Epidemiological studies have identified increased colorectal cancer (CRC) risk with high red meat (HRM) intakes, whereas dietary fibre intake appears to be protective. In the present study, we examined whether a HRM diet increased rectal O6-methyl-2-deoxyguanosine (O6MeG) adduct levels in healthy human subjects, and whether butyrylated high-amylose maize starch (HAMSB) was protective. A group of twenty-three individuals consumed 300 g/d of cooked red meat without (HRM diet) or with 40 g/d of HAMSB (HRM+HAMSB diet) over 4-week periods separated by a 4-week washout in a randomised cross-over design. Stool and rectal biopsy samples were collected for biochemical, microbial and immunohistochemical analyses at baseline and at the end of each 4-week intervention period. The HRM diet increased rectal O6MeG adducts relative to its baseline by 21 % (P< 0·01), whereas the addition of HAMSB to the HRM diet prevented this increase. Epithelial proliferation increased with both the HRM (P< 0·001) and HRM+HAMSB (P< 0·05) diets when compared with their respective baseline levels, but was lower following the HRM+HAMSB diet compared with the HRM diet (P< 0·05). Relative to its baseline, the HRM+HAMSB diet increased the excretion of SCFA by over 20 % (P< 0·05) and increased the absolute abundances of the Clostridium coccoides group (P< 0·05), the Clostridiumleptum group (P< 0·05), Lactobacillus spp. (P< 0·01), Parabacteroides distasonis (P< 0·001) and Ruminococcus bromii (P< 0·05), but lowered Ruminococcus torques (P< 0·05) and the proportions of Ruminococcus gnavus, Ruminococcus torques and Escherichia coli (P< 0·01). HRM consumption could increase the risk of CRC through increased formation of colorectal epithelial O6MeG adducts. HAMSB consumption prevented red meat-induced adduct formation, which may be associated with increased stool SCFA levels and/or changes in the microbiota composition.

Colorectal cancer (CRC) is a leading cause of preventable cancer deaths worldwide, with dietary factors being recognised as key risk modifiers. Foods containing dietary fibre are protective to a degree that the World Cancer Research Fund classifies the evidence supporting their consumption as ‘convincing’. The mechanisms by which fibre components protect against CRC remain poorly understood, especially their interactions with the gut microbiome. Fibre is a composite of indigestible plant polysaccharides and it is emerging that fermentable fibres, including resistant starch (RS), are particularly important. RS fermentation induces SCFA production, in particular, relatively high butyrate levels, and in vitro studies have shown that this acid has strong anti-tumorigenic properties. Butyrate inhibits proliferation and induces apoptosis of CRC cell lines at physiological concentrations. These effects are attributed to butyrate's ability to alter gene transcription by inhibiting histone deacetylase activity. However, the more recent discovery of G-protein coupled receptors that bind butyrate and other SCFA and data obtained from proteomic and genomic experiments suggest that alternative pathways are involved. Here, we review the mechanisms involved in butyrate-induced apoptosis in CRC cells and, additionally, the potential role this SCFA may play in mediating key processes in tumorigenesis including genomic instability, inflammation and cell energy metabolism. This discussion may help to inform the development of strategies to lower CRC risk at the individual and population levels.

Population studies show that greater red and processed meat consumption increases colorectal cancer risk, whereas dietary fibre is protective. In rats, resistant starches (a dietary fibre component) oppose colonocyte DNA strand breaks induced by high red meat diets, consistent with epidemiological data. Protection appears to be through SCFA, particularly butyrate, produced by large bowel carbohydrate fermentation. Arabinoxylans are important wheat fibre components and stimulate large bowel carbohydrate SCFA production. The present study aimed to determine whether an arabinoxylan-rich fraction (AXRF) from wheat protected colonocytes from DNA damage and changed colonic microbial composition in pigs fed with a diet high (30 %) in cooked red meat for 4 weeks. AXRF was primarily fermented in the caecum, as indicated by higher tissue and digesta weights and higher caecal (but not colonic) acetate, propionate and total SCFA concentrations. Protein fermentation product concentrations (caecal p-cresol and mid- and distal colonic phenol) were lower in pigs fed with AXRF. Colonocyte DNA damage was lower in pigs fed with AXRF. The microbial profiles of mid-colonic mucosa and adjacent digesta showed that bacteria affiliating with Prevotella spp. and Clostridial cluster IV were more abundant in both the mucosa and digesta fractions of pigs fed with AXRF. These data suggest that, although AXRF was primarily fermented in the caecum, DNA damage was reduced in the large bowel, occurring in conjunction with lower phenol concentrations and altered microbial populations. Further studies to determine the relationships between these changes and the lowering of colonocyte DNA damage are warranted.

