric acid, which is produced now industrially by fermentation of molasses (Cahn process).
Some molds, mainly belonging to the family Mucoraceae, can saccharify starchy materials and even, under certain conditions, give alcohol. They are at the base of the fermentation processes practiced in the Far East. In this part of the world they play the same role as malt in Europe. [koji]
Particularly noteworthy are the following species:
—Mucor (Amylomyces) Rouxii Wehmer, ferment of the annamite yeast (same), used in the preparation of annamite rice alcohol or Ru’o’on (vulgarly choum-choum). It is also found in the yeast used in China.
—Aspergillus (Eurotium) oryzae Cohn, ferment of Japanese yeast (koji), used in the fermentation of a rice beer (sake).
—Mucor Prainii Chodat et Nechitch, encountered in the yeasts of India (ranu, murcha).
—Mucor oryzae Went, Mucor javanicus Wehmer and Rhizopus oryzae Went and Prinsen-Geerligs, microorganisms of ragi, yeast used in the Dutch Indies, in the preparation of Batavia arak.
These molds, living aerobically, transform the starch of grains and riz into glucose. Some of them (M. Rouzii, M. javanicus, etc.). when grown in anaerobiosis, can also produce alcohol, but low amounts (5 to 7% alcohol maximum) and very slowly. The transformation of sugar into alcohol is especially the work of yeasts, which still exist in abundance in the yeast of the Far East.
The properties of Mucors in distillery of starchy materials have also been used in Europe, and more particularly in the manufacture of grain alcohol (Amylo process, developed by Collette and Boidin). Amylomyces Rourii was first used, followed by various other more active or resistant Mucors (Mucors B, Boulard, Delemar, etc.).
Nitrous fermentation.
Nitrous fermentation is quite common in juice and beet molasses distilleries. It results from the development of reducing bacteria, which decompose the nitrates of the must, giving mainly nitrogen gas and nitrites. The latter are themselves decomposed by the acids with production of nitrogen dioxide, which is transformed in contact with air into vapors sparkling with nitrogen peroxide: the tank is covered with yellowish bubbles. Butyric ferments can also cause, thanks to the hydrogen they emit, the reduction of nitrites to nitrates and the formation of nitrogen peroxide.
Nitrous fermentation has not been reported in Cannes distillery, Bassières (1) however noted in Martinique a case of “nitriform fermentation”, where the vats were covered with large bubbles of a yellow rust and a kind filth of the same color. He could not show the presence of nitrites.
(1) in Kervégant – L’industrie Rhumiere à la Martinique, Bull. Agr. Martinique 11, 29, 1933.
CHAPTER IV
THE YEASTS (1)
(1) Guilliermond (A) et Tanner (F. J.) – The Yeasts, New York, 1919.
We commonly call yeasts all microorganisms which, placed in a sugar solution, give rise to alcohol and carbon dioxide, that is to say determine the alcoholic fermentation.
In the botanical sense of the word, we thus designate unicellular fungi, having oval or round forms, multiplying by budding or fission and producing ascopores [spores] (Guilliermond). Yeasts are the family Saccharomycetaceae, which forms with that of Endomycetaceae, the group of Protoasca, or Ascomycetes inferior. [not sure if the last italicized terms are conjugated properly.]
In addition to these well-characterized yeasts, there are others that never give asci. It is difficult to say whether they are true yeasts that have lost their ability to produce spores, or whether they represent forms derived from higher fungi and fixed in the yeast state. Therefore, they are classified in a provisional group, referred to as Non-Saccharomycetaceae.
Finally, there are mycelial fungi that can give birth to budding cells in the form of yeasts, which can multiply in turn for several generations by budding. In other fungi, the mycelium disarticulates into rectangular cells (anthrospores), capable of continuing to divide by transverse partitioning like Schizosaccharomyces. These organisms are usually referred to as yeast-shaped fungi.
Yeasts are very common in nature: they are found in the air, soil, on the surface of plants (fruits, leaves, etc.), especially when they contain sugar. In cold and temperate regions they overwinter in the soil, but in the tropics they persist year-round on plants. They are found especially on the surface of cane stems, represented by many species (Saccharomyces, Torula, etc.) and accompanied by molds (Aspergillus, etc.) and bacteria (acetic, butyric, lactic).
Morphology, development and composition.
Yeasts are generally isolated cells, very polymorphic (round, oval, elliptical, lemon, pear, baton, etc.) measuring 1 to 9 mus long and 1 to 5 mus broad. Shape and dimensions vary with the age of the yeast and the culture medium acidity for example determines the elongation of the cells, while the oxygen makes them become oval or globular. Therefore, it is not possible to rely solely on morphological characters to identify species. There is, however, often in a crop a predominant form, sometimes characteristic (apiculate yeasts, torulas, etc.).
