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PubMed1987

A simple method for determining extracellular polysaccharide-producing ability of oral streptococci.

A simple method is described for determining the types of extracellular polysaccharides produced by oral streptococci. The method yields quantitative results in which the amount of polysaccharides produced is normalised with respect to DNA. Results correlate well with previously published data for oral streptococci, except in the case of Streptococcus oralis for which there is no previously published data. All species within the mutans group produced both soluble and insoluble glucan; while soluble fructan was only produced by strains of Streptococcus rattus and Streptococcus mutans. The major extracellular polysaccharide produced by Streptococcus salivarius was soluble fructan, although some strains produced soluble and insoluble glucan. Most Streptococcus sanguis strains produced soluble and insoluble glucan. Soluble fructan was produced by some strains of S. sanguis and S. oralis. The latter also produced soluble and insoluble glucan. No detectable extracellular polysaccharide was elaborated by Streptococcus milleri, Streptococcus faecalis or Streptococcus lactis.

Bacteriological Techniques↗

PubMed2002

Transport of 1-kestose across the tonoplast of Jerusalem artichoke tubers.

The capacity for 1-kestose uptake into the vacuole of fructan storing Jerusalem artichoke tubers was investigated. 1-kestose serves both as building block for fructan initiation and as a fructose donor for chain elongation. Tonoplast vesicles were isolated from actively storing tubers, and their vesicles were capable of transporting sucrose in a manner indicative of a sucrose/H(+) antiport. Under similar conditions, 1-kestose was not taken up by vesicles energized by either a pH jump or in the presence of ATP. When added together at 2 mM, sucrose uptake was not affected by the presence of 1-kestose. The data argues against the possible synthesis of 1-kestose in the cytosol and subsequent transport to the vacuole. The data also presents definite evidence for the existence a mechanism for sucrose accumulation in fructan storing vacuoles.

Biological Transport↗

PubMed2002

Cloning of a vacuolar invertase from Belgian endive leaves (Cichorium intybus).

Although a lot of vacuolar invertase (EC 3.2.1.26) cDNAs are available from a diversity of plant species, up to now no sequence information is available on invertases from any dicot fructan-containing species. Therefore, we describe the cloning of vacuolar acid invertase cDNA from etiolated Belgian endive leaves (Cichorium intybus L. var. foliosum cv. Flash), formed throughout the forcing process of the witloof chicory roots. Full-length cDNA was obtained by a combination of RT-PCR, PCR and 5'- and 3' RACE RT-PCR, starting with primers based on conserved amino acid sequences. The cloned chicory acid invertase groups together with vacuolar type invertases and fructan biosynthetic enzymes. A putative role for vacuolar type invertases in fructan synthesizing plants is discussed.

Journal Article↗

PubMed1995

Isolation and enzymic properties of levansucrase secreted by Acetobacter diazotrophicus SRT4, a bacterium associated with sugar cane.

Acetobacter diazotrophicus, a nitrogen-fixing bacterium associated with sugar cane, secretes a levansucrase (sucrose-2,6-beta-D-fructan 6-beta-D-fructosyltransferase; EC 2.4.1.10). This enzyme is constitutively expressed and represents more than 70% of the total proteins secreted by strain SRT4. The purified protein consists of a single 58 kDa polypeptide with an isoelectric point of 5.5. Its activity is optimal at pH 5.0. It catalyses transfructosylation from sucrose to a variety of acceptors including water (sucrose hydrolysis), glucose (exchange reaction), fructan (polymerase reaction) and sucrose (oligofructoside synthesis). In vivo the polymerase activity leads to synthesis of a high-molecular-mass fructan of the levan type. A. diazotrophicus levansucrase catalyses transfructosylation via a Ping Pong mechanism involving the formation of a transient fructosyl-enzyme intermediate. The catalytic mechanism is very similar to that of Bacillus subtilis levansucrase. The kinetic parameters of the two enzymes are of the same order of magnitude. The main difference between the two enzyme specificities is the high yield of oligofructoside, particularly 1-kestotriose and kestotetraose, accumulated by A. diazotrophicus levansucrase during sucrose transformation. We discuss the hypothesis that these catalytic features may serve the different biological functions of each enzyme.

