Microbial fermentation of Fossence™, a short-chain fructo-oligosaccharide, under simulated human proximal colonic condition and assessment of its prebiotic effects-a pilot study.

A short-chain fructo-oligosaccharide (sc-FOS) was tested in a simulator of the human gut microbial ecosystem (SHIME) in vitro model to quantify its prebiotic effects according to Prebiotic Index (PI) and Measure of prebiotic effect (MPE) equations. FossenceTM, (sc-FOS, 0.5%) was fermented in a simulated human proximal colonic condition, using a fecal inoculum from a healthy individual. We analysed the pH reduction, substrate utilization, lactate and short-chain fatty acid (SCFA) production and microbial community modulation. Microbial fermentation of sc-FOS strongly reduced the media pH indicating the production of lactate and SCFA with accumulation of lactate and enhanced levels of acetate (34.38 ± 0.38 mM), propionate (20.93 ± 0.56 mM) and butyrate (4.93 ± 0.03 mM) compared to 18.46 ± 0.20 mM, 6.24 ± 0.10 mM and 3.3 ± 0.06 mM in the blank, respectively. Total SCFA production in test media was 61.91 ± 0.87 mM compared to 33.65 ± 0.36 mM in blank and the contribution of free-sugars present in sc-FOS to SCFAs was negligible. Modulation of the microbial community was analysed through 16S rRNA sequencing and we found that sc-FOS greatly stimulated the beneficial bacteria such as Bifidobacteria and Lactobacillus. We report the PI and MPE values for FossenceTM, as 14.9 and 0.01 respectively at the end of 24 h, which is an indicator of a strong prebiotic effect.

Dietary galactosyl and mannosyl carbohydrates: In-vitro assessment of prebiotic effects.

The prebiotic activities of hydrolyzed guar gum (GMOS, <1 kDa; GMPS, 1-10 kDa), manno-oligosaccharide (MOS, <1 kDa), and galacto-oligosaccharide (GOS, <1 kDa) were evaluated by in vitro fermentation. The tested carbohydrates showed selective prebiotic effects on bacterial growth, short-chain fatty acid (SCFA)-production, and substrate consumption. GOS and GMOS markedly promoted the growth of bifidobacteria and Clostridium butyricum, respectively, whereas MOS showed the strongest butyrogenic effect. Moreover, SCFA production in the hydrolyzed guar gum groups was closely related to the varied molecular weight (Mw) of the hydrolysate. During in vitro fermentation with human fecal inocula, GMOS gave the highest yields of lactate, propionate, and butyrate after 48 h fermentation. Combined application of MOS and C. butyricum increased the abundance of Clostridiaceae_1. Overall, our results indicate that galactosyl and mannosyl carbohydrates have individualized prebiotic effects which are associated with their chemical structures including their glycoside composition and Mw.

Production, Structural Characterization, and In Vitro Assessment of the Prebiotic Potential of Butyl-Fructooligosaccharides.

Short-chain fatty acids (SCFAs), especially butyrate, produced in mammalian intestinal tracts via fermentation of dietary fiber, are known biofunctional compounds in humans. However, the variability of fermentable fiber consumed on a daily basis and the diversity of gut microbiota within individuals often limits the production of short-chain fatty acids in the human gut. In this study, we attempted to enhance the butyrate levels in human fecal samples by utilizing butyl-fructooligosaccharides (B-FOS) as a novel prebiotic substance. Two major types of B-FOS (GF3-1B and GF3-2B), composed of short-chain fructooligosaccharides (FOS) bound to one or two butyric groups by ester bonds, were synthesized. Qualitative analysis of these B-FOS using Fourier transform infrared (FT-IR) spectroscopy, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), nuclear magnetic resonance (NMR) and low-resolution fast-atom bombardment mass spectra (LR-FAB-MS), showed that the chemical structure of GF3-1B and GF3-2B were [O-(1-buty-ß-D-fru-(2→1)-O-ß-D-fru-(2→1)-O-ß-D-fru-O-α-D-glu] and [O-(1-buty)-ß-D-fru-(2→1)-O-ß-D-fru-(2→1)-O-(4-buty)-ß-D-fru-O-α-D-glu], respectively. The ratio of these two compounds was approximately 5:3. To verify their biofunctionality as prebiotic oligosaccharides, proliferation and survival patterns of human fecal microbiota were examined in vitro via 16S rRNA metagenomics analysis compared to a positive FOS control and a negative control without a carbon source. B-FOS treatment showed different enrichment patterns on the fecal microbiota community during fermentation, and especially stimulated the growth of major butyrate producing bacterial consortia and modulated specific butyrate producing pathways with significantly enhanced butyrate levels. Furthermore, the relative abundance of Fusobacterium and ammonia production with related metabolic genes were greatly reduced with B-FOS and FOS treatment compared to the control group. These findings indicate that B-FOS differentially promotes butyrate production through the enhancement of butyrate-producing bacteria and their metabolic genes, and can be applied as a novel prebiotic compound in vivo.