Factors controlling the concentration of resistant starch (RS) in foods are of considerable interest on account of the potential for this type of fibre to deliver health benefits to consumers. The present study was aimed at establishing changes in starch granule morphology as a result of human small-intestinal digestion. Volunteers with ileostomy consumed six selected foods: breakfast cereal (muesli), white bread, oven-baked French fries, canned mixed beans and a custard containing either a low-amylose maize starch (LAMS) or a high-amylose maize starch (HAMS). Analysis showed that digesta total RS (as a fraction of ingested starch) was: muesli, 8·9 %; bread, 4·8 %; fries, 4·2 %; bean mix, 35·9 %; LAMS custard, 4·0 %; HAMS custard, 29·1 %. Chromatographic analysis showed that undigested food contained three major starch fractions. These had average molecular weights (MW) of 43 500 kDa, 420 kDa and 8·5 kDa and were rich in amylopectin, higher-MW amylose and low-MW amylose, respectively. The low-MW amylose fraction became enriched preferentially in the stomal effluent while the medium-MW starch fraction showed the greatest loss. Fourier transform IR spectroscopy showed that absorbance at 1022 per cm decreased after digestion while the absorbance band at 1047 per cm became greater. Such changes have been suggested to indicate shifts from less ordered to more ordered granule structures. Further analysis of amylose composition by scanning iodine spectra indicated that the MW of amylose in ileal digesta was lower than that of undigested amylose. It appears that high-MW amylose is preferentially digested and that MW, rather than amylose content alone, is associated with resistance of starch to digestion in the upper gut of humans.

Himalaya 292 (Hordeum vulgare var. Himalaya 292) is a novel hull-less barley variety lacking activity of a key enzyme of starch synthesis giving a grain containing less total starch, more amylose and higher total dietary fibre. Animal trials have shown that Himalaya 292 contains more resistant starch and has greater positive impact on biomarkers of large-bowel health than comparable wholegrain cereal products. The present study compared the effects of foods made from wholegrain Himalaya 292 with those made from wholegrain wheat on faecal biomarkers of bowel health in human subjects. Seventeen male and female volunteers aged 31–66 years consumed similar quantities of Himalaya 292, whole-wheat or refined cereal foods daily for 4 weeks in a randomised cross-over design. Total dietary fibre intakes from weighed food records were 45, 32 and 21 g/d for the Himalaya 292, whole-wheat and refined cereal periods, respectively. Compared with the refined cereal foods, consumption of Himalaya 292 foods resulted in 33 % higher faecal weight, a lowering of faecal pH from 7·24 to 6·98, a 42 % higher faecal concentration and a 91 % higher excretion of butyrate, a 57 % higher faecal total SCFA excretion and a 33 % lower faecal p-cresol concentration. pH and butyrate concentration and excretion were also significantly different compared with wholemeal wheat. It is concluded that consumption of a diet that included foods made from Himalaya 292 supplied more fibre and improved indices of bowel health compared with refined cereal foods and, for some indices, similar wholemeal wheat foods at equivalent levels of intake.

Non-infectious diseases such as CHD and certain cancers have become major causes of death and disability in affluent countries. Probiotics (principally lactic acid bacteria; LAB) may assist in lowering the risk of these diseases. Experimental studies with probiotics have given generally inconclusive outcomes for infectious disease and for biomarkers for non-infectious disease. In part this situation may reflect their inability to colonise the adult human gut effectively. Prebiotics can assist in promoting colonisation, and resistant starch (RS), as a high-amylose starch, is a prebiotic and synbiotic. This starch exerts its synbiotic action through adhesion of the bacteria to the granule surface. Consumption of RS assists in recovery from infectious diarrhoea in man and animals. A rice porridge, high in RS, appears to modify the autochthonous porcine large-bowel microflora favourably through lowering Escherichia coli and coliform numbers. Many of the beneficial effects of RS on large-bowel function appear to beexerted through short-chain fatty acids (SCFA) formed by bacterial fermentation. In man LAB are found in relatively highest numbers in milk-fed infants where theprofile of fermentation products differs quite markedly from that in adults. It appearsunlikely that ingestion of current probiotics will alter either total SCFA or the proportions of the major acids. More emphasis needs to be given to the investigation of the effects of complex carbohydrates, including RS, on the autochthonous microflora of the human large bowel.

