Dietary curdlan increases proliferation of bifidobacteria in the cecum of rats.

Significant increases in the amounts of short-chain fatty acids and lactate, and in numbers of bifidobacteria were observed in the cecum of curdlan (CD) -fed rats as compared with those of cellulose-fed ones. The in vitro proliferation of 5 species of bifidobacteria was markedly increased in the cultures containing the supernatant obtained from the cecal contents of CD-fed rats. These findings suggest that bifidus factors have been produced in the cecum of CD-fed rats.

Curdlan and gellan gum, bacterial gel-forming polysaccharides, exhibit different effects on lipid metabolism, cecal fermentation and fecal bile acid excretion in rats.

The effects of curdlan (CD) and gellan gum (GG), bacteria-producing polysaccharides, on lipid concentrations of serum and liver, fecal bile acid composition and intestinal fermentation products were studied in rats fed diets containing cellulose powder (CP), CD or GG at 5% for 4 wk. The cecal weight of the CD group increased significantly as compared to that of the other two groups and the pH of its contents was significantly low. The gastrointestinal transit time in the GG group was significantly shorter than that in the CP and CD groups. No significant inter-group differences were observed in the serum concentrations of total cholesterol and HDL-cholesterol, but a significant decrease was observed in the hepatic total cholesterol concentration of the CD group as compared to that of the CP and GG groups. No significant difference in the total bile acid excretion in feces was observed among the groups, but significantly low values were observed in the proportion of secondary bile acids in the CD group as compared to those of the CP and GG groups. Amounts of short-chain fatty acids (acetic, propionic and butyric acid) and lactic acid in the cecal contents were significantly higher in the CD group than in the other two groups. These results reveal that dietary CD is easily degraded and fermented by intestinal bacteria in the cecum and lowers cholesterol concentration in the liver, while dietary GG shortens the gastrointestinal transit time, suggesting the promotion of evacuation.

Secretion and excretion of immunoglobulin A to cecum and feces differ with type of indigestible saccharides.

The study was conducted to elucidate the effects of orally administered indigestible saccharides (IDS) on immunoresponses of the intestinal tracts, especially secretion and excretion of immunoglobulin A (IgA). Male 4-week-old Sprague-Dawley rats were fed diets containing several kinds of IDS (cellulose, corn husk, glucomannan, curdlan and lactulose) at 5% for three weeks. The results indicated that the proportion of IgA-presenting lymphocytes in the cecal mucosa of the tested IDS groups increased significantly or tended to increase compared with that of the cellulose group. No significant differences among the experimental groups were observed in the CD4(+)- and CD8(+)-presenting lymphocytes and the CD4+/CD8+ ratios in the small intestine, cecum and mesenteric lymph nodes. IgA amounts in the cecal contents increased significantly in the glucomannan and curdlan groups as compared with that in the cellulose group. The inconsistent results were observed in the cecal IgA amounts of the lactulose group. Although IgA excretion into feces increased periodically in the cellulose, hardly any changes were observed in the glucomannan and curdlan groups. These results revealed that IgA secretion from cecal mucosa to contents was promoted, and its excretion to feces was decreased by the oral administration of highly fermentable IDS, respectively, while non- or low-fermentable IDS functioned adversely to IgA responses in the intestinal tract. It is suggested that the response of IgA in the intestinal immune system differs with the type of IDS ingested.

Dried green leaf powders of Jew's mellow (Corchorus), persimmon (Diosphyros kaki) and sweet potato (Ipomoea batatas poir) lower hepatic cholesterol concentration and increase fecal bile acid excretion in rats fed a cholesterol-free diet.

