The use of rats associated with a human faecal flora as a model for studying the effects of diet on the human gut microflora.

The activities of four bacterial biotransformation enzymes (beta-glucosidase, beta-glucuronidase, nitrate reductase and nitroreductase) were measured in the caecal contents of conventional flora rats or germ-free rats contaminated with a mixed, human faecal flora and compared with activities present in a fresh human stool preparation. Both the conventional flora rats and the rats inoculated with a human flora exhibited an enzyme profile generally similar to that of human faeces, although the conventional rat flora exhibited negligible nitrate reductase activity. The enzyme profile remained essentially unaltered in both human flora preparations following supplementation of the diet with pectin, whereas the conventional rat flora responded to this plant cell wall carbohydrate with a significant increase in nitrate reductase activity.

The hepatic conversion of some heterocyclic amines to bacterial mutagens is modified by dietary fat and cholesterol.

Groups of Sprague-Dawley rats were fed on diets containing increasing amounts of beef dripping, but having a constant cholesterol content. One group of rats was fed on a diet containing no dripping and no added cholesterol (control). We have studied the ability of individual hepatic S9 preparations to activate the cooked food mutagens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQX) to bacterial mutagens using Salmonella typhimurium TA98 as indicator.

Use of continuous culture to study the gastric microflora of a hypochlorhydric patient.

A method of maintaining the microflora obtained from the hypochlorhydric stomach of a patient suffering from hypogammaglobulinaemia has been developed using continuous culture (chemostat) techniques. The culture was maintained at pH 8.0 (the pH of the original gastric juice) and subsequently at pH 7.

Dietary modification of intestinal bacterial enzyme activities--potential formation of toxic agents in the gut.

The bacterial population colonising the large intestine is able to metabolise a variety of ingested or endogenously produced substances to products, some of which possess toxic, mutagenic or carcinogenic properties. Dietary components, resistant to digestion and absorption in the upper alimentary tract, may influence these reactions by altering the environment of the gut or through the provision of nutrients to the flora. Evidence for the involvement of bacterial enzymes in the formation of toxic products in vivo has come largely from animal studies, particularly where fermentable plant cell-wall components are present in the diet.

Effects of plant-derived flavonoids and polyphenolic acids on the activity of mutagens from cooked food.

The ability of 3 plant flavonoids (morin, myricetin and quercetin) and 4 polyphenolic acids (caffeic acid, chlorogenic acid, ellagic acid and ferulic acid) to inhibit the genotoxic effects of a number of cooked-food mutagens (IQ, MeIQ, MeIQx, Trp-P-1 and Trp-P-2), was investigated in a bacterial mutation assay using Salmonella typhimurium TA98 as indicator and hepatic S9 mixes from either SWR mice or Syrian hamsters as metabolic activating systems. Although the polyphenolic acids failed to have an effect, the flavonoids generally inhibited IQ, MeIQ, MeIQx and Trp-P-1 induced mutagenesis in a dose-dependent manner, irrespective of the source of S9. This was not the case with Trp-P-2 where the flavonoids were only observed to inhibit when SWR mouse S9 but not Syrian hamster S9 was used.

Metabolic conversion of IQ and MeIQ to bacterial mutagens.

The metabolic conversion of 2-amino-3-methyl- and 2-amino-3,4-dimethyl-imidazo[4,5-f]quinoline (IQ and MeIQ respectively) to bacterial mutagens was studied using a bacterial mutation assay. Studies were performed using S9 fractions derived from either corn oil (uninduced) or Aroclor-1254-treated Sprague-Dawley rats. Aroclor 1254 treatment lowered the S9 protein concentration required for optimum levels of mutagenesis, enhanced the numbers of mutants observed and altered the effects of metabolic inhibitors and cofactors added to the assay.

The effect of various dietary fibres on tissue concentration and chemical form of mercury after methylmercury exposure in mice.

The whole-body retention of mercury after exposure of BALB/c mice to methylmercury was measured in animals fed fibre-free, 5% pectin, 5% cellulose or 5, 15 or 30% wheat bran diets. The rate of elimination of mercury was dependent on the diet fed, with dietary bran increasing the rate of elimination. The incorporation of 15 or 30% bran in the diet of the mice decreased the total mercury concentration in the brain, blood and small intestine, although the effects were significant only in those animals on 30% bran diet.

