Microbial utilization and selectivity of pectin fractions with various structures

To evaluate the fermentation properties of oligosaccharides derived from pectins and their parent polysaccharides, a 5-ml-working-volume, pH- and temperature-controlled fermentor was tested. Six pectic oligosaccharides representing specific substructures found within pectins were prepared. These consisted of oligogalacturonides (average degrees of polymerization [DP] of 5 and 9), methylated oligogalacturonides (average DP of 5), oligorhamnogalacturonides (average DP of 10 as a disaccharide unit of galacturonic acid and rhamnose), oligogalactosides (average DP of 5), and oligoarabinosides (average DP of 6). The influence of these carbohydrates on the human fecal microbiota was evaluated. Use of neutral sugar fractions resulted in an increase in Bifidobacterium populations and gave higher organic acid yields. The Bacteroides-Prevotella group significantly increased on all oligosaccharides except oligogalacturonides with an average DP of 5. The most selective substrates for bifidobacteria were arabinan, galactan, oligoarabinosides, and oligogalactosides.

Effect of dextransucrase cellobiose acceptor products on the growth of human gut bacteria

The selective fermentation by human gut bacteria of gluco-oligosaccharides obtained from the reaction between the glucosyl group of sucrose and cellobiose, catalyzed by dextransucrases (DSR) from Leuconostoc mesenteroides, has been evaluated. Oligosaccharides were fractionated according to their molecular weight, and their effect on the growth of different bacterial groups was studied. To determine the structure (position and configuration of glycosidic linkages)�function relationship, their properties were compared to those of DSR maltose acceptor products (DSRMal) and of recognized prebiotic carbohydrates (fructo-oligosaccharides, FOS). Cellobiose acceptor products (DSRCel) showed bifidogenic properties similar to those of FOS. However, no significant differences related to molecular weight or isomeric configurations were found for DSRCel and DSRMal products.

Oligosaccharides of pitaya (dragon fruit) flesh and their prebiotic properties

The major carbohydrates of white and red-flesh pitayas (dragon fruit) were glucose, fructose and some oligosaccharides (total concentrations of 86.2 and 89.6 g/kg, respectively). The molecular weight distribution of the extract was affected by the extraction solvent. The maximum oligosaccharides content (27.40%), which included fractions with molecular weights of 273–275, 448–500 and 787–911 Da, were obtained using 80% ethanol extraction at room temperature (28 ± 2 °C). The low molecular weight fraction, including glucose and fructose, was successfully removed by yeast cultivation. The molecular weights of mixed oligosaccharides (716, 700, 490 and 474 Da) were confirmed by mass spectrometry. The mixed oligosaccharides showed that they were resistant to hydrolysis by artificial human gastric juice and human α-amylase, giving maximum hydrolysis of 4.04% and 34.88%, respectively. The mixed oligosaccharides were also found capable of stimulating the growth of lactobacilli and bifidobacteria.

Identification of metabolites in human hepatic bile using 800 MHz 1H NMR spectroscopy, HPLC-NMR/MS and UPLC-MS

The first application of high field NMR spectroscopy (800 MHz for 1H observation) to human hepatic bile (as opposed to gall bladder bile) is reported. The bile sample used for detailed investigation was from a donor liver with mild fat infiltration, collected during organ retrieval prior to transplantation. In addition, to focus on the detection of bile acids in particular, a bile extract was analysed by 800 MHz 1H NMR spectroscopy, HPLC-NMR/MS and UPLC-MS. In the whole bile sample, 40 compounds have been assigned with the aid of two-dimensional 1H–1H TOCSY and 1H–13C HSQC spectra. These include phosphatidylcholine, 14 amino acids, 10 organic acids, 4 carbohydrates and polyols (glucose, glucuronate, glycerol and myo-inositol), choline, phosphocholine, betaine, trimethylamine-N-oxide and other small molecules. An initial NMR-based assessment of the concentration range of some key metabolites has been made. Some observed chemical shifts differ from expected database values, probably due to a difference in bulk diamagnetic susceptibility. The NMR spectra of the whole extract gave identification of the major bile acids (cholic, deoxycholic and chenodeoxycholic), but the glycine and taurine conjugates of a given bile acid could not be distinguished. However, this was achieved by HPLC-NMR/MS, which enabled the separation and identification of ten conjugated bile acids with relative abundances varying from approximately 0.1% (taurolithocholic acid) to 34.0% (glycocholic acid), of which, only the five most abundant acids could be detected by NMR, including the isomers glycodeoxycholic acid and glycochenodeoxycholic acid, which are difficult to distinguish by conventional LC-MS analysis. In a separate experiment, the use of UPLC-MS allowed the detection and identification of 13 bile acids. This work has shown the complementary potential of NMR spectroscopy, MS and hyphenated NMR/MS for elucidating the complex metabolic profile of human hepatic bile. This will be useful baseline information in ongoing studies of liver excretory function and organ transplantation.

