How specific is the inhibition by methyl fluoride of acetoclastic methanogenesis in anoxic rice field soil?


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Acetoclastic methanogens have been described to be inhibited at much lower concentrations of methyl fluoride, CH3F, than H2/CO2-utilizing methanogens. Therefore, we tested whether CH3F inhibition may be used to determine, in anoxic rice field soil, the contribution of H2/CO2-dependent methanogenesis to the total CH4 production by comparing this technique with the incorporation of 14CO2 into CH4. In general, addition of 0.01–1% CH3F to the gas phase resulted in an immediate partial inhibition of the total CH4 production which lasted for at least 200 h. Inhibition increased with the logarithm of the initial CH3F concentration up to about 0.2–0.6%. The initial CH3F concentration slowly decreased with time, probably due to decomposition. CH4 production sometimes completely recovered during the course of the experiment. The presence of CH3F resulted in the accumulation of acetate, the final concentration of which was usually stoichiometrically related to the deficit in CH4 production and increased with the initial CH3F concentration. In some experiments, acetate accumulation was larger than expected from the CH4 deficit and a substantial incorporation of 14CO2 into acetate was observed. Hydrogen, on the other hand, was only slightly elevated in the presence of CH3F. Addition of increasing CH3F resulted in an increase of the percentage of H2-dependent methanogenesis (measured by conversion of 14CO2 to 14CH4) demonstrating that acetoclastic methanogenesis was preferentially inhibited by CH3F. However, the conversion of 14CO2 to 14CH4 was also slightly inhibited by CH3F. Apparently, CH3F inhibited the H2-dependent methanogenesis to some extent, depending on the concentration of CH3F applied. Indeed, the ratio between the residual CH4 production rate and the fraction of CH4 produced from 14CO2 decreased with an increasing CH3F concentration. A ratio of unity was obtained at initial CH3F concentrations of 0.2–0.6% (58–174 μM). Both methods, i.e. inhibition using 0.5% CH3F and conversion of 14CO2 to 14CH4, were applied to determine the temporal change of the contribution of H2/CO2-dependent methanogenesis to the total CH4 production in two different batches of Italian rice field soil during a 120-days anoxic incubation period. The results of the two methods agreed well within the error of the methods and showed a relatively constant contribution of H2/CO2-dependent methanogenesis of about 25–30% as soon as CH4 was produced at a steady rate and H2 partial pressures had stabilized at about 1.5–2.5 Pa.