Glycerol in the alcoholic fermentation of molasses and sugar cane juice

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Parfait A., Jouret C., 1980. Le glycérol dans la fermentation alcoolique des mélasses et des jus de canne à sucre. Industries alimentaires et Agricoles 97, 721-724.

Le glycérol dans la fermentation alcoolique des mélasses et des jus de cannes à sucre
by PARFAIT A. *, JOURET C. **
with technical collaboration from G. SABIN and Madame G. MIGLIORI

(*) C.R.A.A.G. Station de Technologie, Guadeloupe.
(**) C.R.A. de Toulouse, Laboratoire de Technologie des Produits Végétaux, Auzeville.

Summary

Authors have specified the influence of various factors (mode of conduct, pH, seeding rate, sugar content, yeast species) on the amount of glycerol found in rums.

They have also shown that glycerol can serve as a carbon substrate for various bacteria and give, as a result, derivatives having a negative role on the organoleptic qualities of this eau-de-vie.


Glycerol is a secondary product formed by the metabolism of sugars during alcoholic fermentation.

Depending on the conditions, a more or less important fraction of the sugars is thus transformed into glycerol. This results in different values of the fermentation yield.

Genevois (1936) proposed an equation between various byproducts of alcoholic fermentation

2A + B + 2M + H + 5S = ε = G

A, B, M, H, S and G being respectively the molar concentrations of acetic acid, 2-3 butane diol, acetoin, acetaldehyde, succinic acid and glycerol. This equation has been the subject of much work and has been confirmed by Lafon (1955). It can be considered valid in 90% of fermentations.

Subsequently, Nordstrom (1968) and Oura (1977) showed that there was a correlation between the redox potential of the fermenting medium and the formation of glycerol.

According to Oura, the formation of succinic acid is related to the production of glycerol and has the same purpose: to balance the excess of reduced nucleotides.

Glycerol, whose physiological interest for yeast appears to be small, seems however to play a significant role in the regulation of compounds such as pyruvic and succinic acids that enter, instead, in the formation of constituents of the cell. Similarly, glycerol, via its phosphoric ester, L-α glycerophosphate, combines with activated fatty acids to give phosphatidic acid. This last body leads to lipids. This same ester allows the use of glycerol by many breeds of yeasts as a source of carbon. Although the low volatility of glycerol explains its absence in rums, it has been possible to determine various compounds from its degradation. These generally have a negative effect on the organoleptic qualities of eaux-de-vie.

Since the work of Warcollier and Le Moal (1932) on ciders and those of Serjak et al. (1954), we attribute to the action of lactic acid bacteria the appearance of acrolein in spirits.

Dubois et al. (1973) identified two acrolein derivatives in an abnormal taste rum: ethoxy-3-propanol and ethoxypropane. These compounds are not directly responsible for the unpleasant flavor of the rum studied but can serve as indicators.

Smedt and Liddle (1976) have correlated the presence of allylic alcohol (2propene 1 ol [I think that is correct nomenclature]) with some bad tastes in various types of spirits. They also showed a relationship between the contents of this alcohol and those of ethoxypropane.

Thus, the glycerol which is at the origin of acrolein (and products derived from this aldehyde) following metabolic pathways not completely elucidated, can therefore be degraded in fermenting media based on cane juice and molasses.

Given these biochemical and technological considerations, it seemed interesting to specify some parameters of the production of glycerol in the fermentation of the basic products of the different types of rums.

Experimental Protocol

Glycerol was determined according to the enzymatic technique of Eggstein and Kuhlmann (1974) after defecation of natural media with lead acetate.

Samples of fermented media were taken from industrial plants previously described by Sabin and Parfait (1975). Just remember that the following raw materials cane juice, syrup and molasses, are used respectively for the development of agricultural rums, syrup and industrial.

The results are reported in Table 1.

Table 1.
Glycerol content expressed in g / l in fermentation media of different types of rums

399/5000
Following these measurements, tests were conducted in the laboratory to specify the glycerol formation conditions according to the pH, the fermentable substrate concentration, the yeast seeding rate, the species and the yeast strains.

The growing conditions were as follows:
—Molasses: 300 g / l,
—seeding rate: 1 g /,
—fermentation temperature: 30 ° C.

The seeding was carried out using yeast creams in order to eliminate the glycerol fraction that could be brought by the starter.

The yeast strain saccharomycès cerevisiae used is the No. 493 of our collection, isolated from a natural fermentative medium of agricultural rum.

Influence of pH:
Initial pHs were set at 3.5 – 4.0 – 4.5 – 5.0 – 5.3.

The evolution of the glycerol level during the fermentation was regularly monitored.

For example, for the medium at pH 40, the following figures were noted:

These figures vary very little according to the different pHs tested.

Influence of molasses richness:

The molasses concentrations of 150 g/L, 200 g/L, 250 g/L and 300 g/L were varied, the seeding rate was 1 g/L, the fermentation temperature 30° C and the initial pH set to 5.

The glycerol levels found were in order: 1.5 g/L, 1.7 g/L, 2.3 g/L and 2.5 g/L. They follow the same progression as that of sugars.

Influence of seeding rate:

By changing the seeding rate from 0.25 g/L, 0.50 g/L, 1.0 g/L, 2.7 g/L and 2.5 g/L, 4.0 g/L, 5.0 g/L in a medium similar to the previous one, does not find a statistically valid variation in the final glycerol contents.

Influence of yeast species isolated from natural fermentation media: Hansenula anomala, Saccharomyces cerevisiae, Saccharomyces aceti, Schizzosaccharomyces pombe.

Only these last yeasts have glycerol production curves very different from those of the other yeasts tested.

Influence of the mode of conduct of industrial fermentations:

By examining the manufacturing process generally followed in the industrial production of rum, we realized that glycerol appears during the aerobic growth phase of yeasts. Given the reduced richness of the medium, often less than 200 g/L of molasses or about 100 g/L of fermentable sugars, and the low rate of seeding practiced, it can be said that currently in the French West Indies, a part not negligible sugar is consumed to develop glycerol.

On the other hand, the operation of industrial installations is discontinuous. The canes brought to the distillery may be subject to pre-fermentation, with consequent production of glycerol. The glycerol level reaches 0.8 g/L on average in pipes that are poorly emptied.

Glycerol derivatives

Yeasts and bacteria can use glycerol. For these latter microorganisms, a review was conducted by Lin (1976).

Ganou and Parfait (1980) determined many species of bacteria in the flora of fermentation media leading to the various qualities of rums. As can be seen from Table II, cane juice and molasses, even when preserved, contain few germs. From the first hours of fermentation, a flora of varied origin (installations, water, environment) develops. During the course of fermentation, anaerobiosis causes a reduction in the number of species present in the medium. Lactic acid bacteria and Clostridia are mainly found. Acetobacters may appear at the end of fermentation and degrade the ethanol formed.

If sometimes the intervention is beneficial (some strains of clostridia, among others, for the production of rum aroma) most often it is detrimental to the organoleptic qualities. Acetobacters cause, for example, a detrimental increase in the level of acetic acid and ethyl acetate.

The appearance of volatile derivatives of glycerol is due, for a large part, to the activity of lactic acid bacteria, as we have observed in some distilleries. We searched in the lab, among the species of lactic acid bacteria that we had isolated, those that degraded glycerol. The tests were done aerobically and anaerobically.

Three culture media were used: M1, M2, M3.

TABLE 2

The kinds of bacteria found in fermentative environments. (+) = present, (-) = absent. The number of signs indicates the frequency. A, B and C represent the environments leading to agricultural rum, molasses rum and rum grand arôme

Under the conditions of our tests, some lactic acid bacteria use glycerol as a carbon source. Surely we could identify among them strains of leuconestoc mesenteroid. Other species also having a metabolic activity from glycerol are being identified.

Acrolein and 2propene ol 1 were found among the products formed by gas chromatography using a Tracor 560 with flame ionization detectors. The phase for the 50 foot Scotch column and 0.2 inch diameter used is carbowax 1540; the flow rate of nitrogen, carrier gas, is 3 ml/min. Temperature programming was carried out: 6 minutes at 58° C. and then an increase of 8°C. per minute up to 120°C.

The injection of 1 μL of rum can detect 1 ppm acrolein or 2propene ol 1.

A typical chromatogram is given in Figure 1.

Conclusions

In the production of rum, glycerol may be found in greater or lesser quantities depending on the mode of conduct of fermentation operations.

If the raw material (fresh juice or molasses) has not been the subject of microbial activity, in particular by yeasts, the glycerol contents will be very low.

The use of a leavening tank causes a significant concentration of glycerol from the beginning of the anaerobic phase. Schizzosaccharomyces cause the appearance of significant amounts of glycerol, which can cause problems when there is a risk of contamination by bacteria degrading this substance.

Indeed, lactic acid bacteria (leuconostoc mesenteroid type) can metabolize glycerol to lead in particular to acrolein and 2propene ol 1 found in rums with other products of negative organoleptic character.

These observations should guide the process of fermentation of raw materials to obtain a good quality rum.

BIBLIOGRAPHY

DUBOIS P., PARFAIT A., DE KIMPE J. (Mme), 1973. – Présence de dérivés de l’acroléine dans un rhum à goût anormal. Ann. Technol. Agric., 22, 131-135.

EGGSTEIN M., KUHLMANN E., 1974. – In methods of enzymatic analysis (Bergmeyer H.L.), Vol. 4, 1825-1835, « Verlag Chemie Weinheim ».

GANOU B. (Mme), PARFAIT A., 1980. – Les microorganismes de fermentation de mélasse et de jus de canne (en préparation).

LAFON M. (Mme), 1955. – Contribution à l’étude de la formation des produits secondaires de la fermentation alcoolique. Thèse de Docteur en sciences physiques, Bordeaux, Ann. Techn. Agri., 198 p.

LIN E.C., 1976, – Catabolisme du glycérol et sa régulation chez certaines bactéries. Annal Review of microbiology, vol. 30.

NORDSTROM K., 1968. – Yeast growth and glycerol formation II carbon and redox balances. J. Inst. of brewing, 74, 429.432.

OURA. E., 1977. – Reaction products of yeasts fermentations. Process Biochemistry, 12, 19-35.

SABIN G., PARFAIT A., 1975. — Les fermentations traditionnelles de mélasse et de jus de canne aux Antilles Françaises, lnd. Agric. Alim., 92, 27-34.

SER JAK W.C., DAY W.H., VANLANEN J. M., BORUFF C.S., 1954. – Acrolein production by bacteria found in distillary grain mashes, Applied Microbiology, 2, 14-20.

DE SMEDT P., LIDDLE P.A.P., 1976. – Présence d’alcool allylique (2propène  ol 1) et dérivés dans les eaux-de-vie. Ind. Alim. Agric., 93, .41-43.

WARCOLLIER G., LE MOAL. A., 1932. – Présence accidentelle d’acroléine dans l’eau-de-vie de cidre. C.R. Acad. Science, 194, 394.

Ethyl Esters Of Higher Fatty Acids Of Rhums

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Parfait A., Namory M., Dubois P., 1972. Les esters éthyliques des acides gras supérieurs des rhums. Annales de Technologie Agricoles 21, 2, 199–210.

ETHYL ESTERS OF HIGHER FATTY ACIDS OF RHUMS

A. PARFAIT, M. NAMORY and P. DUBOIS *

Station de Technologie végétale, I. N. R. A.,
Petit-Bourg (Guadeloupe)
*Station de Technologic des Produite vegetaux, I. N. R. A.,
21034 Dijon Cedes

Summary

Rums of good quality, especially in ethyl esters, generally have high fatty acids with a high content of volatile esters, and particularly of ethyl esters of higher fatty acids (from C8 to C16). These esters are by-products of alcoholic fermentation, such as higher alcohols, and behave like them during continuous distillation.

The levels of ethyl esters of the higher fatty acids are higher when the distillation is done on the lees, when cane wax is added to the must before fermentation and by selection of a species, or even of a strain, of yeast.

The ethyl ester contents of the distillate fatty acids were three times higher with Saccharomyces cerevisiae S. 132 than with Saccharomyces cerevisiae Berlin II. The highest content was obtained with a strain of Schizosaccharomyces Pombe yeast from the sugar cane regions.

It has not been possible to establish definitively a correlation between the fatty acid composition of the lipids of the yeast cell and the ethyl esters produced.

Key words: rums, ethyl esters, higher fatty acids, distillation.

I- INTRODUCTION

The eaux-de-vie can be considered as hydroalcoholic solutions of a “non alcohol” which characterizes them and that the chemical analysis makes it possible to separate in dry extract, acids, aldehydes, esters and higher alcohols, These analyzes can be completed by olfactory observations on isolated fractions by distillation. Finally, much finer separations can be obtained by gas chromatography for the determination of volatile constituents and by other chromatographic methods for the study of nonvolatile compounds.

In the case of rums, there may be a relationship between their quality and their ester content (KERVEGANT, 1946) and, in spite of many exceptions, it seems that high ester levels characterize the most aromatic rums. Observations made after fractional distillation even suggest that heavy esters have the greatest influence on the aroma of these eaux-de-vie, as is the case with whiskeys (SALO et al., 1972). The technology of rums should therefore be able to take advantage of recent studies on the formation of esters by yeasts during the fermentation of wines and beers.

A.—Mechanism for the formation of esters

PEYNAUD (1956) has shown that the levels of ethyl acetate in fermented media depend on the yeast species, and that they are always higher than those predicted by the calculation from chemical equilibrium reactions.

According to NORDSTRÖM (1964), the esters are formed by alcoholysis of the acyl-coenzymes A according to the reaction

and their formation depends on the contents of acyl-Co A and alcohols (RAINBOW, 1970) The alcohols react all the better as they are primary, with a linear chain and of lower molecular weight. Since ethyl alcohol is the most abundant, the esters formed are mainly ethyl esters.
On their side, acyl-Co A have three modes of formation.

Activation of fatty acids in the presence of ATP.

In view of the very small quantities of free fatty acids in fermented media, it seems that this reaction can play only a minor role.

Oxidative decarboxylation of α-cetanol acids.

This reaction is thought to be responsible for most of the acetyl-Co A, by oxidative decarboxylation of pyruvic acid. The other α-keto acids, that is to say certain intermediates of the metabolism of sugars and of amino acids, are present in much smaller quantities than pyruvic acid, and very few esters of the corresponding acids are formed. (propionic, isobutyric, methyl-2 and 3-methyl-butyric).

Reaction between an acyt-Co A and malonyl-Co A.

This reaction, which leads to the formation of the fatty acids of the lipids of constitution of the yeast, is also at the origin of the ethyl esters of linear fatty acids with an even number of carbon atoms.

Ester formation is related to yeast growth, as is that of higher alcohols and the formation of higher fatty acid esters is more particularly related to lipid metabolism.

It is therefore, like that of lipids, favored by the presence of pantothenic acid, as constituent of coenzyme A, and of the biotin which participates in the carboxylation of acetyl-Co A in malonyl-Co A, and which therefore, competes with the formation of ethyl acetate.

Factors that limit the development of yeasts have an inhibitory role. This is particularly the case of linear fatty acids having 6 to 10 carbon atoms which are toxic to yeasts.

B.— Fermentations in rhummeries

We know imperfectly the flora that develops in the environments put in fermentation to produce the different types of rums. HARRISSON and GRAHAM (1970) point out, in a development, that the budding yeasts gradually replaced, in Jamaica, a fission yeast, Schizosaccharomyces melacei, which was dominant at the beginning of the century.

Overall, fission yeasts are preferable to budding yeasts, in part because they promote the development of butyric bacteria (Clostridium saccharobulyricum, in particular) that produce very large quantities of esters. These fission yeasts are particularly abundant in the flora used for the elaboration of “grand arôme” rums.

It is also possible that yeasts of the genus Torulopsis play an important role in the formation of esters from sugars.

In the French Antilles, Saccharomyces cerevisiae is the main agent of fermentations. Other yeasts are present, including Pichia, Hansenula, Candida and Schizosaccharomyces. Their role is difficult to assess in practice.

C.— Role of distillation

Esters are poorly soluble in water and behave like head products in low alcohol environments. They behave, at the same time, as bottoms in the distillation columns when the alcoholic degree reaches values ​​of the order of 50 to 60 ° GL. Only ethyl acetate is always in the heads. Butyrate and ethyl hexanoate distill substantially at the same time as ethanol. The ethyl esters of acids that have 8 carbon atoms and more distillate after ethanol and it is these acids that we consider here as superior.

The contents of the rums in the ethyl esters of the higher fatty acids are therefore related to the alcoholic degrees to which they are distilled in the columns. The higher this degree, the poorer the rum obtained in these esters. It is interesting to note that the behavior of higher alcohols is quite similar to that of heavy esters.

In any case, the analytical results obtained on rums, and in particular the quantitative results obtained by LIEBICH et al. (1970) (Table I) shows that the main esters are the ethyl esters of fatty acids number by carbon atoms. Ethyl acetate and butyrate dominate among the light esters, caprate and palmitate among the heavy esters. Certain unsaturated acid esters are also present, but in smaller proportions.

Table I
Ethyl esters reported in rums

After having measured the ethyl esters of higher fatty acids in commercial rums, various factors have been studied that may affect the levels of these constituents in rums: presence of yeasts during the distillation fermentation in the presence of cane wax fermentation by various species of yeast. These different points were completed by the analysis of lipids of yeast constitution.

II- MATERIAL AND METHODS

A.— Culture media

Two media were used, one is based on cane molasses, the other synthetic.

These solutions are brought to pH = 5 with sulfuric acid and sterilized by treatment at 110 ° C. for 35 minutes.

B.— Yeast uses

Apart from Schizosaccharomyces pombe, which was isolated from a fermentation of cane juice medium, the other species came from the Dijon Plant Products Technology Station collection: Saccharomyces cerevisiae strains Berlin II and S. 132, Pichia membranaefaciene, Hansenula anomala and Candida krusei. These yeasts are those which have been reported as participating in the fermentation of musts in rhummeries.

With a young culture of yeasts, flasks containing 100 ml of liquid Wickerham medium are inoculated with malt. After about 24 hours at 28 ° C, in stirred medium, the yeasts are collected by centrifugation, then they are rinsed twice with saline water. All media are inoculated so that they initially contain 5 · 10 ^ 6 yeasts per ml. A magnetic bar agitates the first day and fermentation lasts 3 to 4 days at 28 ° C.

C.— Analysis made on distillates

The apparatus to be distilled is made of glass and includes a balloon, an electric heater, a column of Vigreux of 50 cm of height and a refrigerant.

Ethyl esters of fatty acids of 8 to 16 carbon atoms are not very polar and their most selective solvent must itself be polar. We chose pentane.

A test portion of 100 ml of distillate at 50 ° GL is stirred vigorously with 50 ml of pentane. The addition of 100 ml of water causes an immediate demixtion without emulsion formation. The organic phase is dried over anhydrous sodium sulphate and brought to a tenth of a milliliter by distillation. Two microliters are injected into a Perkin-Elmer model 880 chromatograph under the following conditions: column filled with Chromosorb G 80/100 mesh impregnated with OV 17 at 5 p. 100, length 4 meters, diameter 3 mm. Nitrogen flow 25 ml / min. Temperature program of the injection from 160 to 240 ° C at 2 ° C per minute. Since the rums contain only traces of ethyl pelargonate, this analysis is made quantitative by the addition of one milliliter of an alcoholic solution of this ester at 1 mg / ml to the 100 ml of distillate used. Previous tests performed on synthetic solutions showed that the relative errors were less than 5 percent. The exact nature of the esters was verified in the chromatographic conditions, but in coupling with a Varian *** type CH 5 mass spectrometer (250° C source, 70 eV electron energy). All these ethyl esters are characterized by a rearrangement peak for m / e = 88,

and by their molecular peaks. Mass spectrometry also makes it possible to observe the absence of important interferences.

D.— Analyses faitee sur les levures

After fermentation, the yeasts are separated by centrifugation (10,000 g). They are twice washed with physiological water, and then harvested at the bottom of the tubes using a spatula. At weighing of the wet mass, the dry matter content is determined on an aliquot part. [aliquot was a guess. The scanning is cutting off the edge for this page].

The total lipids of the yeasts are determined by the method of KAHANE and ROUS (1961) and then saponified by the alcoholic potash. Unsaponifiable matter is extracted with petroleum ether. After acidification, the fatty acids are extracted with the same solvent, then they are esterified with hydrochloric methanol and analyzed by gas chromatography on an impregnated column [guess] of diethylene glycol succinate (impregnation rate 15%). 3 meters, inside diameter 3 mm, Chromosorb G 80/1000 mesh, temperature 190 ° C.

