A comparison of indomethacin with ibuprofen on gastrointestinal mucosal integrity in conventional and germ-free rats.

The effects of indomethacin and ibuprofen on gastrointestinal mucosal integrity were studied in conventional and germ-free rats. Only ibuprofen induced significant gastric erosion formation in both conventional and germ-free animals, demonstrating that the presence of micro-organisms is not required in this form of damage. Both indomethacin and ibuprofen caused significant intestinal damage and blood loss in germ-free animals. However, in the conventional counterparts, damage due to indomethacin was enhanced whereas that induced by ibuprofen was not. The results from the present work would suggest that non-steroidal anti-inflammatory drugs such as indomethacin, which are secreted largely in the bile, unlike ibuprofen, may act in concert with bacteria and the constituents of bile to induce, in part, intestinal damage and blood loss.

Nitrate reduction, gastro-intestinal pH and N-nitrosation in gnotobiotic and conventional rats.

The reduction of nitrate in germ-free, gnotobiotic and conventional rats was investigated using blood methaemoglobin values as indicative of nitrite formation. Nitrate reduction was found to occur in the absence of a microbial flora, and throughout the experiment the blood content of methaemoglobin was higher in germ-free than in conventional rats. In vitro incubations of the gastric and small-intestinal mucosae of germ-free rats confirmed the presence of a heat-labile nitrate-reducing system. Measurement of the gastro-intestinal pH of germ-free and conventional rats revealed a generally higher pH value throughout the germ-free gastro-intestinal tract, with highly significant differences in the luminal pH of the forestomach, jejunum/ileum and caecum and a significant difference in the pH of the glandular stomach. Although some formation of N-nitrosoproline from proline and nitrate occurred in germ-free and gnotobiotic rats, nitrosation proceeded more readily in conventional rats. This effect may have been due to the lower gastric pH in the conventional rats although a more direct role for the flora cannot be discounted.

Influence of dietary protein and gut microflora on endogenous synthesis of nitrate and N-nitrosamines in the rat.

Groups of four germ-free (GF) and conventional (CV) rats were given purified diets containing either 50 or 200 g lactalbumin/kg for 2 wk and their urinary excretion of nitrate was measured. Urinary excretion of N-nitrosoproline was also measured in one of the three experiments. Both GF and CV rats given the high-protein diet excreted significantly more nitrate and N-nitrosoproline than those given the low-protein diet. On both diets GF rats excreted more nitrate than their CV counterparts but N-nitrosoproline excretion was not affected by environment. Groups of 11 GF and CV rats given diets containing sesame meal with or without a supplement of lysine-HCl for 2 wk, excreted similar amounts of nitrate on both diets, but more nitrate was excreted by GF rats than by their CV counterparts. N-nitrosoproline excretion by rats given the lysine supplement was higher in both environments. It is concluded that endogenous synthesis of nitrate is mediated by mammalian tissues rather than microflora and that dietary protein is an important source of nitrogen for the synthesis, although surplus amino acids from an imbalanced protein source do not act as precursors of endogenously formed nitrate. Some of the synthesized nitrate or its precursors appears to be metabolized by the microflora in the CV rat.

Influence of dietary protein and gut microflora on endogenous synthesis of nitrate induced by bacterial endotoxin in the rat.

Germ-free (GF) rats were maintained on a diet marginally adequate in protein, with and without a supplement of NH4Cl. Their urinary excretion of total nitrogen, nitrate, urea and creatinine was measured before and for 4 days after injection of Escherichia coli endotoxin (lipopolysaccharide; LPS). Although more nitrogen was excreted by rats on the diet supplemented with NH4Cl, nitrate excretion was increased to a similar extent in rats on both diets. This suggests that oxidation of ammonia released by deamination of amino acids is an unlikely pathway of nitrate synthesis. In a second experiment, nitrate excretion before and after injection of LPS was measured in GF and conventional (CV) rats given high- or low-protein diets. Urinary 3-methylhistidine (3MH) was measured as an index of breakdown of tissue protein. In both environments, nitrate excretion was significantly greater, before and after LPS administration, by rats on the high-protein diet than by their counterparts on the low-protein diet, and was generally greater by GF than by CV rats. Since only small, non-significant rises in urinary 3MH were observed after LPS treatment, it was concluded that the bulk of the nitrogen required for nitrate synthesis in response to endotoxin is derived from dietary protein rather than from nitrogenous products of tissue breakdown.

