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Effect of dietary oligofructose and inulin on colonic preneoplastic aberrant crypt foci inhibition.

Oligofructose and inulin, naturally-occurring fermentable chicory fructans, have been shown to stimulate the growth of bifidobacteria which are regarded as beneficial strains in the colon and inhibit colon carcinogenesis in the laboratory animal models. The present study was designed to determine the effect of oligofructose and inulin on the azoxymethane (AOM)-induced preneoplastic lesions such as aberrant crypt foci (ACF) formation in the colon of male F344 rats. At 5 weeks of age, groups of animals were fed the AIN-76A (control) and the experimental diets containing 10% oligofructose or inulin. At 7 weeks of age, all animals received s.c. injection of AOM dissolved in normal saline at a dose rate of 15 mg/kg body wt, once weekly for 2 weeks. The animals were necropsied 7 weeks after the last AOM injection, and the ACF were visualized under light microscopy in the formalin-fixed, unsectioned methylene blue-stained colons. They were distinguished by their increased size, more prominent epithelial cells and pericryptal space. The feeding of oligofructose or inulin significantly inhibited the ACF formation and the crypt multiplicity in the colon. The degree of ACF inhibition was more pronounced in animals fed inulin than in those fed oligofructose. The findings suggest that chicory fructan supplements inhibit ACF formation, an early preneoplastic marker of malignant potential in the process of colon carcinogenesis.

Animals↗

Availability of cereal fructans and inulin in the rat intestinal tract.

The fate of cereal fructans and inulin in the gastrointestinal tract was investigated by using a rat model. Male Wistar rats received diets containing 5.0% of a preparation of cereal fructans or 4.7 or 9.4% inulin. Each diet was tested with or without Nebacitin, an antibiotic drug given to suppress the intestinal microflora. The availability of fructans in the upper gastrointestinal tract was calculated from the recovery in feces in Nebacitin-treated rats. Animals receiving cereal fructans developed diarrhea, which made it impossible to measure the bioavailability of this substrate. With inulin, the extent of digestion and absorption appeared to be 18-26%. This might, at least to some degree, be explained by hydrolysis by gastric juice in the stomach. Thus, in an in vitro assay, mild acid hydrolysis (0.05 M HCl, 37 degrees C, 120 min) converted about 8% of inulin to free fructose. The apparent digestibility of the fructans was calculated from its recovery in feces from untreated rats. Only minute amounts of fructans could be detected, hence, cereal fructans as well as inulin reaching the hindgut appeared to be almost completely fermentable.

Animals↗

Inulin and oligofructose: what are they?

Inulin is a term applied to a heterogeneous blend of fructose polymers found widely distributed in nature as plant storage carbohydrates. Oligofructose is a subgroup of inulin, consisting of polymers with a degree of polymerization (DP) </=10. Inulin and oligofructose are not digested in the upper gastrointestinal tract; therefore, they have a reduced caloric value. They stimulate the growth of intestinal bifidobacteria. They do not lead to a rise in serum glucose or stimulate insulin secretion. Several commercial grades of inulin are available that have a neutral, clean flavor and are used to improve the mouthfeel, stability and acceptability of low fat foods. Oligofructose has a sweet, pleasant flavor and is highly soluble. It can be used to fortify foods with fiber without contributing any deleterious organoleptic effects, to improve the flavor and sweetness of low calorie foods and to improve the texture of fat-reduced foods. Inulin and oligofructose possess several functional and nutritional properties, which may be used to formulate innovative healthy foods for today's consumer.

Colon↗

Presence of inulin and oligofructose in the diets of Americans.

