SUMMARYFermentation balances determined for different substrates in batch and continuous cultures of Lactobacillus casei revealed two pathways of pyruvate conversion by this organism, a reduction to lactate and the phosphoroclastic cleavage. Pyruvate formed anaerobically from mannitol and citrate was split by the phosphoroclastic enzyme. Lactate was the main fermentation product formed during aerobic growth on mannitol and anaerobic and aerobic growth on glucose. In glucose-limited continuous cultures pyruvate conversion was dependent on the dilution rate. At low dilution rates glucose was fermented exclusively to acetate, ethanol and formate. At high rates only small amounts of acetate, ethanol and formate were formed and lactate production was maximal. Lactate dehydrogenase of L. casei had an absolute requirement for fructose-I,6-diphosphate and manganous ions. The specific activity of lactate dehydrogenase did not differ significantly at different dilution rates. It was concluded that the intracellular level of fructose-I ,6-diphosphate controlled the pathway of pyruvate conversion. In batch cultures YATP values were between 18-2 and 20.9. No evidence for oxidative phosphorylation was found. In continuous cultures YdTP values varied from 18.7 at low dilution rates to 23'5 at high dilution rates. From the dependence of YATP on the dilution rate, a maintenance coefficient of 1-52 x I O -~ was calculated. The YATP value corrected for energy of maintenance was 24.3. The possibility that the molar growth yields were erroneously high because of assimilation of growth substrate into intracellular polysaccharides, or because of energy yield from components of the medium other than the added energy source, was excluded.

Cell-free extracts of 17 strains of Bifidobacterium bifidum (Lactobacillus bifidus) were examined for the presence of aldolase, glucose-6-phosphate dehydrogenase, and fructose-6-phosphate phosphoketolase. All strains turned out to lack aldolase, an enzyme unique to glycolysis, and glucose-6-phosphate dehydrogenase, characteristic of the hexosemonophosphate pathway. In all strains, fructose-6-phosphate phosphoketolase could be demonstrated. It can be concluded that bifidobacteria ferment glucose via a pathway which is different from those found in members of the genus Lactobacillus. The results strengthen the previous suggestions that classification of the bifidobacteria in the genus Lactobacillus is not justified.

For six strains of Bifidobacterium bifidum (Lactobacillus bifidus), fermentation balances of glucose, lactose, galactose, mannitol, and xylose were determined. Products formed were acetate, L(+)-lactate, ethyl alcohol, and formate. L(+)-Lactate dehydrogenase of all strains studied was found to have an absolute requirement for fructose-1 , 6-diphosphate. The phosphoroclastic enzyme could not be demonstrated in cell-free extracts. Cell suspensions fermented pyruvate to equimolar amounts of acetate and formate. Alcohol dehydrogenase was shown in cell-free extracts. Possible explanations have been suggested for the differences in fermentation balances found for different strains and carbon sources. By enzyme determinations, it was shown that bifidobacteria convert mannitol to fructose-6-phosphate by an inducible polyol dehydrogenase and fructokinase. For one strain of B. bifidum, molar growth yields of glucose, lactose, galactose, and mannitol were determined. The mean value of Y (ATP), calculated from molar growth yields and fermentation balances, was 11.3.

S U M M A R YEnzyme determinations in bacteria-free extracts and dual-wavelength experiments with membrane suspensions established that Propionibacteriunz freudenreichii converted glycerol into triose phosphate via glycerol kinase and NAD-independent glycerol I -phosphate dehydrogenase which is closely linked to cytochrome b. Glycerol I-phosphate dehydrogenase uses fumarate as a final hydrogen acceptor. The enzyme system catalysing fumarate reduction with glycerol I-phosphate as a hydrogen donor, is membrane bound and is strongly inhibited by 2-n-heptyl-4-hydroxyquinoline-N-oxide (HOQNO). Fumarate reduction with reduced benzyl-viologen is not inhibited by HOQNO. Cytochrome b is therefore probably involved in the anaerobic electron transport from glycerol I -phosphate to fumarate. Molar growth yields and fermentation balances were determined for P. freudenreichii and P. pentosaceuin growing on glucose, fructose, glycerol and lactate and ATP yields (mol of ATP formed/mol of substrate fermented) were calculated assuming that I mol ATP is formed in the electron transport from glycerol I-phosphate and lactate to fumarate, and that 2 rnol ATP are formed in the electron transport from NADH to fumarate. Mean YaTp values (g dry wt bacteria/mol ATP) were 15-2 and 12-9 for P. freudenreichii, and 16.4 and I 1.8 for P. pentosaceurn each growing on complex or synthetic medium respectively. The observation that for each strain YATP values were constant for the same medium, supported our assumptions on energy generation in propionic acid bacteria.

No abstract