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The human large intestine contains a microbiota, the components of which are generically complex and metabolically diverse. Its primary function is to salvage energy from carbohydrate not digested in the upper gut. This is achieved through fermentation and absorption of the major products, short chain fatty acids (SCFA), which represent 40-50% of the available energy of the carbohydrate. The principal SCFA, acetate, propionate and butyrate, are metabolized by the colonic epithelium (butyrate), liver (propionate) and muscle (acetate). Intestinal bacteria also have a role in the synthesis of vitamins B and K and the metabolism of bile acids, other sterols and xenobiotics.The colonic microflora are also responsive to diet. In the presence of fermentable carbohydrate substrates such as non-starch polysaccharides, resistant starch and oligosaccharides, bacteria grow and actively synthesize protein. The amount of protein synthesis and turnover within the large intestine is difficult to determine, but around 15 ...

Supplementation of the human diet with prebiotic substances such as inulin and non-digestible oligosaccharides (NDO), e.g., galacto-oligosaccharides (GOS), has been associated with various health benefits. However, little information is available regarding the spatial location of their metabolism in human gut bacterial ecosystems. Therefore, the present study investigated the metabolism of inulin and GOS with respect to bacterial growth, bifidobacterial stimulatory properties and anti-mutagenicity potential, in a three-stage continuous culture model of the colon which reproduces the physicochemical characteristics of the proximal (V1) and distal (V2, V3) colons. Fermentation of both carbohydrates was rapid, and occurred primarily in V1, as evidenced by acid formation. Inulin metabolism was associated with 10-fold stimulation of lactobacillus populations, together with smaller increases in bifidobacterial cell counts in V1. However, peptostreptococci, enterococci and Clostridium perfringens also increased in this fermentation vessel. In contrast, GOS was only weakly bifidogenic in V1, although these bacteria did proliferate in V2. GOS also increased lactobacilli by an order of magnitude in V1. However, overall changes in microbial populations resulting from inulin or GOS addition were minimal in V2 and V3. Potential beneficial effects of inulin metabolism included minor reductions in beta-glucosidase and beta-glucuronidase, whereas GOS strongly suppressed these enzymes, together with arylsulphatase (AS). Growth of putatively health promoting micro-organisms was not only associated with reductions in enzymes linked to genotoxicity. For example, both carbohydrates stimulated synthesis of nitroreductase and azoreductase, throughout the fermentation system, while inulin increased AS. Colonic transit time is an important factor in bacterial metabolism in the large bowel, and these data suggest that, in some circumstances, NDO fermentation will occurprincipally in the proximal colon.

In both health and disease, the colonic microbiota plays an important role in several areas of human physiology. This complex assemblage of microorganisms endows great metabolic potential on the large intestine, primarily through its degradative abilities. Many hundreds of different types of bacteria, varying widely in physiology and biochemistry, exist in a multitude of different microhabitats in the lumen of the large gut, the mucin layer and on mucosal surfaces. Both microbiota and host obtain clear benefits from association. For example, growth substrates from diet and body tissues, together with a relatively stable environment for bacteria to proliferate are provided by the host, which in turn has evolved to use butyrate, a bacterial fermentation product, as its principal source of energy for epithelial cells in the distal bowel. The main sources of carbon and energy for intestinal bacteria are complex carbohydrates (starches, non-starch polysaccharides). Carbohydrate metabolism is of great importance in the large intestine, since generically, and in terms of absolute numbers, the vast majority of culturable microorganisms are saccharolytic. The amounts and types of fermentation products formed by colonic bacteria depend on the relative amounts of each substrate available, their chemical structures and compositions, as well as the fermentation strategies (biochemical characteristics and catabolite regulatory mechanisms) of bacteria participating in depolymerization and fermentation of the substrates. Protein breakdown and dissimilatory amino acid metabolism result in the formation of a number of putatively toxic metabolites, including phenols, indoles and amines. Production of these substances is inhibited or repressed in many intestinal microorganisms by a fermentable source of carbohydrate. Owing to the anatomy and physiology of the colon, putrefactive processes become quantitatively more important in the distal bowel, where carbohydrate is more limiting.

