Rapid determination of short-chain fatty acids in colonic contents and faeces of humans and rats by acidified water-extraction and direct-injection gas chromatography

Abstract

Short-chain fatty acids (SCFAs) have attracted much attention recently because of their positive physiological effects. In this work, a rapid and reliable gas chromatographic method for determination of eight SCFAs, in colonic and faecal samples from rats and humans has been developed and validated. The methodology involves extraction of the SCFAs in water before a direct injection procedure on a FFAP capillary column. A stock standard solution containing acetic acid, propionic acid, n-butyric acid, i-butyric acid, n-valeric acid, i-valeric acid, n-caproic acid and n-heptanoic acid was prepared and used. A high linearity (r2 > 0.9990), low quantification limit (2.38-30.14 microm) and high recovery for most acids were obtained. Acidification of faecal samples was found to be crucial for quantitative determination of the SCFAs, and adjustment of pH to 2-3 was regarded as necessary. Glass wool inserted in the glass liner of the injection port proved effective in preventing the contamination of the column by non-volatiles, and 12% formic acid reduced the ghost peak that appeared gradually after several injections. After validation, the methodology was applied on two faecal samples from rats fed diets containing different amount of dietary fibre and one faecal sample from human fed a normal diet to test the accuracy of the developed method.

Substituting whole grains for refined grains in a 6-wk randomized trial has a modest effect on gut microbiota and immune and inflammatory markers of healthy adults

“…Undiluted stool samples were removed from storage at 2808C and freeze dried for 5-7 d. Stool samples were ground to a fine powder with the use of a mortar and pestle. Because the extraction yield of SCFAs can be increased by decreasing the stool homogenate pH (48), each stool sample (0.1 g) was suspended in 1% phosphoric acid and homogenized by shaking for 30 min, which was followed by centrifugation for 15 min at 7000 3 g at 48C. The liquid fraction was decanted and filtered through a 0.2-mm polysulfone membrane (Sigma-Aldrich) and stored at 2808C for later extraction.…”

A human volunteer study to assess the impact of confectionery sweeteners on the gut microbiota composition

“…Dose period 3 (days [30][31][32][33][34][35][36][37][38][39][40][41][42][43][44]. Volunteers consumed 100 g of their treatment chocolate, and kept daily diaries during the period.…”

Determination of Short-chain Fatty Acids in Rat and Human Feces by High-Performance Liquid Chromatography with Electrochemical Detection

“…The determination of SCFAs has been carried out by gas chromatography (GC) [13][14][15][16][17][18] and high-performance liquid chromatography with ultraviolet detection (HPLC-UV), fluorescent detection (HPLC-FL), or mass spectrometric detection (LC-MS). [19][20][21][22][23][24] The fecal analysis by GC required the complicate cleanup of the sample, and it was hard to detect NVFAs such as lactic acid.…”

“…In particular, the composition of lactic acid, called non-volatile fatty acid (NVFA), increased in comparison with that of acetic, propionic, and butyric acids, called volatile fatty acid (VFA), in feces of ulcerative colitis patients. [7][8][9][10][11][12] Thus, the determination of fecal SCFAs is significant to examine homeostasis, characteristics, and metabolism of intestinal bacterial flora, and it would be useful for monitoring of and diagnostic strategy against diseases in the colon.The determination of SCFAs has been carried out by gas chromatography (GC) [13][14][15][16][17][18] and high-performance liquid chromatography with ultraviolet detection (HPLC-UV), fluorescent detection (HPLC-FL), or mass spectrometric detection (LC-MS). [19][20][21][22][23][24] The fecal analysis by GC required the complicate cleanup of the sample, and it was hard to detect NVFAs such as lactic acid.…”