Feeding higher levels of dietary animal protein (as casein or red meat) increases colonic DNA damage and thins the colonic mucus barrier in rats. Feeding resistant starch (RS) reverses these changes and increases large bowel SCFA. The present study examined whether high dietary dairy (casein or whey) or plant (soya) proteins had similar adverse effects and whether dietary RS was protective. Adult male rats were fed diets containing 15 or 25 % casein, whey or soya protein with or without 48 % high amylose starch (as a source of RS) for 4 weeks. DNA damage was measured in isolated colonocytes using the comet assay. Higher dietary casein and soya (but not whey) increased colonocyte DNA damage. DNA damage was highest with soya when fed at 15 or 25 % protein without RS. Dietary RS attenuated protein-induced colonocyte DNA damage in all groups but it remained significantly higher in rats fed 25 % soya compared with those fed 15 % protein. Dietary protein level did not affect colonic mucus thickness overall but the barrier was thinner in rats fed high dietary casein. This effect was reversed by feeding RS. Caecal total SCFA and butyrate pools were higher in rats fed RS compared with digestible starch. Caecal and faecal SCFA were unrelated to genetic damage but correlated with mucus thickness. The present data confirm that higher dietary protein affected colonocyte DNA and colonic mucus thickness adversely but that proteins differ in their effects on these indices of colon health. The data show also that these changes were reversed by RS.

Four groups of young pigs (n 6) were fed a diet containing 50 % maize starch as either a highly digestible waxy starch (control; 0 % amylose) or one of three resistant starch (RS) diets, namely a high-amylose maize starch (HAMS; 85 % amylose), this starch subjected to hydrothermal treatment (HTHAMS; 85 % amylose), or a blend of HAMS and HTHAMS included in equal amounts, for 21 d. Food intake and live weight at the end of the study were similar among the four groups. Ileal starch digestibility was lower in pigs fed the three RS diets but was greater for HAMS (88 %) than for HTHAMS (70 %; P < 0·05). Faecal output and large bowel digesta mass, and concentrations and pools of individual and total SCFA were higher (by about two- to threefold; all P < 0·05) and digesta pH lower (by about 1 unit, all P < 0·001) in pigs fed either HAMS or HTHAMS compared to the controls. These differences in biomarkers were seen along the length of the large bowel. Colon length was 0·5–0·9 m longer (19–35 %) in pigs fed the high-RS diets relative to those fed the highly digestible starch diet (P < 0·05). Faecal and proximal colonic lactobacilli and bifidobacteria numbers were higher (by 1 and 3 log units; P < 0·05) in pigs fed the HAMS or HTHAMS diets. Although both high-amylose starches promoted fermentation throughout the large bowel, the data suggest that the effects of HTHAMS may be more pronounced in the distal region compared to those of HAMS.

Large-bowel fermentation of resistant starch produces SCFA that are believed to be important in maintaining visceral function. High-amylose maize starch (HAMS) and acylated starches are sources of resistant starch and are an effective means of increasing colonic SCFA. Cooking increases digestibility of starches but its effects on the capacity of these starches to raise large-bowel SCFA are unknown. We have examined the effects of cooking of HAMS and butyrylated HAMS (HAMSB) on amylolysis in vitro and their capacity to raise caeco-colonic SCFA in rats. The starches were boiled in excess water and microwaved, followed by drying at 100°C. Cooking increased in vitro glucose release for both starches but significantly less from HAMSB. Rat growth rates were unaffected when fed cooked resistant starch. Digesta pH was increased in the caecum and proximal colon of rats fed cooked HAMS. Distal colonic pH was highest in rats fed cooked HAMSB. Factorial analyses (2×2) of caecal SCFA pools showed significant differences between HAMS and HAMSB, and that cooking significantly lowered caecal butyrate pools. Portal venous butyrate concentrations were higher in both HAMSB groups than those fed HAMS. The data suggest that HAMSB is less susceptible to in vitro amylolysis than HAMS following cooking and delivers more butyrate to rat caecum than HAMS. This attribute may be useful in food applications for specific delivery of SCFA to the colon. Preparation of carbohydrates to simulate human food in animal experiments may be important to assess nutritional and physiological effects accurately.