The reproduction of yeasts is usually done by budding, but it can also be done by scissiparity [fission] or by sporulation.
In multiplication by budding, a small bud is formed on the surface of the yeast globules, which grows little by little and soon takes on the size of the mother cell. The nucleus divides by amitosis. Young cells detach themselves before they have reached their full development, or remain adherent for some time; then each cell starts to proliferate in turn. When the globules remain united in bundles or rosaries which may have 15-20 cells, on a top fermenting yeast, which rises to the surface of musts, raised by carbon dioxide, and forms a sort of hat. When, on the contrary, the globules are isolated or united in pairs, they constitute a bottom fermenting yeast, which remains at the bottom of the fermentation vats. [ale versus lager]
The yeasts of the genus Schizosaccharomyces are characterized by their multiplication by scissiparity: the cell elongates and, having reached a certain size, forms a median partition, diversely directed, which divides it into two daughter cells. These may separate immediately or remain united for some time, constituting excrescences in the form of very elongated skins. Under certain conditions (air deficiency in particular), they have a marked tendency to remain adherent and to form branched chains.
When the fermentation is finished and the liquid has become immobile, the yeast globules may continue to develop aerobically: they form on the surface of the liquid a veil, a crown, or a ring on the line of contact of this surface with the receptacle. The conditions of formation of the veil (speed, temperature limits, etc.), as well as the appearance of it, constitute characters making it possible to differentiate the species. In some cases, the veil appears at the beginning of fermentation (Willia, Mycoderma, etc.); in others, it is not formed at all.
Spores are yeast resistance organs against external agents. It is admitted, with Hansen, that they are formed only from young and vigorous cells, placed in unfavorable dietary conditions, in the presence of oxygen and between certain temperature limits, variable with the yeast races. Spores are found in the veils, and can be easily obtained by placing young, well-nourished yeast in some solid media. The number of spores per cell, or asci, varies between 1 and 12, and is often characteristic of a race or species of yeast. Their dimensions range from 1.5 to 5 mus. Ascospores are usually spherical or oval, but sometimes have characteristic forms: hemispherical, triangular, limoniform, etc. The mode of germination of the spores is also sometimes characteristic of the species.
In some yeasts, asci results from copulation of two cells of equal size (isogamy) or unequal (heterogamy). In the first case, two similar cells unite with one another by a thin canal; the spores form in the two bulges of the ascus, which looks like a dumbbell. In the second case, a small cell unites by a channel to a large cell; the contents of the first emigrate in the second, where the ascopores are born.
Isogamic conjugation is found in the genera Shizosaccharomyces and Zygosaccharomyces; heterogamic conjugation in the genera Deburyomyces, Nadsonia and some Zygosaccharomyces. In addition, there are intermediate cases between iso and heterogamy. Some yeasts (Schwanniomyces, Torulaspora) present only remnants of sexuality: the cells can make several attempts to join together, without achieving it, and give rise to several filaments, which radiate around them. Finally, in some species (s. Ludwigii for example), when the conjugate can not take place at the time of the formation of asci, a copulation occurs between ascopores (parthenogamy).
The chemical composition of the yeasts is very variable according to the conditions of culture and the race studied. For a given yeast, it also changes, at least quantitatively, as the fermentation progresses.
The water content is always high and around 70-75%. The composition of the dry matter varies between the following limits:
The most important carbohydrates are glycogen, a reserve substance that can reach 38% at the end of fermentation, and mannane found in the cell membrane.
The fat content generally reaches 2-5% of the dry matter. It can, however, rise to 20% in old degenerate crops and even, for some breeds, up to 50%. The fats of the yeast are mainly composed of the esters of palmitic, lauric, linolic and arachidic acids. There are also small quantities of lecithins and phytosterols (ergosterol).
Nitrogenous or proteinaceous materials are 90% formed by true proteins (cerevisin, zymocasein) and nucleins. Non-proteinaceous nitrogenous materials include peptones, amino acids (leucine, valine, lysine, etc.), and amides (xanthine, hypoxanthine, guanine) Meisenheimer (1), who studied nitrogenous yeast materials by hydrolysis in the presence of toluene, found among the protein degradation products, all common amino acids, including glucosamine, which was not until then to detect the presence. According to this author, the yeast nitrogen would be distributed as follows:
(1) Woch, Brauerei XXXII, 325, 1915.
