The optimization and validation of a gas chromatography-mass spectrometry method to analyze the concentration of acetate, propionate and butyrate in human plasma or serum.

Fermentation-derived short-chain fatty acids (SCFA)4 are potential mediators of the health benefits associated with dietary fiber intake. SCFA affect physiological processes locally in the gut and on distant organs via the systemic circulation. Since SCFA are used as energy source for colonocytes and substrate for the liver metabolism, their concentrations in the systemic circulation are low. Therefore, quantification of systemic SCFA requires sensitive analytical techniques. This article covers the optimization and validation of a gas chromatography-mass spectrometry method to measure systemic SCFA concentrations following derivatization with 2,4-difluoroaniline (DFA)5 and extraction in ethyl acetate. Sample preparation was optimized by varying the amount of DFA, coupling agent 1,3-dicyclohexylcarbodiimide, ethyl acetate and sodium bicarbonate, which is used to quench derivatization. In addition, evaporation of the samples using a vacuum concentrator resulted in less contamination, notably of acetate, compared to drying with N2 gas. The method showed excellent linearity with coefficient of variation (R2) > 0.99 and a good precision (relative standard deviation < 20 %) and accuracy. Finally, systemic concentrations of SCFA in human plasma samples could successfully be determined.

Effect of Mutant and Engineered High-Acetate-Producing Saccharomyces cerevisiae var. boulardii Strains in Dextran Sodium Sulphate-Induced Colitis.

Acetate-producing Saccharomyces cerevisiae var. boulardii strains could exert improved effects on ulcerative colitis, which here, was preclinically evaluated in an acute dextran sodium sulphate induced model of colitis. Nine-week-old female mice were divided into 12 groups, receiving either drinking water or 2.75% dextran sodium sulphate for 7 days, combined with a daily gavage of various treatments with different levels of acetate accumulation: sham control (phosphate buffered saline, no acetate), non-probiotic control (Baker's yeast, no acetate), probiotic control (Enterol®, transient acetate), and additionally several Saccharomyces cerevisiae var. boulardii strains with respectively no, high, and extra-high acetate accumulation. Disease activity was monitored daily, and feces samples were collected at different timepoints. On day 14, the mice were sacrificed, upon which blood and colonic tissue were collected for analysis. Disease activity in inflamed mice was lower when treated with the high-acetate-producing strain compared to sham and non-probiotic controls. The non-acetate-producing strain showed higher disease activity compared to the acetate-producing strains. Accordingly, higher histologic inflammation was observed in non- or transient-acetate-producing strains compared to the sham control, whereas this increase was not observed for high- and extra-high-acetate-producing strains upon induction of inflammation. These anti-inflammatory findings were confirmed by transcriptomic analysis of differentially expressed genes. Moreover, only the strain with the highest acetate production was superior in maintaining a stable gut microbial alpha-diversity upon inflammation. These findings support new possibilities for acetate-mediated management of inflammation in inflammatory bowel disease by administrating high-acetate-producing Saccharomyces cerevisae var. boulardii strains.

The impact of bariatric surgery on macronutrient malabsorption depends on the type of procedure.

Introduction: Bariatric surgery, currently the most effective treatment for morbidly obese patients, may induce macronutrient malabsorption depending on the type of procedure. Macronutrient malabsorption affects the supply of substrates to the colon, subsequent microbial fermentation and possibly colonic health. Methods: Using isotope technology, we quantified the extent of macronutrient and bile acid malabsorption and its impact on colonic protein fermentation in patients after Roux-en-Y gastric bypass (RYGB) and sleeve gastrectomy (SG) and in controls. Participants consumed a single test meal (day 0) that contained intrinsically labeled (13C, 15N, and 2H) egg protein for quantification of protein digestion, malabsorption and fermentation, respectively, together with a transit marker and a marker for bile acid malabsorption. They collected breath samples up to 6 h and all urine and stool for 48 and 72 h, respectively. Food intake was registered from day -3 to day 2. Results: Malabsorption of fat, protein and carbohydrates differed between groups (p = 0.040; p = 0.046; and p = 0.003, respectively) and was slightly higher in RYGB but not in SG patients compared to controls. Protein fermentation was increased in both RYGB and SG patients compared to controls (p = 0.001) and was negatively correlated to 2H-recovery as a marker of transit (ρ = -0.47, p = 0.013). Conclusion: The limited macronutrient malabsorption likely does not affect the nutritional status of the patient. However, the higher protein fermentation may affect colonic health and warrants further investigation.