A cholesterol-free diet containing dried powder of Jew's mellow leaves, persimmon leaves or sweet potato leaves respectively at 5% level as dietary fiber was fed to male Sprague-Dawley rats for about one month. The experiment was conducted twice except for sweet potato leaves. In the groups fed the diet mixed with powders of any of the three different dried green leaves, the hepatic cholesterol concentration significantly decreased. Such lowering was not observed in serum cholesterol concentration compared with the control (cellulose) group. A significant increase in fecal weight was observed in all the groups fed the green leaf samples. All the dried green leaves increased fecal excretion of bile acids per gram or per day compared with the control group in both experiments, but only the dried Jew's mellow leaves showed an increased excretion of neutral sterols. These results suggest that lowering of hepatic cholesterol by powdered green leaves is not necessarily due to the same factor, but to the increased fecal excretion of bile acids due to inhibited enterohepatic circulation in animals given these samples.

Changes in platelet aggregation and lipid metabolism in rats given dietary lipids containing different n-3 polyunsaturated fatty acids.

We compared the effects of different n-3 polyunsaturated fatty acids (PUFA) on platelet aggregation and lipid metabolism in rats. alpha-Linolenic acid (ALA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were used as n-3 PUFA sources. The rats were fed diets containing 10% lipids (polyunsaturated/saturated fatty acid (P/S) ratio = 1.0; n-3/n-6 = 0.02 for the control group, 0.2 for the test groups) for two weeks. The platelet counts, platelet aggregation, and production of thromboxane A2 (TXA2), plasma total cholesterol (TC) and triacylglycerols (TG) were not different between the ALA group and the control group, but showed a decreasing tendency for the EPA group and significant decreases for the DHA group. The production of prostacyclin in the aorta was significantly decreased in all of the n-3 PUFA groups when compared with that in the control group. Liver TC and TG concentrations were significantly decreased in the DHA group when compared with those in the control group. Based on the above, it is assumed that the physiological action exerted by n-3 PUFA differs by type and that DHA is a more effective n-3 PUFA, both for suppressing platelet aggregation and for modulating lipid metabolism in the plasma and liver of rats.

Histochemical observation and cellular distribution of ascorbic acid in persimmon leaves.

This study was carried out to resolve the discrepancy of data for the proportion of ascorbic acid and dehydroascorbic acid in persimmon leaves at the final stage of the season and to clarify their cellular distributions using histochemical and biochemical techniques. Fresh persimmon leaves were collected and used on July 31, September 5 and October 7, 1996. Ascorbic acid and dehydroascorbic acid in subcellular fractions were determined by the HPLC method that was found to be the most reliable for separation. The percent of dehydroascorbic acid in the total leaves was found to be almost constant (between 32 and 37%) in all preparations tested. In all preparations, more than 90% of the ascorbic acid and dehydroascorbic acid was found in the soluble fraction. The histochemical detection of ascorbic acid and an electron micrograph of persimmon leaf cells showed that the reactive color, after the reduction of silver nitrate under acidic conditions, in the leaves of all three preparations was mainly found on the face side of columned-type palisade parenchyma cells where chloroplasts were not rich and large vacuoles were seen. On the inner side of the palisade parenchyma cells where chloroplasts were the richest, only weak color development was observed. This study demonstrates that the percent of dehydroascorbic acid in persimmon leaves did not exceed 40% at least until October 7. It also shows that in persimmon leaf cells, ascorbic acid is mainly localized in the cytosol of palisade parenchyma tissue cells where large vacuoles are seen.

Effect of indigestible saccharides on B lymphocyte response of intestinal mucosa and cecal fermentation in rats.

The effects of water-soluble and -insoluble indigestible saccharides (IDS) on immune responses of the intestinal tract were studied. Male 4-week-old Sprague Dawley rats were fed for three weeks on diets containing several kinds of IDS at 5%. The results revealed that the proportion of kappa-light chain and IgA-presenting lymphocytes in small intestinal and cecal mucosa differed in increased number depending on the type of IDS. The response of colonic mucosa was not pronounced. The amounts of short-chain fatty acid (SCFA) and lactic acid in the cecal contents of the other test groups except the celfur group tended to be higher than those in the cellulose group, particularly in the lactulose group where many acids showed significant increases. The correlation between the proportion of kappa-light chain and IgA-presenting lymphocytes in the cecal mucosa and lactic acid in the cecal contents was significant, but that between the proportion of both lymphocytes and SCFA was not. Based on the above, we concluded that the oral administration of IDS induces the proliferation of kappa-light chain and IgA-producing B lymphocytes in small intestinal and cecal mucosa, but the degree of response differs depending on the type of IDS. It is thus suggested that IDS are involved in the intestinal immune system of rats.