Measurements of polysaccharide-degrading activity in different fractions of human faeces showed that bacterial polysaccharidases and glycosidases were primarily associated with the washed bacterial fractions. Amylase, pectinase and xylanase were the major polysaccharide-hydrolysing enzymes detected, whilst α-L-arabinofuranosidase, β-D-xylosidase, β-D-galactosidase and β-D-glucosidase were the most active glycosidases. Starch and 3 non-starch polysaccharides (NSP; pectin, xylan and arabinogalactan) were fermented by mixed populations of human faecal bacteria in batch culture. Detailed carbohydrate analysis demonstrated that starch and pectin were the most rapidly degraded substrates and that arabinogalactan and the relatively insoluble polysaccharide xylan were broken down more slowly. Free sugars and oligosaccharides did not accumulate in culture media with any polysaccharide tested. Time-course measurements of polysaccharide remaining in the batch culture fermentations showed that the arabinose side chains of pectin, xylan and arabinogalactan were co-utilised with the backbone sugars. In these cultures, polysaccharide-degrading activity was mainly cell-associated, but extracellular polysaccharidase activity increased as the fermentations progressed. Molar ratios of acetate, propionate and butyrate produced in these experiments were dependent upon the polysaccharide substrate tested. Molar ratios of acetate, propionate and butyrate in the starch, arabinogalactan, xylan and pectin fermentations were 50:22:29, 50:42:8, 82:15:3, and 84:14:2, respectively. The presence of starch did not inhibit the breakdown of arabinogalactan, xylan or pectin by faecal bacterial, providing evidence that multicomponent substrate utilisation occurs when complex populations of faecal bacteria are provided with mixed polysaccharide substrates.

The interactions occuring between populations of a nitrate-respiring Vibrio sp. and autotrophic nitrifying bacteria belonging to the genera Nitrosomonas and Nitrobacter have been investigated in a compound bi-directional flow diffusion chemostat at a dilution rate of 0.025 h−1 and a temperature of 25°C. When grown under NO−3 limitation, the Vibrio sp. produced NH+4 as the principal end-product of nitrate respiration, and there was a corresponding significant increase in cell numbers of the Nitrosomonas sp. population, which derived energy by the oxidation of NH+4 to NO−2. Nitrite in turn was used by the Nitrobacter sp. population as an energy source with the concomitant regeneration of NO−3. Under NO−3 excess growth conditions the Vibrio sp. produced NO−2 rather than NH+4 as the major product of NO−3 dissimilation, and growth of the Nitrobacter population was stimulated as increased quantities of NO−2 became available. In contrast, the Nitrosomonas sp. population declined sharply as the energy source NH+4 became limiting. These data demonstrate that defined mixed populations of obligately aerobic nitrifying bacteria and facultatively anaerobic nitrate respiring bacteria can co-exist for extended time periods and operate an internal nitrogen cycle which is energetically beneficial to both populations.

The effects of O2 tension, temperature, salt concentration and organic matter concentration on the growth and nitrifying activity of Nitrosomonas N3 isolated from Tay Estuary sediments have been investigated. Chemostat-grown cultures were able to grow and nitrify at dissolved O2 concentrations as low as 0.1 mg O2· 1−1 (cell population densities were 15% of those obtained in fully aerated cultures). This bacterium was sensitive to reduced temperatures as chemostat-grown cultures washed out at growth temperatures below 15°C, at dilution rates > 0.025 · h−1. Batch-grown cultures of Nitrosomonas N3 were used to study the effects of NaCl and complex organic matter concentration on nitrifying activity. Maximum rates of NH+4 oxidation were recorded at NaCl concentrations of 1% w/v, whilst tryptone soya broth (TSB), nutrient broth (NB), yeast extract broth (YEB) and peptone were inhibitory at concentrations > 10 mg · 1−1.

A comparative study has been undertaken to determine the efficiency of methods for the enrichment and isolation of autotrophic nitrifying bacteria from soils and estuarine and marine sediments. Chemostat enrichments proved to be the most efficient means of isolating autotrophic NH+4 oxidisers whereas NO−2 oxidising bacteria were never successfully enriched by this method. In contrast, gel enrichment and traditional batch culture enrichments of nitrifying bacteria were comparatively time consuming procedures and the degree of enrichment obtained for NH+4 oxidising bacteria never approached that obtained with continuous culture enrichments. Gel enrichments, however, because they have continuous physicochemical gradients provide qualitative advantages in that morphologically distinct types of nitrifying bacteria can be isolated from the same gel.

Mixed populations of human gut bacteria degraded cas casein by producing a variety of cell-bound and extracellular proteolytic enzymes. Casein was initially hydrolysed to TCA soluble peptides which were subsequently broken down to volatile fatty acids, ammonia, dicarboxylic acids and a range of phenolic compounds. Amino acids did not accumulate to any extent during casein breakdown, suggesting that the rate of peptide hydrolysis was the limiting step in protein utilisation by these bacteria. Similar fermentation products were produced from bovine serum albumin, however, the insoluble protein collagen was considerably more resistant to degradation by the colonic microflora, as evidenced by the reduced quantities of fermentation end-products formed.