Gray N D, Matthews J N S, Head I M
School of Civil Engineering and Geosciences and Centre for Molecular Ecology, University of Newcastle, Newcastle upon Tyne, NE1 7RU, UK.
J Microbiol Methods. 2006 Apr;65(1):180-6. doi: 10.1016/j.mimet.2005.07.006. Epub 2005 Aug 11.
A novel stable isotope titration approach was developed to determine the contributions to total methane production made by CO(2) reduction and the disproportionation of acetate in anoxic environments. (13)CH(4), (12)CH(4), (13)CO(2) and (12)CO(2) production rates were measured in the head space of replicate anaerobic microcosms titrated with increasing amounts of (13)C-labelled substrates. The contribution of CO(2) reduction was calculated from the linear relationship between ratios of labelled and total CH(4) production and ratios of labelled and total CO(2) after the addition of (13)C-bicarbonate. In the case of acetoclastic methanogenesis rates of (13)CH(4) and (12)CH(4) production were fitted to a model based on an assumption that the relationship between the concentration of (13)C-labelled acetate and the rates of labelled and unlabelled methane production followed Michaelis-Menten kinetics. A comparison of the raw data with the model supported the assumption and provided both an estimate of the contribution of acetate to methane production and an estimate of the size of the indigenous acetate pool without the need to measure acetate directly. The method was applied to a freshwater sediment in the English Lake District where it was found that 66.3% (se 4.9) of methane production was due to acetate disproportionation and 28.9% (se 1.9) of methane production resulted from CO(2) reduction. This is in agreement with theoretical predictions and other empirical measurements of methanogenesis.
开发了一种新型稳定同位素滴定方法，以确定在缺氧环境中，二氧化碳还原和乙酸盐歧化作用对总甲烷生成的贡献。在用越来越多的¹³C标记底物滴定的重复厌氧微观世界的顶空区域中，测量了¹³CH₄、¹²CH₄、¹³CO₂和¹²CO₂的生成速率。添加¹³C-碳酸氢盐后，根据标记和总CH₄生成比率与标记和总CO₂比率之间的线性关系，计算二氧化碳还原的贡献。对于乙酸裂解产甲烷作用，¹³CH₄和¹²CH₄生成速率被拟合到一个模型中，该模型基于这样一个假设，即¹³C标记的乙酸盐浓度与标记和未标记甲烷生成速率之间的关系遵循米氏动力学。将原始数据与模型进行比较，支持了该假设，并提供了乙酸盐对甲烷生成贡献的估计值以及本地乙酸盐库大小的估计值，而无需直接测量乙酸盐。该方法应用于英国湖区的淡水沉积物，发现66.3%（标准误4.9）的甲烷生成归因于乙酸盐歧化作用，28.9%（标准误1.9）的甲烷生成源于二氧化碳还原。这与产甲烷作用的理论预测和其他实证测量结果一致。
