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Methane is a major greenhouse gas and a key component of global biogeochemical cycles. Microbial methane often deviates from isotope and isotopolog equilibrium in surface environments but approaches equilibrium in deep subsurface sediments. The origin of this near-equilibrium isotopic signature in methane, whether directly produced by methanogens or achieved through anaerobic oxidation of methane (AOM), remains uncertain. Here, we show that, in the absence of AOM, microbial methane produced from deep-sea sediments exhibits isotopolog compositions approaching thermodynamic equilibrium due to energy limitation. In contrast, microbial methane from salt marsh and thermokarst lakes exhibits significant hydrogen and clumped isotopic disequilibrium due to high free-energy availability. We propose that clumped isotopologs of methane provide a proxy for characterizing the bioenergetics of environments for methane production. Together, these observations demonstrate methane clumped isotopes as a powerful tool to better understand the relation between methane metabolisms and the energy landscape in natural environments. 

ABSTRACT The Belfast Ramped Pyroxidation/Combustion (RPO/RC) facility was established at the 14 CHRONO Centre (Queen’s University Belfast). The facility was created to provide targeted analysis of bulk material for refined chronological analysis and carbon source attribution for a range of sample types. Here we report initial RPO results, principally on background material, but also including secondary standards that are routinely analyzed at 14 CHRONO. A description of our setup, methodology, and background (blank) correction method for the system are provided. The backgrounds (anthracite, spar calcite, Pargas marble) reported by the system are in excess of 35,000 14 C years BP with a mean age of 39,345 14 C years BP (1σ = 36,497–43,800 years BP, N=44) with F 14 C = 0.0075 ± 0.0032. Initial results for standards are also in good agreement with consensus values: TIRI-B pine radiocarbon age = 4482 ± 47 years BP (N=13, consensus = 4508 years BP); IAEA-C6 ANU Sucrose F 14 C= 1.5036 ± 0.0034 (N=10, consensus F 14 C = 1.503). These initial tests have allowed problematic issues to be identified and improvements made for future analyses.