Michelle Durand

Biochemistry of Fermentation

The human large intestine is a complex anaerobic ecosystem, composed of numerous different species, which degrade and ferment substrates that have either escaped the digestion in the upper digestive tract or are produced by the host. It is recognised that a significant daily quantity of undigested dietary carbohydrate enters the colon (Edwards and Rowland, 1992; see Chapter 2). In contrast, the amount of carbohydrate fermented from endogenous sources like mucus remains undefined (Cummings and Macfarlane, 1991; Flourie et al.,1991). The microbial degradation of this organic matter in the colon constitutes a fundamental process which requires the contribution of different groups of microorganisms linked in a trophic chain (Wolin and Miller, 1983). These food-chain reactions break macromolecules such as complex polysaccharides down to short-chain fatty acids (mainly acetate, propionate and butyrate) and gases (H2, CO2 and in some case CH4). Polysaccharide degrading bacteria hydrolyse polymers into smaller fragments that can be used by saccharolytic bacteria. This cross-feeding allows maintainence of bacterial diversity in the ecosystem. The fermentation products of hydrolytic and saccharolytic bacteria include intermediates, such as lactate or succinate, that are metabolised by other species and do not accumulate to any significant extent in the colon. Hydrogen, which derives enterely from these fermentative processes, can be re-utilized in situ by hydrogenotrophic microorganisms.

Use of the rumen simulation technique (RUSITEC) to compare microbial digestion of various by-products

Abstract A semi-continuous culture system, adapted from the rumen simulation technique (RUSITEC) was used to compare the rumen micro-organism fermentation of different by-products. The following by-products were studied: (1) concentrates: citrus pulp; beet pulp; corn gluten feed; wheat bran; (2) roughages: untreated straw samples; straw samples treated with either ammonia or sodium hydroxide. The latter were also compared to alfalfa hay. The undigested feed residues from the bag were analysed for organic matter and cell-wall constituents (van Soest detergent procedure); liquid effluent was analysed for volatile fatty acids (VFA) and gas samples for CH 4 , CO 2 and H 2 . The substrates, representing a wide range of fermentability, fell into 4 groups with decreasing values according to OM digestibility and total daily VFA production: pulp by-products (80%, 93 mmoles), cereal by-products (68%, 69 mmoles), NaOH-treated straw and hay (55%, 60 mmoles) and untreated and NH 3 -treated straw (34%, 40 mmoles). However, cellulose digestibility showed intra-group variations; the largest difference observed was between corn gluten feed and wheat bran (40.1 and 15.6%, respectively). In comparison with the other groups, cereal by-products showed a particular VFA pattern with a high molar proportion of propionate (32 vs. 20–27%) and a low proportion of acetate (51 vs. 58–74%), probably because of their residual starch content (16.7% DM in bran). Methane production relative to hexose fermented (mol mol −1 ) was higher for roughage (0.45–0.58) than for concentrate by-products (0.31–0.35). It was concluded that the RUSITEC technique offers a convenient means of comparing the extent of microbial degradation and the fermentation pattern of various by-products but that, in the future, biomass synthesis should also be measured.

Short chain fatty acid and glucose metabolism in isolated pig colonocytes: modulation by NH4+

Short chain fatty acids (SCFA) from bacterial origin, as well as glucose from vascular origin, are among fuel substrates available to the colonic mucosa. The present work investigated the possible modulation by another bacterial metabolite, i.e. ammonia, of the capacities of colonic epithelial cells to metabolize these substrates. Viable colonocytes were isolated from the proximal colon of 40–50 kg pigs fed a standard diet and were incubated (30 min, 37°C) in the presence of a concentration range of 14C-labeled n-butyrate or acetate, or 14C-labeled glucose (5 mm), with or without NH4Cl (10 mM) addition. 14CO2 and metabolites generated were measured. Butyrate utilization resulted in a high generation of ketone bodies (acetoacetate and β-OH-butyrate), in addition to 14CO2 production. However, the net ketone body generation was significantly decreased for butyrate concentrations higher than 10 mM. In contrast to n-butyrate, acetate when given as the sole substrate got preferentially metabolized in the oxidation pathway. Acetate metabolism was not affected by NH4Cl, thus indicating that the tricarboxylic acid cycle was unchanged. Conversely, 4C02 and ketone body production from butyrate were decreased by 30% in the presence of NH4Cl, suggesting that butyrate activation or β-oxidation was diminished. Glucose utilization rate was increased by 20%, due to an increased glycolytic capacity in the presence of NH4Cl. A dose-dependent stimulation of phosphofructokinase activity by NH4+ could account for this effect. It is concluded that ammonia, whose physiological concentration is high in the colonic lumen, can modulate the metabolism of two major substrates, n-butyrate and glucose, in colonic epithelial cells.