The pathway of methanol conversion by a thermophilic anaerobic consortium was elucidated by recording the fate of carbon in the presence and absence of bicarbonate and specific inhibitors. Results indicated that about 50% of methanol was directly converted to methane by the methylotrophic methanogens and 50% via the intermediates H-2/CO2 and acetate. The deprivation of inorganic carbon species [Sigma(HCO3-+CO2)] in a phosphate-buffered system reduced the rate of methanol conversion. This suggests that bicarbonate is required as an electron (H-2) sink and as a co-substrate for the efficient and complete removal of the chemical oxygen demand. Nuclear magnetic resonance spectroscopy was used to investigate the route of methanol conversion to acetate in bicarbonate-sufficient and bicarbonate-depleted environments. The proportions of [1,2-C-13]acetate, [1-C-13]acetate and [2-C-13]acetate were determined. Methanol was preferentially incorporated into the methyl group of acetate, whereas HCO3- was the preferred source of the carboxyl group. A small amount of the added (HCO3-)-C-13 was reduced to form the methyl group of acetate and a small amount of the added (CH3OH)-C-13 was oxidised and found in the carboxyl group of acetate when (CH3OH)-C-13 was converted. The recovery of [C-13]carboxyl groups in acetate from (CH3OH)-C-13 was enhanced in bicarbonate-deprived medium. The small amount of label incorporated in the carboxyl group of acetate when (CH3OH)-C-13 was converted in the presence of bromoethanesulfonic acid indicates that methanol can be oxidised to CO2 prior to acetate formation. These results indicate that methanol is converted through a common pathway (acetyl-CoA), being on the one hand reduced to the methyl group of acetate and on the other hand oxidised to CO2, with CO2 being incorporated into the carboxyl group of acetate.