Low-Cost Method Generating In Situ Anaerobic Conditions on a 96-Well Plate for Microbial Fermentation in Food Research.

Commercial tools and instruments have been developed for a screening study of microbial fermentation, but they are expensive and mostly confined to aerobic fermentation only. There is little development on the generation of anaerobic conditions directly on a 96-well plate. This report proposed a simple and versatile microbial fermentation system known as OVAMO that makes use of Oxyrase, vacuum, and mineral oil to generate an in situ anaerobic environment on a 96-well plate for at least 48 h. The practicality of OVAMO in anaerobic fermentation experiments used for functional food research was validated by a prebiotic screening study of different carbohydrates by Bifidobacterium longum subsp. infantis. The OVAMO system provides a less expensive but effective way to conduct a microbial fermentation screening study that requires anaerobic conditions without the need for atmospheric control by external devices.

In vitro assessment of iron availability from commercial Young Child Formulae supplemented with prebiotics.

PURPOSE: Iron is essential for development and growth in young children; unfortunately, iron deficiency (ID) is a significant public health problem in this population. Young Child Formulae (YCF), milk-derived products fortified with iron and ascorbic acid (AA, an enhancer of iron absorption) may be good sources of iron to help prevent ID. Furthermore, some YCF are supplemented with prebiotics, non-digestible carbohydrates suggested to enhance iron bioavailability. The aim of our study was to evaluate iron bioavailability of YCF relative to prebiotic and AA concentrations. We hypothesised that YCF with the highest levels of prebiotics and AA would have the most bioavailable iron. METHODS: We used the in vitro digestion/Caco-2 cell model to measure iron bioavailability from 4 commercially available YCF with approximately equal amounts of iron, but varying amounts of: AA and the prebiotics fructo- and galacto-oligosaccharides. Caco-2 cell ferritin formation was used as a surrogate marker for iron bioavailability. RESULTS: The YCF with the highest concentration of prebiotics and AA had the highest iron bioavailability; conversely, the YCF with the lowest concentration of prebiotics and AA had the lowest. After the addition of exogenous prebiotics, so that all tested YCF had equivalent amounts, there was no longer a significant difference between YCF iron bioavailability. CONCLUSION: Our results suggest that ascorbic acid and prebiotics in YCF improve iron bioavailability. Ensuring that iron is delivered in a bioavailable form would improve the nutritional benefits of YCF in relation to ID/IDA amongst young children; therefore, further exploration of our findings in vivo is warranted.

Assessment of in Vitro Digestibility of Dietary Carbohydrates Using Rat Small Intestinal Extract.

There are few studies on the assessment of digestibility of nondigestible carbohydrates, despite their increasingly important role in human health. In vitro digestibility of a range of dietary carbohydrates classified as digestible (maltose, sucrose, and lactose), well-recognized (lactulose, fructooligosaccharides (FOS), and two types of galactooligosaccharides (GOS) differing in the predominant glycosidic linkage), and potential (lactosucrose and GOS from lactulose, OsLu) prebiotics using a rat small intestinal extract (RSIE) under physiological conditions of temperature and pH is described. Recognized and potential prebiotics were highly resistant to RSIE digestion although partial hydrolysis at different extents was observed. FOS and lactulose were the most resistant to digestion, followed closely by OsLu and more distantly by both types of GOS and lactosucrose. In GOS, ß(1 → 6) linkages were more resistant to digestion than ß(1 → 4) bonds. The reported in vitro digestion model is a useful, simple, and cost-effective tool to evaluate the digestibility of dietary oligosaccharides.

A Survey of Commercially Available Isomaltooligosaccharide-Based Food Ingredients.

Isomaltooligosaccharides (IMOs) are included in many commercially available food products including protein/fiber bars, shakes, and other dietary supplements. Marketed as "high fiber," "prebiotic soluble fiber," and/or as a "low-calorie, low glycemic sweetener," IMO may be present in significant amounts, for example, more than 15 g/item or serving. Herein, high-pressure anion exchange chromatography with pulsed amperometric detection and high-pressure liquid chromatography with differential refractive index detection are used to compare 7 commercially available IMO-containing bulk food ingredients. The ingredients are typical of those produced either (a) via bacterial fermentation ("fermented" IMO or MIMO) of sucrose in the presence of a maltose acceptor mediated by a glucosyltransferase enzyme (dextransucrase), or (b) via transglycosylation of hydrolyzed starch with α-glucosidase ("industrial" IMO). Analysis of the results with respect to digestibility suggests that the potential glycemic impact of the ingredients and products containing "industrial" IMO may be inconsistent with the product labeling and/or certificates of analysis with respect to overall fiber content, prebiotic fiber content, and glycemic response and are thus inappropriate for diabetic patients and those on low-carbohydrate (for example, ketogenic) diets.