E.— Autres dosages

The yeasts are counted with a cell count on a possible dilution of the medium. The reducing sugars are dosed, after acid hydrolysis, by the method of Bertrand. Finally, the volatile acids are determined after entrainment by steam. [I think I translated that last part correctly.]

III. — RESULTS

A.— Levels of ethyl esters of the higher fatty acids of commercial rums

The contents of these esters of five rums were compared: two agricultural rums obtained from cane juice in a traditional factory where fermentations are most often spontaneous, and three rums obtained from industrial rum distillates at 64 ° GL, a light rum distilled at 94 ° GL, and a rum “grand arôme” distilled at 63 ° GL as industrial rum, but obtained by a fermentation procedure that uses a starter with yeasts of the genus Schizosaccharomyces and bacteria.

The results obtained are shown in Table 2.
The first three rums have very similar contents in these esters, light rum contains only traces, the most abundant being the *** of ethyl whose content is of the order of 0.05 mg / liter of pure alcohol. The numbers obtained for the rum “grand arome” are much weaker than were expected. This rum has a very low content in total esters (about 2 g / liter of pure alcohol).

Table 2

Ethyl esters of various commercial rums
(in mg per liter of pure alcohol)

B.— Role of yeasts during distillation

The synthetic medium and the molasses-based medium were used with yeast Saccharomyces cerevisiae Berlin II. In each test, four liters of medium are prepared and fermented, then they are divided into two equal parts, one of which is centrifuged before distillation to eliminate the yeasts.

The results reported in Table 3 show that distillation in the presence of yeasts leads to a significant gain in esters. These compounds are poorly soluble in low alcohol environments and are likely to be absorbed on the surface of yeasts and suspended particles.

TABLE 3

Role des levures et de la cire de canne sur les teneurs des distillats en esters ethyliques des acides gras superieure
(n-C8 n-C16 en mg par litre d’alcool pur)

C.— Role of cane wax

The wax is the richest part of the higher fatty acids of the cane (*** according to MARTIN and JUNIPER, 1970), and it was interesting to see what could be its influence.

It was added in the form of an emulsion at a dose of 0.300 g per 4 liters of medium. The results (Table 3) show that this addition of wax allows a medium based on cane molasses to double the levels of distilled ethyl esters of fatty acids with 8 and 10 carbon atoms which are among the interesting ones on the olfactory plane ( SALO et al., 1972).

The synthetic media are incompletely fermented and the results obtained are unusable.

D.- Role of the yeast species

EL SHEHATA (1960) has shown that molasses musts are fermented, in practice, by the following yeast species: Saccharomyces cerevisiae, Schizosaccharomyces pombe, Hansenula anomala. Pichia membranaefaciene and Candida krusei. The first two ferment well in anaerobic environment, the last two do not ferment sucrose.

TABLE 4

Ethyl esters of distillates obtained with different yeast species
(in mg per liter of pure alcohol)

Yeast species used:
Pichia membranacfaciens (P. m.), Hansenula anomala (H. a.), Schizosaccharomyces pombe (S. p), Candida krusei (C. k.), Saccharomyces cerevisiae S. 132 (S. c. 132).

We have fermented, under the conditions defined above, five mediums based on molasses with respectively each of the following yeasts:

– Pichia membranaefaciens (P. m.)
– Hansenula anomala (H. a.)
– Schizosaccharomyces pombe (S. p.)
– Candida krusei (C. k.)
– Saccharomyces cerevisiae S. 132 (S. c. 132).

After fermentation, the media are distilled in the presence of yeasts. The results are summarized in Table 4.

Some media have been incompletely fermented because Pichia and Candida do not ferment sucrose.

E.- Fatty acid content of yeasts

The determinations were made on yeasts derived from fermented media based on molasses similar to the preceding ones, but, to promote the multiplication of yeasts, the stirring period was increased to 48 hours instead of 24. Tables 5 and 6 report on the results obtained during various determinations.

TABLE 5

Data on the fermentation by different species of yeasts in molasses medium

TABLE 6

Comparative composition of lipids of various yeast species

There appears to be some correlation between the amount of ethyl esters of the higher fatty acids produced by these yeasts and their levels of saturated fatty acids. However, the methods used do not make it possible to know if the fatty acids doses were glycerol related in the lipids of constitution, or if they were in the form of ethyl esters and associated with the yeast walls.

IV.— DISCUSSION

The levels of ethyl esters of the higher fatty acids of commercial rums are close to those we were expecting to find except for the “grand arôme” rum. On rum of this type Liebich et al. (1970) indicate contents of the order of 170 mg per liter of pure alcohol, thus of much higher contents and which are similar to those obtained when working with Schizosaccharomyces pombe. The poorly defined origin of these products makes it possible to make only findings.

The presence of yeasts during the distillation makes it possible to obtain richer eaux-de-vie in esters. This confirms the work of GUYMON and CROWELL (1969) and partially explains the preference of practitioners for the lees distillation method.

GUYMON and CROWELL (1969) assume that, during continuous distillations, the fatty acids released by the yeasts are esterified with ethanol on the first plats of the column. The results reported here do not seem to be explained in this way since the distillations were performed, in the laboratory, discontinuously and that therefore the higher fatty acids could not be found in the free state presence of a high concentration of ethanol. It seems more probable that esters were linked to yeasts, either on their wall or in their cells and that they were released by heating.

The results concerning the addition of wax are more difficult to interpret since its composition is poorly known. It is known to contain a small amount of free fatty acids with a very high carbon number, which are yeast growth activators, but its biotin and pantothenic acid contents are not known in industrial practice. It will be interesting to know if the simple addition of palmitic acid would not have effect, palmitic acid being the main fatty acid of the lipids of yeast.

Finally, there were important differences between one yeast species and another, even from one strain to another within the same species. Under the same conditions, Saccharomyces cerevisiae Berlin II produced 36.5 mg of these esters per liter of pure alcohol (Table 3) while Saccharomyces cerevisiae S. produced 114 (Table 4).

The highest content was obtained with Schizosaccharomyces pombe, which is the dominant species in the flora of wines prepared for the production of “grand arôme” rum.

V.— CONCLUSION

Although their mixture can not be at the origin of the characteristic aroma of the rums, it seems probable that the ethyl esters of the volatile fatty acids participate in their qualities. Therefore, it was useful to specify the conditions allows rums rich in these constituents.

The most important factor is doubtless the distillation and alcoholic degree to which the alcohols are obtained continuously. The higher the rectification rate, the lower the ester and higher alcohol contents.

On the other hand, three factors seem to be able to be used to increase the levels of esters while keeping low contents of higher alcohols: the addition of wax, the distillation of the turbid substances and the selection of a species, or even of a yeast strain.

Received for publication in October 1972.

SUMMARY

ETHYL ESTERS OF THE RUM HIGHER FATTY ACIDS

Good quality rums have generally a high content of volatile esters and especially of ethyl esters of higher fatty acids (n-C8, to n-C16). These esters are secondary products of alcoholic fermentation, like higher alcohols,and behave like them during a continuous distillation process.

The quality of rums should be improved using our recent knowledges on esters production by yeasts in beer and wine.

Higher contents of ethyl esters of higher fatty acids can be obtained when the yeasts are not removed from the wines before distillation, when sugar cane wax is added to the must before fermentation and when a yeast species, and even a yeast strain, is selected.

The higher fatty acid ethyl ester contents of distillates were three times higher with Saccharomyces cerevisiae S. 132 than with Saccharomyces cerevisiae Berlin II. The highest content was obtained with a strain of Schizosaccharomyces pombe, a native yeast of sugar cane growing Countries.

A correlation between the fatty acids composition of the lipids of the yeasts cells and the ethyl esters produced could not be conclusively established.

BIBLIOGRAPHY

BARAUD J.,MAURICE A., 1963. Les alcools et esters des eaux-de-vie de canne et de pomme. Ind. aliment. agric., (1), 3-7.

EL SHEHATA A. M., 1960. Yeasts isolated from sugar cane and its juice during the production of Aguardente de Cana. Appl. Microbial., (8), 73-75.

GUYMON J. F., CROWELL E. A., 1969. Gas chromatographic determination of ethyl esters of fatty acids in brandy or wine distillates. Amer. J. Enol. Vitic., 20 (2), 76-85.

HARRISSON J. S., GRAHAM J. C. J., 1970. In The Yeasts, vol. 3, Acad. Press, London.

KAHANE E., Rous S., 1961. Nouvelle methode d’extraction des lipides, in Enzymes of lipid metabolism 82-90, Pergamon Press, Oxford.

KERVEGANT D., 1946. Rhums et eaux-de-vie de canne. Les Editions du Golfe, Vannes.

LIEBICH H. M., KOENIG W. A., BAYER E. 1970. Analysis of the flavor of rum by gas liquid chromatography and mass spectrometry. J. Chromatogr. Sci., 8 (9), 527-533.

MAARSE H., ten NOEVER DE BRAUW M. C., 1966. The analysis of volatile components of Jamaica rum. J. Food Sci., 31, 951-955.

MARTIN J. T., JUNIPER B. E, 1970. The cuticles of plants. Ed. Arnold Publishers Ltd, Edinburgh.

NORDSTRÖM K., 1964. Studies on the formation of volatile esters in fermentation with brewer’s yeast. Svensk Kemisk Tidskrft, 76 (9), 510-543.

PEYNAUD E., 1956. Sur la formation d’acetate d’ethyle par les levures du vin. Industr. aliment. agric. 73 (4), 253-256.

RAINBOW C., 1970. Brewer’s yeasts, in The yeasts, vol. 3, Acad. Press, London.

STEVENS  R., 1965. Gas chromatographic identification of ethyl ester of fatty acids in domestic and imported rums. J. Ass. off. agric. Chem., 48 (4), 802-805.

SALO P., NYKANEN L., SUOMALAINEN H., 1972. Odor thresholds and relative intensities of volatile aroma components in an artificial beverage imitating whisky. J. Food Sci., 37 (3), 394-398.

SUOMALAINEN H., PUPUTTI E., NYKANEN L., 1968. Composition of the aroma in some brands of whisky and rum analyzed by customary methods and by gas chromatography. Kemian Teollisuus 25 (5), 399-404.

Karl Micko’s Quantum Leap

Sponsor my distilling work simply by sharing the artisan workshop of the Bostonapothecary on social media. Copy, Paste, Share, Support!

This ends up being the most pivotal paper in all of spirits in the 20th century. It is not readily apparent, but everyone built upon it. No one references it so Micko’s ideas were known but somehow lost and not in anyone’s bibliographies. Arroyo brought the techniques to a whole new level. The French papers from the INRA acknowledge Micko’s central technique even though they focus on microbiology.

I propose taking it further and updating everything for a new generation of off the shelf equipment and productivity needs. I was even beginning to arrive at some of the same techniques myself in my Distiller’s Workbook series before I got clarity from Arroyo and historical clarity from Micko.

The original document is of a rough scanning that OCR hated so I had to do way too much editing. I think mistakes still live in the document, but whatevs, I’m busy. I rolled this out unannotated so people intentionally miss the significance. I may update it slowly to reveal my ideas. My lab is looking like it will cost $25,000. The techniques encompass product development and evolution for distillates from fermentations and well as a full on gin lab for botanical assay and formula development. The gin lab might actually push the price to $30k. University programs will eventually teach these techniques as the core of their curriculum and we’ll eventually get support from distillery trade organizations.

RESEARCHES ON JAMAICA AND ARTIFICIAL RUM.
By Dr. Karl Micko,
Director der Stattlichen Untersuchungaanstalt 
fur Lebensmittel in Graz.

I.—Examination of Jamaica and Artificial Rum.

Jamaica rum is of commerce, one of the most valuable and highly esteemed spirits of commerce,—a distinction which it owes to its characteristic and inimitable flavour. As it is produced in the tropics it is evident that duty and transport charges raise its original cost to a high figure on the European market. The concentrated rum, known as “original rum,” is too dear for the ordinary consumerit is therefore general for the retailing trade to break down the original rum, and by so doing the price falls in proportion to the degree of dilution effected. With many well established firms it is the usual custom to express the content of the broken spirit in degrees of the dilution and to fix the price accordingly. Generally, the original rum is broken down different dilutions for the convenience of the small dealers. It is, to however, preferable for the merchant to import the original rum and break it down himself. In this way he is able to control the dilution, and to be certain of the true content of the diluted spirit in original rummoreover, economy is thus effected, for charges for dilution and other expenses are saved.

In the case of many kinds of rum it is necessary to break down with spirit, for when in the concentrated condition their flavour is not always apparent. It is a particularity of certain rums that the fine aroma is only developed after breaking down; and this was instanced quite recently in our laboratory in the case of a sample of Jamaica rum for which a high price has been paid, but which when added to tea was stated to have a disagreeable taste. We were, however, soon convinced that this was such an example, and that the fine flavour was only hidden, for on breaking the sample down with 60 per cent. spirit the rich aroma and taste of genuine Jamaica rum was developed.

Since genuine Jamaica rum is a costly spirit. it is only to be expected that many attempts are made to imitate it. There are numerous receipts for the cheap manufacture of rum in Europe from molasses; but although it is known that the esters of formic, acetic, butyric, capric, and other acids occur in Jamaica rum, it has been impossible up to the present to make a spirit even approaching the genuine article. How far such attempts have been from successful may be judged by the fact that a rum expert has no difficulty at all in identifying an artificial rum by its flavour and aroma. There are upon the market many spirits which have been imitated much more successfully than Jamaica rum. Brandy, for example, is now imitated with such skill that the figures obtained by chemical analysis do not always indicate the fictitious article. Indeed, a brandy can be prepared to give analytical values the same as those found from a genuine sampleand brandy experts are now in no way so certain of judgment as formerly. The rum taster, on the contrary, has a much easier taskthe analyst. moreover, can not only readily differentiate the genuine from the artificial product, but is in the position to be able to detect what might easily escape the expert, namely, the admixture of even small amounts of genuine Jamaica rum with an artificial spirit.

The reply to the question as to why it has not been found possible after so many attempts to even approximately imitate the peculiarly fine aroma and taste of Jamaica rum is that it is characterized by a special flavouring constituent, which is not to be found in the best rums made in Europe, nor in the artificially made product. The flavour and aroma of potable spirits is not derived from one but from a number of different bodiesthis is true of Jamaica rum, but the basis of its characteristic flavour is an aromatic constituent which is peculiar to it alone.

This constituent can readily be separated by fractional distillation,  even when present in small amounts. If Jamaica rum is distilled, a simple tubular condenser being used, and the distillate collected in eight fractions, the first four are not in any way specific of the genuine rum. The peculiar flavouring constituent comes over mostly in the fifth and sixth fractions; towards the end of the distillation the amount of this body gradually decreases so that in the eighth fraction little or none is present. Generally most of it comes over in the sixth, but the alcohol content and method of distillation have, of course, an influence in determining the particular fraction. Concentration to a definite fraction can obviously be more readily effected by using a still-head.

Together with the flavouring constituent a characteristic body of a terpene-like odour also comes over. That terpenes occur in brandy has been pointed out by K. Windisch in his well-known work on the subject,* [*Arbeiten aus dem Kaiserl. Gesunuheitsamte, 1893, 8, 279.] and he has expressed the opinion that a certain terpene or terpene hydrate is also present in rum and may contribute largely to its characteristic flavour. Whether this particular terpene body is peculiar only to Jamaica rum we are unable to say with certainty, but we can at any rate assert that is is not the principal distinctive substance of Jamaica rum. It is always present in Jamaica rum together with the other flavouring constituents, and we have never found it in the artificial product.

Besides the flavouring bodies, other substances which are less volatile come overamongst those are certain resinous bodies which partly dissolve in sodium hydroxide, from which they can afterwards be precipitated by the addition of acids.

As in the case of other potable spirits, aldehydes and volatile acids are found in Jamaica rum. The content in these bodies is subject to very large variations, and here the sophisticator has an opportunity of adulterating Jamaica rum with artificial spirit without surpassing the limits generally found by analysis. The mixing of artificial rum with original rum cannot be practised to any great extent, because by so doing the true flavour is decreased, and the value of the spirit consequently diminished. The adulteration of broken Jamaica rum with artificial rum, to the contrary, is often done. Of eleven samples which we tested for aldehydes, all were found to give distinct reactions. As we shall see further on, artificial run can have as high a volatile acidity as Jamaica rum.

The difficult of judging rum on the ground of its analytical figures is generally caused by the fact that these figures are incapable of expressing the distinctive feature of Jamaica rum, namely the presence of its peculiar flavouring substance. There are, however, certain qualitative tests for Jamaica rum, and what is not shown by a chemical analysis can without difficulty be detected by a trained sense of smell. The usual analytical figures can, nevertheless, corroborate the judgment of a rum, and information of much value can be learnt from them.

The samples which we have examined were: Original Jamaica rum; Jamaica rum broken down with dilute alcoholJamaica rum mixed with artificial rumand artificial rum.

Original Jamaica rum was not often examined, because the sale of this article is confined to special firms and it is only occasionally customers require it tested. The spirit marked “Jamaica Rum” is as a rule broken down with dilute alcohol, and is sold in this form to the customer by all firms dealing with itit is only to be expected that here it is necessary to exert a careful chemical control.

Since genuine Jamaica is the most expensive rum of commerce it is not surprising that the sophisticator marks his product “Jamaica Rum” it is, moreover, not beneath him to apply this title to a product which is nothing more than artificial spirit. This is borne out by the table of analytical results which is given below.

Artificial rum has obviously only the value of the alcohol contained in it. For its production artificial rum essences are used, the cost of which is comparatively low. These substances must be added in small amounts; a certain limit must not be surpassed, for the taste of the product would otherwise be rendered unpleasant. It is for this reason that the ester content of artificial rum is, as a rule, low. The adulteration of broken Jamaica rum with artificial rum is enticing in view of the fact that such a mixture has more or less the flavour of the genuine product and in consideration of the large profits to be gained from such a procedure. It is only fair, however, to the majority of the trade to point that genuine Jamaica rum is sharply differentiated from the artificial that spirit.

In Austria artificial Rum goes for under the names of “Cuba Rum,” “Façon Rum,” “Wirtschafts Rum,” and “Inlander Rum.” The name “Cuba Rum” for an artificial rum is indeed not a strictly proper one, but it is now so firmly established in the trade that it can scarcely cause confusion. Cuba rum, moreover, always indicates a spirit of inferior value to Jamaica rum, and on this account the cost of this artificial rum is considerably lower than that of even the very highly broken genuine product.

The results of our examination of 38 samples of various rums are summarized on the table given on pages 228-229.

Method of on Examination.—As the table shows, the following determinations were carried out: Specific gravity, free acids in the distillate from 100 c.c., and ethers; the characteristic flavouring constituent of Jamaica rum, and foreign flavouring and colouring bodies were also examined.

The alcohol content was calculated from the specific gravity of the sample. As rum contains soluble substances this method is not strictly correct, but it was sufficiently accurate to approximately indicate the strength of the spirit. For an original rum it would be advisable to establish a standard of not less than 70 per cent. by volume of alcohol. In Austria there are no regulations at all as to the alcohol content of rums.

To determine the free acids in the distillate, 100 c.c. of the sample were rinsed into a distillation flask with 15 c.c. of water and the liquid distilled down to about 10 c.c. The distillate was then neutralized with N/10 sodium hydroxide and the result expressed in terms of acetic acid. Jamaica rum has as a rule a greater volatile acidity than artificial rum. The determination of the free acidity in the distillate bears this out, and it is generally found that this value is with Jamaica rum well above those given by artificial rums. The acid value of artificial rum is often strikingly low; it, however, sometimes happens that this value is as high as that of a Jamaica rum when broken down, and for this reason the acid value cannot be regarded as a certain criterion for the judgment of rum.

The ethers were determined by the cold saponification method. To the neutralized distillate, 30 c.c. of N/10 sodium hydroxide were added, and the liquid allowed to remain in a closed flask for at least 24 hours. In the case of concentrated rums N/2 alkali was used.

With artificial rums it was observed that the ester aroma had completely disappeared the next daybut with concentrated Jamaica rum, and even with broken Jamaica rums having a low ester content, the characteristic aroma could be detected after 24 hours, although to a somewhat less extent. By using, however, stronger alkali, viz., a N/2 solution, this aroma more readily disappeared, and gave place to one resembling the terpenes of coniferae.

From the greater power of resistance against dilute alkali of the flavouring constituent, and from the fact that after the disappearance of the ester aroma scarcely any more alkali is absorbed, we have come to the conclusion that the characteristic flavour of Jamaica rum in hardly to be ascribed to the esters.

