Thermo trace 1310 isq

Colonic feces were collected, frozen in liquid nitrogen, and thawed at room temperature before preparation. Phosphoric acid (50 μL, 15%), 100 mg glass beads, 100 μL 4-methylpentanoic acid (125 μg/mL), and 400 μL diethyl ether were added to each fecal sample, ground twice for 60 s at 60 Hz, and then centrifuged at 12000 rpm for 10 min. The supernatant was used for the analysis. After methodological investigation (Supplementary Figure S1 and Table S1), the SCFAs contents were detected using a Thermo TRACE 1310-ISQ gas chromatography-mass spectrometer (GC-MS) (Thermo Fisher Scientific, Waltham, MA, USA).

The 24 samples were hydrolyzed and extracted as previously described [54 (link)]. A total of 100 μL upper clear liquid was diluted by n-hexane to 1 mL and filtered by a 0.45 μ membrane. The data collection instrument system was Thermo Trace 1310 ISQ (Thermo Fisher Scientific, Waltham, MA, USA), with HP-88 Agilent 100 m × 0.25 mm × 0.20 μM as chromatographic column, injection port temperature at 290 °C, and He as carrier gas. The flow rate was set at 1.0 mL/min. The heating program was initially maintained at 100 °C for 13 min, and the temperature was raised at a rate of 10 °C/min to 180 °C for 6 min, raised at a rate of 1.5 °C/min to 192 °C for 6 min, and then raised at a rate of 3.5 °C/min to 240 °C for 4 min. The integral peak area ratio of all detected samples of fatty acids was taken into the standard curve linear equation for calculation and further incorporated into the calculation formula to obtain the absolute data on fatty acid content. Fatty acid concentrations were expressed on a % dry basis.