Physicochemical properties and intestinal protective effect of ultra-micro ground insoluble dietary fibre from carrot pomace.

Carrot pomace is an abundant, but underutilized, byproduct from the juice industry. In this study, the insoluble dietary fiber from carrot pomace was treated using an ultra-microgrinding process, and the resulting changes in its physicochemical properties and intestinal protective effect against heavy metal damage were examined. The SEM and fluorescence microscopy results showed that the grinding process could significantly decrease the particle size of carrot insoluble dietary fibre and increase its Brunauer-Emmett-Teller surface area from 0.374 to 1.835 m(2) g(-1). Correspondingly, the water-holding capacity, swelling capacity, and oil-holding capacity increased by 62.09%, 49.25% and 45.45%, respectively. The glucose-, nitrite-, and lead ion-adsorbing abilities also improved significantly compared with the raw samples. In addition, apoptosis assessment by AO/EB revealed that the ground fibre could effectively protect Caco-2 cells from lead ion damage. The MTT assay showed that carrot insoluble dietary fibre has no toxicity for Caco-2 cells at a concentration of 10.0 mg L(-1). The findings of this study highlighted the potential of the ultra-microgrinding process to produce a high added-value fibre ingredient from carrot residues.

In vitro assessment of the prebiotic potential of Aloe vera mucilage and its impact on the human microbiota.

Aloe vera mucilage is reported to be rich in acemannan that is a polysaccharide with a backbone of ß-(1→4)-D-mannose residues acetylated at the C-2 and C-3 positions and contains some side chains of galactose and arabinose attached to the C-6 carbon. The evaluation of the prebiotic potential of Aloe vera mucilage was carried out by in vitro fermentation using intestinal microbiota from six healthy donors as the inoculum. The prebiotic activity was assessed through the quantification of short chain fatty acids (SCFA) and the evaluation of dynamic bacterial population in mixed faecal cultures by fluorescence in situ hybridization (FISH). Our findings support the possible incorporation of the Aloe vera mucilage in the development of a variety of food products known as prebiotics aimed at improving gastrointestinal health.

Development and assessment of healthy properties of meat and meat products designed as functional foods.

This review deals with the two major aspects to be considered in the context of meat-based functional foods and human health. One involves the different strategies used to improve (increase or reduce) the presence of bioactive (healthy and unhealthy) compounds in meat and meat products in order to develop potential meat-based functional foods; these strategies are basically concerned with animal production practices, meat processing and storage, distribution and consumption conditions. Since the link between the consumption of those foods and their potentially beneficial effects (improving health and/or reducing the risk of several chronic diseases) needs to be demonstrated scientifically, the second aspect considered is related to intervention studies to examine the functional capacity of meat-based potentially functional foods in humans, discussing how the functionality of a food can be assessed in terms of its effects on health in relation to both target body functions and risk factors.

Modifying the gastrointestinal ecology in alternatively raised poultry and the potential for molecular and metabolomic assessment.

Consumer demand for nonconventional poultry products continues to increase in the United States. In pasture flock and organic poultry production, probiotics and prebiotic feed additives have potential advantages because they are thought to promote intestinal health and may offer a replacement for current intervention strategies that are not considered acceptable for these production systems. Prebiotics have been demonstrated to produce effects on the gastrointestinal tract including modulation of microflora by promoting selective increases in beneficial bacteria concomitant with decreases in undesirable bacteria. In-depth assessment of microbial community changes during host growth and development as well as the establishment of beneficial microbial species by adding biologicals such as probiotics and prebiotics is important to achieve predictable and consistent improvements in chicken health and productivity. To analyze microflora shifts and metabolites produced by bacteria in the gut as well as host responses to biological additives, sophisticated molecular techniques are now available and are becoming more widely used. Polymerase chain reaction assays, denaturing gradient gel electrophoresis, and temperature gradient gel electrophoresis offer approaches for detecting microbial shifts in the gut. Likewise, the employment of microarrays and molecular analysis of gut tissues can reveal insight into gut physiological and responses to dietary and other changes. Recent application of 16S rDNA sequencing and analysis utilizing basic local alignment search tool (BLAST) and FASTA databases on poultry gut samples have the potential to provide a much more in-depth assessment of the gut microbiome. Utilizing ultra pressure liquid chromatography-mass spectroscopy profiling, metabolomic assessment of gut contents will also allow for parallel comparisons of changes in the gut contents with microbiome and physiological responses. Combining all these technologies will provide a plenary understanding of poultry gut health in alternative production systems.