The ester content of artifical rum is, as we have already mentioned, only small, and cannot be appriciably raised without imparting to the product a bad flavour. In the case of broken Jamaica rum the ester content can be considerably diminished by the dilutionstill, as we show further on, such a spirit can nevertheless be recognized as a genuine one. Although the ester content of original Jamaica rum is subject to very great variations we would have no hesitation in stating that sample 21 in the table marked “Original Jamaica Rum” is not an original rum, and this from the ester content alone without judging from other defects which are indicated by the analytical figures.

The samples 36, 37, and 38 marked Original Rum” came from reliable sources and may be taken as genuinetheir ester content varied those between given in 0.378 and 0-799, which are about which are about the same values as given in Konig’s “Chemie der menschlichen Nahrungs- und Genussmittel.”

It may be asked whether the ester content may be considered a criterion for the quality of a Jamaica rum. The ester content can indicate whether the rum is concentrated or dilute. But the quality of the spirit cannot be judged on the ground of the ester determinationfor obviously the quality of the rum depends not upon the amount of esters but upon their nature and relative proportions, as well as upon the other flavouring substances present. The strength of the aroma of Jamaica rum is indeed dependent upon the peculiar flavouring constituentbut the flavouring constituent is not saponifiable, and therefore is not indicated by the ester determination.

It is to be remarked that during the estimation of the esters in Jamaica rum the liquid assumes a more or less yellow colour according to the extent to which the rum has been brokenbut that with artificial rum the liquid often remains colourless, or is but very slightly coloured. This yellow coloration is due to the greater content of the Jamaica rum in aldehydes, including furfural, than in the case of the artificial product.

We have, however, met with samples of artificial rum which, on saponification, assumed a fine yellow colour, which was not surpassed by even original Jamaica rum. In such cases it is probable that the artificial rum manufacturer had added aldehydes to his product in the hope of more nearly imitating the genuine article.

(To be continued.)

RESEARCHES OF JAMAICA AND ARTIFICIAL RUM.
By Dr. Karl Micko,
Director der Staatlichen Untersuchungsanstalt fur Lebenemittel in Graz.(Continued from page 232.)

Fractional Distillation.Fractional distillation forms the most important method of gauging a rum, for it is thereby possible to concentrate the ethers, ethereal oils, and other aromatic constituents to definite proportions, and to identify them by their smell. For this purpose 200 c.c. of rum were mixed with 30 c.c. of water, and the mixture fractionally distilled. Eight fractions were collected, of which seven consisted of 25 c.c. and the eighth comprised the balance of the distillate. As much of the liquid was distilled off as could be removed without burning the concentrated residue. To carry out the smelling test, glass beakers were employed which were filled each with one of the fractions. According as the liquids adhering to the walls evaporated, the different smells arising from the ethers and other volatile bodies passed off. The first two or three fractions contained besides alcohol a very light volatile ether, also formic and acetic acid ethers. The subsequent fractions gave off smells peculiar to artificial rums and not to Jamaica rums. The typical aromas of Jamaica rums are found as a rule in the fifth or sixth fractions; which depends chiefly on the alcohol content of the original sample, being later in a rich alcoholic rum and earlier in a poor one. In the case of original or concentrated rum these aromas are divided amongst two or three fractions, whereas in diluted rum they are only noticeable in one fraction. As already said, the typical aroma of Jamaica rum is accompanied by a body rich in terpenes. Both bodies are entirely wanting in artificial rum. This terpene body has, however, a less pronounced odour and is less characteristic a proof of Jamaica rum, as in other high class spirits similar bodies rich in terpenes are to be found.

Artificial rum often gives off odours which are practically wanting in Jamaica rum, e.g., of strawberries, cassia or vanillin. These will establish the mixing of artificial with Jamaica rum.

It is to be observed that the aroma test must take precedence over the tasting, as otherwise the sensitiveness of the former will be practically destroyed. If the the tasting is however undertaken first, then the mouth must be well rinsed out with water before inhaling the aromas. The smelling tests should preferably be carried out in the morning hours, as then the sense of smell is stronger than in the afternoon, as smokers can testify.

The last, or else the penultimate, fraction appears turbid in the case of Jamaica rum, providing it has not been diluted too much.

This turbidity disappears on the addition of sodium hydroxide, only to reappear more strongly on acidifying. But if the sample be from an artificial rum the last fraction is as a rule clear. The partial solubility of the heavier volatile constituents of rum in sodium hydroxide is however no proof of a Jamaica rum, as it is a feature of other high class spirits also. Hence it happens that spirits derived from wines sometimes contain relatively large amounts of the heavy volatile bodies insoluble in water, which however consist only in a small degree of the higher alcohols such as amyl alcohol. A part of these bodies dissolves in sodium hydroxide and is reprecipitated on the addition of a mineral acidanother part dissolves only when heated in sodium hydroxide, but remains in solution when cooled, and only when acidified assumes a flocculent or oily turbidity. The smell of the sample is altered by the latter treatment. One is dealing here, it should be observed, with bodies of clearly complex composition somewhat akin to ethereal oils, and which are decomposed or otherwise altered when heated.

The heavy volatile bodies from the last fraction of rum can be separated by the aid of chloroform. Here we find before all others vanillin, which is often added to commercial rum essences and so is found in most artificial rums. If the rums to be distilled to as great a concentration as feasible, the vanillin carried over in the vapour appears equally in the distillate. It is most prevalent in the eighth fractionin the case of strong hydrated rum even the seventh fraction may contain it. In rums which contain over 70 per cent. alcohol it is advisable to mix the highly concentrated distillate with 20 to 30 c.c. of water and then to continue the distillation further.

The three last fractions were shaken up in a separating funnel with about 5 c.c. of chloroform. Since the sixth fraction of a rum rich in alcohol can still contain so much spirit that any separation of the fluids is not possible, it is necessary in such a case to add enough water to the fraction to enable the chloroform to separate from the remaining fluids. The chloroform is run into a beaker, and the beaker placed on a hot water bath. The chloroform must not boil however but only slowly evaporate. It is best to expedite the evaporation of the chloroform by frequently rotating the glass and as soon as the last trace of chloroform has disappeared the beaker should be covered with a watch-glass and laid aside to cool. There upon the smell of the residue can be tested.

The sixth fraction of an artificial rum often reveals a smell of cassia oil and other bodies all foreign to Jamaica rum. The seventh, and the eighth especially, contain vanillin providing this was present in the original sample. The smell of vanillin generally does not develop at once but only after an interval of hours or even days. It is therefore necessary when this smell is not immediately forthcoming to cover the last two samples with a watch-glass, leave them for two or three days and test them from time to time for the smell. It may happen when only a trace of vanillin is present that its smell is hidden by the scent of the other aromatic bodies, which however eventually volatilize or lose their smell owing to some influence such as oxidation, while the more stable and heavier volatile vanillin remains behind and then is gradually detected by its characteristic smell.

In the case of Jamaica rums the chloroform solution produces aromatic oleaginous or resinous residues; but the author has so far failed to detect vanillin in them with any certainty.

For the detection of vanillin in rum he employed the following test:150 to 200 c.c. of rum were made distinctly but not excessively alkaline, and while still alkaline were heated on a water bath to volatilize the alcohol, then acidified with HCl, separated with chloroform, and the chloroform solution evaporated at as low a temperature as possible. The small residue was often resinous and gave off smells which hid that of the vanillin, and sometimes hardly let it reveal itself at all, so that the author had to treat the residue with warm water, filter off the liquid from the undissolved portion and again shake up with chloroform. After evaporating the solution and allowing the residue to stand, the smell of vanillin if it was at all present was as a rule easily detected.

Chloroform is better than carbon bisulphide for the detection of small quantities of vanillin, for the latter has to be freshly prepared for the test, since otherwise it will give off an odour which would affect that of the vanillin. Apart from that, carbon bisulphide on account of its inflammability is not conducible to pleasant working.

This experiment has the disadvantage as compared with the distillation method that on shaking up the spirituous rums with chloroform an emulsion is easily formed, and it needs a longer interval before the chloroform will separate from the aqueous liquid. This disadvantage is absent from the distillation test, as in the latter immediately after shaking up of the aqueous distillate with chloroform the two liquids separate sharply, and there is no need for any further cleansing of the residue from the chloroform solution. Finally, the search for vanillin can be combined in one operation with searches for other aromatic essences.

Foreign colouring bodies are frequently present in concentrated rum. Even if it does not happen that the quality of a rum is judged by its colour, the presence of these colours reveals a case of intentional manipulation and as a matter of fact they are often found in imitation Jamaica rums or in mixtures of Jamaica with false rums, also in broken Jamaica rums that are sold as original Jamaica.

The testing of rums for foreign colouring bodies may, however, prevent any further identification of rum samples, such as may be demanded in legal cases. This concerns in particular the estimation of the ethers and the volatile acids in a distillate, but the alcohol content can be ascertained definitely by means of a hydrometer. As an instance one may cite the samples Nos. 17, 18, and 19, in the tables which were obtained from the same dealer, were produced to all appearance from the same recipe, but had been furnished with labels of different origin and with different specifications.

The tests made on the sample of commercial rums that were submitted to the Untersuchungsanstalt for analysis are to be found in the tables (see pages 228 and 229) numbered from to 34. The author deems it unnecessary to give particulars of more samples than these, as it would only lead to needless reiteration of figures. It is, however, clear enough from the instances cited that there is no difficulty in distinguishing artificial rum from Jamaica rum. One is able, even in the case of strongly broken rums and, within reasonable limits, also in the case of a low content in others as in sample 28, to identify the typical aroma of Jamaica rums. In many samples said to be Jamaican, but which were artificial rums, traces of this aroma of Jamaica rum were detected. The writer ascribed this to their being mixed with small amounts of Jamaica rum and in order to confirm the theory of his supposition he interrogated the spirit merchants and their their their answer was invariably that they added some Jamaican rum to their “Wirtschafts,”Cuba,” and artificial rums in order to improve their flavour. It must, therefore, not be overlooked that for similar reasons some Jamaica rum may be added to artificial rum essences whereby the peculiar taste will be imparted to them.

Additions of artificial rums to Jamaica rums yield aromatic bodies not found in the latter. But a more difficult task awaits one when only a small addition is under test, for then the adulteration may be hidden by the aroma of the Jamaica rum. In such a case it is advisable to fractionally distill a larger quantity than 200 c.c., and then to separate the individual fractions by further distillation. The artificial rum is less visible in the first fractions than in later ones, for both in Jamaica and in artificial rums the most volatile constituents consist mainly of the esters of formic and acetic acids and of alcohol. The first distillates of artificial rums have however a more obtrusive smell than have Jamaica rums, and this smell is also of a kind not found in Jamaicas, so that a case of adulteration is easily proved. Besides there are differences in taste and smell in the case of false rums. Many of them are at once detected by their strong smell of artificial ethers and vanillin. Others have a more finished aroma and taste, according as the distiller has the greater skill in the preparation of the artificial rums. In the tables we find No. 25 described as a Porto Rico rum; this was marketed with considerable advertisement. It contained a striking amount of cassia oil, and had also every indication of being a false rum. In the column, “Flavouring Substances foreign to Jamaica Rum,” the only adulterants mentioned are vanillin and cassia oil. But that does not imply that only these and no other foreign matters were present. They have only been cited because they are more easily identified by smell, and, especially vanillin, are very commonly found in false rums, while other aromatic bodies observed, if they are not found  in Jamaica rums too, are much more difficult to identify.

The difficulty of imitating the aroma of rum does not lie so much in the accurate selection of ethers as in the circumstance that the typical smell of Jamaica rums arises from bodies which either belong to the class of ethereal oils, or stand in close relation to them, but have no definite formula. Windisch was right in speaking of “ethereal rum oils,” which he he had obtained by the separation of rums with chloroform.

E. Sell, from his own investigations into the composition of rums, comes to the following conclusion. “The opinion expressed at the conclusion of some investigations on cognac, to the effect that it was impossible to distinguish genuine from fictitious liquors by mere chemical tests, is not a bit less opposite in the case of valuing rums. Here also the preference must be given to such expert opinion as bases its decision on the taste and the smell of the sample.”

It is thus the case that the figures obtained through chemical analysis are not by themselves reliable for distinguishing artificial from Jamaica rums, but must be supplemented by investigations into the taste and smell. Any analyst who has a sensitive nose and palate can not only distinguish Jamaica rum from artificial rum, but also a large proportion of the cases where the two have been mixed. In virtue of his wider knowledge of chemical bodies and through suitable experimentation in the laboratory, the analyst is in a better position to detect the foreign bodies not found in Jamaica rum than is the practical expert. But estimations of price and quality fall necessarily within the latter’s sphere.

[To be continued.)

RESEARCHES ON JAMAICA AND ARTIFICIAL RUM.
By Dr. Karl Micko,
Director der Staatlichen Untersuchungsanstalt fur Lebensmittel in Graz.(Continued from page 414.)

II. The Identification of the typical Flavouring Body of Jamaica Rum.

For the identification and nearer characterization of the peculiar flavouring constituent of Jamaica rum, I proceeded in the following manner:—

1900 c.c. “Original Jamaica Rum” (No. 38, page 229) was fractionally disitlled, eight fractions (I.-VIII.) being collected. Fractions I. and II. possessed a very distinct smell of formic and acetic esters, Fraction IV. a smell of butyric ester. The specific aroma which characterizes Jamaica rum appeared in Fraction V. but only feebly. It was quite distinct, however, in Fraction VI., and strongest in Fraction VII. Fraction VIII had an acid and an aromatic smell; it was cloudy and oily drops floated on its surface.

Fractions V. and VI. were mixed together and the mixture fractionally distilled, four fractions (I.-IV.) being collected. The first two of these fractions contained no typical aroma, and in the third fraction it appeared only to a very small extent. It existed strongly in the fourth fraction together with a body of a terpene-like odour. This fourth fraction was mixed with Fractions VII. and VIII. of the original distillation, and the whole fractionally distilled into five fractions (A-F).

Fraction A had only a little rum aroma; it was much stronger  in Fraction B, but Fraction C contained the chief quantity  of the peculiar flavouring constituent. In all three fractions was present the above mentioned terpene-like body (reminding one perhaps of juniper oil). Dilution of these three fractions produced turbidity.

Fraction D was turbid and watery, and did not possess the characteristic rum aroma, its smell being rather of other aromatic bodies. It was shaken up with chloroform, the chloroform solution separated, and the chloroform carefully evaporated away. It left behind a yellowish, resinous substance of the aromatic smell, which dissolved in caustic soda, and on acidification was reprecipitated. It is questionable whether this resinous substance is an original product of the fermentation. On the other hand many aldehydes incline to condensation, forming resinous substances which behave in the same way as the above with alkalis and acids. It is quite probably, therefore, that aldehydes are concerned in the formation of the aroma of brandy.

At all events, aldehydes react readily with alkalis. Grey says the specific rum aroma is produced by the action of lime on sugar solutions during its manufacture, and it is not impossible, therefore, that aldehydes are concerned in the production of the peculiar rum aroma. By the following experiments it will be seen that the peculiar flavouring constituent of Jamaica rum assumes another smell by the action of caustic soda. The formation of the rum aroma would be best studied by investigations during the different phases of manufacture.

Fraction had only a slight smellit was turbid and was shaken up with chloroform. The chloroform substance resembling that obtained solution left on evaporation from Fraction D. Fraction F had hardly any smell. Fractions A, B, and C all gave the aldehyde reaction on additions of Schiff’s reagent, and the furfurol reaction with aniline acetate. The aldehyde reaction was strongest in Fraction A, and weakest in C, but Fraction C behaved in quite the opposite manner with the two reagents. The following experiments were undertaken with distillates B and C :—

Experiment 1.

Since all signs tend towards the fact that the peculiar flavouring body of Jamaica rum does not belong to the esters, it remained to be proved whether it was not due to an aldehyde or ketone group which may be present. may be present. To decide this question I made use of fraction B and C. treated 5 c.c. therefrom separately with phenylhydrazine, hydroxylamine and semicarbazide. Fraction B was closely observed for change of smell, because it did not contain so much of the typical flavouring body, and any changes would, therefore, be noticed more quickly than Fraction C. But even after one week the typical aroma was recognizable, the phenylhydrazine test only having less smell, yet still quite recognizable, and, therefore, no reaction with either of the three reagents had taken place.

By the negative result of this experiment, it is not probable, therefore, that the typical flavouring body of Jamaica rum belongs either to the aldehydes or ketones.

Experiment 2.

The chief quantity of Fraction B was first treated with a saturated solution of sodium bisulphite, whereby any traces of aldehydes were removed, and the mixture shaken up with ether, whereby the typical flavouring constituent passed into the ether. The separated bi sulphite solution give out no aroma on treatment with dilute sulphuric acid.

The ethereal solution was shaken up with sodium carbonate in order to remove any sulphurous acid present. The typical aroma remained unchanged. The ethereal solution was separated and subjected to careful distillation on a water bath, the ether passed over first containing no typical aroma, then followed the alcohol together with the typical flavouring constituent. The aroma was not very pure or strong since Fraction B contained only a small quantity of the typical flavouring body.

Only a part of the alcohol was distilled off. The alcoholic distillation residue exhibited no distinctive smell of the typical flavouring body of the rumit was distinctly alkaline. Probably a slight trace of sodium carbonate remained in the ethereal solution thus causing the alkaline reaction of the residue. The latter was treated with excess of ether, the ethereal fluid separated the next day and the ether carefully distilled off on the water bath. The small quantity of alcoholic residue remaining from the distillation had a pronounced terpene-like odour reminding one of juniper oil, such as have continually noticed to be present with the peculiar flavouring constituent. The neutral reacting fluid gave only a feeble furfurol reaction, but with a distinct dstinct though no strong reaction with Schiff’s reagent. It still contained, therefore, a trace of aldehyde but the principal amount was at all events removed. the alcoholic residue was mixed with 10 c.c. N/2 NaOH, which produced a turbidity and also a yellow coloration on standing. The smell of terpene was preserved for some days. By titration with acid it was found that only 0.10 c.c. of NaOH had been used.

The titrated fluid was again made alkaline and shaken up with ether. The ethereal solution left behind after the evaporation of the ether a yellowish, aromatic terpene smelling like oil. The other part of the liquid turned cloudy on acidification with dilute hydrochloric acid. On shaking up this cloudy solution with ether and separating them, evaporating off the ether, only a trifling quantity of a brownish yellow oil was left whose aroma was not aromatic.

From this experiment it follows that the typical flavouring constituent does not enter into combination with sodium bisulphite as the aldehydes do to form an oxysulphonic acid. The terpene-like body which is present with the typical flavouring constituent is not soluble in dilute sodium hydroxide, and suffers no loss of smell through prolonged contact with the same. This body belongs at all events neither to the esters nor to the aldehydes.

Experiment 3.

This was conducted on Fraction C, which contained the chief quantity of the typical aroma. It was to distillation until a little alcohol passed over. The distillation residue is called (a), the distillate (b). Both (a) and (b) showed strongly the typical aroma, but the aroma from the distillate was much purer than that arising from the residue which smelt besides of the before mentioned terpene- like body and also of other aromatic bodies.

The residue (a) was cloudy and on the surface thereof swam drops of oil. It was very was carefully neutralized with barium thereof hydrate and diluted with water, shaken up with ether, separated and the ethereal fluid carefully distilled, so that nearly all the ether and only traces of the typical flavouring body distilled over.

The residue left amounted to only a few c.c.it reacted feebly acid, and a strong but not pure smell of the typical flavouring constituent. The trifling quantity of acid was neutralized with very dilute sodium hydrate and then 30 c.c. N/2 caustic soda was added. The alkaline solution with the oil drops after a time turned strongly yellow. After four days the smell of the typical flavouring constituent disappeared, and a peculiar aromatic smell took its place. The terpene smell was, however, very distinct. On titration with acid the consumption of N/2 caustic soda was found to be 0.9 c.c. The titrated fluid was again made alkaline and shaken up with ether. On separation and evaporation of the ethereal layer an oil was left behind, similar in smell to that obtained in Experiment 2. The other layer of liquid was submitted to distillation in a current of steam in order to see whether it contained any volatile acids.

About 400 c.c. of distillate was collected which was slightly cloudy only and reacted almost neutral. Evidently it contained volatile acids in only small quantities. It was made alkaline with barium hydrate, evaporated to dryness, the residue extracted with hot water, then filtered and the filtrate evaporated to dryness. The residue left was very small, and on the addition of two drops of dilute sulphuric acid a smell resembling that of butyric ester was formed.

