Hydrogen and formate are important electron carriers in methanogenic degradation in anoxic environments such as sediments, sewage sludge digestors and biogas reactors. Especially in the terminal steps of methanogenesis, they determine the energy budgets of secondary (syntrophically) fermenting bacteria and their methanogenic partners. The literature provides considerable data on hydrogen pool sizes in such habitats, but little data exist for formate concentrations due to technical difficulties in formate determination at low concentration. Recent evidence from biochemical and molecular biological studies indicates that several secondary fermenters can use both hydrogen and formate for electron release, and may do so even simultaneously. Numerous strictly anaerobic bacteria contain enzymes which equilibrate hydrogen and formate pools to energetically equal values, and recent measurements in sewage digestors and biogas reactors indicate that - beyond occasional fluctuations - the pool sizes of hydrogen and formate are indeed energetically nearly equivalent. Nonetheless, a thermophilic archaeon from a submarine hydrothermal vent, Thermococcus onnurineus, can obtain ATP from the conversion of formate to hydrogen plus bicarbonate at 80°C, indicating that at least in this extreme environment the pools of formate and hydrogen are likely to be sufficiently different to support such an unusual type of energy conservation.

Pool sizes of short-chain fatty acids (formate, acetate, propionate, and butyrate), hydrogen, and carbon monoxide were assayed in digesting sludge from four different methanogenic reactors degrading either sewage sludge or agricultural products and wastes at pH 8.0 and 40 or 47°C. Free reaction energies were calculated for the respective degradation reactions involved, indicating that acetate, propionate, and butyrate degradation all supplied sufficient energy (−10 to −30 kJ per mol reaction) to sustain the microbial communities involved in the respective processes. Pools of formate and hydrogen were energetically equivalent as electron carriers. In the sewage sludge reactor, homoacetogenic acetate formation from H 2 and CO 2 was energetically feasible whereas syntrophic acetate oxidation appeared to be possible in two biogas reactors, one operating at enhanced ammonia content (4.5 g NH 4 + -N per l) and the other one at enhanced temperature (47°C). Maximum capacities for production of methanogenic substrates did not exceed the consumption capacities by hydrogenotrophic and aceticlastic methanogens. Nonetheless, the capacity for acetate degradation appeared to be a limiting factor especially in the reactor operating at enhanced ammonia concentration.

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The energetic situation of terminal fermentations in methanogenesis was analyzed by pool size determinations in sediment cores taken in the oligotrophic Lake Constance, Germany. Distribution profiles of fermentation intermediates and products were measured at three different water depths (2, 10, and 80 m). Methane concentrations were constant below 10 cm of sediment depth. Within the methanogenic zone, concentrations of formate, acetate, propionate, and butyrate varied between 1 and 40 μM, and hydrogen was between 0.5 and 5 Pa. From the distribution profiles of the fermentation intermediates, Gibbs free energy changes for their interconversion were calculated. Pool sizes of formate and hydrogen were energetically nearly equivalent, with -5 ± 5 kJ per mol difference of free energy change (ΔG) for a hypothetical conversion of formate to hydrogen plus CO The ΔG values for conversion of fatty acids to methanogenic substrates and their further conversion to methane and CO were calculated with hydrogen and with formate as intermediates. Syntrophic propionate oxidation reached energetic equilibrium with formate as the sole electron carrier but was sufficiently exergonic if at least some of the electrons were transferred via hydrogen. The energetic consequences of formate versus hydrogen transfer in secondary and methanogenic fermentations indicate that both carrier systems are probably used simultaneously to optimize the energy yields for the partners involved. In the terminal steps of methane formation in freshwater lake sediments, fermenting bacteria cooperate syntrophically with methanogens and homoacetogens at minimum energy increments via interspecies electron transfer. The energy yields of the partner organisms in these cooperations have so far been calculated based mainly on hydrogen partial pressures. In the present study, we also analyzed pools of formate as an alternative electron carrier in sediment cores of an oligotrophic lake. The formate and hydrogen pools appeared to be energetically nearly equivalent and are likely to be used simultaneously for interspecies electron transfer. Calculations of reaction energies of the partners involved suggest that propionate degradation may also proceed through the pathway, which converts propionate via butyrate and acetate to three acetate residues, thus circumventing one energetically difficult fatty acid oxidation step.