Enzyme activities of the hindgut microflora of laboratory animals and man.

A comparison was made in six species of animal (rat, mouse, hamster, guinea-pig, marmoset and man) of five enzyme activities associated with the hindgut microflora. Marked differences were found in the caecal activities of azoreductase, beta-glucosidase, beta-glucuronidase, nitrate reductase and nitroreductase in the four rodents, with no one species exhibiting consistently higher or lower enzyme activity. None of the laboratory animals, including the marmoset, provided an approximation of the enzyme profile associated with human faecal flora.

The use of continuous flow systems for studying the metabolic activity of the hindgut microflora in vitro.

To investigate the involvement of bacterial enzyme activities in the biotransformation of xenobiotic compounds, we have developed a simulation of the rat hindgut microflora in vitro. This mixed bacterial population exhibits many similarities to the native rat flora, and the diversity of bacterial species and the activity of a number of hydrolytic and reductive enzymes (e.g.

Induction of unscheduled DNA synthesis in rat and hamster hepatocytes by cooked food mutagens.

The genotoxicity of the cooked-food mutagens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) was studied by monitoring the induction of DNA repair (unscheduled DNA synthesis; UDS) in primary cultures of rodent hepatocytes. The hepatocytes were derived from male Sprague-Dawley rats or Syrian hamsters by collagenase perfusion and the cells were cultured for 4 hr before being exposed to various concentrations of the mutagens. DNA repair was determined by measuring incorporation of [3H]thymidine into extracted DNA over 17 hr using beta-scintillation counting.

Effect of mixtures of dietary fibres on the enzyme activity of the rat caecal microflora.

The enzyme activity of the caecal microflora from weanling rats was determined after feeding 1 of 3 basal diets (purified fibre-free; purified plus cellulose; and stock), with or without additional dietary fibre (pectin, i-carrageenan or carboxymethylcellulose 5% w/w). The wet weight of caecal contents and total bacterial numbers were similar for the purified fibre-free and purified plus cellulose diets, yet were significantly higher in animals fed the stock diet. Pectin supplementation of the basal diets had no effect of caecal bacterial numbers, but significantly increased total nitrate reductase activity per caecum except when added to stock diet.

Influence of wheat bran on some reductive and hydrolytic activities of the rat cecal flora.

For 30 days, male weanling rats were fed a semipurified, fiber-free diet or a diet that contained 5, 15, or 30% (wt/wt) wheat bran. The activities of four cecal microbial enzymes were determined. Wheat bran significantly increased the wet weight content of the cecum and total bacterial count per cecum at the intermediate- and high-treatment levels, but it had no effect on bacterial concentration per gram wet weight of cecal contents.

The influence of the host on expression of intestinal microbial enzyme activities involved in metabolism of foreign compounds.

The activities of four enzymes (beta-glucuronidase, nitrate reductase and nitroreductase) in selected intestinal bacteria (Escherichia coli, Clostridium sp., Streptococcus sp., Bacteroides sp.

Metabolic adaptation of rat faecal microflora to cyclamate in vitro.

Previous studies have demonstrated that the rat faecal microflora maintained in vitro under conditions of continuous flow possesses bacteriological and metabolic characteristics similar to those of the native bacterial population of the caecum. Addition of sodium cyclamate (75 mM) to the culture concurrent with the progressive dilution of the growth medium promoted metabolism of cyclamate to cyclohexylamine (sulphamatase activity) within 4 wk. The maximum formation of cyclohexylamine was attained in about 8 wk and was equivalent to a 2-3% molar conversion of cyclamate to cyclohexylamine.

The role of oral nitrate in the nitrosation of [14C]proline by conventional microflora and germ-free rats.

The urinary excretion of N-nitroso-L-[U-14C]proline by conventional microflora and germ free rats was used to assess the role of the gut bacteria and oral nitrate in the endogenous formation of N-nitroso compounds. The formation of nitrosoproline was qualitatively similar in conventional and germfree rats (equivalent to nitrosation of approximately 0.01-0.