In vitro fermentation of lactulose-derived oligosaccharides by mixed fecal microbiota

Fermentation properties of oligosaccharides derived from lactulose (OsLu) and lactose (GOS) have been assessed in pH-controlled anaerobic batch cultures using lactulose and Vivinal-GOS as reference carbohydrates. Changes in gut bacterial populations and their metabolic activities were monitored over 24 h by fluorescent in situ hybridization (FISH) and by measurement of short-chain fatty acid (SCFA) production. Lactulose-derived oligosaccharides were selectively fermented by Bifidobacterium and lactic acid bacterial populations producing higher SCFA concentrations compared to GOS. The highest total SCFA production was from Vivinal-GOS > lactulose > OsLu > GOS. Longer incubation periods produced a selective fermentation of OsLu when they were used as a carbon source reaching the highest selective index scores. The new oligosaccharides may constitute a good alternative to lactulose, and they could belong to a new generation of prebiotics to be used as a functional ingredient for improving the composition of gut microflora.

Antigenotoxicity of probiotics and prebiotics on faecal water-induced DNA damage in human colon adenocarcinoma cells

Six strains of lactic acid producing bacteria (LAB) were incubated (1 x 10(8)cfu/ml) with genotoxic faecal water from a human subject. HT29 human adenocarcinoma cells were then challenged with the resultant samples and DNA damage measured using the single cell gel electrophoresis (comet) assay. The LAB strains investigated were Bifidobacterium sp. 420, Bifidobacterium Bb12, Lactobacillus plantarum, Streptococcus thermophilus, Lactobacillus bulgaricus and Enterococcus faecium. DNA damage was significantly decreased by all bacteria used with the exception of Strep. thermophilus. Bif. Bb12 and Lact. plantarum showed the greatest protective effect against DNA damage. Incubation of faecal water with different concentrations of Bif. Bb12 and Lact. plantarum revealed that the decrease in genotoxicity was related to cell density. Non-viable (heat treated) probiotic cells had no effect on faecal water genotoxicity. In a second study, HT29 cells were cultured in the presence of supernatants of incubations of probiotics with various carbohydrates including known prebiotics; the HT29 cells were then exposed to faecal water. Overall, incubations involving Lact. plantarum with the fructooligosaccharide (FOS)-based prebiotics Inulin, Raftiline, Raftilose and Actilight were the most effective in increasing the cellular resistance to faecal water genotoxicity, whereas fermentations with Elixor (a galactooligosaccharide) and Fibersol (a maltodextrin) were less effective. Substantial reductions in faecal water-induced DNA damage were also seen with supernatants from incubation of prebiotics with Bif. Bb12. The supernatant of fermentations involving Ent. faecium and Bif. sp. 420 generally had less potent effects on genotoxicity although some reductions with Raftiline and Elixor fermentations were apparent.

Survival of freeze dried Lactobacillus plantarum in instant fruit powders and reconstituted fruit juices

