Contribution of methanol to the production of methane and its 13C-isotopic signature in anoxic rice field soil

Conversion of methanol to CH4 has a large isotope effect so that a small contribution of methanol-dependent CH4 production may decrease the delta(CH4)-C-13 of total CH4 production. Therefore, we investigated the role of methanol for CH4 production. Methanol was not detectable above 10 mu M in anoxic methanogenic rice field soil. Nevertheless, addition of C-13-labeled methanol (99% enriched) resulted in immediate accumulation of (CH4)-C-13. Addition of 0.1 mu M C-13-methanol resulted in increase of the delta(CH4)-C-13 from -47 to -6 parts per thousand within 2 h, followed by a slow decrease. Addition of 1 mu M C-13-methanol increased delta(CH4)-C-13 to +500 parts per thousand within 4 h, whereas 10 mu M increased delta(CH4)-C-13 to +2500 parts per thousand and continued to increase. These results indicate that the methanol concentrations in situ, which diluted the C-13-methanol added, were <= 0.1 mu M and that the turnover of methanol contributed only about 2% to total CH4 production at 0.1 mu M. However, contribution increased up to 5 and 17% when 1 and 10 mu M methanol were added, respectively. Anoxic rice soil that was incubated at different temperatures between 10 and 37 degrees C exhibited maximally 2-6% methanol-dependent methanogenesis about 1-2 h after addition of 1 mu M C-13-Methanol. Only at 50 degrees C, contribution of methanol to CH4 production reached a maximum of 10%. After longer (7-10 h) incubation, however, contribution generally was only 2-4%. Methanol accumulated in the soil when CH4 production was inhibited by chloroform. However, the accumulated methanol accounted for only up to 0.7 and 1.2% of total CH4 production at 37 and 50 degrees C, respectively. Collectively, our results show that methanol-dependent methanogenesis was operating in anoxic rice field soil but contributed only marginally to total CH4 production and the isotope effect observed at both low and high temperature.