Quantification of Fecal Metabolites

Quantification of amino acids, derived biogenic amines, ammonium, and GABA was carried out by UHPLC following a derivatization with diethyl ethoxymethylenemalonate (DEEMM, Sigma-Aldrich, St. Louis, MO, USA) in 100 µL of fecal water, by the procedure described in Redruello et al. [33 (link)], adapted and optimized to human fecal cell-free supernatants by Salazar et al. [34 (link)]. Samples were filtered through 0.22 µm pore diameter PTFE membranes (VWR International, Radnor, PA, USA) prior to injection of 1 µL into the UHPLC chromatographic system. Concentrations of the analyzed compounds in fecal supernatants were calculated in millimolar (mM) and referred as concentration in fecal samples.