Isolation of prebiotic carbohydrates by supercritical fluid extraction. Scaling-up and economical feasibility.

Production of prebiotic carbohydrates at competitive prices is a challenge nowadays since the well-established production processes involve many purification steps which are labour intensive and require important amounts of reagents and products thus increasing prebiotic's price. Several processes have been studied in our laboratory involving the use of Supercritical Fluid Technology to fractionate and purify carbohydrate solid mixtures. Research carried out at laboratory scale using theoretical mixtures (lactose/lactulose and galactose/tagatose), commercially available carbohydrate mixtures and carbohydrate mixtures produced by enzymatic transglycosylation and isomerized with complexating reagents demonstrated that purification of prebiotic carbohydrates was technically possible by supercritical fluid extraction. In the present work, the process optimized at laboratory scale to fractionate carbohydrate mixtures produced by enzymatic transglycosylation has been scaled-up to an industrial level and its economic feasibility has been simulated employing AspenONE(®) V7.3 software to obtain consistent data supporting the interest of a potential investment for prebiotics production at large scale using supercritical fluids.

Synthesis and characterization of a potential prebiotic trisaccharide from cheese whey permeate and sucrose by Leuconostoc mesenteroides dextransucrase.

The production of new bioactive oligosaccharides is currently garnering much attention for their potential use as functional ingredients. This work addresses the enzymatic synthesis and NMR structural characterization of 2-α-D-glucopyranosyl-lactose derived from sucrose:lactose and sucrose:cheese whey permeate mixtures by using a Leuconostoc mesenteroides B-512F dextransucrase. The effect of synthesis conditions, including concentration of substrates, molar ratio of donor/acceptor, enzyme concentration, reaction time, and temperature, on the formation of transfer products is evaluated. Results indicated that cheese whey permeate is a suitable material for the synthesis of 2-α-D-glucopyranosyl-lactose, giving rise to yields around 50% (in weight respect to the initial amount of lactose) under the optimum reaction conditions. According to its structure, this trisaccharide is an excellent candidate for a new prebiotic ingredient, due to the reported high resistance of α-(1→2) linkages to the digestive enzymes in humans and animals, as well as to its potential selective stimulation of beneficial bacteria in the large intestine mainly attributed to the two linked glucose units located at the reducing end that reflects the disaccharide kojibiose (2-α-D-glucopyranosyl-D-glucose). These findings could contribute to broadening the use of important agricultural raw materials, such as sucrose or cheese whey permeates, as renewable substrates for enzymatic synthesis of oligosaccharides of nutritional interest.

Drivers of liking for yogurt drinks with prebiotics and probiotics.

Several studies have addressed the sensory properties of yogurt. However, as the market for yogurt continues to expand and new varieties of yogurt with novel ingredients emerge, additional sensory tests are needed to ensure the quality of the products. Three selected prebiotics, soluble corn fiber, polydextrose, and chicory inulin, were each added at an excellent source of fiber (5 g fiber/serving) or a good fiber source (2.5 g fiber/serving) levels into a yogurt drink base. Three additional yogurt drinks contained 5 g of each of the separate prebiotics along with a mixture of probiotics (Bifidobacterium lactis Bb-12 and Lactobacillus acidophilus LA-5). A control sample with no prebiotics or probiotics was also included in the experimental design. Yogurt drinks were evaluated by 110 consumers for overall acceptance, acceptance of aroma, appearance, taste, and texture, and purchase intent. Demographic information pertaining to consumer knowledge of prebiotics and probiotics was collected. Consumer data were correlated with previously obtained descriptive analysis data to identify drivers of liking. Data were analyzed by analysis of variance (ANOVA), Fisher's least significant difference (LSD), cluster analysis, internal preference mapping, and external preference mapping. Total variance explained by the internal and external preference maps were 32.2% and 64.6%, respectively, which showed higher levels of the prebiotics with probiotics drove consumer liking compared to lower levels without probiotics. In terms of ingredients added, chicory inulin and polydextrose were preferred over soluble corn fiber. Yogurt drinks with these prebiotics included and probiotics were characterized by a medium level of sweetness and high viscosity. Development of new prebiotic and probiotic containing drinkable yogurts should strive for a medium level of sweetness and high viscosity for maximum consumer acceptance.