The U.S. Department of Agriculture 1994-1996 Continuing Survey of Food Intakes by Individuals was used to estimate the intake of naturally occurring inulin and oligofructose by the U.S. population. Two nonconsecutive 24-h dietary recalls from >15,000 Americans of all ages were conducted, and a special database of inulin and oligofructose was developed specifically for the analyses. American diets provided on average 2.6 g of inulin and 2.5 g of oligofructose. Intakes varied by gender and age, ranging from 1.3 g for young children to 3.5 g for teenage boys and adult males. When standardized for amount of food consumed, the intakes showed little difference across gender and age. Significant differences in intake of these components were seen between categories within region of the country, season, income, and race and origin; however, the actual differences were relatively small. Major food sources of naturally occurring inulin and oligofructose in American diets were wheat, which provided about 70% of these components, and onions, which provided about 25% of these components. The estimation of the presence of inulin and oligofructose in the diets of Americans has not been published to date.

Adolescent↗

Inulin, oligofructose and intestinal function.

Inulin and oligofructose have attracted much attention recently as nonabsorbable carbohydrates with prebiotic properties. When inulin and oligofructose were added to a controlled diet, significant increases were noted in colonic bifidobacterial populations, and it has been proposed that these changes promote both colonic and systemic health through modification of the intestinal microflora. Inulin and oligofructose are rapidly and completely fermented by the colonic microflora with the production of acetate and other short-chain fatty acids. As with lactulose, they may also result in the growth of the fecal biomass, and in doing so, entrap ammonia for bacterial protein synthesis or conversion to the ammonium ion. As with dietary fiber and other nonabsorbable carbohydrates, there is also interest in inulin and oligofructose from the standpoint of inhibition of colonic carcinogenesis, blood cholesterol reduction, immune stimulation and enhanced vitamin synthesis. In these areas, the influence of their molecular weight is also an issue, with the longer chain length providing a more sustained fermentation pattern. More human studies are now required, including studies on the long-term effects of inulin and oligofructose consumption on colonic health, in particular on markers of cancer risk such as reduction in colonic polyp recurrence.

Animals↗

Effects of inulin on lipid parameters in humans.

Convincing lipid-lowering effects of the fructooligosaccharide inulin have been demonstrated in animals, yet attempts to reproduce similar effects in humans have generated conflicting results. This may be because of the much lower doses used in humans as a result of the adverse gastrointestinal symptoms exhibited by most subjects consuming daily doses in excess of 30 g. Two studies that fed either oligofructose (20 g/d) or inulin (14 g/d) observed no effect on fasting total, LDL or HDL cholesterol, or serum triglycerides. Two other studies that fed inulin either in a breakfast cereal (9 g/d) or as a powdered addition to beverages and meals (10 g/d) reported similar reductions in fasting triglycerides (-27 and -19%, respectively). In one of these studies, total and LDL cholesterol concentrations were also modestly reduced (5 and 7%, respectively). Because animal studies have identified inhibition of hepatic fatty acid synthesis as the major site of action for the triglyceride-lowering effects of inulin, and because this pathway is relatively inactive in humans unless a high carbohydrate diet is fed, future attempts to demonstrate lipid-lowering effects of inulin should consider the nature of the background diet as a determinant of response.

Animals↗

Database mining and transcriptional analysis of genes encoding inulin-modifying enzymes of Aspergillus niger.

As a soil fungus, Aspergillus niger can metabolize a wide variety of carbon sources, employing sets of enzymes able to degrade plant-derived polysaccharides. In this study the genome sequence of A. niger strain CBS 513.88 was surveyed, to analyse the gene/enzyme network involved in utilization of the plant storage polymer inulin, and of sucrose, the substrate for inulin synthesis in plants. In addition to three known activities, encoded by the genes suc1 (invertase activity; designated sucA), inuE (exo-inulinase activity) and inuA/inuB (endo-inulinase activity), two new putative invertase-like proteins were identified. These two putative proteins lack N-terminal signal sequences and therefore are expected to be intracellular enzymes. One of these two genes, designated sucB, is expressed at a low level, and its expression is up-regulated when A. niger is grown on sucrose- or inulin-containing media. Transcriptional analysis of the genes encoding the sucrose- (sucA) and inulin-hydrolysing enzymes (inuA and inuE) indicated that they are similarly regulated and all strongly induced on sucrose and inulin. Analysis of a DeltacreA mutant strain of A. niger revealed that expression of the extracellular inulinolytic enzymes is under control of the catabolite repressor CreA. Expression of the inulinolytic enzymes was not induced by fructose, not even in the DeltacreA background, indicating that fructose did not act as an inducer. Evidence is provided that sucrose, or a sucrose-derived intermediate, but not fructose, acts as an inducer for the expression of inulinolytic genes in A. niger.