Changes in blood coagulation, platelet aggregation, and lipid metabolism in rats given lipids containing docosahexaenoic acid.

In order to identify an adequate intake level of docosahexaenoic acid (DHA), changes in various parameters related to health benefits were studied in rats fed on diets containing 10% test lipids at different n-3(DHA)/n-6 ratios for two weeks. An evaluation of the critical level of the dietary n-3/n-6 ratio which had a significant effect on the parameters of several tissues indicated that the response to the dietary ratios differed according to the parameter, the variation in ratio ranging approximately from 0.20 to 1.77 with either a positive or negative effect on the health benefit. These results suggest that a suitable intake level of DHA would be within this range. In view of safety, however, the critical level for the dietary n-3/n-6 ratio may be around 0.56, as shown by a detailed analysis on the lower limit level of the harmful parameters. We thus propose that the dietary intake of DHA should not be more than 0.56 in terms of the n-3/n-6 ratio.

Accelerating effect of chitosan intake on urinary calcium excretion by rats.

The effect of chitosan on calcium (47Ca) metabolism was investigated in rats. The whole-body retention of 47Ca by rats fed on a 5% chitosan diet was significantly decreased when compared with that of rats fed on a cellulose diet, but showed no significant difference from that of rats fed on a fiber-free diet. Although there was no significant difference in the fecal excretion of 47Ca between the chitosan group and the cellulose or fiber-free group, the urinary excretion of 47Ca was significantly increased in the chitosan group when compared with the cellulose group. These results suggest that dietary chitosan would affect the calcium metabolism in animals.

Effect of dietary n-3/n-6 fatty acid ratio on the total count, fatty acid composition, and histamine and leukotriene concentrations of mast cells in tunica mucosa bronchiorum of type I allergic guinea pig.

To search for the most effective dietary n-3/n-6 ratio to suppress the type I allergic response, we performed basic experiments that applied parameters, associated with the type I allergy. Guinea pigs fed on diets containing lipids with the n-3/n-6 ratio at different levels and the polyunsaturated fatty acid/saturated fatty acid ratio of a fixed level were sensitized with ovalbumin and reared for two weeks. The lowest or critical level of the n-3/n-6 ratio which produced a significant difference in the parameters was as follows: about 2.0 for the response of mast cells and eosinophils; 0.5 and 1.0, respectively, for the uptake of n-3 and n-6 polyunsaturated fatty acids and decreased histamine production; and 0.2 for decreased leukotriene B4 and total leukotrienes 4, and increased leukotrienes 5/leukotrienes 4. The critical level of the n-3/n-6 ratio thus differed widely according to the parameter. Overall, the upper limit for the dietary n-3/n-6 ratio to suppress antigen-induced type I allergic responses is suggested to be around 1.0.

Effects of different types of dietary fiber preparations isolated from bamboo shoots, edible burdock, apple and corn on fecal steroid profiles of rats.

This study was conducted to evaluate the effects of different types of dietary fibers (DF) under the conditions with or without cholesterol (Chol) loading on the amount and composition of steroids in rat feces. Rats were fed Chol-unsupplied diets containing 10% lard and 5% DF preparation isolated from four kinds of food, bamboo shoots, edible burdock, apple and corn, for three weeks. The respective diets were supplemented with 0.5% Chol and then given to the rats for a further two weeks. The excretion of total bile acid (BA) and several major BAs increased significantly in the apple group with or without Chol loading when compared with that in the cellulose (CP) or other DF groups. The tendency in the apple group was more noticeable when the diet was supplemented with Chol. This is presumably a major reason for the tendency of decrease in serum and liver Chol concentrations in the apple group. The ratio of secondary BAs to total BA in the feces was significantly low in the apple group. Although the lithocholic acid (LCA)/deoxycholic acid (DCA) ratio, a risk index for colorectal cancer, was significantly lower in the bamboo, burdock and apple groups than in the CP or corn groups when given the diet without Chol, the differences disappeared with the addition of Chol. The proportion of coprostanol, a secondary metabolite of Chol, was smaller in the former three groups than in the CP or corn groups. These results suggest that the intake of some DF by host animals works beneficially for the microbial conversion of BA and Chol in the large intestine but that the addition of Chol acts to cancel such beneficial effects.