The quantity of this substance was so small that it at all events did not correspond to the residue which required 0.9 c.c. N/2 caustic soda for saponification. The residue left from the extraction with hot water did not dissolve completely in hydrochloric acid. A white turbidity of the hydrochloric acid solution was caused through a fine flocculent substance, which did not dissolve in ether, but in alcohol. The quantity was so small, however, that it could not be put to further test.

The distillate (b) contained the typical flavouring body in its purest form. For the purification and isolation of the typical flavouring constituent the distillate (b) was strongly diluted with water, then shaken up with ether, and the ethereal solution further shaken with water. The typical flavouring body always remained in the ether which was distilled off very carefully at a temperature of 30°C so that ether and practically none of the flavouring constituent distilled over. The alcoholic residue amounting to about 50 c.c. was diluted with water several times in order to remove the alcohol, then mixed with ether, the ethereal solution washed several times more with water  and separated. It amounted finally to approximately 30 c.c.

5 c.c. of this left behind, after evaporation of the ether at room temperature, colourless drops of a fluid which possessed the typical aroma of Jamaica rum very intensely, and which in one hour completely volatilized and filled the laboratory with the characteristic aroma of Jamaica rum.

The aroma is more characteristic in the dilute than in the concentrated condition, and its boiling point certainly is higher than that of ethyl alcohol, yet it evaporated at ordinary temperature fairly quickly. When rum is rubbed on the palm of the hand the typical aroma can be detected for a fairly long time, and it seems, therefore, that Jamaica rum contains more difficult volatile bodies than the typical one, this being held in solution longer, thus preventing quicker volatilization. When the rum is fractionally distilled the typical constituent is concentrated in one or two fractions, and the aroma from these fractions is much stronger than in original rum, also evaporating more quickly out of the palm of the hand than in the case of the original rum.

A further 5 c.c. of the same ethereal solution was taken and shaken with 5 c.c. of N/10 sodium hydrate for some minutes, the alkaline fluid separated off, and the remainder washed with water until neutral. The typical flavouring body remained unaltered and is, therefore, not soluble in sodium hydrate. On treating this ethereal solution with 10 c.c. N/10 sodium hydrate and leaving for four days in a lightly corked flask with periodical shaking up, the smell at the end of this time was decisively altered, being certainly aromatic, but not corresponding with the typical flavouring body. The consumption of N/10 NaOH was found to be only 0.15 c.c. on titration with acid.

The same experiment was repeated only with this difference that the saponification was carried out by heating for half hour on the water-bath under reflux condenser. Only 0.1 c.c. of N/10 NaOH was used in this case, and the smell was altered as in the preceding case. 

The titrated fluid was again made alkaline, shaken up with ether, the ethereal layer separated, and after gently warming the remaining fluid on the water-bath to expel traces of ether it was acidified with dilute H2SO4. From both tests a scarcely perceptible turbidity was produced, and no smell was apparent.

For the third experiment 10 c.c. of alcohol (previously distilled over caustic soda), 10 c.c. N/10 caustic soda and c.c. of the ethereal solution which had been shaken with dilute caustic soda, were mixed together. After half an hour’s heating on the water bath under reflux condenser, the titre of the fluid remained almost the same. The smell was, however, altered as by the first two tests. From all investigations with distillate (b) no yellow coloration took place by the action of alkali, as was the case with the residue (a). The yellow coloration produced in (a) would probably be due to aldehydes or furfurol in small quantities, but the amount of alkali used was so small that it could hardly be attributed to aldehydes. When an alkaline solution of furfurol is allowed to stand, however, a yellow colour is produced at first, afterwards turning cloudy. By the acidification of this turbid alkaline solution, a reddish brown flocculent precipitate is thrown out.

As the investigation with Fraction B proves, the typical flavouring body does not combine with alkali. The absence of a yellow colour points to the fact that it cannot belong to the aldehydes. We have from the earlier experiments seen that no combination is effected with ether phenylhydrazin, hydroxylamine and semicarbazide. Neither does it combine with sodium bisulphite. On the other hand, caustic soda alters the smell of the typical flavouring constituent slowly in the cold, but more quickly in the warm, but no saponification takes place however.

Summary of Results

From the above investigations the following conclusions may be drawn:—

1. Jamaica rum contains an aromatic constituent peculiar to it alone, which is the basis of its characteristic flavour. This constituent is found neither in high class European spirits nor in artificial rum.:

2. This typical flavouring body of Jamaica rum is a colourless not difficultly volatile fluid of a delicate aromatic smell and its boiling point lies higher than that of ethyl alcohol.

3. This typical belongs neither to the esters, ketones, or aldehydes. It has the general characteristics of an ethereal oil, and it is not improbable that it stands in nearer relation to the terpenes.

4. The typical flavouring body does not dissolve in caustic soda, but on prolonged contact with it, it assumes an aromatic but more resinous smell.

5. In Jamaica rum as in other high class spirits is a body possessing a terpene-like aroma which is entirely absent from artificial rum. But it is less characteristically a proof of Jamaica rum as in other high class spirits similar bodies rich in terpenes are found.

6. In Jamaica rum there occurs in the last distillation fraction an aromatic smelling, resinous substance, which dissolves in caustic soda and is precipitated by the addition which of acids. It is questionable whether this substance is a primary fermentation product. For we can produce such substance easily from aldehydes.

7. The analyst with sensitive nose and palate can easily distinguish artificial from Jamaica rum. He is also in the position to be able to detect mixtures of Jamaica with artificial rum.

8. From chemical analysis alone, however, no thorough conclusion is possible but when used in conjunction with the smelling test it is extremely valuable. The ester number is of especial value for deter mining whether the given sample is of a concentrated or a diluted rum.

THE CHEMICAL EXAMINATION OP VARIOUS RUMS.
By Karl Micko.

In a previous article (this Journal, 1909, 225, 410, and 446) we have dealt with the examination of rum, and have attributed the great difference between Jamaica and artificial rums to the presence of a typical aromatic constituent in the former. We have now extended our studies to other kinds of rum, especially Cuba and Demerara rums, and to different arracks. The special object in view was to determine whether Cuba and Demerara rums contain the same typical aromatic constituent, and to find out in what way these two kinds differ from one another.

Our previous work has shown (loc. cit.) that the typical aromatic constituents of Jamaica rum is more resistant towards alkali than the esters; for whilst the esters are completely saponified by an alcoholic N/10 caustic soda solution in 24 hours at ordinary room temperature the typical aromatic odour persists to finally give place to another more resinous smell.

It was inferred that by carefully saponifying the esters it would be possible to separate the peculiar aromatic constituent, and this was found to be actually so. First the ester content of a portion of distillate was determined using an excess of alkali, then to a second portion of distillate less alkali was added than sufficed for the saponification of the esters. The amount of caustic soda added was less than that actually necessary to effect saponification on account of the fact that the aldehydes readily decompose
in alkaline solution forming acid products. In the case of this happening the odour would be affected; but our main object in working with as small amount of alkali as possible was to avoid attacking the non-ester constituent.

The distillate thus treated was alkaline, completely but the excess of  alkali was slight as the esters were almost completely saponified. It was then acidified with tartaric acid, and submitted to fractional distillation.

Working in this way we examined not only the above-mentioned tropical spirits but also some European products; and our conclusions may be summarized as follows:–

All the spirits examined possess a peculiar aromatic constituent which does not belong to the esters. It is possible aromatic by means of the above-described method to readily differentiate between artificial spirits flavoured with esters, etherial oils, and other substances and the genuine product. This peculiar aromatic constituent is of great value in judging the purity of spirits, and is in this connection of greater significance than the esters. It
imparts most of the specific aroma to the spirit. It is a general criterion, and has not been imitated up to the present time. It can be separated by carefully saponifying the esters, and has an extremely delicate fruity odour.

The tropical spirits which were examined, Cuba, Demerara, and Jamaica rums, and Batavia arracks, contain in addition to other flavouring constituents the same, or a very similar aromatic constituent, as the one we obtained from Jamaica rum (this Jl., 1910, 225 et seq.), but in Jamaica rum it is present in much greater amount.

In Jamaica rum it can be detected by fractional distillation of the original rum, or even by strongly breaking down; whilst in the other spirits, since
it occurs in much smaller amount and in admixture with other specific aromatic bodies and esters, it can only be recognised by carefully saponifying the esters and fractionating the ester-free liquid thus obtained.

Besides the fragrant-smelling bodies another body of a characteristic odour which generally appeared in the last fractions of the fractional distillation was found in the tropical spirits.

As to the origin of the typical aromatic constituent, it may either be formed during fermentation, or may be present as such in the primary material.
In the case of Jamaica rum the first supposition is probably true. Perhaps there are present in the sugar cane certain bodies which during fermentation give rise to the aromatic substances; or again possibly the aldehydes, ketones, &c, react during the production of the spirit with the formation of the aromatic bodies.

The results of our analytical examination of the various rums are given in the following table:—

Arroyo’s Invitation: The Problem of the Ripening of Crude Rum

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There are no technical revelations here, but this is important for the historic record and in describing the market and need for new ideas in 1940. The translation was a little challenging. Hopefully I did not kill it. It also begs the question, where are we now? Are established producers making easy to age crudes and are any new American producers or are we building and aspiring brick house rums like Hampden? Where is Lost Spirits in all this?

Arroyo R. 1940. The Problem of the Ripening of Crude Rum. Revista de Agricultura del Puerto Rico, volume 32, pages 588-591.

El Problema de la Curación del Ron Crudo
by RAFAEL ARROYO, Ch. E. & S. E.,
Especialista en Fermentaciones Industriales; Jefe Division de Quimica Industrial de la Estacion Experimental Agricola de la Universidad de Puerto Rico.

THERE ARE two different schools in terms of the method to follow to impart the desirable maturity stage to raw rum.

One of these two groups favors the methods of accelerated curing, which aim to try to impart the characteristics of a ripe rum to the raw distillate in the shortest possible time, using physical or chemical methods, or combination of these. It is presumed that the use of aging barrels is ruled out in these methods called accelerated or rapid aging. The other group continues to place their trust in the classic method of slow aging, through natural aging in the oak barrel.

Both the followers of one school and those of the other have incurred a common error, which consists of not having recognized the true value of the quality of the crude distillate, and the transcendental role it plays in the problem of aging.

A lot of ingenuity, time and money has been spent trying to make a ripe rum quickly, starting from any distillate, lacking in most of the times the inherent qualities to be turned into a ripe rum of quality. But, on the other hand, no attempt has been made to exercise that same ingenuity, and to use time and money in the search for methods and apparatus for the obtaining of raw rums worthy of such a name. On the one hand, the supporters of aging in the barrel believed that without taking into account the quality and conditions of the distillate with which it was filled, at the end of aging the barrel would in all cases return a ripe rum of excellent qualities. For this the barrel was the whole in the production of quality rum. Why worry about the conditions and characteristics of the crude distillate when the barrel is there to correct all the errors, clean up all the roughness, and finally return the distilled product to the distiller in perfect conditions? On the other hand we had the supporters of accelerated aging methods, fanatical believers in their secret methods and formulas, with which it did not matter what kind of raw rum reached their hands, and they would know how to get ready to get a product of incomparable quality out of it.

And in this way, the ones with their blind faith in the wonders of the barrel, and the others infatuated by the incomparable virtues and magical results of their recipes, formulas and secret methods, completely neglected the production of crude rum, relegating to secondary position the only factor able to solve the problems inherent to the economical curing of raw rum. The key to success in the rapid attainment of maturity consisted mainly in the production of true quality raw distillates capable of becoming a quality rum in a relatively short time of treatment in a barrel; or to respond effectively to the indicated accelerated treatment.

It is to be noted, however, that although behind the scenes the rum confection is carried out by using one of the other rival methods, or combinations of both; All the rums are presented to the consuming public as old rums cured in the aging barrels.

It seems that the followers of the methods of quick aging prefer that their rums get presented as products of the classic method. In this way, they are, of course, granting supremacy to this method. But do they recognize this in their hearts because they do not adopt all of the classic methods? Well simply because this method is, according to consensus opinion, so expensive that it only adapts to be used by entities with large capital. And the main cost lies precisely in those long years of aging in barrels that is necessary within the usual technique for the raw distillate to acquire a characteristic of ripe rum. They did not think that such objection present that perhaps there would be means to shorten those long years of aging without leaving the norms of classic method? Was it really necessary to go to such extremes to pretend in a few days, and even in a few hours, what you saw so far cost many years of painful and expensive waiting to get? It would not be a step forward to cut those years in half or reduce them to a third part? Was not it more feasible and prudent to modify a method already proven to be good, than to try to jump precipitously to the discovery of something new and of doubtful success? But let’s renew our theme.

We said that in any way that the commercial rums are made, these are announced to the public as rums properly placed in carefully selected barrels. We have not yet had occasion to hear an announcer on the radio referring to a rum in these terms: “This rum, dear listeners, is not an old rum, but a rum prepared in a few days, but it is a good rum , with all the characteristics of goodness inherent to a rum in full maturity of first quality.” And for the public, the phrase Ron Viejo is synonymous with that of ripe rum, with rum possessing those organoleptic qualities of taste and smell that are so appetizing for the consumer. However, in order not to hide the age of a rum from the public, it would be enough to show them that maturity and old age in a rum are not necessarily synonymous attributes; well, a rum can be old without it having reached maturity, as a rum can be ripe without necessarily being qualified as old.

The consumer does not prefer the old rum for the mere fact of the years it has spent in a barrel, but for the modification it acquires during that time, because his past experience tells him that rum usually means ripe and good rum. To date, age has been the only index of kindness … but, if that degree of kindness, excellence of bouquet and taste can be imparted without the need for many years of aging… do my readers believe that it would matter a great deal to that public the method used to reach such a result? Would we refuse to accept that a rum could be of exquisite bouquet and good taste for the mere fact of not having long years of aging? Is aging an end, or a means that takes us to that end?

Among the arguments presented by the facilitators of the fast practices for rum elaboration we will mention some:

1. Eliminate the inconveniences, expenses and risks involved in the storage of thousands of gallons of rum during its maturing period.

2. In the terrible, sterile competition based on price levels only, rums of long years of aging would have to be sold within a scarce and always doubtful margin of profit for the manufacturer, or else they could not compete with those made by other methods.

3. The vast majority of rum consumers do not know how to appreciate or distinguish subtle gradations of goodness and exquisiteness between different rums.

4. The process is cheaper and therefore yields greater benefits.

5. Much less capital investment is needed in the business.

6. There are no official “standards” to catalog and classify the different rums according to quality, it happens that in the practice of the market the values are confused, receiving very similar prices rums manufactured by one and another method. Under these conditions the method of manufacture that is easier and gives a cheaper result should be chosen.

The supporters of the classic method answer these arguments, such as the following:

1. Flavor, body and aroma superior, and above all more consistent.

2. The retrogradations in the quality are eliminated; for this (the quality) increases and improves as time passes. In this way a permanent clientele is created.

3. The purity in the constitution of rum is superior.

4. In the case of export rum there is a big saving by not paying the tax of 30 cents per rectified gallon.

5. Actually there are better prices for this class of rums both locally and in the United States.

6. Guarantee of the existence and economic stability of the industry through those changes and legal requirements that may occur in the future affecting the current state of affairs in the manufacture and sale of rum.

We now ask ourselves—which of these two movements is better oriented? We believe that when this question is submitted to a group of presumed rum manufacturers, we would find that the consensus of opinion is in favor of accelerated methods. And it is not that this majority cease to state that the rums cured by the classical procedure are in truth and in fact superior to date to those of artificial maturing, but that they fear in an extraordinary way those long years of aging in barrels. If the rums matured in the barrels in a few months instead of years, then we could give all the supporters of artificial methods of maturing as defectors. Will this be possible? We believe that yes; At least our experimental work has been proving it.

Until now, the biggest obstacle to the general adoption of the classic cure system has been the belief that it is necessary to cure a raw rum for four or more years to turn it into a truly quality and meritable drink. We believe that with the crude rum currently manufactured that is the case; but we are sure that by elaborating better crude rums we can cut the barrel curing time by half or a third of the time that is now considered indispensable. Our research work has fully verified this.

In other words up to the present, the general opinion is that the manufacture of rum by the classical method yields the best products; but given its scarcity and complexity excludes all those interested in the business except those with large capital. Only a millionaire, according to the opinion, could put on the market rums that need three to four years of barrel aging to give signs of having acquired the state of desirable maturity.

Those of us who, as the author of these lines, are especially interested in the technical and scientific part of the question, believe that both movements are healthy for the present and future development of this industry. There are good arguments in both trends, because it is undeniable that the superior qualities of goodness, taste and aroma imparted to the crude distillate by time and barrel are difficult, very difficult to equal, or even imitate by the artificial methods for the acquisition of maturity hitherto known and put into practice; but it is also necessary to accept the economic arguments in favor of quick methods, and in this sense there is no doubt that a rum that has to be aged for four or more years to reach the acquisition of the necessary maturity, is almost prohibitive manufacturing unless you have great capital. And even with the necessary capital, the question would always remain pending… will the consumer pay for this kind of rum with a price difference necessary for the business to be profitable for the manufacturer distiller?

On the other hand, we have already contemplated as mark after mark of the rums elaborated by artificial procedures they have a more or less fleeting and transitory life, and a more fleeting root still in the appreciation of the consumer people; to then fall into the most lamentable oblivion and abandonment.

This state of affairs in the problem of curing raw rum can be solved as indicated above by the search for crude distillate manufacturing methods that guarantee us the rapid acquisition of maturity for these distillates once subjected to the classic treatment of curing. In other words, it is necessary to manufacture better crude rums. Let’s start by packing a distillate in the barrel that can be called a genuine raw rum and not any alcoholic distillate that we’re used to calling raw rum. We can not trust to simply barrel “anything” that we call crude rum, and hope that it is doing miracles and returns to us turned into a rum full of quality in a short time.

The study carried out by us on this problem puts us in a position to offer those who fear the classical method for the time and money it takes, a method that allows them to use it without the need for large capital investments. How? As we have already said it, again and again, putting more love and care in the preparation of the crude distillate; there lies the success of the cure in a short time. Our research has shown us fully that the acquisition of maturity in a rum is not the product of a single factor, such as the barrel in which it is aged or the still in which it is distilled. On the contrary, it is the result of a whole series of factors that starts with the selection of yeast and raw material, and ends with bottling for the market. But all these contributing factors to the same end can be condensed into two groups:

1. Those factors that combine and support each other for the production of a good raw rum.

2. Those other factors that combine to help a good crude rum to acquire its desirable maturity.

Now, if the newly distilled rum (which we call crude) lacks the necessary characteristics, then the action of the second group of factors can not fulfill its mission with efficiency and full satisfaction. It is as if a good teacher was trying to make a prodigy of a child lacking in natural intelligence.

Therefore, the success of its cure depends on the quality of the crude, especially as regards the time it takes (the cure) to be taken. Do not our readers believe, and especially those of knowledge and experience in the aches of rum cure, that it would be worth producing a good crude if this implies shortening the healing time by half or a third of its normal duration?

Once the elaboration of a true quality crude rum is achieved, we will find the problem of the classic cure solved, since in a period of one to two years in some cases, and even a few months in others, the distillates thus produced will be turned into ripe rums, ready for the market.

And to those manufacturers who can not age for a long time, but who would settle for producing a genuine and good rum without trying to enter into competition with the finest and most exclusive of the market, we tell them that they could do it through an aging of no more than six to nine months, if always taking care in the production of quality crude rum, or that they knew how to choose these, in the case of not being distillers of their own raw material.

To those of our readers of practical minds who like more of objective and palpable demonstrations than of long spoken and written dissertations and arguments, we extend a cordial invitation to visit our laboratories and witness and feel the facts that have induced us to write this article. Native rum producers are especially invited.

Pineapple Disease & Rum

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Recently, a great friend of the blog turned in an English annotated version of Hubert Olbrich’s chapter on rum.

The most interesting thing to come out of it was this bit (remember this is annotated, not translated):

What are the influences of Raw and auxiliary materials on the rhum quality? The flowery, sugary bouquet of molasses is not very involved. It exists a link with the sugarcane. It should be grown on the right soil: in Jamaica the cane is grown on flat coastal areas, for the production of common rhum. For high ester rhum they use irrigated cane from the mountainous south of the ile.
Iod taste is supposed to come from cane which is grown near the sea, and has absorbed iodic organic compounds. This has not been proven analytically yet.
The flowery Ananas type comes from an infection by a black mold (ananas sickness), which is grown in tank filled with broken sugarcanes.

