2.2. Preparation of Methane-Rich Saline (MRS) and Methane-Rich Medium (MRM)

Methane was dissolved in 0.9% saline and DMEM (with 10% FBS) for 4 h at a 0.4 MPa pressure to produce supersaturated MRS and MRM. The prepared MRS and MRM were stored and sterilized as previously described [19]. MRS and MRM were freshly prepared 1 day before the experiments. The methane levels in the saline and medium were estimated by gas chromatography (Gas Chromatography-9860, Qiyang, Shanghai, China). The concentrations of the MRS and MRM were approximately 1.2–1.5 mmol/l and 1.4–1.8 mmol/l, respectively.