Activation of the food mutagens IQ and MeIQ by hepatic S9 fractions derived from various species.

The ability of hepatic S9 mixes derived from different rodent species (rat, mouse, Syrian and Chinese hamster) to activate the mutagens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) was investigated using Salmonella typhimurium strain TA98. In general, the mutagenicity of IQ and MeIQ was greatest in the presence of S9 fractions from Swiss albino mice and least from fractions derived from Chinese hamsters. However, treatment of rats or hamsters with Aroclor 1254 had little or no effect on the activation of IQ or MeIQ to mutagens.

Changes in the distribution of histochemically localized mercury in the CNS and in tissue levels of organic and inorganic mercury during the development of intoxication in methylmercury treated rats.

The development of neurotoxicity in rats after exposure to methylmercuric chloride was monitored using behavioural indices. At selected time points the cellular localization of mercury and the relative amounts of organic and inorganic mercury were determined in several regions of the CNS, and in some non-neural tissues. The CNS showed an affinity for organic mercury, the levels of inorganic mercury remaining low throughout symptomatic intoxication.

Modification of rat caecal microbial biotransformation activities by dietary saccharin.

Male Sprague-Dawley rats were fed a purified fibre-free diet containing 5% (w/w) sodium saccharin for 4 weeks or 20 weeks and changes in caecal bacterial numbers and enzyme activities (endogenous ammonia production, beta-glucosidase, beta-glucuronidase, nitrate reductase, nitroreductase, aryl sulphatase) determined in vitro. Saccharin treatment gave marked caecal enlargement but had no effect on bacterial concentration at either treatment period, and significantly decreased beta-glucuronidase, nitrate reductase and sulphatase activities/g caecal contents. The incubation of a suspension of caecal contents from control rats with saccharin (75 mM) in vitro inhibited beta-glucuronidase and nitrate reductase activities, and ammonia production from endogenous substrates.

Influence of dietary carrageenans on microbial biotransformation activities in the cecum of rodents and on gastrointestinal immune status in the rat.

Rats, mice, and hamsters were fed iota-carrageenan incorporated in a fiber-free, purified diet for 30 days, and the activities of a number of cecal microbial enzymes were determined in vitro. Carrageenan treatment produced cecal enlargement in all species, yet significantly decreased the concentration of bacteria per gram of cecal content. Azoreductase, beta-glucosidase, beta-glucuronidase, nitrate reductase, and nitroreductase activities per gram of cecal content were significantly decreased in the rat, although less consistent effects were found in these enzymes in the mouse and hamster.

The role of DNA-repair processes in N-nitrosopyrrolidine-induced mutagenesis.

The cytotoxic and mutagenic effects of increasing concentrations of N-nitrosopyrrolidine (NPYR) were studied using various DNA repair mutants of Escherichia coli together with rat-liver S9 activation system. Irrespective of which strain was used, the cytotoxic effects of NPYR were similar to those observed in the parent strain. Mutagenicity studies revealed that the uvrA- derivative was more mutable than its repair proficient parent.

The resistance of E. coli cultivated in low concentrations of dichlorvos to N-methyl-N'-nitro-N-nitrosoguanidine induced mutagenesis.

Cultivation of E. coli B/r strain WP2 in low concentrations of either 4-nitroquinoline N-oxide (4NQO) or N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) had no effect on the mutagenic or cytotoxic consequences of subsequent challenge with dichlorvos (DCV). However, although the sensitivity of E.

The effect of diet on the mammalian gut flora and its metabolic activities.

The review will encompass the following points: A brief introduction to the role of the gut flora in the toxicology of ingested food components, contaminants, and additives, including known pathways of activation and detoxication of foreign compounds and the implication of the flora in enterohepatic circulation of xenobiotics. The advantages and disadvantages of the various methods of studying the gut flora (classical bacteriological techniques, metabolic and enzymological methods) will be critically discussed with special reference to their relevance to dietary, toxicological, and biochemical studies. Sources of nutrients available to the gut flora will be described including host products (mucus, sloughed mucosal cells, hormones, proteins) and exogenous nutrients derived from diet.