An important auxiliary is “dunder”: The nutritious base for yeast growth, and main source for the rich microflora and is metabolism essential for the aroma production.
This alcohol free residue of preceding production, has treated in multiple ways.
One example (2597): Some producers develop their yeasts, bacteria and fungi in clay pits. In these pits, the dunder, sugarcane straw, and sand, are layered. Then, the pit is closed.

At the end we see the classic “pit”, but up paragraph we see a new concept which is a tank full of broken canes floating in water and infected by a black mold. The black mold is also referred to as Pineapple disease (Ceratocystis paradoxa).

So, like, what the fuck? Does this make it into anything we drink? Was this one eccentric producer Olbrich collected an anecdote from? Does this not make drinking heavy bodied rums exciting to anyone else?

Olbrich even slips a bibliographic reference to his mention of the pit but sadly I think its an internal document from the Pott-Kompass importer. Nicht paginiert translates to not page numbered. This does mean producers knew what they were getting.

Pott-Kompass 1967, H.2, nicht paginiert [S. 12]

One of the other things to note in the Olbrich files is that, very much like the claims of Felton & Son’s New England rum pre prohibiton, there were many rums being produced that were just too heavy to drink. These were just bio reactors of rando-aromas for the confection, perfume, and tobacco industries. It is not completely clear where the line gets drawn between rum concentrates to be cut for drinking and the stuff solely for the nape of the neck. Olbrich does provide the best new clues.

A search for pineapple disease does yield some new clues to follow.

Fungus maladies of the sugar cane;: With notes on associated insects and nematodes (Report of work of the Experiment station of the Hawaiian sugar … of pathology and physiology. Bulletin) 1906 by Nathan August Cobb.

This is the most interesting reference so far, but sadly it is from Hawaii who barely produced rum so Cobb is not likely to have any distillery anecdotes. We are simply looking for references to being aroma-beneficial.

V. THE PINEAPPLE DISEASE.

(Thielaviopsis ethaceticus, Went.) This disease was first studied by Dr. F. Went, in Java. He first investigated and classified the fungus causing the disease. Since that time (1893) it has been observed in the West Indies and in Hawaii.

I’m going to pull out only the fun parts.

It is commonly asserted that this disease of the sugar-cane receives its common name on account of the fact that its presence in the tissues of the cane gives rise to an odor resembling that of pineapple. The specific name of the fungus, ethaccticus, refers to the same fact, its translation into English reading, “acetic ether,” so that we may call the species the acetic-ether-producing fungus.

It is true that, in some of its stages, and especially in some varieties of cane, the growth of the fungus gives rise to an odor reminiscent of ripe pineapple, but a delicate nostril would seldom, I think, mistake one odor for the other. More often the odor is that which we associate with fermenting fruit juice, due no doubt to a mixture of the vapors of various alcohols, acids, and ethers, prominent among which may be, and probably is, acetic ether. This odor of fermentation is the usual characteristic of most of the stages of the pineapple fungus as it occurs in cane. Only the later stages of the fungus attack are devoid of this odor, or if present, it is overpowered by others.

There is another reason why the name pineapple disease may be applied to this malady of cane, and that is that the same disease attacks the pineapple, as well as some other fruits. This fungus is, in fact, one of the serious diseases of the pineapple in some places. It is prevalent in the Hawaiian Islands on pineapples and does no small damage on some plantations.

The fact that the disease can be present in a most pronounced form without the odor of pineapple being noticeable renders the name a little unfortunate from the first point of view, but nothing can b’e said against the name from the second point of view, that is to say in view of the fact that the disease also attacks the pineapple. It is well to know that in fields where the disease is common one may often dig up and examine scores of cuttings without once detecting a pronounced odor of pineapple. As before stated, the variety of cane is one factor in the production of this odor. YellowCaledonia is one of the varieties that even when suffering acutely usually gives off merely an odor of fermentation. I have found the ethereal odor most pronounced in such varieties as the Striped Singapore, and in such canes the odor is sometimes much stronger than that of the most highly scented pineapple.

Okay, there are you go. We see that its aroma is also most pronounced in certain varieties. These are simply dimensions of rum connoisseurship and the rebooting of fine production we have yet to explore.

This differs from Arroyo’s mold which really turned out to be the ethyl tiglate producing yeast Saprochaete Suaveolens.

Also, if you remember Paraguay, and Empresa Azucarera, I would not be surprised if their uncrushed canes were simply pineapple disease infected canes tossed into a typical ferment. Good prices in Germany implies that it was a heavy rum.

A superior quality of rum is distilled in Paraguay, being made from uncrushed cane. Small amounts of this spirit have been shipped to Germany and, it is said, obtained good prices.

Traditional Fermentations of Molasses and Cane Juice in the French Antilles

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Parfait A., Sabin G., 1975. Les fermentations traditionnelles de mélasse et de jus de canne aux Antilles françaises. Industries alimentaires et Agricoles 92, 27–34.

 

Traditional Fermentations of Molasses and Cane Juice in the French Antilles

by A. PARFAIT et G. SABIN
institut National de la Recherche Agronomlque
Station de Technologie des Produits Végétaux – Petit-Bourg – GUADELOUPE

SUMMARY [their English]

A review is given here of studies which are in progress on rum fermentations in French Antilles. The influence of pH, yeast strain and sugar and nitrogen compounds concentrations are discussed.

Summary

This article reviews the ongoing studies of rum fermentation at French Antilles. The influence of pH, yeast strain and concentrations of sugar and nitrogen compounds is discussed.

Introduction

The fermentations of molasses and cane juice lead to wines which after distillation give rums. From 1900 to 1945, they were the subject of many observations which Kervegant (1946) summarized. Subsequently, there has been a lot of concern about rum analysis; but the results found have not always been related to the circumstances of manufacture. Under these circumstances, it was difficult to arrange or methodically modify fermentation techniques, either to improve the quality of finished products or to develop new types of rums.

1. FERMENTATION IN THE MANUFACTURE OF RHUMS

Fermentation is a stage in the production of rums from different raw materials. Figure 1 shows the main processes. In addition, when making rum “grand arôme” and in case we want to reduce the consumption of water, vinasse can enter the composition of musts. Tables 1, 2 and 3 provide a number of data on fermentations and rums.

Table 1
Some data on fermentations in the French West Indies

Table 2
Analysis of different samples of rum. For the agricultural rum, the figures represent the average of 30 examples, for the rum of molasse: 6 examples. The dosing methods used correspond to those of the official French method

1.1. RAW MATERIALS

The characteristics of the raw materials have variable values. They depend on the variety, the state of maturity of the cane and the conditions of harvesting and extraction of sugar. The average composition of the cane is as follows: water: 70%, ligneous material: 14%, sucrose: 14%, impurities: 2%. The woody fraction comprises the cellulosic material serving as a support for the plant. The impurities in the juice comprise crystallizable or non-crystallizable reducing sugars, organic materials and inorganic materials. Ammonia nitrogen is in the form of traces; the pH of the juice is of the order of 5. Molasses are viscous liquids with a density of 1.4 to 1.5 and whose pH is of the order of 6. They contain a significant amount of reducing sugars, i.e. more than 25% of the sugar fraction. Ashes can represent 5 to 6% molasses.

Table 3
Production statistics for Guadeloupe and Martinique
The production of cane is expressed in tons, that of rum in hectoliters of pure alcohol

1.2. LA FERMENTATION STRICTLY SPEAKING

1.2.1. The microflora in fermentation media.
Work in pure culture with given microorganisms is not practiced. Yeasting is carried out from a stock tank stocked with natural flora. An average value is given to the cell population by generally adding baker’s yeast. Several authors have commented on the strains that can be found in the rum industry. Pairault (1899), Kaiser (1916) identified Saccharomyces and Schizosaccharomyces. We have made an inventory of the yeast flora on the one hand to identify all the species present and on the other hand to select later those with good rum skills. Samples were taken at three levels: on the raw material, in the tank in full fermentation and on the wine. With suitable dilutions, two Petri dishes containing Wickheram medium at pH 4.5 were seeded. The different colonies were taken. The identifications were made following the method of Lodder and Van Rij. The survey was conducted during the 1970 campaign. It involved ten facilities on molasses and sixteen on cane juice. Two hundred colonies were analyzed. In addition to cryptococci, which do not ferment sugars and are contributed by earthy particles, the main recognized species are the same as those that El Tabey Shehata (1960) has shown in Brazil. Saccharomyces aceti and Saccharomyces chevalieri are important wild yeasts because of the number of times they are reported. Schizosaccharomyces pombe is present in musts leading to ‘grand arôma’ rum. In all other cases, Saccharomyces cerevisiae is the main fermentation agent for molasses and cane juice. The results of this survey are shown in Table 4.

TABLE 4
Presence of yeast species in fermentative environments. The number in the column represents the number of times that the yeast is corresponding (*) (*) Is reported in the medium considered.
Isolated yeasts            Raw material Must          Wine fermentation wine

In fermentative media, yeast concentrations are low. The corresponding dry matter often represents less than 1 g/l. The aeration systems in the tanks are poorly adapted to give a good multiplication of the agent chosen for the fermentation. Also, on cane juice, wild yeasts sometimes present in abundance, use some of the sugars and ammoniacal nitrogen. More seriously, they can supplant Saccharomyces cerevisiae in poorly maintained facilities.

The information available on bacteria in fermentations of molasses and cane juice are fewer. Kervegant (1946) reports the presence of acetic bacteria and butyric ferments. Maurel (1965) noted a high relative propyl alcohol content and traces of secondary butanol, facts that may suggest the intervention of bacteria in the manufacture of “grand arôma” rums. Reported sugar losses were linked by Tilbury (1970) to lactobacilli activity. We have with Dubois (1973) highlighted, in an agricultural rum acrolein derivatives that come from the metabolism of bacteria.

1.2.2. The pH.

The pH of the molasses is about 6, that of the cane juice 5. The pH values found in the fermentations are of the order of 4.5. They are acquired by addition of sulfuric acid and by the action of the acids contained in the yeast starter. Molasses has a high buffering capacity. In the laboratory, we found that for molasses concentrations of 200 g/l, 2 g/l of pure, concentrated sulfuric acid was required to increase the pH of the solution from 5.8 to 4.6. At the end of the fermentation, the ph drop is about one unit. These values of ph theoretically should protect fermentations against bacterial attack. This is not the case, as is shown by the amounts of volatile acids produced during the fermentation and the microscopic presence of many bacteria, even if they are not all active.

1.2.3. Total acidity and volatile acidity.

On rums from Martinique analyzed by gas chromatography, Nykanen (1968) found that acetic acid accounted for 75-85% of the volatile acids. For the remaining fraction, the propionic, butyric, caprylic and capric acids each represent about 3%. The other saturated fatty acids are in varying proportions. Unsaturated fatty acids are in trace amounts.

Table 5 shows the average acid values of molasses and cane juice.

The inhibition of yeasts by acetic acid results in decreases in the fermentation rate, the number of yeasts produced and the amount of fermented sugar. This is a well-known fact since the Muller-Thurgau experiments in 1885. Table 6 shows similar results obtained on a molasses fermentation at 100 g/l of sugar (Parfait, 1970).

 

TABLE 5
Average values of acidities in fermentations of molasses and cane juice

TABLE 6
Influence of acetic acid on fermentation of molasses
Culture medium: molasses q.s.p. 100 g/l sugar, ammonium sulphate 1 g tap water q.s.p. 1.000 L
Initial population in yeast (S. cerevisiae strain Berlin l) 2 X 10 ^ 6 / ml initial pH: 5 – Fermentation temperature: 28° C Fermentation time ten days
The fermentation rate is expressed in grams of fermented sugar in 24 hours in one liter of medium
The volatile acidity of the must without the addition of acetic acid is 8.2 mg/l.

Many factors and in particular the ecological conditions, the stage of maturity and the variety considered affect the nitrogen content of canes. Almost half of this amount remains in the bagasse after grinding. In cane juice 0.015 to 0.5% is found in nitrogenous matter. The mineral part and in particular the ammoniacal nitrogen is in the form of traces.

On molasses from Guadeloupe from different plants and collected at different times, we did the determination of total nitrogen (Kjeldahl method) and ammoniacal nitrogen (displacement method by magnesia). On average, the molasses used have 48 mg of ammonia nitrogen and 3.28 g of total nitrogen per 100 g of product.

The amount of nitrogen supplied by the raw material must be supplemented with a source derived from either mineral (phosphate and ammonium sulphate) or organic (urea) sources. Measurements made in factories show that in less than 24 hours the ammoniacal nitrogen has disappeared. We studied in the laboratory the use of some nitrogen compounds. The tests were conducted with Candida utilis. The samples are removed from the yeasts by centrifugation. Ammonium ions are assayed after being displaced by magnesia. Urea is assayed enzymatically. In a Warburg apparatus, in buffered medium at pH 5, the release of carbon dioxide produced by urease is measured. The results are shown in Table 7.

TABLE 7
Use of some nitrogen compounds, yeasts.
Culture medium: yeast carbon base 11.7 g, sucrose 100 g, nitrogen source, water q-s-p. 1,000 ml.
Fermentation temperature 28 ° C. Both ammonium polyphosphate I and II have different origins.

We have also found that the sharp and sudden drop in pH with ammonium sulphate hinders cell multiplication. If after 24 hours the pH is reduced to 5.2 with potash, the cell multiplication after a new latency phase resumes an exponential pace.

1.25. Sugar concentration and ethanol formation.

The Creole columns accept wines from 4 to 5° GL. Under these conditions, must concentrations are often less than 100 g/l in total reducing sugars.

There are three phases in the formation of ethanol. The formation speed gradually increases, it then takes an exponential pace, finally, it gradually decreases. After 30 hours, almost all the ethanol is formed. The by-products and in particular the higher alcohols form in parallel curves to those of ethanol. The non-alcohol of the rums has a great importance in the constitution of the aroma; it leads to choosing certain working conditions. To lower the higher alcohols, low sugar concentrations have been adopted. The figures giving the yields of the fermentation reflect the double desire to obtain the maximum of alcohol from the sugar and a non-alcohol corresponding best to the desired product. According to the Gay Lussac equation, 51.11 g of pure alcohol are obtained from 100 g of sugar.

Pasteur has established that secondary products are also produced and that the alcohol figure should be reduced by 5%. The theoretical yield is then 48.55.

We have estimated the performance of fermentation operations in an agricultural distillery. Figures range from 35.82 to 41.77 grams of pure alcohol per 100 grams of sugar. On average, the fermentation efficiency is 75 to 80 % in agricultural distillery.

1.3 DISTILLATION

There have been three generations of distillers in the French West Indies. The first stills used double distillation with recycling. In the following Systems, a bubbler or a condenser was introduced to obtain an alcoholic beverage on the first run. The continuous columns were modified in the French West Indies to give Creole columns accepting a wine of 4 to 5o GL. The losses in the vinasses are normally low. The characteristics of the column and the flavor of the product during distilling enable the operator to define and control the quality of the product obtained.

CONCLUSION

The fermentations of molasses and cane juice present particular aspects to the French West Indies. Cane juice occupies an important place among the raw materials. The production of traditional rum tends to increase while that of cane decreases for various reasons. Organoleptic analysis can recognize the characteristic flavor of rums, but it is not used rigorously to distinguish different types. The chemical analysis reveals a fairly great heterogeneity in the values taken by the non-alcohol constituents. The quality of raw materials, the conditions of distillation and especially those of fermentation are partly responsible for the aroma.

This traditional production of the French Antilles gives an important place to the art of the operator. The parameters of the fermentation were fixed either according to the natural conditions (temperature, flora) or according to the characteristics of the installation (apparatus to be distilled) or according to the quality of the product (addition of ammonium salts, low concentration in sugar) or according to simple bacteriological rules (acidification of the medium). However, it can be seen that all this does not guarantee the obtaining of a given product under regular conditions. These parameters must be defined according to more stringent criteria. Non-alcohol components are formed during fermentation. They each have a particular importance in the aroma. It will be necessary to study the mechanisms of their formation and to determine the factors which are responsible for their rate of presence in rums. From there, we will set the values ​​of certain parameters of fermentations. Yeast plays a key role in the development of aroma. The selection of strains, fairly good producers of ethanol and responsible for a particular aroma is to be made. Seeding at high levels of population under good aseptic conditions should be sought. Some products at very low concentrations in the rums contribute greatly to the aroma. The various means of analysis, in particular those of gas chromatography associated with mass spectroscopy, will have to be used to make their inventory. We will then study their origin and the circumstances of their presence at a certain rate in rums.

This traditional production of French West Indian rums represents only a small share of the world market. Light rums predominate largely on the latter. Generally, the production of these light rums uses: the treatment of the raw material, the use of selected yeasts, fermentations with must rich in sugars (130 to 150 g / l), distillations following precise techniques in continuous columns. There is more and more problems of manufacturing these products in the French West Indies. Several solutions are studied, the choice of which will be best suited to the French West Indies can be facilitated by the knowledge acquired in the field of traditional fermentations. We think in particular of the precise inventory of the substances of the aroma and the control of the contents with higher alcohols. We have accordingly oriented our research work in these directions.

 

BIBLIOGRAPHY

EL TABEY SHEHATA A.M. (1960). – Yeast isolated from sugar cane and its juice during the production of arguardente de cana. Applied microbiology, 8, 73-75.

KAISER A. (1916). — Contribution à l’étude des ferments alcooliques. Annales sciences agronomiques, 297, 322.

KERVEGANT D. (1946). — Rhums et eaux-de-vie de canne. Les éditions du golfe. Vannes, 512 pages.

MAUREL A., SANSONNET O. (1965). — Etude chimique et examen chromatographique en phase gaZeuse des rhums. Annales de falsification et d’expertise chimique, 868, 291-303.

NYKANEN L., PUPUTTI E., SUOMALANEN H. (1968). – Volatile fatty acids in some brands of Wisky cognac and rum. Journal of food science, 33, 8892.

PAIRAULT E.A. (1899). — Notes sur la fabrication du rhum à la Guadeloupe. Bulletin association des chimistes, 17, 246-255.

PARFAT A. (1970). — Observations sur l’acidité volatile des moûts servant à la fabrication du rhum aux Antilles Françaises. Notes et informations du C.T.C.S., 5, 1-9.

PARFAIT A., DUBOIS P. (1973), DEKIMPE J. —
PARFAIT A., DEKIMPE J., DUBOIS P. (1973). — Présence de dérivés de l’acroléine dans un rhum à goût anormal. Annales de Technologie Agricole – INRA (sous presse).

TILBURY R.H. (1970). – The ecology of Leuconostoc mesenteroides and control of post harvest by deterioration of Sugar cane in Jamaïca. Rapport du Tate and Lyle LTD. Research center.

 

Technology and Typical Elements of French Antilles Rums

Technology and typical elements of French Antilles rums

by Louis Fahrasmane, Berthe Ganou-Parfait, Francius Bazile, Paul Bourgeois

Summary
Rum technology and typicality factors in the French West Indies L. Fahrasmane, B. Ganou-Parfait, F. Bazile, P. Bourgeois

Rum has been produced in the French West Indies since the 17th century. The changes in production since then have been influenced by technical, economic and qualitative factors. Rum must not only have its typical organoleptic qualities but also be competitive on the international market, and this requires technological progress. Through yeast-strain selection, we have contributed towards improving alcoholic fermentation in Cane-sugar-based media. Rum production in the French West Indies is typified by the raw materials (molasses, cane syrup or juice), the microbiology of the fermentation media allowing bacterial activity, and the distilllation equipment with its so-called a Creole columns ) producing a range of aromatic strengths. Until the end of the 19th century, slops and froth were used in making the musts. Following Pasteur’s work, a new understanding of hygiene led to these substances being replaced by Water, with the result that Saccharomyces yeasts replaced the Schizo saccharomyces as alcoholic fermentation agents. Distillation equipment has also progressed. That used today depends on the type of rum produced (figure 1).

A rum technology involves several unit operations (1): the preparation of the must (more singularly called “composition”), fermentation, distillation and maturation. In the 17th century, the control of alcoholic distillation as a production tool became a key factor in the emergence of rum production, which appeared as a way of using the by-products of the sugar factory, especially those derived from sugar byproducts.

In the middle of the nineteenth century, came the agricultural rum, whose particularity is the use of musts based on sugar cane juice. This type of rum became, in the French West Indies, an export product, a production in its own right, distinct from the sugar factory, keeping, in some aspects, an artisanal character and having markers of recognition of its typicality related to the practices and production conditions.