Effects of dietary n-3/n-6 and polyunsaturated fatty acid/saturated fatty acid ratios on platelet aggregation and lipid metabolism in rats.

We studied the effects of dietary lipids on platelet aggregation and lipid metabolism in rats by varying the n-3/n-6 ration while maintaining the polyunsaturated fatty acid/saturated fatty acid (P/S) ratio fixed, and vice versa. After two weeks, the platelet counts decreased as the dietary n-3/n-6 ratio rose, and platelet aggregation was sufficiently suppressed at the ratio of 0.2. Differences in the dietary P/S ratio, however, did not affect either the platelet counts nor platelet aggregation. As the dietary n-3/n-6 ratio rose, the proportion of arachidonic acid (AA) in the plasma and the phospholipids (PL) of the platelets and aorta decreased gradually, whereas the proportion of eicosapentaenoic acid (EPA) in each tissue increased gradually. The proportion of EPA was higher in the platelets than in the aorta, while that of docosahexaenoic acid (DHA) was higher in the latter. The production of platelet thromboxane A2 (TXA2) and aortic prostacyclin (PGI2) showed sharp declines, from the values for the n-3/n-6 ratio of 0.02 (control) to those for 0.5. These results suggest that the n-3/n-6 ratio of dietary fats necessary to ensure the suppression of platelet aggregation in normal rats would be at least 0.2 and no more than 0.5.

Effects of semi-purified dietary fibers isolated from Lagenaria siceraria, Raphanus sativus and Lentinus edodes on fecal steroid excretions in rats.

Rats were fed diets, each of which contained 5% of the semi-purified dietary fiber (DF) preparations from three kinds of foods, continuously for a total of 7 weeks. The fat level was varied, and cholesterol (Chol) was added toward the end of the experimental period. After 7 weeks, decreases of serum and liver Chol concentrations were observed in the Shiitake (Lentinus edodes) group when compared to the cellulose (CP) group. In the absence of Chol, fecal excretions of several secondary bile acids and total bile acids were affected by the type of DF, showing an increase particularly in the Shiitake group. Total neutral steroid excretions were decreased in the test DF groups compared to the CP group at both fat levels, and were increased by Chol supplementation in all of the test DF groups. It was demonstrated that the effects of the three DF preparations on fecal steroid excretions somewhat differed depending on the dietary factors. In the absence of Chol, the lithocholic acid (LCA)/deoxycholic acid (DCA) ratio was significantly lowered in the test DF groups compared to the CP group by the fat level elevation, and was affected by the interaction of fiber with fat. This ratio responded more intensely to the dietary factors than the two other indices (composition ratios of fecal bile acids). However, this response was lost in the presence of Chol. These results suggest that further studies are warranted to examine if the intense response of the LCA/DCA ratio to the tested DF preparations and fat in the present study will also be shown to other DF and what significance it has in evaluating the function of DF.

Water-soluble viscous substance of Jew's mellow leaves lowers serum and liver cholesterol concentrations and increases fecal steroid excretion in rats fed a high cholesterol diet.