Hordeum vulgare var. Himalaya 292 is a new barley cultivar with altered starch synthesis and less total starch but more amylose, resistant starch (RS) and total and soluble NSP including β-glucan. To determine its nutritional potential, young pigs were fed diets containing stabilised wholegrain flours from either Himalaya 292, Namoi (a commercial barley), wheat bran or oat bran at equivalent dietary NSP concentrations for 21 d. Serum total cholesterol was significantly lowered by the Himalaya 292 diet relative to wheat bran, indicating that Himalaya 292 retained its hypocholesterolaemic potential. In all groups SCFA concentrations were highest in the proximal colon and decreased towards the rectum. Digesta pH was lowest in the proximal colon and highest in the distal colon. Large-bowel and faecal pH were significantly lower in the pigs fed the barley and oat diets, indicating greater bacterial fermentation. Caecal and proximal colonic pH was lowest and SCFA pools highest in the pigs fed Himalaya 292. Total and individual SCFA were lowest in the mid- and distal colon of the pigs fed Himalaya 292 or oat bran. These data suggest the presence of more RS in Himalaya 292 and suggest that its fermentation was rapid relative to transit. Differences in faecal and large-bowel anaerobic, aerobic, coliform and lactic acid bacteria were relatively small, indicating a lack of a specific prebiotic action. These data support the potential of this novel barley cultivar to improve health through plasma cholesterol reduction and increased large-bowel SCFA production.

Adult male pigs were fed on a diet containing (% of energy) fat 25 starch 55 from white rice and providing 20 g fibre/pig per d (diet WR). In two other groups rice bran was added to the diet to provide 43 g fibre/d. One group received the diet unmodified (diet RB), but in another (diet RO) heat-stabilized unrefined rice oil replaced the palm oil. In a further group brown rice replaced white rice and provided 37 g fibre/pig per d (diet BR). Plasma cholesterol concentrations were similar with diets WR, RB and BR. With diet RO the concentration was significantly lower than with diets WR and BR but was not different from diet RB. Plasma high-density-lipoprotein-cholesterol and plasma triacylglycerols were unaffected by diet. In all groups, digesta mass rose from the caecum to the proximal colon but fell in the distal colon. Diet WR gave the lowest digesta mass while diet BR gave a significantly higher mass along the large bowel length. RB- and RO-fed pigs had equal masses of digesta which were intermediate between BR- and WR-fed pigs at all sampling sites. Pools of individual and total volatile fatty acids (VFA) in the proximal large bowel were unaffected by diet. Pools of total and individual VFA in the median and distal colon were lowest with diets WR and RB and significantly higher with diet BR. In these regions of the colon pools of acetate in RO-fed pigs did not differ from those in the BR-fed group but were higher than in other groups. However, pools of propionate and butyrate with the RO diet were significantly lower than with diet BR and the same as with diets WR and RB. Portal venous VFA concentrations were unaffected by diet. The higher large bowel digesta masses and VFA with diet BR may reflect the escape of starch from the smallintestine.

Male rats were fed on either a non-purified rodent diet (JS) or cholesterol-free purified diets containing wheat bran (WB) or oat bran (OB). Some animals were allowed normal access to their faeces for coprophagy (coprophagy +), while in others coprophagy was prevented by placement of a plastic cup over the anus (coprophagy −). Direct ingestion of faeces from the anus was observed in the former groups. Food intake was unaffected by diet or coprophagy status and body weight gain was unchanged with OB − and JS − but was significantly lower with WB −. Plasma cholesterol was highest with WB and equally lower with OB and JS and was unaffected by coprophagy status. Plasma triacylglycerols were highest with OB and were unaffected by coprophagy status. Caecal digesta mass was highest with JS, intermediate with OB and lowest with WB. Digesta mass was unaffected by coprophagy status with WB and JS but was higher with OB −. Digesta moisture content was lowest with WB + but highest with WB −. Digesta volatile fatty acid (VFA) concentrations were similarly lower with OB + and OB −, but were significantly lower with JS − and WB − than in the corresponding coprophagy + group. In all groups digesta butyrate concentrations were reduced by coprophagy prevention. Pools of total VFA, acetate and butyrate in the digesta were highest with JS. Pools of total VFA in digesta were highest with JS +, OB + and OB − and lowest with WB + and WB −. The propionate pool was highest with OB −, intermediate with OB +, and equally low in all other groups. The pool of butyrate was highest with JS + and lowest with OB −. Effects of oats and wheat on plasma cholesterol in the rat do not seem to be mediated through faecal re-ingestion.