In Europe rum has been defined since 1989 by the Community Regulation on spirit drinks (R. (EEC) No 1 576/89): It is the spirit drink obtained exclusively by alcoholic fermentation and distillation either molasses or syrups derived from the manufacture of cane sugar, i.e. sugar cane juice itself, and distilled at less than 96% vol, so that the product of the distillation has a perceptible specific organoleptic characters of rum. The minimum acquired alcoholic strength by volume is fixed at 37,5% (vol.).

The French national regulation (Decree of 22 April 1988 on Appellation of Origin rums) distinguishes “agricultural rum” from cane juice, “traditional rum” from molasses and syrup, and “rum grand arôme” which is a variant of traditional rum, more loaded with aromatic substances (Table 1). “Light rum”, once defined by national regulations, is no longer so today [2]. Distillates of different types can be delivered for consumption either in the form of untouched eau-de-vie or after ripening and dilution to the commercial level. They can also be matured longer or aged in wooden casks with a maximum capacity of 650 liters and for at least three years (Decree of 25 July 1963). Volatile elements other than ethanol must be at least 225 grams per hectolitre of pure alcohol.

Origins of rum production

The art of distillation dates back more than three thousand years, and it is thought that the Persians had discovered and implemented it to make rose water. The first stills were designed by Christians of Egypt, in 700 B.C.

The appearance of eaux-de-vie seems to have been preceded by that of alcoholic perfume, which began with the physician, philosopher and Arab alchemist, Rhases (864: 932). Around 1360, Hungarian water made from rosemary appeared. Perfumery developed later with Jean-Marie Farina (1685-1766), an Italian chemist who made the eau de Cologne created by his uncle Jean-Paul Feminis in 1690.

Wine brandy appeared in Europe, as medicine and the elixir of life, with Arnaud de Villeneuve (1235-1313) and Raymond de Lulle (1233-1315). Under the influence of the navies of Northern Europe (in particular Dutch), the distillation of white wines of the Charente became a common rural activity, leading to the development and marketing of Cognac and Armagnac in 1630 In 1624, the Corporation of Distillers was organized in France for the manufacture and sale of brandies. From the 18th century, the distillation of wine became a prosperous activity in France.

The appearance of rum production followed the development of cane-based sugar production (Saccharum officinarum L.) with natural hybrids on the American continent in the 17th century. The migration of sugar production from the Mediterranean area to the New World is linked to the capture of Constantinople by the Turks in 1453 and the expulsion of the Moors from Spain in 1492. There was a decline in the cultural influence of Arab origin and, with it, to that of cane, after about seven centuries of cultivation and sugar production in the islands and around the Mediterranean Sea [3, 4].

Holders of Genoese and Venetian capital, in search of new areas suitable for growing sugar cane, followed Christopher Columbus to the Americas. The expansion of sugar cane on the American continent triggered rum production as an annex to the sugar factory.

One of the first authors to talk about cane alcohol is the Father du Tertre who stayed in the Caribbean between 1640 and 1657. Father Labat, who arrived in the West Indies in 1694, describes at length the manufacture of the guildive in his New Journey to the Islands of America [title translated], The main characteristics of the rum production of the seventeenth and eighteenth centuries are:

– the use of byproducts, scums and molasses, resulting from crystallizing sugars (foams and deposits produced during the defecation of cane juice in sugar and syrupy residues of non-crystallizable sugar from the manufacture of sugar) as sources of fermentable sugar. The composition of musts at the beginning of the last century is shown in Table 2 [5];

– the spontaneous alcoholic fermentation due to the microbial germs having withstood the various operations of candying (concentration, cooked syrups) and those provided by the wooden bins used for fermentation. Fermentation lasted one to two weeks in the presence of abundant bacterial flora associated with yeasts of the genus Schizosaccharomyces. This microbial complex, of low productivity and generating very tasty products, was favored by the supply of pre-fermented vinasses (residues of distillations) during the storage, which lasted several weeks, and during which acidifying bacterial fermentations took place. The musts thus obtained were acidic, with a high osmotic pressure. Only yeasts of the genus Schizosaccharomyces were active under such conditions of the medium.

The first distillation apparatuses installed were discontinuous stills operating by recycling [my interpretation of what they mean by “repasse”] (figure 1a). Most of them consisted of a copper boiler, surmounted by a head also of copper; the time under heat of the fermented musts was important, which favored esterification. These devices allowed the elimination of negative volatile compounds of “heads”, sulfur and amines, and some of the very heavy compounds constituting the “tails”. The quality of the products obtained was often mediocre or frankly bad, because of the inferior quality of the raw materials used, the little care given to the fermentations, the non-rectification of the distillates which would have been necessary to eliminate the substances responsible for bad tastes. The best quality rum “is the one made only with molasses; but not the one in whose fermentation one leaves the debris of the sugar cane, the foam, etc., always preserves an unpleasant acid point and often contracts the taste of empyreanism, which causes it to be rejected from commerce. [6] ” During the eighteenth century, devices were used to obtain a sellable eau-de-vie on the first pass. [I think the end of this paragraph is translated wrong.]

Rum technology, from the beginnings of the industrial era to today.

Towards the end of the nineteenth century, the cane was selected for a better adaptation of the raw material to sugar technology. Saccharum officinarum carries many factors related to the richness of sucrose, fiber content and stalks vigorous and mechanically resistant.

The first artificial hybrid was produced by Fairchild in 1708. In 1880, the rediscovery of cane fertility led to scientific initiatives, with Slotwedel in Java in 1888 and Harrison and Bovell in Barbados. 1889, through intra and inter-specific crossings. The success of cane hybridization in Java, Barbados and Demarara, Guyana, has led to the proliferation of hybrid breeding stations around the world. Modern varieties derived almost exclusively from hybridizations arrived at the commercial stage eight to twelve years later, with selection criteria including agronomic traits, sucrose richness, disease resistance, specific locality characteristics, and so on. In 1921, Jeswiet obtained a hybrid clone nicknamed “the marvelous” (POJ 2878), far superior to the natural hybrid noble canes by its resistance to pathogens and its agricultural and industrial yields.

Increasing alcohol consumption among the working classes, economic liberalism, and the phylloxera crises that hit the wine liquors catalyzed, during the nineteenth century, the important development of the rum industry. Production, which had remained relatively low until the beginning of the nineteenth century (an average of 3 to 4 million liters per year for Martinique, Guadeloupe and Guyana combined), exceeded 21 million liters in 1892. This increase brought profound changes affecting the structure of the rum industry and manufacturing techniques.

Sugar plants appeared from 1865 and annexed distilleries for the treatment of molasses. Price and quantity constraints led to the establishment, in 1818 in Saint-Pierre in Martinique, in addition to stills without iron, single columns (Figure 1b), Creole type, to increase productivity. The Creoles columns are used to distill wines, or fermented musts, containing 4 to 5% (vol.) of ethanol. Ordinarily, they comprise three to five trays in concentration, which makes it possible to obtain distillates at 60-70% (vol.) of ethanol. Rum that is distilled too high loses its aromatic qualities. The equipment must have at least fifteen trays so that there is no loss of alcohol in the vinasse [stillage / dunder]. All the equipment parts (basement, sections and trays) can be made of stainless steel but it is very important that the concentration parts (trays and Goose neck) are made of copper. The oxidative catalysis of copper with respect to sulfur products has been demonstrated. This type of distillation device became widespread in the French Antilles around 1880; it no longer allowed the extraction of heads and tails significantly. Thus, the resulting eau-de-vie reflected the quality of the fermented must, with no possibility of correcting organoleptic defects. Later improvements to the apparatus, in particular the optimization of fractionation, multi-column device (FIG. 1c), made it possible to obtain products of a light nature, free from bad tastes, but stripped of volatile esters of interest.

Alongside the sugar plants, distilleries called “agricultural” were developed in the French West Indies and French Guiana, whose products became quite important from 1883 onwards. Some owners of old homes far from the sugar plants, rather than sell them their cannes, burdened with high transport costs or trying to obtain substandard sugar, found it more advantageous to turn their crops into rum by fermenting the juice either directly or after concentrating, which gave birth to agricultural rum.

The post-Pasteurian hygienist wave of the beginning of the century also concerned the rum industry and caused a stir. The need to replace spontaneous fermentations with pure fermentations [7] was concluded, and in 1913 a detailed study of rum yeasts [8] led to the promotion of pure fermentation with selected yeasts. The concern to improve productivity was decisive for researchers who thought that bacterial flora was detrimental to rum fermentation, whereas chemists attributed to bacteria an important part in the formation of the bouquet of high-flavored rums [9, 10].

The application of pure fermentations led to changes in conduct. First of all, the operating conditions of the spontaneous yeasts were reduced by lowering the density of the must and adding Sulfuric acid to lower the pH in order to limit the bacterial activity, and ammonium sulphate to complement the nutrient nitrogen medium. Then, the use of yeasts acclimated to certain antiseptics spread. Yield improvements were obtained, but the aromatic quality of the products was significantly reduced as they became more and more neutral. Most of the producers subsequently gave up the use of the selected yeasts and concluded, around 1920, the superiority of mixed spontaneous fermentations which made it possible to obtain rums that were more full-bodied, with a more intense and characteristic bouquet.

The use of scums and vinasse [dunder/stillage] in the composition of musts was gradually abandoned and replaced by water as a means of dilution. Currently, the vinasse is no longer used, except in the preparation of musts for the manufacture of rums of the high flavor type. For about fifteen years, the new element in rum technology is a supplement of the yeast flora of alcoholic fermentation, by supply of baker’s dry yeast, cheap and very available. We selected a strain of yeast for rum (Box I).

[Box 1]
Saccharomyces cerevisiae var. cerevisiae 493 EDV, a rum specific yeast.
The main technical characteristics of this strain are:
– optimal pH = 4.5; | – optimum temperature = 33° C;
– yield of alcoholic fermentation = 0.595 liters of alcohol pure / kg glucose equivalent (1 ap / kg glucose);
– ethanol productivity = 3.0 g / l / h.

This yeast improves the productivity and fermentative yield, compared to spontaneous fermentations and those carried out with supplemental yeast or bakery yeast supply and shows, with respect to this, a good occupation of the environment and a better rate of living cells. Possessing the “Killer” character, it inhibits certain types of yeasts. It keeps a good activity at a temperature of 36° C and does not cause the appearance of bad tastes in the products.

Yields of sugar-alcohol processing | usually obtained in distillery are relatively low (0.52 || ap / kg glucose on molasses and 0.47 || ap / kg glucose on cane juice), while the optimum yield is of the order of 0.60 | | ap / kg glucose.

We are working to improve the fermentative efficiency of yeast, by adding sterol extracts and candy defecation mud to the must [11].

Yeasts constitute above all a factor of technical efficiency of the sugar-alcohol transformation while having a part in the synthesis of the components and the precursors of aromas. This is the case, for example, in the formation of volatile fatty acids, the synthesis of which is modulated, depending on the strain, by the citric acid content of the raw material [12].
Saccharomyces cerevisiae var. cerevisiae 493 EDV, a rum yeast

Elements of typicity

White rums are presented under four types, three of which are defined in French regulations, by their non-alcohol content (or TNA) and the type of raw material used (table 1). If typicity is what characterizes a product and allows us to recognize it, we must look for elements of the typicality of rums in non-alcohol, or all compounds other than the water and ethanol that constitute it. The TNA of the aromatic rums of the French West Indies is generally higher than that of the light rum, the last of the four types, where the bacterial presence and activity are weak or non-existent. This parameter, however, remains lower than that of the rum aroma which is an archetype where the bacterial activity reaches a high level.

The bacterial flora is at the origin of the production of volatile acidity and precursors of aromatic compounds such as esters. Rums from environments where bacterial activity exceeds acceptable product quality limits have a high volatile acidity (> 15 mEg/l) [13] and contain undesirable substances such as acrolein [14] and butanol-2, markers of bacterial problems. The level of formic acid in rums can also be an element of appreciation of possible bacterial problems and, therefore, of quality [15].

The chemistry of the rums reveals propionic acid as singularizing the rum within the eaux-de-vie, because of the relatively high contents observed [16]. The level of propionic acid formation is related to fermentation yeast activity, which appears to be specific in cane-derived media [17]. Bacteria inventoried in distillery media contribute to the formation of propionic acid. They are Propionibacterium, Bacillus and Clostridium.

The alkylpyrazines seem to be of interest for the analytical differentiation between white and agricultural white rum. Given the thresholds of perception of these compounds, we think that they participate in the aroma of some rums with olfactory notes of brulee, caramel and leather [18].

The use of the Creole distillation column, determines the quantitative level of the TNA (it decreases when the degree of alcohol rises) by the low number of trays in concentration (three to ten ) which, itself, affects the degree of proof of the distillate by limiting it (60 to 80% (vol.) of ethanol); regulations allow distilling up to 96% (vol.) of ethanol.

Damascenone is present in molasses [19]. It has been shown that an isomer of this compound, at the same mass spectrum, has a characteristic odor of rum [20]. This ketone and its presumed isomer, identified in other products of plant origin, could, on the basis of quantitative considerations, be a differentiating factor of rums within eaux-de-vie.

The production conditions of rums (fermentation microbiology and distillation) as well as the raw material thus contribute to the development of their analytical typicity. There is a sensory analysis work to do to describe this typicity, by characterizing the components that will have to be looked after to improve the competitiveness of rums.

Perspectives and conclusion

In the French West Indies, rum production must, in order to remain competitive, adopt fermentation methods that leave no room for spontaneous fermentations. The aim of producing aromatic rums, whether from molasses or cane juice, is to follow rigorous protocols that take into account the following aspects: use of selected yeasts, control of the bacterial flora, rational choice of distillation parameters, quality management of raw materials, products and by-products (Box 2).

The reasoned conduct of fermentations by acidification of the musts (in order to contain, within appropriate limits, the presence and the activity of the bacterial flora), the control of the temperature and the use of selected yeasts for the rum production allow an active fermentation with a reduced latency. This way of operating gives way to a positive bacterial expression with regard to the quality and authenticity of the products.

The potential evolution of separation techniques – increasing the number of trays (twenty to thirty) in the Column Concentration, use of vacuum distillation, pervaporation and reverse osmosis – should allow the selective extraction of certain volatile compounds which, present in too large quantity, mask the expression of the notes of typicity. In this way, the latter could be better expressed, despite the decrease in TNA, according to pre-established product profiles in which ethanol becomes more and more a vector of aromas [22,23].

We must, moreover, seek to make the most of the quality signs (AOC, label …) related to the regulations. Although the rum was originally an annex to the sugar refinery, the same is not true of the agricultural distillery in the French West Indies, which is a full-fledged production structure whose qualitative requirements for raw materials could be considered in a specific way, depending on fermentation considerations and the aromatic properties of the products. Indeed, the production of agricultural rum could benefit from a raw material better adapted to its peculiarities than the hybrids used in sucrose production, by selection of varieties richer in non-sugar (nitrogen, phosphorus, magnesium …) and in aromatic precursors, in order to better meet the nutritional needs of fermentation agents and to reinforce the typicity associated with the raw material.

Acknowledgements

They are addressed to R. Pichy, M.L. Saint-Marc and C. Galas, from INRA Pointe-à-Pître, whose technical collaboration has been invaluable

[Box 2]
Wastewater treatment and environmental protection in the rum

The rum production generates residues (vinasse) with a high pollutant load (250 kg of COD / m * of pure alcohol in agricultural distillery, 1500 to 1900 kg of COD / mol of pure alcohol in molasses distillery). This sector of activity is more and more likely to integrate, downstream of its work plans, measures for the protection of the environment. To treat wastewater from rum, various methods can be used: evaporation-incineration, irrigation-spreading, anaerobic lagooning, biomass production and anaerobic digestion. The latter way, while reducing the organic pollutant load, produces combustible biogas. It allows, in the case of a molasses rum production unit, to decontaminate the effluent to 65% with biogas production, providing 60% of the energy necessary for the operation of the distillery. The wastewater treatment of the agricultural distillery, whose COD varies from 15 to 25 g / l with a BOD / COD ratio of 0.5, is easier than that of the waters of the molasses distillery, whose COD varies from 90 at 120 g / l with a BOD / COD ratio of 0.2 to 0.4 (21). Indeed, purification rates of the organic load of more than 90% are obtained.

Treatment of wastewater and environmental protection

Références

[1] Fahrasmane L. Rum. In : Encyclopedia of Food Science Food Technology and Nutrition. Londres : Academic Press, 1993 : 394 1-6.

[2] Borghese T. Rhum, rhum agricole, rhum traditionnel. Définitions légales. In: Actes du ColJoque Sur les rhums traditionnels. Pointe-à- Pitre : CRITT-BAC, 1994 : 51 – 4.

[3] Meyer J. Histoire du Sucre. Paris : éditions Desjon quère, 1989 ; 335 p.

[4] Fahrasmane L. Canne à Sucre et rhum à travers le temps. In: Actes du Colloque sur les rhums traditionnels. Pointe-a-Pitre : CRITT-BAC, 1994 : 330-5.

[5] Porter GR. The nature and properties of the Sugar Cane. Londres : Smith, Elder & Co., 1830 : 93-102.

[6] Le Normand. L’art du distillateur des eauxde-vie. Paris : éditions Chaignieau, 1817 ; 112 p.

[7] Pairault E. A. Le rhum et Sa fabrication. Paris : C. Naud, 1903 ; 292 p.

[8] Kayser E. Contribution à l’étude des ferments et de la fermentation des rhums. Ann Sci Agric 1917, 34:297-322.

[9] Allan C. Report on the manufacture of Jamaica. Sugar exp. Stat. Report, for 1905, 119-140. VVest Ind Bull 1906 7 : 141–2.

[10] Ashby SF. Studies of fermentation in manufacture of Jamaica rum. Inst Sug J 1909 ; 7 ; 243-51 et 300-7.

[11] Fahrasmane L, Bourgeois P. Apport d’extraits stéroliques de cire de Canne à sucre en fermentation alcoolique. In : Actes du Colloque Sur les rhums traditionnels. Pointe-à-Pitre : CRITT-BAC, 1994 : 128-35.

[12] Fahrasmane L, Parfait A, Jouret C, Galzy P. Production of higher alcohols and short chain fatty acids by different yeast used in rum fermentations. J Food SC 1985, 50: 1427-36.

[13] Fahrasmane L, Parfait A, Jouret C, Galzy P. Etude de l’acidité Volatile des rhums des Antilles francaises. Indus Alim Agr 1983, 100: 297-301.

[14] Lencrerot P, Parfait A, Jouret C. Rôle des Corynébactéries dans la production d’acroléine (2-propénal) dans les rhums. Indus Alim Agr 1984; 101: 763-5.

[15] Jouret C, Pace E, Parfait A. L’acide formique COmposant de l’acidité volatile des rhums. Indus Alim Agr 1990; 107 : 1239-41.

[16] Suomalainen H. Ouelques aspects généraux de l’arôme des boissons alcooliques. Ann Techno Agir 1975 ; 24 : 453-67.

[17] Fahrasmane L. Contribution à l’étude de la formation des acides gras courts et des alcools Supérieurs par des levures de rhumerie. Thèse de doctorat de troisième cycle. Université des Sciences et Techniques du Languedoc. 1983; 171 p.

[18] Jouret C, Pace E, Parfait A. Différenciation analytique des rhums agricoles et industriels par les alkylpyrazines. Annales des falsifications de l’expertise chimique et toxicologique 1994 ; 87: 85-90.

[19] Godshall MA. Minor constituents identified in the sugarcane plant and sugarcane products. SPRI short report 1984; 3; 9 p.

[20] Dubois P, Rigaud J. Étude qualitative et quantitative des constituants volatils du rhum. Ann Techno Agir 1975 ; 24 : 307 – 15.

[21] Bories A, Bazile F, Lartigue P. Traitement anaérobie des vinasses de distillerie en digesteurs à micro-organismes fixés. In : Actes du Colloque sur les rhums traditionnels. Pointe-à-Pitre : CRITT-BAC, 1994: 219-42.

[22] Escudier JL. La distillation des rhums : typicité et récupération d’arômes. In : Actes du Colloque sur les rhums traditionnels. Pointe-à-Pitre : CRITT-BAC, 1994; 187-205.

[23] Cogat P. Technologies applicables à l’atelier de distillation pour éliminer les composés négatifs, pour composer le profil aromatique. In : Actes du Colloque sur les rhums traditionnels. 1994 : 163–86.