Wheat bran with reduced particle size increases serum SCFAs in obese subjects without improving health parameters compared with a maltodextrin placebo.

BACKGROUND: Wheat bran (WB) has been associated with improved gastrointestinal health and a reduced risk of metabolic disorders. Reducing the particle size of WB might increase its fermentability and facilitate cross-feeding between the gut bacteria and in this way produce health effects. OBJECTIVES: We investigated the impact of WB with reduced particle size (WB RPS) on colonic fermentation and host health in normal-weight (NW) and obese (OB) participants compared with placebo (PL). METHODS: During 1 mo, 36 NW and 14 OB participants daily consumed 20 g WB RPS or PL (maltodextrin). Before and after the intervention, fasting serum and fecal SCFAs, fecal metabolite profiles, and microbiota composition were measured as fermentation parameters. Fecal output, fecal dry weight (%), fat excretion, transit, stool consistency, intestinal permeability, and serum total cholesterol, triglyceride, and C-reactive protein concentrations were measured as health parameters. The impact of WB RPS on the fermentation of other carbohydrates was assessed by quantifying postprandial cumulative serum 13C-SCFA after a challenge with 13C-inulin. RESULTS: WB RPS increased fasting serum acetate (P < 0.05) and total SCFA (P < 0.05) concentrations in OB participants. Fasting serum propionate concentrations were lower in OB than in NW participants at baseline (NW: 1.57 ± 0.75 µmol/L; OB: 0.89 ± 0.52 µmol/L; P < 0.01), but not after WB RPS (NW: 1.75 ± 0.77 µmol/L; OB: 1.35 ± 0.63 µmol/L; P = not significant). WB RPS did not enhance colonic fermentation of 13C-inulin and did not affect microbiota composition. Health parameters were not affected by the WB RPS intervention, either in NW or in OB participants. CONCLUSIONS: WB RPS increased fasting serum SCFA concentrations in OB participants. These changes were not associated with beneficial effects on host health.

Wheat Bran Does Not Affect Postprandial Plasma Short-Chain Fatty Acids from 13C-inulin Fermentation in Healthy Subjects.

Wheat bran (WB) is a constituent of whole grain products with beneficial effects for human health. Within the human colon, such insoluble particles may be colonized by specific microbial teams which can stimulate cross-feeding, leading to a more efficient carbohydrate fermentation and an increased butyrate production. We investigated the extent to which WB fractions with different properties affect the fermentation of other carbohydrates in the colon. Ten healthy subjects performed four test days, during which they consumed a standard breakfast supplemented with 10 g 13C-inulin. A total of 20 g of a WB fraction (unmodified WB, wheat bran with a reduced particle size (WB RPS), or de-starched pericarp-enriched wheat bran (PE WB)) was also added to the breakfast, except for one test day, which served as a control. Blood samples were collected at regular time points for 14 h, in order to measure 13C-labeled short-chain fatty acid (SCFA; acetate, propionate and butyrate) concentrations. Fermentation of 13C-inulin resulted in increased plasma SCFA for about 8 h, suggesting that a sustained increase in plasma SCFA can be achieved by administering a moderate dose of carbohydrates, three times per day. However, the addition of a single dose of a WB fraction did not further increase the 13C-SCFA concentrations in plasma, nor did it stimulate cross-feeding (Wilcoxon signed ranks test).