Simultaneous HS-SPME GC-MS determination of short chain fatty acids, trimethylamine and trimethylamine N-oxide for gut microbiota metabolic profile.

Trimethylamine (TMA), trimethylamine-N-oxide (TMAO) and short chain fatty acids (SCFAs), as acetic, propionic, butyric and valeric acids are among the most important products of the gut microbiota (GM) metabolism. The present study is aimed at the determination of TMA, TMAO and SCFAs by a double step headspace-solid phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC-MS) analysis, allowing the simultaneous quantitation of both the acidic and basic metabolites in faecal samples. TMAO amount was evaluated after its reduction to TMA by using Fe(II)-EDTA complex as a reagent. Under the fully validated experimental conditions, adequate sensitivity (LOQ 0.011-0.23 µmol g-1), good accuracy (79 - 110%) and precision (CV% < 11%) were achieved for all the target analytes. The presented method is successfully applied to the quantitation of the considered gut metabolites in faecal samples from Italian healthy volunteers.