The aim of this study was to investigate the survival of freeze dried Lactobacillus plantarum cells mixed with several freeze dried instant fruit powders (strawberry, pomegranate, blackcurrant and cranberry) during storage for 12 months as well as after reconstitution with water each month. Inulin and gum arabic were also added to the instant fruit powders at two levels (10% and 20% w/w of dry weight) to improve the cell survival and functional properties of the product. The best cell survival over the 12 months of storage was observed for the blackcurrant powder (almost no decrease) followed by strawberry (~ 0.3 log decrease), pomegranate (~ 0.9 log decrease), whereas the worst survival was obtained in cranberry powder (~ 4.5 logs). To explain these results multiple regression analysis was conducted with the log decrease [log10N0 month − log10N12 months] as the dependent variable and water activity, pH, citric acid, dietary fibre and total phenol as the independent variables. The results indicated that among all the examined factors, the [log10N0 month − log10N12 months] depended only on the water activity (P < 0.05). Inulin and gum arabic demonstrated a substantial protective effect on cell survival (1–1.5 log) in the case of cranberry, which was likely due to a physical interaction between the cells and the carbohydrates. After reconstituting the dried fruit powders at room temperature and measuring cell viability for up to 4 h, it was shown that in the case of strawberry juice there was no decrease, and very little in the case of pomegranate and blackcurrant juices (< 0.5 log). On the other hand, a significant decrease was observed for cranberry juice (P < 0.05), which increased as the storage time of the dried cranberry powder increased, indicating that the cells became more susceptible with prolonged storage. Multiple regression analysis indicated that the main factors influencing cell survival were water activity and pH, while citric acid, dietary fibre and total phenol did not have an effect. Furthermore, inulin and gum arabic addition did not have a significant (P > 0.05) effect upon reconstitution of the dried fruit powder. This study showed that instant juice powders are very good carriers of probiotic cells and constitute good alternatives to highly acidic fruit juices.

Fermentable carbohydrate alters hypothalamic neuronal activity and protects against the obesogenic environment

Obesity has become a major global health problem. Recently, attention has focused on the benefits of fermentable carbohydrates on modulating metabolism. Here, we take a system approach to investigate the physiological effects of supplementation with oligofructose-enriched inulin (In). We hypothesize that supplementation with this fermentable carbohydrate will not only lead to changes in body weight and composition, but also to modulation in neuronal activation in the hypothalamus. Male C57BL/6 mice were maintained on a normal chow diet (control) or a high fat (HF) diet supplemented with either oligofructose-enriched In or corn starch (Cs) for 9 weeks. Compared to HF+Cs diet, In supplementation led to significant reduction in average daily weight gain (mean ± s.e.m.: 0.19 ± 0.01 g vs. 0.26 ± 0.02 g, P < 0.01), total body adiposity (24.9 ± 1.2% vs. 30.7 ± 1.4%, P < 0.01), and lowered liver fat content (11.7 ± 1.7% vs. 23.8 ± 3.4%, P < 0.01). Significant changes were also observed in fecal bacterial distribution, with increases in both Bifidobacteria and Lactobacillius and a significant increase in short chain fatty acids (SCFA). Using manganese-enhanced MRI (MEMRI), we observed a significant increase in neuronal activation within the arcuate nucleus (ARC) of animals that received In supplementation compared to those fed HF+Cs diet. In conclusion, we have demonstrated for the first time, in the same animal, a wide range of beneficial metabolic effects following supplementation of a HF diet with oligofructose-enriched In, as well as significant changes in hypothalamic neuronal activity.

Development of antimicrobial synbiotics using potentially-probiotic faecal isolates of Lactobacillus fermentum and Bifidobacterium longum

The aims of the present study were to investigate in vitro the antimicrobial activity of Lactobacillus fermentum and Bifidobacterium longum, isolated from faeces of healthy elderly individuals, against enterohaemorrhagic Escherichia coli (E. coli O157:H7) and enteropathogenic E. coli (E. coli O86), to determine the capability of the selected strains to tolerate acid and bile in vitro, to select suitable carbohydrates in order to enhance the growth and maximise antimicrobial activity of the putative probiotic organisms and examine the adhesion properties of the synbiotics. Antimicrobial activity of the putative probiotics and synbiotics was investigated by a microtitre method using cell-free culture supernatants (CFCS). Results of the antimicrobial assay showed that both putative probiotic strains produced compounds at pH 5 that lead to higher lag phases of both E. coli O157:H7 and E. coli O86. When half the quantity of cell-free culture supernatants of both probiotic strains was used at pH 5, B. longum maintained the same antimicrobial effect against both strains of E. coli, whereas L. fermentum lead to a higher lag phase of E. coli O86 only. Neutralization of the culture supernatants with alkali reduced the antimicrobial effect with only cell-free supernatant of L. fermentum causing lower maximum growth rates of E. coli O157:H7 and E. coli O86. L. fermentum appeared to be acid tolerant whereas B. longum was more susceptible to acid and both isolates were bile tolerant. A short chain fructooligosaccharide (scFOS) and an isomalto-oligosaccharide (IMO) proved to be the most effective substrates, enhancing antimicrobial activity for L. fermentum and B. longum respectively. The adhesion of the synbiotic combinations showed that L. fermentum, exhibited higher percentage of adhesion when grown on glucose and as a synbiotic combination with scFOS whereas B. longum exhibited lowest percentage of adhesion when grown on both glucose and IMO.