Gastrointestinal pH measurement in rats: influence of the microbial flora, diet and fasting.

The pH of the rat intestinal tract was decreased by the presence of a microbial flora, but its influence in the forestomach is less clear. Stomach pH values varied according to the amount of food present at the time of measurement. Fasting increased the pH of the gastrointestinal tract in conventional rats but had little effect in germfree rats. In the conventional rat, feeding a purified diet compared with a commercial diet resulted in a lower pH in the forestomach and a higher pH in the caecal contents. Magnesium trisilicate promoted gastric emptying in conventional rats and its antacid effect was observed only in the caecum and colon.

Dietary fat and N-nitrosation in the rat.

1. Groups of four conventional (CV) rats ate natural or purified diets either with or without 100 g fat/kg and drank 0.235 M-sodium nitrate. The fats tested were butterfat, coconut oil, olive oil, maize oil and safflower oil. 2. Decreased urinary excretion of N-nitrosoproline (NPRO) was observed in rats fed on fat-supplemented diets compared with those fed on low-fat diets, with butterfat having the greatest effect of the fats tested. 3. Reduced excretion of NPRO was not the result of inhibition of the intragastric N-nitrosation reaction or absorption of nitrosamine from the gastrointestinal tract. 4. The availability of nitrite in aqueous solution was decreased by the fat diets but the effect was similar in all the fats tested. 5. Nitrate reductase activity was present in the forestomach contents of CV rats at pH greater than 4 and was apparently inhibited by feeding a fat diet. No nitrate reductase activity was detected in stomach contents of germ-free rats. 6. Nitrate reductase activity in stomach and small intestinal tissue was not altered by feeding a fat diet. 7. It was concluded that nitrate reductase activity in stomach contents was of microbial origin and the decreased urinary excretion of NPRO on feeding the fat diets was mainly due to the inhibition of nitrate reductase activity in stomach contents.

Effect of dietary fats on endogenous formation of N-nitrosamines from nitrate in germ-free and conventional rats and rats harbouring a human flora.

Urinary excretion of N-nitrosoproline (NPRO) was measured in groups of four germ-free (GF) and conventional (CV) rats given a purified diet with or without inclusion of 100 g butterfat, coconut oil or maize oil/kg, and drinking water containing 0.235 M-NaNO3. In the CV environment rats given the fat-supplemented diets excreted significantly less NPRO than those on the low-fat diet. No corresponding decrease in NPRO excretion occurred in the GF environment. Nitrate reductase activity was measured in stomach contents and homogenates of stomach tissue from GF and CV rats given the different diets. No activity was detected in any of the contents from GF rats. Nitrate reductase activity was significantly reduced in contents from all the CV rats given the fat-supplemented diets, the effect being most marked in those given butterfat. Activity was much lower in tissue homogenates from GF rats than in those from their CV counterparts, but was not affected by diet in either environment. Groups of four CV rats, or rats harbouring a human faecal flora (HF), were given the purified diet with or without addition of 100 g butterfat or maize oil/kg and drinking water containing 0.235 M-NaNO3. All groups given the fat-supplemented diet with the exception of the HF rats given butterfat. It is concluded that the reduced excretion of NPRO by rats given diets containing fat was mainly due to inhibition of microbial nitrate reductase activity in the foregut. The smaller effect of butterfat on the HF rats accords with earlier findings in human subjects.