The effect of Jew's mellow leaf powder and its water soluble viscous substance on cholesterol metabolism in rats fed a high cholesterol diet was examined. When compared to the controls, total serum and liver cholesterol concentrations were significantly decreased or tended to decrease in the groups given dry powder of fresh Jew's mellow leaves, dry powder purchased from the market or residual powder after extracting with ethanol, whereas no difference was observed in those given residual powder after extracting with water. There were significant increases or increasing tendencies in the fecal excretion of bile acids, total neutral sterols and cholesterol in those fed the experimental diets when compared to the control group. Rats fed a diet containing a water-soluble viscous substance (1.7%, about 1% as dietary fiber) obtained from the dry powder of Jew's mellow leaves showed significant decreases in serum and liver cholesterol concentrations and increases in fecal excretions of bile acids and neutral sterols. Based on the above, the component of dry powder of Jew's mellow leaves that is effective in decreasing serum and liver cholesterol concentrations was found to be a soluble dietary fiber, and the mechanism was assumed to be largely due to the increased excretion of bile acids and neutral sterols.

Effects of dietary casein and soy-protein on metabolic conversion of eicosapentaenoic acid to docosahexaenoic acid in the liver of rat.

The differential effects of dietary proteins on the metabolic process of eicosapentaenoic acid (EPA, 20:5n-3) via docosapentaenoic acid (DPA, 22:5n-3) to docosahexaenoic acid (DHA, 22:6n-3) were studied using parameters calculated from the proportion of n-3 polyenoic acid in liver phospholipid (PL) of rats. Rats were given casein or soy-protein isolate (SPI) diet containing 3% EPA with or without methionine (Met) supplementation for 3 weeks. The (22:5 + 22:6)/20:5 and 22:5/20:5 ratios of the PL fractions in the liver of rats given SPI were evidently elevated compared with those given casein. These ratios were also elevated when linoleic acid (LA, 18:2n-6) was added to the diet. There were no significant differences in the 22:6/22:5 ratio of the same PL fractions between the casein and SPI groups without the addition of LA, but the ratio was lowered by the addition of LA. It was found that these parameters for metabolic conversion were not affected by supplementation of Met or by a decrease in body weight. The above results suggest that the elongation step from EPA to DPA is affected by the type of protein, i.e., it is accelerated by SPI and is not affected by the presence of LA, while the metabolic process from DPA to DHA is not affected by the type of proteins without the addition of LA but is suppressed when LA is added.

Effects of casein and soy-protein on alpha-linolenic acid metabolism in rats.

In order to study the effects of different proteins on alpha-linolenic acid (alpha-LnA) metabolism, rats were given the diet added respectively with milk casein and soy-protein isolate (SPI) as sources of proteins and perilla oil as a source of lipid. The results obtained are as follows. The ratio of (C20:3 + C20:4)/C18:2 in liver microsomal PL, liver PE fraction, and kidney PE and PC fractions was significantly lowered by the SPI treatment when compared to the casein treatment, similarly to the already established results. In the liver microsomal PL and PE and PC fractions of liver and kidney in rats treated with SPI, there was also observed a significant decrease or a decrease tendency in the (C20:4 + C20:5)/C18:3 ratio. A similar tendency was again shown in the sigma (n-3)M/C18:3 ratio indicating metabolic conversion from C18:3(n-3) to C22:6. On the other hand, contrary to the ratios of (C20:3 + C20:4)/C18:2, sigma (n-3)M/C18:3, and (C20:3 + C20:5)/C18:3, the (C22:5 + C22:6)/C20:5 ratio which is the parameter for metabolic conversion of C20:5(n-3) was elevated in the PE and PC fraction of liver, heart and kidney in the SPI group compared to the casein group. Then, further analysis of the metabolic process from C20:5 to C22:6 showed that the C22:5/C20:5 ratio increased while the C22:6/C22:5 ratio decreased in the SPI group compared to the casein group. Based on these results, it is assumed that the metabolic process from C18:3(n-3) to C20:5(n-3) and from C22:5 to C22:6 is affected by SPI but that the elongation process from C20:5(n-3) to C22:5(n-3), on the contrary, is rather accelerated by SPI.

