Investigating the Human Gut Microbial Activity on Exopolysaccharides

Begin with the supernatant produced by human gut microbiota during in vitro fermentation of exopolysaccharides. The supernatant contains deprotonated short-chain fatty acids (SCFAs) and other metabolites.

Add metaphosphoric acid to lower the pH of the supernatant and vortex to mix thoroughly. This converts SCFAs into their protonated forms, suitable for gas chromatography analysis.

Centrifuge to separate debris and collect the supernatant containing SCFAs and other metabolites.

Filter the supernatant to eliminate residual debris and avoid interference with chromatography.

In a high-performance gas chromatography system, inject the supernatant into the heated inlet to vaporize it. Then, mix the vapor with an inert gas and pass it through a polar stationary-phase column for separation.

SCFAs interact with the column’s stationary phase and separate based on their volatility, with shorter-chain acids eluting earlier than longer ones.

Identify the concentrations of different SCFAs by comparing the results with internal standards. This reflects microbial degradation of exopolysaccharides under gut-like conditions

To study the effects of EPS on SCFA production, add 1 milliliter of the fermented