Acetobacter↗

PubMed2004

Cold acclimation and abscisic acid induced alterations in carbohydrate content in calli of wheat genotypes differing in frost tolerance.

The effect of cold and abscisic acid (ABA) treatment on soluble carbohydrate content was compared in callus cultures of wheat genotypes differing in frost tolerance. The effect of 5A chromosome substituted from the frost tolerant < > to the sensitive < > on cold-induced carbohydrate accumulation was also determined. Following cold hardening, the increase in sucrose and fructan level in calli of tolerant varieties was significantly higher than those of the sensitive ones. In 5A substitution line higher sucrose and fructan content was detected than in recipient < >. Tendentiously, cold stress caused higher degree of changes in carbohydrate content than the exogenously applied ABA did. Comparing the accumulation pattern of the components of WSC measured in vitro to the previously published in vivo results it can be concluded that in the case of sucrose and fructans it was similar, while for the reducing sugars it was different. The regulatory role of chromosome 5A either in the development of freezing tolerance or carbohydrate accumulation was confirmed in dedifferentiated calli, as well.

Abscisic Acid↗

PubMed2005

Impact of inulin and oligofructose on gastrointestinal peptides.

In the present paper, we summarise the data supporting the following hypothesis: dietary inulin-type fructans extracted from chicory root may modulate the production of peptides, such as incretins, by endocrine cells present in the intestinal mucosa, this phenomenon being involved in the regulation of food intake and/or systemic effects. To test this hypothesis, male Wistar rats received for 3 weeks either a standard diet or the same diet supplemented with 10 % inulin-type fructans with different degrees of polymerisation. All the effects were most pronounced with the diet containing oligofructose, and consisted of (i) a decrease in mean daily energy intake and in epididymal fat mass; (ii) a higher caecal pool of the anorexigenic glucagon-like peptide-1 (7-36) amide (GLP-1), and peptide YY (PYY), due to caecal tissue proliferation; (iii) an increase in GLP-1 and of its precursor - proglucagon mRNA - concentrations in the proximal colon; (iv) an increase in portal serum level of GLP-1 and PYY; (v) a decrease in serum orexigenic peptide ghrelin. Moreover, oligofructose supplementation improved glucose homeostasis (i.e. decreased glycaemia, increased pancreatic and serum insulin content) in diabetic rats previously treated with streptozotocin, a phenomenon that is partly linked to the reduction in food intake and that correlates with the increase in colic and portal GLP-1 content. Based on these results it appears justified to test, in human subjects, the hypothesis that dietary inulin-type fructans could play a role in the management of obesity and diabetes through their capacity to promote secretion of endogenous gastrointestinal peptides involved in appetite regulation.

Animals↗

PubMed1999

Fiber, inulin and oligofructose: similarities and differences.

The biological, chemical and physical properties of dietary fibers are associated with physiologic actions in the small and large intestine that have important metabolic implications for health. These properties of fiber include dispersibility in water, bulk, viscosity, adsorption and binding of compounds and fermentability. Dietary fructans share some of the properties of dietary fiber and thus are likely to have similar metabolic effects. Within the small intestine, properties such as dispersibility in water, bulking and viscosity are associated with slowing the digestion and absorption of carbohydrate and lipid and promoting nutrient absorption along a greater length of the small intestine. Both of these actions are related to cholesterol reduction and blunting of alimentary gylcemia. Although fructans are dispersible in water and will provide some bulk because they are nondigestible in the small intestine, they do not appear to be associated with significant increases in viscosity. Thus one would predict that any immediate effects on alimentary glycemia or on cholesterol reduction are likely to be modest compared with more viscous polysaccharides. Fermentability and bulking capacity of nondigestible carbohydrates define an essential role of fiber in maintaining gastrointestinal health. Within the large intestine, carbohydrates that are not digested in the small intestine are available for fermentation by the microflora present. Carbohydrates that are dispersible in the aqueous phase are more readily digested by microbes. A large body of evidence indicates that dietary fructans are digested in the large intestine, resulting in an increase in microbial mass and production of short-chain fatty acids.