[24] Maldonado O, Espinosa R, Rolz C, Humphrey AE. Technical details of a process to manufacture industrial alcohol from sugar cane. Ann Technol/ Agir 1975 ; 24 : 335-42.

Role of Corynebacteria in the Production of Acrolein (2 propenal) in Rums

Lencrerot P., Parfait A., Jouret C., 1984. Rôle des corynebacteries dans la production d’acroléine (2-propenal) dans les rhums. Industries alimentaires et Agricoles 101, 579–585.

Role of corynebacteria in the production of acrolein (2-propenal) in rum

by P. LENCREROT*. A. PARFAIT*, C. JOURET** avec and the technical collaboration of B. GANOU* and E. PACE**

* Station de Technologie des Produits Végétaux – Centre de Recherches lNRA Antilles-Guyane – 97170 Petit-Bourg (Gpe) ** Laboratoire de Technologie des Produits Végétaux – Centre de Recherches Agronomiques de Toulouse – 31320 Castianet-Tolosan

Summary

Some rums have an accidental taste alteration (“peppery” or pungent flavor) due to the presence of 2 propenal.

Among the bacteria isolated from an altered medium of fermented cane juice, a corynebacterium is able to degrade glycerol to give propenal and, at least in some cases, participates in this serious organoleptic defect in rums.

SUMMARY [their English]

Some rums present an accidental taste damage (pepper or purgent flavour) due to 2 propenal.

Among the bacteria isolated from an altered must medium, a corynebacterium is able to degrade the glycerol to produce 2 propenal and therefore, in some cases at least, it can play a part in the indesirable flavour in rums.

Certain eaux-de-vie, rums and cider brandies in particular, sometimes have an accidental taste change Known as “bitterness”, associated with a pungent “peppery” or “pharmaceutical” smell . RENTSCHLER and TANNER (1951) studying the organoleptic defects of these eaux-de-vie have shown that there is a relation between the impression of bitterness and the presence of an aldehyde: acrolein.

In wines having undergone various bacterial attacks, DUCLAUX (1874) had made a distinction between “turning” and “bitterness”; the former decreasing the total acidity, the latter increasing it by the formation of fixed acidity and volatile acidity. This observation already made it possible to advance the hypothesis that the precursor of the aldehyde responsible for this second fermentation accident is a neutral substance. Since then, the role of glycerol has been demonstrated and various diagrams of its biochemical degradation to give 2-propenal have been proposed among others by GOLFINE and STADTMANN (1960) AKADO et al, (1981), FREND, as well as SOBOLEW AND SHILEY Cited by WOOD (1961) RIBEREAU-GAYON (1977).

2-propenal often leads to derived compounds, including acetals, of which MISSELHORN (1975) specified the formation in rums.

DUBOIS et al. (1973) studying “acrolein” rums showed the presence of products derived from this aldehyde: 1-ethoxy-3-propanol and 1-ethoxy-1-1-2 propane. DE SMEDT and LIDDLE (1975) identified ethoxy 1-1 propane. It has been shown that even when present in trace amounts in eaux-de-vie, these compounds have a negative effect on the organoleptic quality.

The work of WALCOLLIER and LE MOAL (1932) on cider, those of SERJACK (1954) and DE MILLS (1954) on cereal fermentation media, led to the attribution of the action of lactic acid bacteria, the appearance of 2 propenal in the eau-de-vie. VOISSENET (1910) attributes, for its part, the presence of 2 propenal to the action of another bacterium: Bacillus amaracrylus.

PARFAIT and JOURET (1980) in their study on the formation of glycerol during the fermentation of sugar cane conclude that certain lactic acid bacteria and in particular a strain of Leuconostoc mesenteroïds, could metabolize glycerol to lead to 2 propenal; however, BIDAN (1967), PEYNAUD (1967) and RIBEREAU-GAYON et. al. (1977) consider that lactic acid bacteria only partially attack glycerol and that this degradation does not lead to propenal.

In the work presented here, we have tried to isolate microorganisms responsible for this fermentation by creating a so-called “acrolein” tank.

Experimental Protocol

We used a selective culture medium (G medium) in which glycerol is introduced as a carbon source. This environment has the following composition:
– glycerol: 20 g
– yeast extract: 10 g
– ammonium sulphate: 5 g
– distilled water – qsp. ; 1,000 ml

The pH is brought to 4.5 with concentrated sulfuric acid and then the medium is sterilized at 120° C for 15 minutes. Samples are taken aseptically, and then diluted with sterile physiological saline, and suitable dilutions are used to inoculate the Petri dishes containing the culture medium.

After petri dish development, we have been able to isolate a number of strains, several of which produce acrolein from glycerol when grown in selective medium G.

The microorganisms were identified in the medium according to the BERGEY method (1974) as being corynebacteria. The identification is completed in some cases using A.P.I galleries. We were able to group the different isolated strains among the following genera: Arthrobacter, Microbacterium, Brevibacterium, Corynebacterium.

Subsequently, we conducted further tests using a “Co” strain with more pronounced abilities to produce propenal; this strain belongs to the genus Corynebacterium.

In our fermentation trials, we used the “Co” strain either alone or in combination with a bacterium and / or yeast. The lactic acid bacterium used is a Lactobacillus fermenti (BERGEY) from the Pasteur Institute collection. The strain of Saccharomyces cerevisiae No. 493 from the INRA collection was isolated from a fermentative medium based on sugarcane (GANOU-PARFAIT B. 1978).

All microorganisms employed are first cultured separately on a liquid medium; Corynebacterium sp. on medium G, the yeast on WICKERHAM malt medium and the lactic acid bacterium on ROGOSA and SHARP medium.

Not having been able to control all the development parameters of Corynebacterium sp. on cane juice, we were led to use the glycerol-based synthetic culture medium (medium G).

Materials and Methods

The cells of the microorganisms from 500 ml of medium are harvested by centrifugation (3000 g for 10 minutes), washed twice with sterile physiological saline and resuspended in another 20 ml volume of physiological saline.

Aliquots are used to inoculate 500 ml of G medium and fermentation takes place for 72 hours at 36° C.

The analysis focused on the fermentation must and the distillate.

We measured 2-propenal and 2-propenol according to the method indicated by PARFAIT and JOURET (1980) as well as fatty acids and higher alcohols according to FAHARASMANE et al. (1983).

 

Quantities of certain alcohols and short fatty acids produced by Sacch yeast. cer. and the mixture of the 3 microorganisms on fermentation medium G.

 

Results

In Table 1, we reported the results of analysis of 2-propenal and 2-propenol on the 7 fermentation combinations. In Table 2, the interesting elements are indicated, only on two very different types of fermentation: using pure yeast, and using the combination of the three micro organisms retained.

It can be noted that the presence of Corynebacterium sp. is essential for the production of 2-propenal and 2-propenol;
with the Coryneform bacteria, the concentration of ethanol and propanol is half as high as in its absence; if the overall acidity varies little, as the level of butyric acid, on the other hand, the concentration of propionic acid is almost four times higher, and the C3/C4 ratio goes from 0.5 to 1.28 when the Corynebacterium intervenes.

Conclusion

Isolated, with various other microorganisms, from a cane wine with the characteristics of a basic product of rhum acroléine, Corynebacterium sp. tested degrades glycerol to give 2-propenal and 2-propenol and also modifies the composition of the short fatty acids of the medium.

Although it was not possible to reconstruct the fermentation accident in the laboratory with cane juice, we can, however, think that this bacterium is an agent, responsible in some cases at least, for the appearance of “spicy” rum.

This fermentative diversion can be clearly attenuated or even avoided by ensuring the sanitary quality of the sugar cane and by paying attention to the medium.

Bibliography

AKADO M., COONEY CI., SINSKEY A.J., (1981) Bioconversion of propionate to acrylate by Clostridium propionicum. Ad biotechnol (proc. Int, Ferment symp) 6th 1980 (pub. 1981)

BERGEY’S. Manual of determinative bacteriology. 8th édition. 1974. Ed. William and Wilkins.

B9DAN P. (1967), es facteursode la croissance des bactéries lactiques du vin. Fermentation et vinifications, 1, 165-213, 2° symposium international d’oenologie (Bordeaux).

DUBOIS P. PARFAIT A., DEKIMPE J. 1973. Présence de dérivés de l’acroléïne dans un rhum à goût anormal. Ann. Techno. Agric. 22, 131-135.

DUCLAUX (1874). Ann. Chim. Phys., 2, 233 et 3, 108 cité pa RIBEREAU-GAYON et al (1977).

FAHRASMANE L. (1983). Etude de la formation de quelques produits de la fermentation par les levures de rhumerie. (Publication en cours)

GANOU-PARFAIT B., 1978. La flore des milieux fermentaires en rhumerie. Nouv. Agron, Antilles-Guyane, 4, 161-273.

GOLDFINE H., STADTMAN E.R. 1960, Propionic acid metabolism. V – The conversion of Clostridium propionicum. The journal of biological chemistry vol., 235 n°8 2238-224.5

MILLS. D.E., BAUGH N.D. CONNEA H.A., 1954. Studies on the formation of acrolein in distillery mashes. Appl. Microbiol, 2, 913.

MISSELHOFRN K. 1975 – Formation of acetals in rum. A Kinetic study. Ann. Technol. Agric. 24, 371-382.

PARFAIT A., JOURET C. 1980. Le glycérol dans la fermentation alcoolique des mélasses et des jus de canne à sucre. Ind. Alim. Agric., 97, 721-724.

PEYNAUDE., 1967, Etudes récentes sur les bactéries lactiques du vin. Fermentation et vinifications, 1, 219-262. 2° symposium international d’œnologie Bordeaux-Cognac 13-14 Juin 1967. Vol. 1.

RENTSCLER H. TANNER H., (1951). Mitt gebiete lebens unters. hyg, 42, 463 cité par RIBEREAU-GAYON et al. 1977.

SERJEK W.C. DAY W.H., VAN LANEN J.M. BORUFF C. S., 1954, Acrolein production by bacteria found in distillery grain mashes.

RIBEREAU-GAYON J., PEYNAUD E. SUDRAUD P, RIBEREAU-GAYON P., 1977 – Dégradation du glycérol : maladie de l’amertume. Sciences et techniques du vin, tome 2 (pages 493495) ed. DUNOD.

de SMEDT P., LIDDLE P., 1976. Présence d’alcool allylique (propène 2 ol 1) et dérivés dans les eaux-de-vie. Ind. Alim. Agric. 93, 41-43

VOISSENET E. 1910, CR Acad, Sci 150, 40 et 1614, 151,518 et 522 (1911) Ibid, 153, 363 et 398, (1913) Ibid, 156, 1181 et 1410 cité par WOOD 1961.

WOOD W.A., 1961. In the bacteria. | Metabolism by Gunsalus l.C. Stanier R.Y. pages (59 à 149).

Problems Posed by the use of Schizosaccharomyces Pombe in the Making of Rums

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Ganou-Parfait B., Parfait A., 1980. Problèmes posés par l’utilisation de Schizosaccharomyces pombe dans la fabrication des rhums. Industries alimentaires et Agricoles 97, 575-580.

Problems posed by the use of Schizosaccharomyces pombe in the making of rums

by B. GANOU-PARFAIT and A. PARFAIT
Station de Technologie des Produits Végétaux
Centre de Recherches des Antilles et de la Guyane, INRA
97.17O PETIT-BOURG -Guadeloupe 

SUMMARY

Schizosaccharomyces pombe can be used like Saccharomyces cerevisiae in rum technology. Strains of S. pombe have been selected for microbiological and biochemical studies. A medium with cane juice is proposed. The rates of the fermentation can be increased with yeast concentration. According to the formation of the major volatile components S. pombe seems better than Saccharomyces; nevertheless other studies are necessary to confirm potentialities.

[this was supplied in English the French which I translate below comes out noticeably different.]

Summary
In the production of rums, strains of Schizosaccharomyces pombe and those of Saccharomyces cerevisiae are used. A selection was conducted to have a collection of Schizosaccharomyces pombe on which microbiological, biochemical and technological studies were conducted. A culture medium based on cane juice is proposed. Fermentation rates are generally low, but we want to accelerate the fermentations using significant seeding rates. The level of formation of the major components among the volatiles should give preference to Schizosaccharomyces pombe. It turns out that further work is needed to allow the reintroduction of Schizosaccharomyces pombe under the best conditions in fermentation media.

————————

In general, the use of the selected yeasts have several advantages in fermentations leading to alcoholic beverages. The choice of corresponding yeast species and strains obeys a certain number of criteria which are fixed, but it can also be the consequence of a given industrial situation. These particular considerations are for molasses, syrups and cane juices that are raw materials in fermentations leading to rums.

Kervégant (1946) collected a series of observations on schizosaccharomyces pombe in rum. This yeast was present in fermentations, especially molasses and syrup. Several species and several strains were known. In the production of rums, budding yeasts of the Saccharomyces cerevisiae type have been preferred to Schizosaccharomyces pombe because the former are in general faster.

The use of gas chromatography, alone or in combination with other techniques, makes it possible more and more to make a complete analysis of rums under conditions that are generally quite easy. It is therefore possible to propose quality criteria for rums: absence or presence of certain compounds at given concentrations. To meet these requirements it is sometimes necessary to resort to technological innovation.

Following our observations in the French West Indies, Parfait et al. 1975 and in view of current techniques used elsewhere in the world – Kampen (1975) – it is likely that the rum industry will experience such a situation. It is therefore reasonable to envisage the reintroduction of Schizosaccharomyces pombe in fermentative media.

SELECTION OF STRAINS OF SCHIZOSACCHAROMYCES POMBE

These yeasts are common in tropical environments. Several authors have reported them in fermentations of derivatives of sugar cane. Like all Schizosaccharomyces, the essential physiological characteristic is division by fission. The spherical to cylindrical cells are often larger than those of other yeasts and in particular those of Saccharomyces cerevisiae. We first made microscopic observations. The identification of colonies obtained after plating a colony on Petri dishes is done by the method of Lodder and Van Rij. The Schizosaccharomyces pombe cells are practically absent in fermentative environments in the French West Indies, except in the case where heave-flavored rums are manufactured. These same cells are found in certain soils where sugar cane is cultivated, but there they are in very small numbers. They are much larger in the fermented musts of small distilleries in Haiti. For decades, they have not changed their manufacturing conditions, and they are often isolated in the middle of the countryside. We can therefore estimate that the modifications of the flora have been practically nil. In all cases, to facilitate the selection of strains of Shizosaccharomyces, different properties are used Ganou-Parfait (1979).

Some are mentioned below:

Table 1

Use of citric acid by S. pombe. (+) low growth, (-) no growth The concentration of citric acid is 0.5%

Table 2

The influence of butyric acid on a mixture of yeasts. The seeding rate, 1 × 10 6 / ml for each yeast, Count of revivifiable cells after 76 hours of culture at 30° C. in a medium with malt extract containing 150% of sucrose.

Sensitivity to Acids.

In rums, in general, acetic acid is the most important constituent of the acid fraction, of which it accounts for nearly 80% of the total. For a type of rum represented by the large aroma rums, the butyric acid fraction is also significant. We compared the behavior of Schizosaccharomyces pombe and Saccharomyces cerevisiae strains in the presence of varying amounts of different acids. The comparison was made either aerobically or anaerobically, and the determination of the number of total germs by the Malassez cell made it possible to measure the sensitivity of yeasts to acids.

In the case of citric acid, 10 ml of medium are placed in test tubes. In each case, citric acid is added at a concentration of 0.5%. The results are obtained on strains of Schizosaccharomyces pombe, they are shown in Table 1, and are not better if the citric acid is replaced by malic acid. Note that some authors have found that in the case of the latter acid, there is a sharp reduction in the growth of Saccharomyces cerevisiae for concentrations ranging from 0.2 to 0.4% malic acid.

The influence of increasing amounts of acetic acid on yeast growth is well known. There is a slowdown in the fermentation rate and a decrease in the amount of sugar used. The results are identical with butyric acid.

To compare the influence of the latter on a mixture of Saccharomyces cerevisiae and Schizosaccharomyces pombe, we made a count of total germs after sixteen hours. The medium used is the following: 10 ml of malt extract supplemented with sucrose at a rate of 150 g/l. The seeding rate is 1 X 10 6/ml for each yeast. The butyric acid slows down according to the concentration used, the growth of Saccharomyces cerevisiae and has a variable effect on that of Schizosaccharomyces pombe. In case we want to make a selective medium to isolate Schizosaccharomyces pombe in view of previous results the addition of 0.25% butyric acid could be a formula. However, experience shows that the results obtained on a liquid medium are not transposable in a solid medium. In all our experiments, the growth of Saccharomyces cerevisiae has always superseded that of Schizosaccharomyces pombe.

Search for a favorable environment for Schizosaccharomyces pombe

During microscopic observations, it was found that in natural samples, Schizosaccharomyces pombe cells had a granular appearance which disappeared after multiplication of the cells in a favorable medium. We have sorted in a large number of culture media used for yeasts. During this operation the following conclusions were reached:

Peptones represent a better source of nitrogen than ammonium salts. In fact, when comparing the ammonium salts with each other, it is found that the acidity induced by the anion is an essential factor. The optimum pH is of the order of 5, but the pH range is from 4 to 6. Sucrose is better assimilated than glucose.

We compared several synthetic media: Wickerham malt, Czapek, Dox Agar, Davis Yeast Salt Agar, malt extract. This last medium supplemented with sucrose gives the best results. We made different media formulas from cane juice and molasses. It is with cane juice that we have the best results. We therefore propose the following medium:

— Peptone = 1 g
— Ammonium Sulfate = 2 g
— Cane Juice = 1.000 ml.

The pH is adjusted to 5, sterilized at 120 ° C for 15 minutes. In order to reduce the importance of flocculation during sterilization, peeled sugar canes are used.

Selection Results

Of all the samples we have studied, we have extracted a strain of yeast to make rums of great aroma, and sixty strains from different media collected in Haiti. Beside these last strains, we also found about ten strains of Schizosaccharomyces malidevorans. This species is easily distinguished because it is the only Schizosaccharomyces that does not use maltose. It should be noted that Schizosaccharomyces sporulate with difficulty, whatever the medium used.

GROWTH OF STRAINS OF SCHIZOSACCHAROMYCES POMBE

Generally, Schizosaccharomyces pombe is considered to have a low growth rate. In order to get closer to the industrial criteria, we used the procedure below to compare the strains.

The cells proliferate for 72 hours at 30° C on agitated Wickerham malt medium. They are recovered by centrifugation 3,000 tr/15 minutes, and then washed. After counting, a molasses-based medium is inoculated at 1 X 10 6 yeast / ml. The fermentation is carried out in 125 ml flasks closed with a rubber stopper crossed by a tapered glass tube at one end, and plugged at the other end with carded cotton.

The environment is as follows:

— Molasses 300 g
— Ammonium Sulfate = 1 g
— Water q.s.p. = 1.000 m)
— pH = 5.2, sterilization 15 minutes at 110°C.

The fermentation curves are plotted in Figure 1. From this examination it appears that the lag phase is longer for Schizosaccharomyces pombe and that overall the fermentation rate is lower than that of Saccharomyces cerevisiae.

But fermentation rates can be varied by increasing seeding rates. The tests are conducted under the same conditions as above, with different seeding rates determined by the dry matter. Fermentation rates are conventionally represented by the mass losses of each vial after 24 hours.

Table III Fermentation speed for increasing rates of seeding with Schizosaccharomyces pombe

We find in Table III results similar to those we found with S. cerevisiae. It can therefore be estimated that for large seeding rates (2 to 5 g/L) the behavior of these two yeast species is close.

Use in Industrial Fermentation

For twenty years, there has been an interest in the use of Schizosaccharomyces pombe to deacidify wines, Bidan (1974). During this operation, the malic acid is converted into ethanol.

In his important work on rums, Arroyo found that both species Schizosaccharomyces pombe and Saccharomyces cerevisiae could both provide good products. In making rums, he advocated the second because it fermented faster. Recently, Rose (1976) has selected S. pombe strains from yeasts that can produce from molasses musts an alcohol content of 11° to 12° GL. Such a concentration of ethanol makes it possible, compared with conventional methods, to reduce the quantities of energy required during distillation relative to wines of 4-5 ° GL.

Today, S. pombe is a fermentative agent of cane molasses next to several Clostridia including Clostridium acetobutylicum in the manufacture of grand arôme. This type of rum in the French West Indies is characterized by a high level of non-alcohol (800-1,800 g/hl pure alcohol). In detail, there is a significant fraction of ethyl acetate and acetic acid, about 300 grams for each term and a small amount of higher alcohols — less than 100 g — with a abundance of n-propanol. The musts are composed with vinasses [stillage or dunder] that have surely undergone the phenomena of pre-fermentation. They are rich in volatile acids and in fixed acids. Fermentations are slow and must involve different metabolic pathways that have not yet been fully elucidated.