As for mineral substances, they consist mainly of phosphoric acid (45 to 60% of ash), potash (30 to 40%) and magnesia (4 to 8%). Lime, soda, silica, iron, sulfur and chlorine are also present in small quantities. The proportion of the various substances above varies, moreover, within wide limits, according to the chemical composition of the medium in which the yeast has developed. This observation applies especially to lime and magnesia, which may be very abundant or in very small quantities.
Yeast Nutrition.
Like all living beings, yeast needs, for its maintenance and development, nitrogen, hydrocarbon and mineral matter. It is important, moreover, to distinguish the action of these substances on the multiplication of the cell on the one hand, and on the zymatic function, that is, on fermentation, on the other. There is frequently antagonism between plant function and ferment function.
Mineral matter.
The role played by mineral materials in the multiplication of yeasts was first studied by Mayer (2). This author has found that the most favorable nutritive medium had the following composition, which corresponds, with regard to the equilibrium of the various mineral elements, to that of the ashes of the yeast:
(2) Lehrbuch der Gärungschemie. 5° éd. 1902.
Phosphates and potash are of paramount importance in the nutrition of yeast. Sulfur is also essential.
Lime and magnesia, which can not be replaced by each other, do not seem absolutely necessary, but nevertheless play a role in the internal chemistry of the cell which renders them extremely useful. In fact, it has been found that yeast degenerates rapidly in lime-free environments, Hayduck and Kenneberg have also observed that, placed in a solution of pure sugar, the yeasts of beer die very quickly, but that if one adds in the middle a small quantity of salts of lime, their vitality is much greater. It is likely that the death of the cells is determined by the formation within them of the acids, which are neutralized in the presence of lime or other alkalis. [always be buffering!]
Mayer considered iron salts useless. However other authors (Molich, Wehmer) consider that they exert a favorable action on the multiplication of the yeast.
Phosphates and sulfur also have a considerable influence on the zymatic function. Elion has been able to observe that monopotassium phosphate and neutral phosphate of ammonia produce an increase in the release of carbonic acid, which varies, for a given time, between 23 and 63% according to the yeasts. Harden and Young have shown that soluble phosphates play an essential role in the action of alcoholic zymase.
Stern has observed that in a nutrient medium completely deprived of sulfur, complete fermentation of the sugar can not be achieved. But if Ca sulphate and Mg sulphate are added, the transformation of the sugar is complete.
When they exceed a certain proportion in the culture medium, the mineral elements do not intervene any more usefully in the nutrition of the yeast, nor in the fermentation. Stern observed that beyond 250 mgr. per liter of sugar solution (added with 250 mg of nitrogen in the form of asparagine), there is no longer any increase in the quantity of nitrogen assimilated, the weight of yeast nor the sugar consumed.
Independently of the above elements, recognized as essential for the development and the zymatic function of yeast, there are other mineral salts which, in moderate doses, exert a favorable action. It has thus been recognized that the ferment power is increased by small doses of manganese salts (Kayser and Marchand), tin protochloride and bismuth sub-nitrate (Gimel), copper sulphate and sodium cyanide (Hildebrandt and Boyce), etc. … Aluminum salts also have a slight stimulating action on fermentation and cell multiplication (Sikes), while lithium salts are harmful.
Nitrogenous substances.
The proteins themselves (albumins, casein, fibrin), not diffusible through the cell membranes, constitute bad nitrogenous foods. According to Pasteur and Mayer, yeasts can not assimilate egg white albumin or blood fibrin. Under certain conditions, however, complex nitrogenous materials could be used. Thus, Boullanger found that the milk, seeded with certain yeasts, coagulates little by little and that after a few months the coagulum is liquefied, with formation of ammoniacal salts, tyrosine and leucine: there was thus dissolution and digestion of casein.
On the other hand, albumins and peptones are good foods for yeast. Duclaux (1) has shown that the nitrogenous substances contained in the yeast water, and which consist mainly of peptones, would even have more favorable action on the multiplication of the cells than the ammoniacal salts. Hayduck, experimenting with asparagine and peptone, found that the latter favored the multiplication of yeast much more than asparagine.
(1) Traité de Microbiologie – t. III. La fermentation alcoolique. Paris, 1905.
The more advanced degradation products of albuminoids (amides, amino acids, etc.) are assimilated more easily than albumins. However, assimilability varies with the nature of these substances. Some amino acids, such as leucine, isoleucine, adenine, aspartic acid, are easily absorbed, while others (histidine, choline, thymine, hypoxanthine) are difficult to absorb. Of the amides, allantoin, asparagine and urea are assimilated, but not creatine, creatinine or succinamic acid.
