SCFA and BCFA levels were determined in the fecal supernatants by means of gas chromatography, as described by Moris et al.77 (link) Briefly, 250 μl of cell free-supernatants were mixed with 100 μl methanol, 50 μl internal standard solution (2-ethylbutyric 1.05 mg/ml), and 50 μl of 20% v/v formic acid. The mix was then centrifuged, and the supernatant was injected into a system composed of a 6890NGC injection module (Agilent Technologies Inc., USA) connected to a flame injection detector (FID) and a mass spectrometry detector (MS, 5973 N) (Agilent) for quantification of both SCFA and BCFA.
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