Aspergillus niger↗

Dietary inulin alleviates hepatic steatosis and xenobiotics-induced liver injury in rats fed a high-fat and high-sucrose diet: association with the suppression of hepatic cytochrome P450 and hepatocyte nuclear factor 4alpha expression.

Inulin enzymatically synthesized from sucrose is a dietary component that completely escapes glucide digestion. Supplementing inulin to a high-fat and high-sucrose diet (HF) ameliorated hypertriglycemia and hepatic steatosis in 8-week-fed rats by suppressing elevated levels of serum triacylglycerols, fatty acids, and glucose, and the accumulation of hepatic triacylglycerols and fatty acids. Inulin intake prevented phenobarbital (PB)- and dexamethasone-induced liver injuries in the HF group. No significant alteration in the baseline expression of CYP2B, CYP2C11, CYP3A, and NADPH-cytochrome P450 (P450) reductase mRNAs and proteins was found. In contrast, baseline and PB-treated expressions of CYP2E1 mRNA were reduced in HF-fed rats. The induction of P450s in response to PB was affected by the nutritional status of the rats; mRNA levels of CYP2B1 and CYP3A1 after PB treatment, as assessed by quantitative real-time polymerase chain reaction analysis were reduced in the inulin-supplemented HF (HF+I) group, compared with those in the HF group. Western blot analysis detected the corresponding changes of CYP2B and CYP3A proteins. These alterations were correlated with changes in hepatic thiobarbituric acid-reactive substances. Furthermore, no significant difference in the expression of nuclear receptors constitutive androstane receptor, pregnane X receptor, and retinoid X receptor alpha and coactivator peroxisome proliferator-activated receptor-gamma coactivator 1alpha proteins was found in the hepatic nucleus between the HF and HF+I groups, but the expression of hepatocyte nuclear factor alpha (HNF4alpha) protein was significantly reduced in the HF+I group. Taken together, these results indicate that inulin intake ameliorates PB-induced liver injury, associated with a decline in lipid accumulation and PB-induced expression of CYP2B and CYP3A, which may be related by a reduction in the nuclear expression of HNF4alpha.

Alanine Transaminase↗

Effect on components of the intestinal microflora and plasma neuropeptide levels of feeding Lactobacillus delbrueckii, Bifidobacterium lactis, and inulin to adult and elderly rats.

The aim of this study was to compare the effects of the mixture of Lactobacillus delbrueckii subsp. rhamnosus strain GG, Bifidobacterium lactis Bb12, and inulin on intestinal populations of lactobacilli, bifidobacteria, and enterobacteria in adult and elderly rats fed the same (in quality and quantity) diet. The portal plasma levels of two neuropeptides, neuropeptide Y (NPY) and peptide YY (PYY), were also evaluated to assess the physiological consequences of the synbiotic treatment for the gastrointestinal (GI) tracts of rats of different ages. Adult (n = 24) and elderly (n = 24) male rats were fed the AIN-93 M maintenance diet. After 2 weeks of adaptation, the diet of 12 rats of each age group was supplemented with 8% inulin and with strains GG and Bb12 to provide 2.2 x 10(9) CFU of each strain g(-1) of the diet. Blood and different regions of the GI tract were sampled from all rats after 21 days of the treatment. Treatment with the mixture of strain GG, strain BB12, and inulin induced significantly different changes in the numbers of lactobacilli, bifidobacteria, and enterobacteria of the stomach, small intestine, cecum, and colon microflora. Moreover, the GG, BB12, and inulin mixture increased the concentrations of NPY and PYY for adult rats. For the elderly animals, the PYY concentration was not changed, while the NPY concentration was decreased by treatment with the GG, BB12, and inulin mixture. The results of the present study indicate that the physiological status of the GI tract, and not just diet, has a major role in the regulation of important groups of the GI bacteria community, since even the outcome of the dietary modification with synbiotics depends on the ages of the animals.