According to Lindner (1), compounds whose hydrocarbon groups are in long open chains (leucine, adenine, lysine) are more easily absorbed than those with a closed chain (histidine, thymine, choline). The yeast race is also of great importance: the most aerobic yeasts use nitrogen compounds that are difficult to assimilate more easily.
(1) Chem. Ztg XXXIV. 1144, 1910.
The carbon of amino acids and amides can not be used for yeast nutrition. Ehrlich showed that the nitrogen was absorbed, probably in the form of ammonia, and transformed into proteic matter, while the rest of the molecule was released into the medium in the form of higher alcohols. Effront has also reported the existence of amidase, which acts on amides, giving rise to ammonia and volatile acids.
The ammoniacal salts are very well absorbed by yeasts, as found by many experimenters (Pasteur, Duclaux, Mayer, etc.)
In the presence of a mixture of ammoniacal nitrogen and organic nitrogen, the yeasts preferably first attack the ammoniacal salts. But these, as Ducluaux has shown, have a much more favorable effect on the functioning of zymase than on cell multiplication. Here are the increases in dry yeast obtained with various nitrogenous materials (Bokorny) (2):
(2) Chem. Ztg. XL, 366, 1916.
In contrast to higher plants, nitrates are poorly used by yeast. According to Laurent, they could not be assimilated and, by transforming themselves into nitrites, they would exert a toxic action on the yeasts. However, the experiments of Kayser (3) and those of Ferebach and Lanzerberg (4) have shown that if nitrates mediate cell multiplication, they activate the action of zymase and promote fermentation. According to Nicoleau (5), nitric nitrogen shows its accelerating action on yeast ferment from a nitrate concentration of about 5%, with a variable optimum depending on the medium, the initial quantity of seed, the vigor yeast, etc.
(3) C. R. CXLIV, 574 1907, CLI 816. 1910.
(4) C.R CLI. 726, 1910.
(5) Les nitrates dans la vie de la levure. Paris 1924.
As a final result of the studies that have been done on the subject, the foods of choice for the development and multiplication of the yeast are the complex nitrogenous materials in the form of peptones, while for the exercise of the functions of fermentation, this the nitrogenous matter is deeply degraded and as close as possible to the form of ammonia.
Above a certain proportion, the nitrogenous materials have an unfavorable effect on the fermentation. According to Pringsheim, the quantity of nitrogen giving the best results in terms of alcohol yield varies, depending on the yeast races and the conditions of the environment, between 0.004 and 0.008%.
Hydrocarbon materials or carbohydrates.
The hydrocarbon metabolism of yeasts is different, depending on whether they live in aerobiosis (plant function) or in anaerobiosis (fermentative or zymatic function).
In aerobic life, yeast can use many organic compounds as maintenance foods. According to Laurent’s research (1), it is capable of assimilating the carbon of organic salts (acetates, lactates, citrates, tartrates, malates, succinates), organic acids (citric, tartaric, lactic succinic), ethyl alcohol and polyalcohols (glycerol, mannite), C6 and C12 sugars, and bodies capable of giving sugars, glucosides or dextrins. On the other hand, esters, fatty acids (in the form of acids), amides, glycine, hydroquinone, cellulose, etc. could not feed the yeast.
(1) Ann. Inst. Pasteur TII, 1889.
Bokorny came to similar conclusions. It seems, however, that under certain conditions and for certain species, yeasts may use ethyl alcohol (Trillat, Kayser, Lindner).
Maltose appears to be the sugar most easily assimilated by aerobic yeast; sucrose, glucose, levulose, raffinose are less easily, while lactose and dextrins are used only in special cases. There is no relation between the fermentability of a sugar and its assimilability in aerobiosis: Schizosaccharomyces. Pombe, for example, which actively ferment sucrose, glucose and levulose, is unable to assimilate them (Linder and Saito) (2).
(2) Woch. Braurei XXVII, N° 41. 1910.
In an aerobic or fermentary life, yeast attacks only certain sugars and the polysaccharides giving rise to them by hydrolysis.
According to Fischer and Thierfelder (3), only sugars containing 3 carbon atoms (or a multiple of 3) are fermentable. However, there appear to be exceptions to this rule: pentoses (arabinose, xylose), for example, may be attacked by certain yeasts (Lindner). On the other hand, the trioses appear to be transformed into hexoses before any fermentation. Virtually only the following 3 hexoses: d-glucose, d-fructose (levulose) and d-mannose, which are directly and easily attackable. The fermentation of d-galactose is only obtained with yeasts acclimated to this sugar.
(3) Ber. Deut. Chem. Ges. XXVII, 2114, 1894.
