The light in the dark: in-situ biorefinement of crude oil to hydrogen using typical oil reservoir Thermotoga strains

H₂ is a CO₂ free energy carrier that can be produced biologically through dark fermentation using specific bacteria. In general, biological production of H₂ needs a carbon source and is more efficient at higher temperatures. Mature petroleum reservoirs have the required high temperatures for H₂ production, and they contain a significant amount of organic matter in form of residual hydrocarbons. In this work, we evaluated whether indigenous microorganisms isolated from hydrocarbon reservoirs are able to biorefine hydrocarbons to H₂. We observed that two Thermotoga strains, Pseudothermotoga hypogea DSM-11164 and Pseudothermotoga elfii DSM-9442, are able to convert hydrocarbons to H₂. DSM 9442 produced 0.47 and 1.02 mmol H₂ per liter of growth medium from 20 ml/L of n-hexadecane or a crude oil, respectively. DSM 11164 only produced H2 from n-hexadecane (0.94 mmol/L). Addition of 25 mg/L Tween 80, to reduce phase separation, together with 1 g/L glucose increased H₂ production from hydrocarbons up to 12-fold. Via an energy analysis we show that bioconversion of crude oil into H₂ can be more efficient than conversion of crude oil to gasoline. Therefore, we suggest dark fermentation as a promising alternative to biorefine crude oil and unlock the energy trapped in hydrocarbon reservoirs after abandonment.