The stable carbon isotope compositions of methane and CO2 in the sampled gas phase were analysed as described previously62 . Methane concentrations were measured by GC (GC-4000, GL Science) using a Shincarbon ST 50/80 column (1.0 m × 3.0 mm inner diameter; Shinwa Chemical Industries) and a flame ionization detector with nitrogen as a carrier gas.
Amino acid concentrations in pure co-cultures of MK-D1 and Methanogenium were quantified through a previously described method63 (link),64 . In brief, we processed the acid hydrolysis with 6 M HCl (110 °C, 12 h) for the culture liquid samples after filtration using a 0.2-μm pore-size polytetrafluoroethylene filter unit (Millipore). The amino acid fraction was derivatized to N-pivaloyl iso-propyl esters before GC using a 6890N GC instrument connected to the nitrogen phosphorus and flame ionization detectors (Agilent Technologies). For cross-validation of qualitative identification of amino acids, GC–MS on the 7890 system (Agilent Technologies) was used61 (link).
Amino acid concentrations in pure co-cultures of MK-D1 and Methanogenium were quantified through a previously described method63 (link),64 . In brief, we processed the acid hydrolysis with 6 M HCl (110 °C, 12 h) for the culture liquid samples after filtration using a 0.2-μm pore-size polytetrafluoroethylene filter unit (Millipore). The amino acid fraction was derivatized to N-pivaloyl iso-propyl esters before GC using a 6890N GC instrument connected to the nitrogen phosphorus and flame ionization detectors (Agilent Technologies). For cross-validation of qualitative identification of amino acids, GC–MS on the 7890 system (Agilent Technologies) was used61 (link).
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