Abstract. An anaerobic enrichment culture degraded 1 mol of acetone to 2 tool of methane and 1 tool of carbon dioxide. Two microorganisms were involved in this process, a filament-forming rod similar to Methanothrix sp. and an unknown rod with round to slightly pointed ends. Both organisms formed aggregates up to 300 gm in diameter. No fluorescing bacteria were observed indicating that hydrogen or formate-utilizing methanogens are not involved in this process. Acetate was utilized in this culture by the Methanothrix sp. Inhibition of methanogenesis by bromoethanesulfonic acid or acetylene decreased the acetone degradation rate drastically and led to the formation of 2 mol acetate per mol of acetone. Streptomycin completely inhibited acetone degradation, and neither acetate nor methane was formed. 1*CO2 was incorporated exclusively into the C-1 atom of acetate indicating that acetone is degraded via carboxylation to an acetoacetate residue. It is concluded that acetone is degraded by a coculture of an eubacterium and an acetateutilizing methanogen and that acetate is the only intermediate transferred between both. The energetical problems of the eubacterium converting acetone to acetate are discussed.

Abstract.From anaerobic digestor sludge of a waste water treatment plant, a gram-negative, strictly anaerobic sulfate-reducing bacterium was isolated with acetone as sole organic substrate. The bacterium was characterized as a new species, Desulfocoecus biacutus. The strain grew with acetone with doubling times of 72 h to 120 h; the growth yield was 12.0 (+_2.1) g . [ m o l acetone] -1. Acetone was oxidized completely, and no isopropanol was formed. In labelling studies with 14CO2, cell lipids (including approx. 50% PHB) of acetone-grown cells became labelled 7 times as high as those of 3-hydroxybutyrate-grown cells. Enzyme studies indicated that acetone was degraded via acetoacetyl-CoA, and that acetone was channeled into the intermediary metabolism after condensation with carbon dioxide to a C4-compound, possibly free acetoacetate. Acetoacetyl-CoA is cleaved by a thiolase reaction to acetyl-CoA which is completely oxidized through the carbon monoxide dehydrogenase pathway. Strain K M R A c t S was deposited with the Deutsche Sammlung yon Mikroorganismen, Braunschweig, under the number DSM 5651.

The pathway of anaerobic acetone degradation by the denitrifying bacterial strain BunN was studied by enzyme measurements in extracts of anaerobic acetone-grown cells. An ADP-and MgC12-dependent decarboxylation of acetoacetate was detected which could not be found in cell-free extracts of acetate-grown cells. It is concluded that free acetoacetate is formed by ATP-dependent carboxylation of acetone. Acetoacetate was converted into its coenzyme A ester by succinyl-CoA: acetoacetate CoA transferase, and cleaved by a thiolase into acetyl-CoA. The acetyl residue was completely oxidized in the citric acid cycle. The ADP-dependent decarboxylation of acetoacetate was inhibited by EDTA, but not by avidin. High myokinase activities led to equilibrium amounts of ATP, ADP, and AMP in the reaction mixtures, and prevented determination of the decarboxylase reaction stoichiometry, therefore.

A mixed culture, WoAct, growing on acetone, consisted of two dominant morphotypes: a rod-shaped acetone-fermenting bacterium producing acetate, and an acetate-utilizing Methanosaeta species. Dense cell suspensions, largely free of the aceticlastic methanogen and supplemented with bromoethanesulfonate, were able to degrade acetone and grow in small volumes in membrane-separated culture devices in which the acetate produced could diffuse into a large volume of medium. Acetone degradation and growth halted when the acetate concentration reached about 10 to 12 mM. Cell suspensions were able to degrade acetone in the absence of active methanogenesis, but the addition of 10 mM acetate inhibited acetone metabolism. Addition of an active culture of Methanosaeta sp. greatly stimulated the rate of acetone degradation. The results show that acetate removal in the mixed culture is not a prerequisite for growth and acetone degradation by the acetone-fermenting bacterium.

Five strains of Gram-negative denitrifying bacteria that used various ketones as sole carbon and energy sources were isolated from activated sludge from a municipal sewage plant. Three strains are related to the genus Pseudomonas; two non-motile species have not yet been affiliated. All strains grew well with ketones and fatty acids (C, to C7), but sugars were seldom utilized. The physiology of anaerobic acetone degradation was studied with strain BunN, which was originally enriched with butanone. Bicarbonate was essential for growth with acetone under anaerobic and aerobic conditions, but not if acetate or 3-hydroxybutyrate were used as substrates. An apparent K , value of 5.6 mM-bicarbonate was determined for growth with acetone in batch culture. The molar growth yield was 24.8-29.8 g dry cell matter (mol acetone consumed)-', with nitrate as the electron acceptor in batch culture; it varied slightly with the extent of poly-P-hydroxybutyric acid (PHB) formation. During growth with acetone, 4C02 was incorporated mainly into the C-1 atom of the monomers of the storage polymer PHB. With 3-hydroxybutyrate as substrate, 14C02 incorporation into PHB was negligible. The results provide evidence that acetone is channelled into the intermediary metabolism of this strain via carboxylation to acetoacetate.