Sesquiterpenoids lactones: benefits to plants and people

Sesquiterpenoids, and specifically sesquiterpene lactones from Asteraceae, may play a highly significant role in human health, both as part of a balanced diet and as pharmaceutical agents, due to their potential for the treatment of cardiovascular disease and cancer. This review highlights the role of sesquiterpene lactones endogenously in the plants that produce them, and explores mechanisms by which they interact in animal and human consumers of these plants. Several mechanisms are proposed for the reduction of inflammation and tumorigenesis at potentially achievable levels in humans. Plants can be classified by their specific array of produced sesquiterpene lactones, showing high levels of translational control. Studies of folk medicines implicate sesquiterpene lactones as the active ingredient in many treatments for other ailments such as diarrhea, burns, influenza, and neurodegradation. In addition to the anti-inflammatory response, sesquiterpene lactones have been found to sensitize tumor cells to conventional drug treatments. This review explores the varied ecological roles of sesquiterpenes in the plant producer, depending upon the plant and the compound. These include allelopathy with other plants, insects, and microbes, thereby causing behavioural or developmental modification to these secondary organisms to the benefit of the sesquiterpenoid producer. Some sesquiterpenoid lactones are antimicrobial, disrupting the cell wall of fungi and invasive bacteria, whereas others protect the plant from environmental stresses that would otherwise cause oxidative damage. Many of the compounds are effective due to their bitter flavor, which has obvious implications for human consumers. The implications of sesquiterpenoid lactone qualitiesfor future crop production are discussed.

Natural caprine whey oligosaccharides separated by membrane technology and profile evaluation by capillary electrophoresis

The functional food market is growing rapidly and membrane processing offers several advantages over conventional methods for separation, fractionation and recovery of bioactive components. The aim of the present study was to select a process that could be implemented easily on an industrial scale for the isolation of natural lactose-derived oligosaccharides (OS) from caprine whey, enabling the development of functional foods for clinical and infant nutrition. The most efficient process was the combination of a pre-treatment to eliminate proteins and fat, using an ultrafiltration (UF) membrane of 25 kDa molecular weight cut off (MWCO), followed by a tighter UF membrane with 1 kDa MWCO. Circa 90% of the carbohydrates recovered in the final retentate were OS. Capillary electrophoresis was used to evaluate the OS profile in this retentate. The combined membrane-processing system is thus a promising technique for obtaining natural concentrated OS from whey. Powered