Animals↗

PubMed2000

Fructo-oligosaccharide malabsorption: benefit for gastrointestinal functions.

Fructo-oligosaccharides or inulin-type fructans are mixtures of oligomers that are composed primarily of beta-D-fructose monomers linked by beta2-1 osidic bonds. They are natural constituents of many edible plants. They resist digestion and are not absorbed in the upper part of the gastrointestinal tract, but they are fermented in the colon. That fermentation produces short-chain fatty acids, which acidify the colonic content; increase bacterial biomass and, consequently, fecal mass; and modify the composition of the microflora, especially by stimulating the growth of bifidobacteria. Fructo-oligosaccharides are thus prebiotics and a method of the Association of Official Analytical Chemists exists to measure them in food products. Because of this behavior in the gastrointestinal tract, inulin-type fructans have a low caloric value compared with carbohydrate absorbed in the small intestine, they improve Ca absorption and Ca balance, and, at least in experimental models, they inhibit the development of chemically induced aberrant crypt foci. Moreover, at a systemic level, they may contribute to modulating lipogenesis and reducing triglyceridemia. The data reviewed in this article support the classification of inulin-type fructans as functional food ingredients for which claims will or should become authorized when data for humans become available.

Journal Article↗

PubMed2007

Review article: fructose malabsorption and the bigger picture.

Fructose is found widely in the diet as a free hexose, as the disaccharide, sucrose and in a polymerized form (fructans). Free fructose has limited absorption in the small intestine, with up to one half of the population unable to completely absorb a load of 25 g. Average daily intake of fructose varies from 11 to 54 g around the world. Fructans are not hydrolysed or absorbed in the small intestine. The physiological consequences of their malabsorption include increasing osmotic load, providing substrate for rapid bacterial fermentation, changing gastrointestinal motility, promoting mucosal biofilm and altering the profile of bacteria. These effects are additive with other short-chain poorly absorbed carbohydrates such as sorbitol. The clinical significance of these events depends upon the response of the bowel to such changes; they have a higher chance of inducing symptoms in patients with functional gut disorders than asymptomatic subjects. Restricting dietary intake of free fructose and/or fructans may have durable symptomatic benefits in a high proportion of patients with functional gut disorders, but high quality evidence is lacking. It is proposed that confusion over the clinical relevance of fructose malabsorption may be reduced by regarding it not as an abnormality but as a physiological process offering an opportunity to improve functional gastrointestinal symptoms by dietary change.

Adolescent↗

PubMed2005

Fermentation of fructooligosaccharides and inulin by bifidobacteria: a comparative study of pure and fecal cultures.

The utilization of fructooligosaccharides (FOS) and inulin by 55 Bifidobacterium strains was investigated. Whereas FOS were fermented by most strains, only eight grew when inulin was used as the carbon source. Residual carbohydrates were analyzed by high-performance anion-exchange chromatography with pulsed amperometric detection after batch fermentation. A strain-dependent capability to degrade fructans of different lengths was observed. During batch fermentation on inulin, the short fructans disappeared first, and then the longer ones were gradually consumed. However, growth occurred through a single uninterrupted exponential phase without exhibiting polyauxic behavior in relation to the chain length. Cellular beta-fructofuranosidases were found in all of the 21 Bifidobacterium strains tested. Four strains were tested for extracellular hydrolytic activity against fructans, and only the two strains which ferment inulin showed this activity. Batch cultures inoculated with human fecal slurries confirmed the bifidogenic effect of both FOS and inulin and indicated that other intestinal microbial groups also grow on these carbon sources. We observed that bifidobacteria grew by cross-feeding on mono- and oligosaccharides produced by primary inulin intestinal degraders, as evidenced by the high hydrolytic activity of fecal supernatants. FOS and inulin greatly affected the production of short-chain fatty acids in fecal cultures; butyrate was the major fermentation product on inulin, whereas mostly acetate and lactate were produced on FOS.