Animal Feed↗

Culture-independent microbial community analysis reveals that inulin in the diet primarily affects previously unknown bacteria in the mouse cecum.

Inulin is a well-known fructose-based prebiotic which has been shown to stimulate the growth of bifidobacteria, a bacterial group generally considered beneficial for intestinal health. In the present study, we analyzed inulin-associated shifts in the total bacterial community of wild-type mice and mice carrying a genetically inactivated adenomatous polyposis coli tumor suppressor gene by using DNA-based approaches independent of bacterial culturability. Mice were fed a high-fat, nonfiber diet with or without inulin inclusion at a 10% (wt/wt) concentration. Cecal contents were analyzed after 0, 3, and 9 weeks on the experimental diets. Inulin inclusion significantly affected the total bacterial community structure of the cecum as determined by both a nonselective percent-guanine-plus-cytosine-based profiling analysis and a more specific 16S ribosomal DNA sequence analysis. The shifts included stimulation of bifidobacteria and suppression of clostridia, but sequence comparison revealed that the major shifts were within previously unknown bacterial taxa. Concomitantly, significantly higher bacterial densities, determined by flow cytometry, were observed with the inulin-amended diet, and the metabolism of the cecal bacterial community was altered, as indicated by higher levels of residual short-chain fatty acids, particularly lactic acid. With regard to all of the microbiological parameters measured, the wild-type mice and mice carrying a genetically inactivated adenomatous polyposis coli tumor suppressor gene were essentially identical. Studies of the implications of pre- and probiotics may need to be expanded to include careful analysis of their effects on the entire microbial community, rather than just a few well-known species. Further studies are needed to increase our understanding of the possible roles of currently unknown gastrointestinal bacteria in health and disease.

Animal Feed↗

Tissue uptake of insulin and inulin in red and white skeletal muscle in vivo.

By employing a tissue uptake technique, the kinetics of the plasma-interstitial equilibration of radiolabeled insulin, inulin, and albumin were followed in four muscles of differing capillarity in anesthetized rats. The soleus muscle (SOL), and the red portion of the gastrocnemius muscle (RG), as well as the extensor digitorum longus muscle (EDL) and the white portion of the gastrocnemius muscle (WG) were investigated. After constant intravenous tracer infusions and repeated plasma sampling under euglycemic clamp conditions, animals were killed at varying time intervals and the muscles mentioned above were dissected out. The radioactivity of tracer per gram of tissue in each muscle divided by the plasma activity of tracer per milliliter of plasma, i.e., "the plasma equivalent space" of tracer, thus could be followed as a function of time. From this function the permeability-surface area (PS) of inulin as well as the distribution volumes at time 0 (V0) of inulin and insulin and their equilibrium distribution volumes (VE) were assessed. The PS for inulin (in ml.min-1.100 g muscle-1) was 0.52 +/- 0.10 (mean +/- SE) in WG, increasing with more red fibers to 1.37 +/- 0.18 in SOL. Also the inulin interstitial distribution volume (at blood-tissue tracer equilibrium; VE) increased in this order (in ml/100 g) from 7.30 +/- 0.91 in WG to 12.93 +/- 0.89 in SOL. The V0 for insulin was found to be approximately fivefold larger than the plasma volume in each muscle sample, indicating a high degree of binding of insulin to structures within the vascular compartment, conceivably to the vascular endothelium.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Variations in serum complement following inulin infusion in man.