Products of Fermentation

Among the compounds found in rums, some are already present in molasses as a result of various more or less advanced prefermentations. But yeast is mainly responsible for their formation during the alcoholic fermentation. In various previous works, Parfait (1977-79), we have studied certain products of the molasses fermentation by S. pombe, which can be referred to for the various procedures.

a. Ethyl esters of higher fatty acids

Pombe produces more of these compounds than most baker’s yeasts of the species Saccharomyces cerevisiae, but some good yeasts in our collection belonging to this species have equivalent productions to S. pombe. This production is related (FIG. 2) with the cell growth yield that can be appreciated by the ratio of final yeast to initial yeast. For seeding rates between 0.1 and 5 g /L yeast dry matter, there is a correlation between the amount of ester produced and the cell yield. This proportionality is also checked for each ester in the series.

b. Ethyl Acetate

In quantity, ethyl acetate is the main ester of rums made with Saccharomyces cerevisiae. Under the same conditions of fermentation and distillation S. pombe brings a double production, 100 g/hl of pure alcohol instead of 50 g/hl pure alcohol. In industrial rums of high aroma type, the production is very strong, more than 300 g/ hl pure alcohol without the production of ethyl esters of higher fatty acids is affected. Esterification is primarily a biochemical phenomenon, distillation in the presence of yeasts can increase the levels of ethyl esters of rums, but their formation involves acetyl CO A. If we are inoculating a must of molasses with a mixed culture of Schizosaccharomyces pombe and Clostridium acetobutylicum, the presence of the bacterium has the effect of increasing the amount of ethyl acetate formed. One may wonder if under certain culture conditions, especially in musts leading to rums of high aroma type where the medium is already rich in acetic acid, there is no different functioning of esterase.

c. Higher Fatty Acids

Caprylic and capric acids are the major constituents of this fraction of rums made with Saccharomyces cerevisiae or Schizosaccharomyces pombe. Temperature and pH affect the total production of higher fatty acids, just as they affect the general activity of yeast. Depending on the sugar concentration, fatty acid concentrations increase in rums made from Schizosaccharomyces pombe, except for caprylic (Table IV).

Table IV: Influence of molasses concentration on the formation of higher fatty acids. The results for the fatty acids are expressed in mg/l of pure alcohol. The initial seeding rate is 3 g/l

d. Acetic Acid

In pure culture, the productions of acetic acid are comparable for Schizosaccharomyces pombe and Saccharomyces cerevisiae. The high levels of acetic acid found in the aroma of rums originate from the vinasses which enter into the composition of the must and the further extraction during the distillation.

e. The Higher Alcohols

We have already reviewed the mechanisms of formation of higher alcohols, Parfait (1975). The Schizosaccharomyces pombe strains generally provide fewer higher alcohols than those of Saccharomyces cerevisiae, and this with a predominance for n-propanol.

DISCUSSION

A number of questions arise when using Schizosaccharomyces pombe in the production of rums.

The growth of yeast is lagging at low seeding rates. It is possible to accelerate the fermentations by increasing this rate. This technological device must not obscure the different physiological behaviors of Saccharomyces cerevisiae and Schizosaccharomyces pombe. In the study of a suitable medium for the culture of this last yeast, we found that sucrose was better than glucose. This result was explained by Hayashibe (1973). The growth curves are not the same for glucose and mannose on the one hand, and sucrose on the other hand; but fermentation rates are the same when using cell extracts. It can therefore be linked to sugar transport phenomena. Billon-Grand (1977) demonstrated the existence of intracellular enzymes 1α and β glucosidases and invertase or β fructofuranosidase, capable of degrading these sugars. As often in this case, the transport of sugars is facilitated by the addition of NH4 + ions in the medium. It should be noted that Schizosaccharomyces pombe does not use glycerol and ethanol. This difference with Saccharomyces may partly explain the high levels of glycerol. Many studies have been done on the influence of oleic acid and sterols in the anaerobic metabolism of several yeasts. Very little data has been established on the fatty acid and lipid composition of yeasts of the genus Schizosaccharomyces. Their obtaining will thus explain the importance of the lag phase for these microorganisms. Bush (1977) has confirmed the absence of mannan which plays a role in the budding process of several yeasts, but the presence of galactomannan raises the question of the nature of the compounds that play a role in the fission process. Similarly, there is a difference in the plasticity of the cell wall and its protective role vis-à-vis the cellular content. Ultimately, the composition of the cell membrane and its impact on Schizosaccharomyces pombe metabolism are important enough to explain the differences in physiological behavior with Saccharomyces.

Of the volatile compounds produced during fermentation by Schizosaccharomyces pombe, special mention must be made of ethyl acetate and higher alcohols. Part of the ethyl acetate arises as a result of the oxidative decarboxylation of pyruvic acid and an alcoholysis reaction:
(1) CH3CO COOH — NAD — CoA v SH –———>

CH3CO v SCoA + NADH2 + CO2

[not sure about this notation and what the italicized “v” stands for]

(2) CH3CO SCOA + CH3CH2OH ——->
CH3 COOC2H5 + HS v CoA

But the high concentration of acetic acid that exists in some musts may explain the formation of ethyl acetate by shifting the equilibrium during the reaction.

(3) CH3 COOH + CH3CH2OH ⇔ CH3 COOCH2
CH2 + H2O

We will undertake enzymatic and kinetic studies of these three reactions to justify the different levels of ethyl acetate found in rums.

The amounts of each higher alcohol manufactured by Schizosaccharomyces pombe are quite remarkable: low levels of methyl-3-butanol. 1, methyl 2 – butanol 1, and isobutanol, against a higher propanol content. The latter is manufactured in the following way:

Thréonine –» amino acid – 2 butenoïque –» acide
thréonine déhydratase
deaminase σ cétobutyrique –> CO2 — τn propanaldéhyde –>
n propano
décarboxylation déhydrogénase

[I’m insecure about translating this section. Any help? and background on it?]

This route for propanol is specific, even though it contains σ ketobutyric acid, which is a key intermediate in the biosynthetic formation of other higher alcohols. We have done a nearly complete study of the formation of higher alcohols in rums. It appears necessary, in the case of Schizosaccharomyces pombe, to determine the variations of the amino acid pool, taking into account the ambient factors and in particular the nitrogen diet during the fermentation.

Without waiting for its results, our first observations – Pafait, (1977) – showed that by means of the acceleration of fermentations can ferment with Schizosaccharomyces pombe molasses musts containing 150 to 180 g/l of sugar under conditions as well as can Saccharomyces cerevisiae. For this last yeast, it appears that the choice of the strain is determining in the level of formation of the volatile products and in the fermentative efficiency. This is also the case for Schizosaccharomyces pombe and some strains show, in particular, a very low fermentative efficiency. The properties of these yeasts begin to be explained through different biochemical studies.

Here we have specified a number of pathways (cell membrane formation and metabolite transport, kinetics of ethyl acetate formation, composition and amino acid pool variation) that are promising. Besides this, an organoleptic study of rums is needed. We chose the technique of Micko – Parfait, (1979) – for the tasting of rums and cane spirits. Equivalent fractions may have a different flavor depending on the yeast and the strain that served as the fermentation agent. The perception thresholds of each constituent are not the same depending on whether they are used alone or in association with other bodies. Nowadays, the distillates obtained from Schizosaccharomyces pombe and Saccharomyces cerevisiae have different contents, at least for the main constituents: higher alcohols, aldehyde and ethyl acetate. Finally, it is as a result of various technological operations, fermentation, distillation, assembly, maturation that the rums obtained from Saccharomyces cerevisiae present a composition and a favor given. The reintroduction of Schizosaccharomyces pombe in fermentation media will allow these different operations to be carried out under other conditions to obtain products equivalent to those which now exist.

CONCLUSION

Different studies have shown that the current compositional criteria for rums can be more easily achieved with Schizosaccharomyces pombe as a fermentative agent, rather than Saccharomyces cerevisiae. The selection of strains of this first yeast, even in favorable ecological environments, has only been made possible by a study of some of its microbiological and physiological properties. The use of Schizosaccharomyces pombe in musts made from molasses and sugar cane juice poses a series of biochemical, technological and organoleptic problems whose solution lies in a better knowledge of the metabolic pathways. This preliminary work made it possible to determine the axes that will be the subject of future research.

BBLOGRAPHE

G. BILLON-GRAND (1977). – Recherche d’enzymes intracellulaires dans le genre Schizosaccharomyces, lmplications systématiques. Mycopathology, 61 (2), 111-115 

P. Bidan (1974). — Les Schizosaccharomyces en CEnologie.
Bull. OIV, 47 (523), 682-706.

D.A. BUSH, M. HORISBERGER, I. HORMAN, P. WURSCH, 1977. – The wall structure of Schizosaccharomyces pombe. Nestlé research News, 73-77.

M. HAYASHIBE, N. SANDO, Y. OHBA, K. NAKAMURA, K. DKA., K. KONNO, M. GOYO (1973). — Utilisation of hexoses in fission yeast. Proceedings of the 3rd international specialized symposium on yeast OTANIEN. Helsinki Part, II, 91-102.

B. GANOU, PARFAIT, 1979. — Les microorganismes des fermentations de mélasse et de jus de cannne. 1979 (en préparation).

D. KAMPEN (1975). – Technology of the rum industry. Sugar y azucar, 70 (8), 36-43.

KERVEGANT (1946). – Rhums et eaux-de-vie de canne. Les Editions du Golfe, Vannes.

A. PARFAIT (1972). — Les esters éthyliques des acides gras supérieurs de rhums. Ann. Technol. Agric.., 21 (2), 199-210.

A. PARFAIT (1975). – Formation des alcools supérieurs dans les rhums. Ann. Technol. Agric., 24 (3-4), 421-436.

A. PARFAIT et G. SABIN (1975). — Les fermentations traditionnelles de mélasses et de jus de canne aux Antilles françaises. Industries alimentaires et agricoles, 2 (1) 27-30.

A. PARFAIT (1977). — La fabrication des rhums. Rapport d’un contrat DGRST, No 74-7-09-06.

A. PARFAIT (1979). — Suite de l’étude sur la fabrication des rhums. Rapport d’un contrat DGRST, No 77-7-03-55.

D. ROSE (1976). – Yeasts for Molasses alcohol. Process Biochemistry, 12 (2), 10-16.

Fermentation Properties of Rhumerie Yeasts

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Fahrasmane L., Parfait A., Galzy P., 1986. Propriétés fermentaires des levures de fermentation. Industries alimentaires et Agricoles 103, 125-127.

Fermentation Properties of Rhumerie Yeasts
by L. Fahrasmane*, A. Parfait*, P. Galzy**

* Station de Technologie INRA-Antilles – Domaine Duclos 97170 Petit-Bourg
** Laboratoire de la chaire génétique ENSAM, Place Viala, 34060 Montpellier

Introduction
Fermentations of molasses and sugar cane juice take place in the West Indies, in a non-sterile environment (Parfait and Sabin, 1975). Formerly, the dominant yeast species was Schizosaccharomyces pombe Lindner. This yeast is osmophilic and often gives rums of quality in association with an abundant bacterial flora. She was most often supplanted by Saccharomyces cerevisiae Hansen. The latter species, baker’s yeast, was commercially available in bulk and at low prices. It was therefore tempting for manufacturers to regulate and accelerate fermentations by massive sowing of baker’s yeast.

The purpose of this note is to compare the fermentative properties of these two species which are still the pivot of rum fermentations. We will not present here the result of a particular experiment, but rather a synthesis of several independent studies carried out on laboratory strains in sterile medium (Parfait et al., Perfect et Jouret, 1975, 1979, Fahrasmane, 1983, Fahrasmane et al., 1985); these results are discussed in the light of numerous industry observations and long experience in making rums acquired by some of us.

Material and Methods

1. Biological Materials
Most of the works summarized or cited here have been done with a large number of Strains. However, to simplify the presentation we have limited ourselves voluntarily to give results of a strain of each species considered representative. These two strains are:

– Saccharomyces cerevisiae listed 493,
– Schizosaccharomyces pombe listed G.

2. Culture Media
We used a cane juice (vesou) from natural and health canes, diluted to 100 g/l of sugar; a molasses-based medium also reduced to 100 g/l of sugar and a synthetic medium according to Oura (1974) supplemented with the main organic acids of cane juice according to Fahrasmane (1983).

3. Analysis Techniques
We used the Classic methods of rums study, including:

-the official method of assaying the higher alcohols in the eaux de vie (Fraud Control, Anonymous, 1973).
-Jouret’s method for the determination of short chain fatty acids described by Fahrasmane et al. (1983).
-The method described by Parfait et al. (1972) for the determination of ethyl esters of higher fatty acids.

Experimental Results

I. Biomass and ethanol production

Schizosaccharomyces pombe generally gives slow growth and a relatively small amount of biomass, much lower than that obtained with Saccharomyce cerevisiae (table 1). The difference between the two species fades in the case of a mixed culture. It Seems that Schizosaccharomyces pombe has special nutritional requirements that it does not find on synthetic medium or on cane juice (Vesou); on the contrary, it finds them in the much richer environment constituted by molasses. This result suggests difficulties in all industrial uses of Schizosaccharomyces pombe. The addition in a synthetic medium of the organic acids of the cane juice, in particular of cis-Aconitic acid, causes an abundant cell multiplication. This result suggests that these acids activate cell multiplication by probably intervening in the Krebs Cycle. He also explains that the yeast populations observed in crops on cane products are still exceptionally abundant. Correlatively, the yield of ethanol is not very good in rum fermentation.

The yield of ethanol expressed as a percentage of the Pasteur yield is always higher, in pure culture, for Schizosaccharomyces pombe than for Saccharomyces cerevisiae. This observation very largely explains the current craze of certain distillers who recommend the use of Schizosaccharomyces pombe.

The fermentation times are always very long for Schizosaccharomyces pombe. As a result, the fermentation medium is always more sensitive to bacterial contamination. The duration of the fermentation become extremely long on synthetic medium; the use of Schizosaccharomyces pombe for fermenting new substrates in relatively poor environments certainly has an indisputable randomness.

It should be noted that Schizosaccharomyces pombe produces significant amounts of glycerol (8 to 10 g/l per 100 grams of fermented sugar); under the same conditions, Saccharomyces cerevisiae produces only 2-3 g/l (Parfait and Jouret, 1980). Given the large bacterial flora able to attack glycerol in rum fermentation, this character is certainly a serious problem for the use of Schizosaccharomyces роmbe.

II. Formation of Higher Alcohols

Using the same culture media we studied the higher alcohols produced by both strains (Table II).

Schizosaccharomyces pombe produces far fewer higher alcohols than Saccharomyces cerevisiae. However, it appears again here that Schizosaccharomyces pombe is more sensitive to environmental conditions than Saccharomyces cerevisiae. While the latter species gives total higher alcohol concentrations substantially independent of the culture conditions, Schizosaccaromyces pombe produces twice as much higher alcohols in molasses culture than in the other Crop Conditions tested.

III. Formation of Volatile Fatty Acids

Again (Table 3), Schizosaccharomyces pombe produces much less short-chain fatty acids, important constituents of the aroma of rums, than Saccharomyces cerevisiae. It is worth mentioning that both species produce propionic acid on cane juice medium. Only Schizosaccharomyces pombe produces acrylic acid; it is probable that propionic acid is the precursor of acrylic acid. It is also likely that sugarcane media contain a propionic acid precursor for use by both yeasts.

In cultures on product derived from sugar cane (molasses) it also appears in the medium of long chain fatty acids C8 to C16 as well as the corresponding ethyl esters. Fermentation of 100 g of sugar yields about 80 to 100 mg/l of these esters regardless of the yeast species used (Parfait et al., 1972).

Conclusion
Schizosaccharomyces pombe presents in the laboratory the considerable advantage of giving a high yield of ethanol; it also has the advantage of giving relatively few higher alcohols and fatty acids. In fact, it seems obvious that these two advantages are related. Low cell growth, partly indirectly responsible for good ethanol yield, is not only beneficial; a slow and slow growth of the yeasts largely leaves room for bacterial developments. The abundant production of glycerol is also a favorable factor for the development of many germs, some aroma beneficial, other sources of manufacturing flaws. These general properties should make Schizosaccharomyces pombe a good strain of rum fermentation: it is able to give very aromatic rums with a good bacterial flora; it could give very light rums, particularly sought after, as long as one manages to control the flora; unfortunately manufacturing flaws can occur.

In recent years, it has been sought by industrialists for new substrates for the production of ethanol. Schizosaccharomyces pombe could a priori be suitable for the production of alcohol for chemical use or rectified alcohol as only a few secondary products are formed. The results we have presented show that this species is very demanding from the point of view of growth needs. This can result in a significant over-cost related to the need to complement the new fermentation media. The relative fragility of the fermentative medium with respect to bacterial contamination is also a disadvantage that should not be underestimated.

Saccharomyces cerevisiae gives higher amounts of higher alcohols and fatty acids; the yield of ethanol is a little lower than that observed in Schizosaccharomyces pombe. But the growth is fast and abundant, the occupation of the ground is good, the danger of serious bacterial accidents is reduced. This species ultimately makes it possible to obtain relatively light rums. For fermentations of new products, this species has definite advantages, provided that the substrate to be fermented is accessible (hexose, sucrose or maltose).

The characteristics of these two species explain fairly well the evolution of the rum fermentation technique. In the past, rum was prepared almost exclusively from molasses.

Vinasses [dunder or stillage] were recycled as a means of diluting molasses. Thus the fermentation medium was rich in mineral salts, nitrogenous matter. Osmotic pressure was important. This medium was favorable to Schizosaccharomyces pombe which was naturally selected. This system also favored the preferential proliferation of heat-resistant, sporulated, anaerobic bacteria. This resulted in a very particular type of rum. The sugar crisis helped, it was made more and more of direct fermentation of Vesou [fresh cane juice]. The osmotic pressure became much weaker here. The medium was poorer in biotic elements and lacked nitrogen feed for the yeast. Under these conditions, it was inevitable that Saccharomyces cerevisiae would replace Schizosaccharomyces pombe. In the same way, the dominant bacteria flora became naturally present on sugar cane: aerobic Coryneform bacteria, aerobic Bacillus, and lactic flora. The Yeast defend better against this type of flora, it resulted in a lighter rum and better suited to current consumption. It seems clear to us that the lessons learned from a reflection on the rum industry are not without interest for other ethanol manufacturing industries be it alcohol, alcohol for industrial use or alcohol fuel.

It would be useful to better understand the nutritional requirements and the general metabolism of the fermentation strains of these two species. This work becomes more and more necessary as the variety of used substrates expands. Let’s mention in the case of rum the range of raw materials: vesou, juice defecated, syrup and molasses at various stages including molasses final.

[The vesou here as opposed to defecated juice may refer to what Cape Verde uses which isn’t centrifuged and strained.]

Bibliography

FAHRASMANE L. – 1983 – Contribution à l’étude de la formation des acides gras Courts et des alcools supérieurs par des levures de rhumerie. Thèse de 3° cycle. USTL Montpellier.

FAHRASMANEL, PARFAITA., JOURETC., GALZY P. – Production of higher alcohols and short chain fatty acids by different yeats used in rum fermentation. Accepte pour publication le 22 avril 1985 par Journal of Food Science.

OURA E. – 1974 – Some aspects of aeration intensity on the biochemical composition of baker’s yeast. 1. – Factors affecting the type of metabolism. Biotechnol-Bioeng. 16, 9, 1197.

PARFAITA., NAMORY M., DUBOIS P. — 1972 — Les esters éthyliques des acides gras supérieurs des rhums. Ann. Technol. Agric., 21, 2, 199-210.

PARFAITA., SABIN G. — 1975 – Les fermentations traditionnelles de mélasse et de jus de canne aux Antilles Françaises. Ind. Agric. Alim., 92, 1, 27-34.

PARFAITA, JOURET C. — 1979 – Rapport fin de Contrat DGRST. Décision d’aide n° 74 70906 et 74 7O 907.

PARFAITA., JOURETC. – 1980 – Le glycérol dans la fermentation alcoolique des mélasses et des jus de canne à sucre. Industries alimentaires et agricoles, 7-8, 721-724.

Répression des fraudes – 1973 – Méthodes officielles d’analyse des alcools et eauxde-vie. J.O. de la République Française du 2.10, no 73-231.