Bifidobacterium↗

PubMed1989

Biochemical characterization and evaluation of virulence of a fructosyltransferase-deficient mutant of Streptococcus mutans V403.

The Streptococcus mutans extracellular fructosyltransferase (FTF) enzyme may play a role in the formation of dental caries by synthesizing a fructan polymer that serves as an extracellular storage polysaccharide. We sought to determine if an FTF-deficient strain of S. mutans was less virulent than wild-type cells in a rat animal model system. Cloned ftf gene sequences from S. mutans GS5 were used to generate a defective copy of the ftf gene by inserting into the ftf coding region a DNA fragment which encoded erythromycin resistance. The plasmid which carried the defective ftf construct was introduced into S. mutans V403 by using genetic transformation. This defective construct replaced, by allelic exchange, the wild-type copy of the ftf gene carried on the V403 chromosome. FTF activity assays indicated that the recombinant strain, V1741, was deficient in fructan synthesis. However, extracellular protein preparations from this strain displayed an increased ability to generate glucose polymers (glucans) compared with V403 preparations. Levels of adherence to glass and rat tooth surfaces by strain V1741 were similar to those of the V403 strain. Both strains caused moderate decay on rat tooth surfaces; however, the FTF-deficient strain was less pathogenic compared with the wild-type strain. These results suggest that FTF activity contributes to the pathogenicity of S. mutans V403, possibly by generating extracellular fructans which serve as storage compounds.

Animals↗

PubMed1992

Fructose and related food carbohydrates. Sources, intake, absorption, and clinical implications.

It is possible to point out subjects consuming considerable quantities of fructose and sorbitol, and the intake seems to be increasing both from added and natural sources. Studies of the absorption of fructose in animals are inconsistent, and the mechanisms of fructose uptake seem to vary in accordance with the species. In most species fructose absorption takes place by a specific carrier (facilitated transport), but it may be active in the rat. In vitro studies of human intestine are very scarce; there is no evidence of active intestinal fructose transport in the human intestine. By means of hydrogen breath tests, a very low absorption capacity for fructose given as the free monosaccharide has been found in humans. Fructose given as sucrose or in equimolar combinations with glucose is well absorbed, and only fructose in excess of glucose is malabsorbed. On this basis it is hypothesized that two different uptake mechanisms for fructose are present in the human intestine. One of these may be a disaccharidase-related uptake system. Sorbitol ingestion may aggravate malabsorption of fructose given as the monosaccharide; it is not known whether a specific mechanism is involved. In children and adults with functional bowel distress the absorption capacities for fructose may not differ from those of healthy individuals, but malabsorption of fructose and/or sorbitol may be the cause of or aggravate abdominal symptoms. Fructose polymers (fructans) are also subject to increasing nutritional interest. Fructans are not absorbed in the small intestine but are strongly fermented in the large bowel. Fructans may be of potential benefit for large-bowel function and blood glucose regulation.

Breath Tests↗

PubMed2006

RNA interference (RNAI) as a tool to engineer high nutritional value in chicory (Chicorium intybus).