Complement levels were determined in 12 normal human volunteers while receiving intravenous infusions of inulin for determination of standard inulin clearances. Significant decreases in beta IC globulin and total hemolytic complement activity were observed when inulin was infused, but not following infusion of saline as a control. These effects were noted only when steady-state inulin levels were greater than 26 mg%, suggesting a dose-dependent response. No evidence of increased coagulation was noted as measured by fibrinogen and fibrin degradation products. Since C4 levels remained largely unchanged following inulin infusion, it was concluded that the results most likely occurred through activation of the alternate complement pathway.

Adult↗

A new enzymatic method for the determination of inulin.

A new enzymatic method for the determination of inulin in plasma and urine, using inulase (EC 3.2.1.7), fructokinase (EC 2.7.1.4), phosphoglucoisomerase (EC 5.3.1.9) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) is described. The assay is based on the hydrolysis of inulin or Inutest (INutest which is the injectable form of inulin), by inulase and the determination of fructose released. The assay was linear up to 2 g/L of Inutest. The within-batch and between batch coefficients of variation were 2.3% and 2.2%, respectively. Recovery of added Inutest from plasma and urine was 98-102%. There was no interference from glucose (27.7 mmol/L), fructose (1.7 mmol/L) or mannose (1.7 mmol/L). When inulin clearance (using this method) and thiosulphate clearance were compared in 37 patients the inulin clearance was 9.3 mL/min (12%) lower than the thiosulphate clearance. We conclude that this enzymatic method is a simple and specific method.

Anthracenes↗

Glomerular filtration rate by single-injection inulin clearance: definition of a workable protocol for children.

The total serum clearance of inulin after a single intravenous injection is an established method for the determination of the glomerular filtration rate (GFR). To optimize this procedure in children, we analyzed 117 serum inulin time-decay curves in 59 children (age range 1.3-15.2 years)with renal disease. It was found that the two-compartment model fitted statistically better to the disappearance curves of serum inulin than the one-compartment model. The sampling frequency could be reduced from 12 to a minimum of seven data points (t=0, 10, 20, 30, 65, 120 and 240 min) in 4 h without a relevant change in the fit of the two-compartment model to the date or in the calculated clearance of serum inulin. We conclude that, by using a standardized 'seven-sample schedule', followed by a two-compartment analysis, the single-injection serum clearance of inulin is a suitable compromise between practical convenience and clinical accuracy for the determination of the GFR in children.

Adolescent↗

Inulin and oligofructose are part of the dietary fiber complex.

Dietary fiber has been defined as the remnants of plant cells resistant to hydrolysis by human alimentary enzymes. Its main chemical constituents are hemicelluloses, celluloses, lignin, pectins, gums, and waxes. The U.S. Food and Drug Administration and the U.S. Department of Agriculture determine compliance with nutritional labeling regulations for dietary fiber by use of the existing AOAC INTERNATIONAL methods for total dietary fiber. The above compounds are readily detected by these methods. However, some oligo- and polysaccharides are resistant to human alimentary enzymes and do not precipitate in 78% ethanol, the usual reagent for precipitating dietary fiber in analytical procedures. Some of these saccharides, termed fructans, are inulin and oligofructose. They possess many physiological attributes normally associated with dietary fiber. Inulin is a mixture of oligo- and polysaccharides composed of fructose moieties joined by beta(2-->1) linkages in linear chains. Almost each chain ends with a glucose moiety. Oligofructose is a synonym for fructo-oligosaccharides, with fructose moieties joined by beta(2-->1) linkages, as in inulin. Not all molecules have a glucose unit, and the chain length is less than 10 units. A method for inulin and oligofructose was developed and approved official first action by AOAC INTERNATIONAL in early 1997. It involves extraction of sample and treatment of the extract with amyloglucosidase followed by fructozyme (Fructozyme Enzyme Process Division, Novo Nordisk, Novo Industry, Copenhagen, Denmark). The sugars released in each of the 3 steps are measured by anion-exchange chromatography. The concentration of fructans is calculated as the difference of sugars, glucose and fructose, after the enzymatic treatments and the initial sample. The repeatability standard deviations for inulin and oligofructose ranged from 2.9 to 5.8% and the reproducibility standard deviations ranged from 4.7 to 11.1%. The method was accepted by AOAC INTERNATIONAL.

