Db ffap chromatographic capillary column

Fecal SCFAs were analyzed by using gas chromatography (Shimadzu G2010Plus, Kyoto, Japan) equipped with a DB-FFAP chromatographic capillary column (30 m × 0.530 mm × 1.00 μm; Agilent, Santa Clara, CA, USA). The pre-treatment of samples was based on a method described before with some modifications [64 (link)]. In brief, fresh feces (100–150 mg) were dissolved in 0.8 mL 0.2 M HCl (0.02 mg/mL 2-ethylbutyric acid as internal standard) and 0.2 mL 0.15 M oxalate. The mixture was vortexed for 1 min and centrifuged at 12,000× g and 4 °C for 15 min. The supernatant was filtrated using a 0.22 μm filter for further analysis. N2 was supplied as carrier gas at a flow rate of 10 mL/min. The flow rates of air, H2, and N2 as make up gas were 260, 30, and 30 mL/min, respectively. The oven temperature was increased from 100 °C to 160 °C at 5 °C/min and then held at this temperature for 4 min. The SCFAs contents, including acetate, propionate, isobutyrate, butyrate, valerate, and caproate were quantified using standard curves.

Colonic contents were diluted, mixed, and centrifuged, and then, 25% (w/v) metaphosphoric acid solution was added to the supernatant to extract SCFA. SCFA analysis was performed according to the method previously described using gas chromatography (Agilent 6890, CA, USA) with a DB-FFAP chromatographic capillary column (30 m × 0.25 mm × 0.5 μm; Agilent) [20 (link)].