Yeast shows a marked preference for one or the other of the fermentable hexoses (phenomenon of the elective fermentation). In a mixture of glucose and levulose, for example, it is found that the glucose usually disappears more rapidly than the levulose at the beginning of the fermentation. Subsequently, the phenomenon is reversed, so that finally, there remains in the liquid an excess of glucose. Some breeds, however, are an exception: Sauternes yeasts, described by Dubourg, make levulose disappear more rapidly than glucose from the beginning to the end of fermentation. The elective property also depends on the constitution of the culture medium and the temperature. Thus, in the presence of manganese salts, levulose disappears faster than glucose (Kayser).
Disaccharides (sucrose, maltose, lactose, trehalose) and trisaccharides (raffinose) are attacked only if the yeast possesses the necessary diastases for their hydrolysis in fermentable hexoses. Most species ferment sucrose, much maltose, but little attack raffinose and trehalose.
As for the polysaccharides (starch, dextrins, inulin, etc.), they can undergo alcoholic fermentation under the action of certain fungi (Mucors), but they are only attacked quite exceptionally by the yeasts. It should be noted, however, that Schizosaccharomyces Pombe and Schizosaccharomyces mellacei, for example, ferment dextrin and inulin.
Ferment power and yeast activity.
The term “ferment power” designates the ratio of the quantity of sugar consumed to the quantity of yeast produced, that is to say the quantity of sugar which the unit of weight of this yeast is capable of making disappear. This power varies according to the race of yeast and the conditions of the culture medium. Certain breeds, especially those used in the manufacture of industrial alcohol, can thoroughly push the fermentation of must rich in sugar while others stop before having processed all the sugar.
The ferment power represents, as Lindet (1) points out, the sum of the vegetable power and the zymase power, that is to say, the quantities of sugar used by the yeast for its maintenance and its development on the one hand; for the fermentation of alcohol under the action of zymase on the other hand. These two powers are complementary, so that the weight of alcohol formed decreases when the amount of sugar used for the functioning of plant life increases. Lindet has shown that the part of the vegetable function is of so much more importance, and that of the zymatic function all the weaker, that the fermentation is slower. The lower the amount of yeast at the beginning, the longer the fermentation is, and the lower the yield of alcohol. However, when we exceed a certain limit (1 per 1000 yeast supposedly dry), we obtain a very rapid fermentation, but then the maintenance and breathing of an excessive number of cells determine an exaggerated consumption of sugar in the vegetable phase.
(1) C. R. CLXIV, 58, 1917 : CLXVI, 910, 1918.
The activity of a yeast is the amount of sugar that the unit weight of the yeast makes disappear in the unit of time. This activity also varies with yeast races and with environmental conditions.
Finally, a yeast is said to have low or high attenuation limit, depending on the amount of fermentable sugars remaining in the must at the end of the fermentation is more or less important. If we compare their action in sweet liquids of various compositions, the races of yeast always rank in the same order. This made it possible to group them industrially in yeasts with low attenuation (Saaz type), medium attenuation (Frohberg type) and high attenuation (Logos type, Pombe yeast). [schizosaccharomyces]
The attenuation power of a yeast is not only due to its ability to ferment carbohydrates, but also to its resistance to increasingly unfavorable environmental conditions as and when that the fermentation advances (increase of acidity, depletion of the must, etc.). “In short, the attenuation limit of a must does not correspond to the disappearance of any fermentable material, the stopping (at least apparent) of the fermentation is due to a set of factors that slow down the phenomenon more and more energetically” (Van Laer).
In the distilleries of industrial alcohol, preference is given to yeasts whose ferment power and activity are high, that is to say which give in a short time a high production of alcohol, It also seeks to achieve the conditions that bring power to a close and activity to their maximum. This is not always the case in the manufacture of eaux-de-vie, where, in order to obtain aromatic products, it is often advantageous to slow down the fermentation. Sometimes even in the fermented beverages industry, it is advisable to address low attenuation yeasts, leaving in the liquid a certain quantity of unprocessed sugar (making sweet cider, for example).
Diastases.
The living cell’s chemical reactions (hydrolyses, oxidations and reductions, various condensations) are made by means of “biochemical catalysts”, called diastases or enzymes.
These are divided into two major groups: digestion diastases or hydrolases, which hydrolyze food, and respiration or fermentation diastases, desmolases, which carry out the oxidation and reduction phenomenon, and disruption of organic molecules.
In the first group, there are esterases, which cause esterification and saponification, and glucidases, which hydrolyze carbohydrates and nitrogenases, which hydrolyze proteins. The best known of the desmolases is the alcoholic zimase.