Fecal output, gastrointestinal transit time, frequency of evacuation and apparent excretion rate of dietary fiber in young men given diets containing different levels of dietary fiber.

Five healthy young male subjects were given meals containing different levels of dietary fiber (DF) for one week each, and the effects on the fecal output, gastrointestinal transit time, frequency of evacuation, and apparent excretion rate of dietary fiber in feces were studied. The experiments were conducted twice by varying the subjects and the DF levels. The following results were obtained. 1) With an increase in the intake of DF, feces increased significantly in both wet weight and volume. The fecal specific gravity did not show any clear-cut relation with the intake level of DF. 2) The gastrointestinal transit time tended to shorten in each DF period compared to the DF-free period. However, it did not shorten with an increase in the intake level of DF. 3) Frequency of evacuation tended to be high in any of the DF periods compared to the DF-free period. However, the differences in the intake level of DF produced no changes in the frequency. 4) The DF content in feces increased significantly with an increase in the intake of DF, indicating that the larger the fecal output, the larger the fecal DF content. The apparent mean excretion rate of DF in feces remained in the range of 60-75% throughout the two experimental phases. Residues in feces other than DF increased with an increase in the DF intake. 5) The results obtained here indicate that the DF intake of at least 20 g per day produces 140-150 g of fecal output which can be easily excreted daily by a normal adult person.

Effect of polychlorinated biphenyls on tissue distribution and intestinal absorption of alpha-tocopherol in rats.

We conducted experiments to elucidate the mechanism by which alpha-tocopherol (alpha-toc) concentration in the liver remarkably increases in rats treated with polychlorinated biphenyls (PCB) when excess amounts of DL-alpha-tocopheryl-acetate (alpha-toc-ac) were given. First, diets containing 5 and 50 mg of alpha-toc-ac per 100 g were respectively added with 0.05% of PCB and given to rats for 10 days, and alpha-toc distributions in tissues were determined. Next, the disappearance of alpha-toc from a ligated intestine of vitamin E (E)-deficient rats was then studied. There were no significant differences between the control and the PCB groups in alpha-toc concentrations in the liver and other tissues except a few tissues when a lower level of alpha-toc-ac (5 mg) was given. Conversely, when a higher level of alpha-toc-ac (50 mg) was given, alpha-toc concentration in the liver of the PCB group significantly increased compared to that of the control group. This agrees with the result previously reported, showing a remarkable reproducibility. In addition, alpha-toc concentrations in tissues other than the liver were also significantly higher in the PCB group than in the control group. Transmigration of alpha-toc to the liver from other tissues was therefore excluded. The alpha-toc absorption test using the ligated intestine of E-deficient rats revealed that alpha-toc disappears from the intestine at a higher rate in the presence of PCB compared to the untreated control group. This may indicate that alpha-toc absorption by passive diffusion is accelerated in the coexistence of PCB.

Effect of viscous indigestible polysaccharides on pancreatic-biliary secretion and digestive organs in rats.

Effects of viscous indigestible polysaccharides on the pancreas exocrine function were investigated in growing rats. Rats were fed a nonfiber diet or a diet containing approximately 5% of one of the following fibers: apple pectin, lambda-carrageenan, locust bean gum, gum xanthan, guar gum or sodium (Na) alginate. Pancreatic-bile secretion was found to be elevated in rats fed for 2 wk the highly viscous polysaccharides, sodium alginate, locust bean gum, gum xanthan and guar gum. The polysaccharides may have interfered with the digestion and absorption of nutrients, resulting in a decreased digestibility and an enlargement of digestive organs. When alginic acid and calcium alginate, insoluble polysaccharides that did not contribute to viscosity, were given to rats, they had no effect on pancreatic and biliary secretion compared with sodium alginate. The results demonstrate that consumption of viscous indigestible polysaccharides leads to changes in the exocrine pancreatic-biliary function and may depress the process of digestion and absorption. Rats may compensate for the inefficiency of digestion and absorption with a hyperplasia/hypertrophy of digestive organs and an increased secretion of digestive juice.