Human microbiota in health and disease

Each human body plays host to a microbial population which is both numerically vast (at around 1014 microbial cells) and phenomenally diverse (over 1,000 species). The majority of the microbial species in the gut have not been cultured but the application of culture-independent approaches for high throughput diversity and functionality analysis has allowed characterisation of the diverse microbial phylotypes present in health and disease. Studies in monozygotic twins, showing that these retain highly similar microbiota decades after birth and initial colonisation, are strongly indicative that diversity of the microbiome is host-specific and affected by the genotype. Microbial diversity in the human body is reflected in both richness and evenness. Diversity increases steeply from birth reaching its highest point in early adulthood, before declining in older age. However, in healthy subjects there appears to be a core of microbial phylotypes which remains relatively stable over time. Studies of individuals from diverse geopraphies suggest that clusters of intestinal bacterial groups tend to occur together, constituting ‘enterotypes’. So variation in intestinal microbiota is stratified rather than continuous and there may be a limited number of host/microbial states which respond differently to environmental influences. Exploration of enterotypes and functional groups may provide biomarkers for disease and insights into the potential for new treatments based on manipulation of the microbiome. In health, the microbiota interact with host defences and exist in harmonious homeostasis which can then be disturbed by invading organisms or when ‘carpet bombing’ by antibiotics occurs. In a portion of individuals with infections, the disease will resolve itself without the need for antibiotics and microbial homeostasis with the host’s defences is restored. The administration of probiotics (live microorganisms which when administered in adequate amounts confer a health benefit on the host) represents an artificial way to enhance or stimulate these natural processes. The study of innate mechanisms of antimicrobial defence on the skin, including the production of numerous antimicrobial peptides (AMPs), has shown an important role for skin commensal organisms. These organisms may produce AMPs, and also amplify the innate immune responses to pathogens by activating signalling pathways and processing host produced AMPs. Research continues into how to enhance and manipulate the role of commensal organisms on the skin. The challenges of skin infection (including diseases caused by multiply resistant organisms) and infestations remain considerable. The potential to re-colonise the skin to replace or reduce pathogens, and exploring the relationship between microbiota elsewhere and skin diseases are among a growing list of research targets. Lactobacillus species are among the best known ‘beneficial’ bacterial members of the human microbiota. Of the approximately 120 species known, about 15 are known to occur in the human vagina. These organisms have multiple properties, including the production of lactic acid, hydrogen peroxide and bacteriocins, which render the vagina inhospitable to potential pathogens. Depletion of the of the normal Lactobacillus population and overgrowth of vaginal anaerobes, accompanied by the loss of normal vaginal acidity can lead to bacterial vaginosis – the commonest cause of abnormal vaginal discharge in women. Some vaginal anaerobes are associated with the formation of vaginal biofilms which serve to act as a reservoir of organisms which persists after standard antibiotic therapy of bacterial vaginosis and may help to account for the characteristically high relapse rate in the condition. Administration of Lactobacillus species both vaginally and orally have shown beneficial effects in the treatment of bacterial vaginosis and such treatments have an excellent overall safety record. Candida albicans is a frequent coloniser of human skin and mucosal membranes, and is a normal part of the microbiota in the mouth, gut and vagina. Nevertheless Candida albicans is the most common fungal pathogen worldwide and is a leading cause of serious and often fatal nosocomial infections. What turns this organism from a commensal to a pathogen is a combination of increasing virulence in the organism and predisposing host factors that compromise immunity. There has been considerable research into the use of probiotic Lactobacillus spp. in vaginal candidiasis. Studies in reconstituted human epithelium and monolayer cell cultures have shown that L. rhamnosus GG can protect mucosa from damage caused by Candida albicans, and enhance the immune responses of mucosal surfaces. Such findings offer the promise that the use of such probiotic bacteria could provide new options for antifungal therapy. Studies of changes of the human intestinal microbiota in health and disease are complicated by its size and diversity. The Alimentary Pharmabiotic Centre in Cork (Republic of Ireland) has the mission to ‘mine microbes for mankind’ and its work illustrates the potential benefits of understanding the gut microbiota. Work undertaken at the centre includes: mapping changes in the microbiota with age; studies of the interaction between the microbiota and the gut; potential interactions between the gut microbiota and the central nervous system; the potential for probiotics to act as anti-infectives including through the production of bacteriocins; and the characterisation of interactions between gut microbiota and bile acids which have important roles as signalling molecules and in immunity. The important disease entity where the role of the gut microbiota appears to be central is the Irritable Bowel Syndrome (IBS). IBS patients show evidence of immune activation, impaired gut barrier function and abnormal gut microbiota. Studies with probiotics have shown that these organisms can exert anti-inflammatory effects in inflammatory bowel disease and may strengthen the gut barrier in IBS of the diarrhoea-predominant type. Formal randomised trials of probiotics in IBS show mixed results with limited benefit for some but not all. Studies confirm that administered probiotics can survive and temporarily colonise the gut. They can also stimulate the numbers of other lactic acid bacilli in the gut, and reduce the numbers of pathogens. However consuming live organisms is not the only way to influence gut microbiota. Dietary prebiotics are selectively fermented ingredients that can change the composition and/or activity of the gastrointestinal microbiota in beneficial ways. Dietary components that reach the colon, and are available to influence the microbiota include poorly digestible carbohydrates, such as non-starch polysaccharides, resistant starch, non-digestible oligosaccharides (NDOs) and polyphenols. Mixtures of probiotic and prebiotic ingredients that can selectively stimulate growth or activity of health promoting bacteria have been termed ‘synbiotics’. All of these approaches can influence gut microbial ecology, mainly to increase bifidobacteria and lactobacilli, but metagenomic approaches may reveal wider effects. Characterising how these changes produce physiological benefits may enable broader use of these tactics in health and disease in the future. The current status of probiotic products commercially available worldwide is less than ideal. Prevalent problems include misidentification of ingredient organisms and poor viability of probiotic microorganisms leading to inadequate shelf life. On occasions these problems mean that some commercially available products cannot be considered to meet the definition of a probiotic product. Given the potential benefits of manipulating the human microbiota for beneficial effects, there is a clear need for improved regulation of probiotics. The potential importance of the human microbiota cannot be overstated. ‘We feed our microbes, they talk to us and we benefit. We just have to understand and then exploit this.’ (Willem de Vos).