In adult male rats fed on a cholesterol-free synthetic diet, plasma cholesterol concentrations were lowest with oat bran, intermediate with cellulose and highest with wheat bran. Plasma triacylglycerols (TAG) were similar with wheat bran and cellulose but higher with oat bran. The concentrations and pools of caecal volatile fatty acids (VFA) were lowest with cellulose and equally higher with oat bran and wheat bran. Plasma VFA concentrations in the hepatic portal vein reflected those in caecal digesta and were unrelated to plasma cholesterol. Feeding oat bran after extraction with n-pentane gave plasma cholesterol concentrations similar to that found with wheat bran. Reconstitution of oat bran with extracted lipids did not restore the cholesterol-lowering effect. Addition of the extracted material to a wheat-bran diet had no effect on plasma cholesterol. Plasma TAG were higher with the oat bran and reconstituted-oatbran diets than with wheat-bran or cellulose diets. However, extracted oat bran+safflower oil gave similar TAG concentrations to that with wheat bran. These extractions and additions did not change caecal bile acid or neutral sterol concentrations. Effects of these diets on plasma cholesterol were unrelated to their tocotrienol or tocopherol content. Addition of n-pentane to oat bran followed by evaporation of solvent gave plasma cholesterol concentrations that were significantly higher than untreated oat bran but lower than similarly treated wheat bran. It is concluded that oat bran affects cholesterol metabolism through a pentane-soluble component as well as non-starch polysaccharides. It appears that the activity of this lipid is not transferable by simple addition of the solvent extract to the whole diet.

1. Adult male rats were maintained on diets containing 80 g methylcellulose/kg of low (25 cP), medium (400 cP) and high (1500 cP) viscosity.

2. After 10 d, the viscosity of stomach and caecal contents was found to have increased in proportion to that of the dietary fibre. Concentrations of volatile fatty acids in caecal digesta were lowest with the high-viscosity fibre but acetate was the major acid present with all three diets. Acetate was the only acid found in significant quantities in hepatic portal venous plasma and concentrations of this acid were unaffected by diet.

3. Concentrations of glucose in arterial blood were low with the medium-and high-viscosity diets while the content of liver glycogen was high. These effects of fibre were not directly on glucose absorption as the intestines were net removers of the hexose at the time of sampling.

4. Hepatic lipogenesis and plasma triacylglycerol concentrations were both higher in rats fed on the low-viscosity fibre. Plasma cholesterol concentrations, hepatic cholesterol synthesis and faecal bile acid excretion were not altered by dietary fibre viscosity.

5. We conclude that the effects of dietary fibre on carbohydrate absorption and storage and fatty acid synthesis are a function of the viscosity of the fibre in solution, high viscosity slowing the digestion and absorption of nutrients in the small intestine. Large-bowel microbial fermentation is not of direct significance to these events. In contrast, effects of fibre polysaccharides on sterol metabolism seem not to be related to their rheological properties.

1. Adult male rats were fed on diets containing 100 g dietary fibre/kg either as α-cellulose or wheat bran or the pericarp-seed coat or aleurone layers prepared from that bran by sequential milling and air elutriation and electrostatic separation.

2. After 10 d, concentrations of total volatile fatty acids (VFA) in caecal fluid were significantly different between groups and fell in the order: aleurone > wheat bran > pericarp-seed coat > cellulose. This ranking probably reflected the ease of fermentation of fibre polysaccharides by colonic bacteria which also resulted in a considerably higher faecal bacterial mass in the aleurone group.

3. Because of the differences in the volume of caecal digesta, the mass of caecal VFA was considerably the highest in the aleurone group, intermediate with wheat bran and equally low in the pericarp-seed coat and cellulose groups.

4. The diet based on aleurone gave a relatively higher proportion of propionate but with both pericarp-seed coat and wheat bran the contribution of butyrate was raised.

5. VFA concentrations in hepatic portal venous plasma were proportional to caecal concentrations with very high (> 3 mM) values being recorded in the aleurone group.

6. The findings are discussed in relation to the apparent susceptibility of the morphological components of wheat bran to fermentation by large bowel bacteria.