Alternative method for gas chromatography-mass spectrometry analysis of short-chain fatty acids in faecal samples

Short-chain fatty acids are the major end products of bacterial metabolism in the large bowel. They derive mostly from the bacterial breakdown of carbohydrates and are known to have positive health benefits. Due to the biological relevance of these compounds it is important to develop efficient, cheap, fast, and sensitive analytical methods that enable the identification and quantification of the short-chain fatty acids in a large number of biological samples. In this study, a gas chromatography-mass spectrometry method was developed and validated for the analysis of short-chain fatty acids in faecal samples. These volatile compounds were extracted with ethyl acetate and 4-methyl valeric acid was used as an internal standard. No further cleanup, concentration, and derivatization steps were needed and the extract was directly injected onto the column. Recoveries ranged between 65 and 105%, and no matrix effects were observed. The proposed method has wide linear ranges, good inter- and intraday variability values (below 2.6 and 5.6%, respectively) and limits of detection between 0.49 μM (0.29 μg/g) and 4.31 μM (3.8 μg/g). The applicability of this analytical method was successfully tested in faecal samples from rats and humans.