Fecal SCFAs
Flash frozen stool will be transferred into plastic tubes; 2,2-dimethylbutyric acid will be added at 1 mmol/L as internal standard, vortexed and centrifuged, and supernatant filtered through a Millex-GS filter (Millipore, Burlington, MA, USA) and refiltered with a microconcentrator (Millipore, Burlington, MA, USA). The final filtrate will be analyzed using an Agilent Technologies 6890N Network GC System (Santa Clara, CA, USA) with a flame ionization detector for SCFAs. A mixed-SCFA standard solution will be prepared using high purity (0.99%) reagents (Sigma, St. Louis, MO, USA). Fecal SCFA concentrations will be computed by using a peak area ratio of the sample profile relative to the internal standard. Analysis will be conducted by the UIC Mass Spectrometry, Metabolomics and Proteomics Facility.