Dietary Fiber↗

Determination of inulin in foods.

A method was developed for determining fructan inulin in various foods (yogurts, honey cakes, chocolates). Warm water was applied for extraction of samples, and mono- and dissacharides were determined by a thin-layer chromatographic densitometric method. A portion of the test solution was hydrolyzed 30 min with 1% oxalic acid in a boiling water bath. Fructose was determined in the hydrolysate. The amount of inulin in a sample was calculated as the difference between the amount of fructose in the sample before and after hydrolysis. The fructose from sucrose formed during the hydrolysis was also considered. The mean recovery from yogurt fortified with 4% inulin was 95.5 +/- 4.5% (mean +/- standard deviation); from honey cakes extract fortified with 10% inulin, 97.3 +/- 5.5%; and from chocolate extract fortified with 30% inulin, 98.6 +/- 6.6% (6 replicates in all cases). Determination of glucose is not necessary for analyzing fructans with the composition expressed shortened to GFn-1 (G, glucose; F, fructosyl) with the average degree of polymerization 8 < or = n < or = 15.

Animals↗

Adjuvant effect of gamma-inulin is mediated by C3 fragments deposited on antigen-presenting cells.

The adjuvant effect of gamma-inulin, a strong activator of the alternative complement pathway, is well-known, but its exact mechanism is not revealed yet. Here, we show that macrophages, isolated from the peritoneal cavity of gamma-inulin-injected mice and used as antigen-presenting cells, enhance the proliferation of antigen-specific T-cells up to 2.5-fold when compared with macrophages of non-treated animals. This effect is abrogated by the presence of anti-C3 F(ab')2 fragments and by prior decomplementation of the donor animals with CVF. It is demonstrated that treatment of mice with the adjuvant results in deposition of C3-fragments onto the surface of peritoneal macrophages, as does in vitro incubation of the cells with gamma-inulin in the presence of fresh autologous serum. Prior incubation of macrophages with gamma-inulin plus serum in vitro enhances subsequent C3 production. Because it has been shown earlier that CR1/2 expressed on activated T-cells and interacting with covalently bound C3-fragments plays an important role in the augmentation of the adaptive response, our present results reveal a mechanism that contributes to the adjuvant effect of gamma-inulin and point to a further link between innate and adaptive immunity.

Animals↗

Indirect measurement of lymphatic absorption with inulin in continuous ambulatory peritoneal dialysis (CAPD) patients.

To elucidate the importance of possible trapping of macromolecules in peritoneal tissue on the calculation of peritoneal lymphatic drainage, we compared the transport of inulin administered i.v. and i.p. in nine continuous ambulatory peritoneal dialysis (CAPD) patients on two separate days. In the intraperitoneal study inulin (5 g) was added to the dialysate and in the intravenous study inulin (5 g) was given i.v. 3 h before the test. No differences were found in the mass transfer area coefficients (MTC) of urea, creatinine, and glucose between the two tests. The MTC after inulin i.p. was 3.2 +/- 0.7 mL/min (mean +/- SD) and after inulin i.v. 1.8 +/- 0.5 (p less than 10(-5]. As the difference in transport kinetics between i.v. and i.p. administration is likely to be caused by lymphatic absorption, a mean lymphatic flow of 1.4 mL/min could be calculated. This value corresponds to the data obtained with macromolecules. Our results therefore favor the hypothesis that no local accumulation of macromolecules in the peritoneal tissues takes place and that their disappearance from the peritoneal cavity represents lymphatic absorption.

Biological Transport↗