Quantification of Colonic Short-Chain Fatty Acids

SCFAs (i.e., lactate, acetate, propionate, butyrate) in colonic contents were quantified the procedures of Wang et al. [29 (link)]. Briefly, pig samples were thawed on ice and approximately 0.5-g samples were added to 8 mL of deionized water (For mouse samples: 50 mg of samples was added to 1 mL of deionized water). The mixture was thoroughly homogenized by vortexing for 1 min and centrifuged at 13,000 × g for 5 min. The supernatant was diluted 50-fold for pig samples or 25-fold for mouse samples and filtered through a 0.22-μm filter (Millipore, USA). The solution was transferred into the vial for quantitative determination using ion chromatography (ICS-3000, Dionex, USA) with a Dionex Ionpac™ AS11 analytical column (4 × 250 mm). To determine SCFA concentrations of the bacterial media, 1 mL media was collected at each time point and centrifuged at 13,000 × g for 5 min. The supernatant was diluted 100-fold and filtered through a 0.22-μm filter for ion chromatography analysis.