The major component of chicory (Chicorium intybus) root is inulin, which is a polymer of fructose. Inulin production from chicory is hampered by the enzyme fructan 1-exohydrolase (1-FEH) that degrades inulin and limits its yield. Increased FEH activity results in massive breakdown of fructan and production of Fructose and inulo-n-oses. The latter phenomena are to be avoided for industrial fructan production. RNA silencing, which is termed post-transcriptional gene silencing (PTGS) in plants, is an RNA degradation process through sequence specific nucleotide interactions induced by double-stranded RNA. For genetic improvement of crop plants, RNAi has advantages over antisense-mediated gene silencing and co-suppression, in terms of its efficiency and stability. We are generating a transgenic chicory plants with suppressed FEH (exohydrolas) genes using RNAi resulting in supressed inulin degradation. A small but important part of the construct is a sequence unique for the target gene (exons) or genes,which were cloned. The hairpin constructs were made and chicory was transformed by Agrobacterium tumifaciense, strain (C58C1). The transgenics should be select and check by means of molecular techniques.

Carbohydrate Metabolism↗

PubMed1998

Prebiotics and synbiotics: concepts and nutritional properties.

The main role of diet is to provide enough nutrients to meet the requirements of a balanced diet, while giving the consumer a feeling of satisfaction and well-being. The most recent knowledge in bioscience supports the hypothesis that diet also controls and modulates various functions in the body, and, in doing so, contributes to the state of good health necessary to reduce the risk of some diseases. It is such an hypothesis which is at the origin both of the concept of 'functional food' and the development of a new scientific discipline of 'functional food science'. In the context of this paper the potential 'functional foods' to be discussed are the prebiotics and the synbiotics. The prebiotics developed so far are the non-digestible oligosaccharides and especially the non-digestible fructans among which chicory fructans play a major role. The chicory fructans are beta (2-1) fructo-oligosaccharides classified as natural food ingredients. They positively affect various physiological functions in such a way that they are already or may, in the future, be classified as functional food ingredients for which claims of functional effects or of disease risk reduction might become authorized. They are classified as prebiotic and have been shown to induce an increase in the number of bifidobacteria in human faecal flora. As part of a synbiotic-type product, they are already bifidogenic at a dose of 2.75 g/d and the effect lasts for at least 7 weeks. The other potential functional effects are on the bioavailability of minerals, but also, and more systemically, on the metabolism of lipids. Potential health benefits may concern reduction of the risk of intestinal infectious diseases, cardiovascular disease, non-insulin-dependent diabetes, obesity, osteoporosis and cancer. However, except for the prebiotic effect, and tentatively the improvement of calcium bioavailability, the evidence to support such effects is still missing in humans though hypotheses already exist to justify nutrition studies.

Bifidobacterium↗

PubMed1990

Production, thermal stability and immobilisation of inulinase from Fusarium oxysporum.

Fusarium oxysporum produced maximum extracellular inulinase after 9 days of its growth at 25 degrees C on a medium (pH 5.5) containing 3% fructan and 0.2% sodium nitrate. The level of this enzyme decreased on the addition of either glucose, fructose, galactose or sucrose to F. oxysporum already growing on a fructan-containing medium. A significant increase in invertase production which resulted in an increase of the invertase/inulinase (S/I) ratio, was observed on addition of inulin to this fungus growing on other carbon sources. Glycerol (10%) gave better protection to inulinase against thermal denaturation at 50 degrees C compared to ethylene glycol and sorbitol. Inulinase immobilised in polyacrylamide gel retained 45% of its original activity. The immobilised enzyme showed a higher optimum temperature (45 degrees C) compared to free enzyme (37 degrees C). The immobilised enzyme after storage at 25 degrees C for 96 h showed 58% activity. Thermal stability of entrapped inulinase increased in the presence of inulin.

Alcohols↗

PubMed2005

Carbohydrate partitioning between upper and lower regions of the crown in oat and rye during cold acclimation and freezing.