Sucrose transport in the phloem: integrating root responses to phosphorus starvation

Sugars in plants, derived from photosynthesis, act as substrates for energy metabolism and the biosynthesis of complex carbohydrates, providing sink tissues with the necessary resources to grow and to develop. In addition, sugars can act as secondary messengers, with the ability to regulate plant growth and development in response to biotic and abiotic stresses. Sugar-signalling networks have the ability to regulate directly the expression of genes and to interact with other signalling pathways. Photosynthate is primarily transported to sink tissues as sucrose via the phloem. Under phosphorus (P) starvation, plants accumulate sugars and starch in their leaves. Increased loading of sucrose to the phloem under P starvation not only functions to relocate carbon resources to the roots, which increases their size relative to the shoot, but also has the potential to initiate sugar-signalling cascades that alter the expression of genes involved in optimizing root biochemistry to acquire soil phosphorus through increased expression and activity of inorganic phosphate transporters, the secretion of acid phosphatases and organic acids to release P from the soil, and the optimization of internal P use. This review looks at the evidence for the involvement of phloem sucrose in co-ordinating plant responses to P starvation at both the transcriptional and physiological levels.

Sugar signaling in root responses to low phosphorus availability

Over the last decade, major advances have been made in our understanding of how plants sense, signal, and respond to soil phosphorus (P) availability (Amtmann et al., 2006; White and Hammond, 2008; Nilsson et al., 2010; Yang and Finnegan, 2010; Vance, 2010; George et al., 2011). Previously, we have reviewed the potential for shoot-derived carbohydrate signals to initiate acclimatory responses in roots to low P availability. In this context, these carbohydrates act as systemic plant growth regulators (Hammond and White, 2008). Photosynthate is transported primarily to sink tissues as Suc via the phloem. Under P starvation, plants accumulate sugars and starch in their leaves. Increased loading of Suc to the phloem under P starvation primarily functions to relocate carbon resources to the roots, which increases their size relative to the shoot (Hermans et al., 2006). The translocation of sugars via the phloem also has the potential to initiate sugar signaling cascades that alter the expression of genes involved plant responses to low P availability. These include optimizing root biochemistry to acquire soil P, through increased expression and activity of inorganic phosphate (Pi) transporters, the secretion of acid phosphatases and organic acids to release P from the soil, and the optimization of internal P use (Hammond and White, 2008). Here, we provide an Update to the field of plant signaling responses to low P availability and the interactions with sugar signaling components. Advances in the P signaling pathways and the roles of hormones in signaling plant responses to low P availability are also reviewed, and where possible their interactions with potential sugar signaling pathways.

How do plants respond to nutrient shortage by biomass allocation?

Plants constantly sense the changes in their environment; when mineral elements are scarce, they often allocate a greater proportion of their biomass to the root system. This acclimatory response is a consequence of metabolic changes in the shoot and an adjustment of carbohydrate transport to the root. It has long been known that deficiencies of essential macronutrients (nitrogen, phosphorus, potassium and magnesium) result in an accumulation of carbohydrates in leaves and roots, and modify the shoot-to-root biomass ratio. Here, we present an update on the effects of mineral deficiencies on the expression of genes involved in primary metabolism in the shoot, the evidence for increased carbohydrate concentrations and altered biomass allocation between shoot and root, and the consequences of these changes on the growth and morphology of the plant root system.