In vitro sulphate reduction with methane

The simultaneous determination of methane utilisation and sulphide production to investigate the stoichiometry of the process can be carried out in an incubation experiment without gas phase. A relatively large gas phase of methane compared to the aqueous phase would not allow reliable measurement of methane consumption, particularly at the beginning of the experiment when the decrease in the total amount of methane is still relatively small. On the other hand, a small gas phase of methane would result in a slow supply of methane into the medium and sediment. Therefore, a special glass tube is used (modified from Alperin and Reeburgh, 1985) that allows head space-free incubation of sediment samples with dissolved methane concentrations above the saturation limit given at ambient pressure. At one end, the glass tube (length, 180 mm; inner diameter, 17 mm) is tapered towards a regular orifice (inner diameter, 9 mm) with a stopper (Viton rubber, fixed by screw cap with hole) that allows withdrawal of aliquots with a syringe. At the other end, the tube is tapered towards a smaller elongated glass tube (length, 65 mm; inner diameter, 9 mm) that harbours a gas-tight piston (silicon-lubricated Viton rubber); the piston is held in position by a screwing device. If a sample is withdrawn via the opposite stopper, the piston is simultaneously pushed into the tube so as to avoid any underpressure and formation of gas bubbles. High dissolved methane concentrations are achieved via an initial small (5 ml) head space in which 0.4 MPa (4 atm) of methane is kept for some hours under shaking. The gas phase is finally replaced by anoxic medium, and remaining bubbles are allowed to escape via an inserted hypodermic needle while the piston is slightly moved. This device, which is provided with 6 ml sediment slurry in a total volume of 40 ml, is incubated as the tubes described above.

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