Hydrogenotrophic activity: A tool to evaluate the kinetics of methanogens

Anaerobic-digestion-based technology is key to achieving sustainable water management and resource recovery. It is essential to understand the material flux and kinetics involved in methanogenesis to optimize the organic matter removal and methane production. In this sense, specific methanogenic activity is a cost-effective tool to characterize the biological activity of anaerobic biosludge, to monitor the performance of reactors, and study the kinetics of acetate and H₂ conversion to methane. Established protocols are applied for the acetoclastic activity test. However, hydrogenotrophic activity assay remains less widespread and is not standardized. In this work, the assay design for hydrogenotrophic activity is discussed and full calculation is presented, based on the kinetics for the H₂/CO₂ conversion to methane. An equation to calculate the inoculum size is proposed, suitable for a wide variety of types of biosludge: from a wastewater treatment plant to solid digesters, from a high-rate reactor to lagoons. The applied zero-order model fitted adequately to data for pilot-scale and full-scale anaerobic reactors: the p-values from the ANOVA F-test were below 1E-03; standard deviations for triplicate experiments were between 3 and 12%, coherent with the values found in the literature. Microbial growth during the test was negligible, below 1.2% of the biomass dosed in the vial. As a complement, acetoclastic activity was determined for each sample. The use of both acetoclastic and hydrogenotrophic activity is relevant for the study of the methanogenesis and gives a better characterization of the performance of the biosludge in anaerobic reactors rather than only using the specific acetoclastic methanogenic activity.