Carbohydrates have long been recognized as an important aspect of freezing tolerance in plants but the association between these two factors is often ambiguous. To help clarify the relationship, the allocation of carbohydrates between specific tissues within the over wintering organ (crown) of winter cereals was measured. A winter-hardy and non-winter-hardy oat (Avena sativa L.), and a rye (Secale cereale L.) cultivar were grown and frozen under controlled conditions. Crown tissue was fractionated into an upper portion, called the apical region, and a lower portion, called the lower crown. These tissues were ground in liquid N and extracted with water. Extracts were analyzed by HPLC for the simple sugars, sucrose, glucose, fructose, and for fructan of various size classes. After 3 weeks of cold acclimation at 3 degrees C, carbohydrates accounted for approximately 40% of the dry weight of oats and 60% of the dry weight of rye. The apical region, which is the tissue within the crown that acclimates to the greatest extent, was generally 10% higher in total carbohydrates than the lower crown. During a mild freeze, various carbohydrates were allocated differently between specific tissues in the three genotypes. When frozen, fructan generally decreased to a greater extent in the lower crown than in the apical region but sugars increased more in the apical region than in the lower crown. Results suggest that to understand how carbohydrates relate to freezing tolerance, regions of the crown that endure freezing stress differently should be compared.

Acclimatization↗

PubMed2005

Cranberry high molecular weight constituents promote Streptococcus sobrinus desorption from artificial biofilm.

Dental biofilm harbouring oral bacteria is highly correlated with the progression of dental diseases. Disruption of biofilm formation via anti-adhesion agents is an alternative means to the antibacterial approach. Previous studies have shown that high molecular weight non-dialysable material (NDM) derived from cranberry juice inhibits the adhesion of Escherichia coli and the coaggregation of a variety of oral bacteria. In addition, it inhibits the formation of glucans and fructans synthesised by GTF and FTF. In the present study, we examined the anti-adhesion effect of NDM on S. sobrinus. NDM promoted desorption of S. sobrinus from biofilm in the presence and absence of extracellular glucans and fructans, although the effect was more pronounced in the absence of these polysaccharides. Precoating of the bacteria with NDM reduced their ability to form biofilm. Our results indicate that NDM could be exploited as an anti-biofilm agent.

Bacterial Adhesion↗

PubMed2005

Effect of membrane lateral pressure on the expression of fructosyltransferases in Lactobacillus reuteri.

The effect of environmental conditions on the production of homo-polysaccharides and oligosaccharides from sucrose and the regulation of glycosyltransferase genes responsible for biosynthesis of homo-polysaccharides was determined in Lactobacillus reuteri TMW1.106 (reutericyclin-producer) and LTH5448 (reutericyclin-negative). Strain L. reuteri TMW 1.106 harbours the glycosyltransferase genes gtfA and inu, strain LTH5448 harbours a fructosyltransferase, ftfA. Fructan and fructose-oligosaccharide (FOS) production in both strains was inducible by reutericyclin, trans-isohumulone, and nigericin at the levels of their minimum inhibitory concentrations (MIC) as well as phenylethanol (6mM) and elevated growth temperatures (45 degrees C), but not by nisin, CCCP or gramicidin. Elevated temperature (45 degrees C), reutericyclin or trans-isohumulone but not CCCP furthermore increased enhanced inu and ftfA transcription in L. reuteri TMW1.106 and LTH5448, respectively. Generally, effects of the various agents on fructosyltransferase transcription corresponded to their effect on formation of poly and oligosaccharides from sucrose. The effect of membrane-active agents on fructosyltransferase expression was compared to their effect on membrane biophysical parameters. The ability of chemical and physical agents to induce expression of fructosyltransferases correlated to their effect on the membrane lateral pressure as measured by pyrene-labelled phospholipids in membrane vesicles. Dextran, levan and fructose-oligosaccharides added at 50gL(-1) protected L. reuteri towards the membrane-active inhibitors nisin, reutericyclin, and CCCP. The induction of glycosyltransferases by membrane stress indicates a protective role of fructans and FOS to lactobacilli exposed to physical and chemical environmental stressors.

Cyclopentanes↗