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).

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