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The mechanism of ethanol production in bacterial fermentations has not been elucidated. Among the alcohol-producing bacterial systems available for study, the mixed ethanol-lactate fermentation characteristic of the heterofermentative lactic acid bacteria presents an interesting and apparently relatively simple case for experimentation. The major products of glucose fermentation by these organisms are lactic acid, ethanol, and carbon dioxide. Gayon and Dubourg (1901), Peterson and Fred (1920), and Nelson and Werkman (1935) have shown the occurrence of two fermentation patterns among the heterofermentative lactobacilli. With glucose as substrate, these are: (1) the production of lactate, ethanol, and C02, with occasional traces of acetate and (2) the formation of glycerol along with lactate, acetate, and C02. Among the heterofermentative cocci, genus Leuconostoc, Pederson (1929) found only the first type-glucose fermentation yielded equimolar quantities of lactate, ethanol, and C02. Friedemann (1939) confirmed this fermentation balance using Leuconostoc dextranicum as did Bang (1945) for Leuconostoc citrovorus. With one nucleotide-linked dehydrogenases for ethanol, lactate, triosephosphate, and 2,3butylene glycol. The lactic dehydrogenase is specific for D(-)lactic acid, the

The strictly aerobic nature of the Marburg strain of Bacillus subtilis has been amply demonstrated (Knaysi and Gunsalus, 1944; Blackwood et al., 1947; Hyndman, 1948; Knight and Proom, 1950), the organism being readily cultivated aerobically in an inorganic nitrogen-carbohydrate medium and apparently deriving energy wholly from the operation of an oxidative mechanism. The organism does not grow appreciably anaerobically in any medium thus far tested if an external hydrogen acceptor, such as nitrate, is lacking. It, therefore, was of interest to find that growth of this organism in complex medium containing tryptone, yeast extract, and glucose yields cells which dissimilate glucose anaerobically although anaerobic growth does not occur. Moreover, cultivation in simple inorganic nitrogen medium containing glucose yields cells practically incapable of fermentation but which dissimilate glucose aerobically according to a mechanism different from cells grown in complex medium. A comparison of these two metabolic types, a study of the dissimilatory mechanisms involved, and an examination of the nutritional basis for these differences were initiated; the results of some of these investigations are presented herein.

Glucose fermentation by Leuconostoc me8enteroides has been shown to proceed via a mechanism which differs from the classical Embden-Meyerhof glycolytic scheme. DeMoss, Bard, and Gunsalus (1951) found no evidence in cell-free extracts of this organism for the key enzyme aldolae but showed that cellular suspensions ferment glucose to equimolar quantities of ethanol, C02, and lactate as obligatory products. The diphosphopyridine nucleotide linked dehydrogenases for D-3-phosphoglyceraldehyde, D(-) lactic acid, and ethanol were demonstrated, indicating some resemblance of the Leuconostoc mechanism to the classical glycolytic scheme. Fermentation of glucose-1-C'4 by cellular suspensions was reported by Gibbs and DeMoss (1951) and Gunsalus and Gibbs (1952) to yield C14 exclusively in C02 at six times the specific activity of the glucose fermented, whereas glucose-3,4-C04 yielded carboxyl labeled lactate, carbinol labeled ethanol, and unlabeled C02. These results support the data obtained with enzyme extracts and, in addition, demonstrate that both C02 and ethanol must arise by a previously undescribed fermentative pathway(s). The observation of a glucose-6-phosphate dehydrogenase in extracts of L. menteroide8, reported in preliminary form by DeMoss, Gunsalus, and Bard (1951), suggested the operation of an anaerobic counterpart of the oxidative hexosemonophosphate pathway via phosphogluconic acid as the fermentative mechanism involved. Such an oxidative system, previously recognized only in aerobic organisms and studied by Warburg and Christian (1932), Lipmann (1936), and Dickens (1938), has been described recently in detail by Horecker and co-workers