In vitro fermentation of bacterial cellulose composites as model dietary fibers

Plant cell walls within the human diet are compositionally heterogeneous, so defining the basis of nutritive properties is difficult. Using a pig fecal inoculum, in vitro fermentations of soluble forms of arabinoxylan, mixed-linkage glucan, and xyloglucan were compared with the same polymers incorporated into bacterial cellulose composites. Fermentation rates were highest and similar for the soluble polysaccharides. Cellulose composites incorporating those polysaccharides fermented more slowly and at similar rates to wheat bran. Bacterial cellulose and cotton fermented most slowly. Cellulose composite fermentation resulted in a different short-chain fatty acid profile, compared with soluble polysaccharides, with more butyrate and less propionate. The results suggest that physical form is more relevant than the chemistry of plant cell wall polysaccharides in determining both rate and end-products of fermentation using fecal bacteria. This work also establishes bacterial cellulose composites as a useful model system for the fermentation of complex cell wall dietary fiber.     

Effect of species variation and processing on phenolic composition and in vitro antioxidant activity of aqueous extracts of Cyclopia spp. (Honeybush Tea)

The in vitro antioxidant activity of aqueous extracts prepared from four Cyclopia spp. (unfermented and fermented) was assessed using radical (ABTS *+) scavenging, ferric ion reduction, and inhibition of Fe2+-induced microsomal lipid peroxidation as criteria. Aqueous extracts of unfermented and fermented Aspalathus linearis (rooibos) and Camellia sinensis teas (green, oolong, and black) were included as reference samples. Qualitative and quantitative differences in phenolic composition were demonstrated for the Cyclopia spp. The xanthone glycoside, a.k.a. mangiferin, was the major monomeric polyphenol present in the Cyclopia extracts, with both unfermented and fermented C. genistoides extracts containing the highest quantities. Fermentation resulted in a significant reduction in extract yields and their total polyphenolic and individual polyphenol contents. Unfermented plant material should preferentially be used for preparation of extracts, as fermentation significantly ( P < 0.05) lowered antioxidant activity of all species, except in the case of C. genistoides, where the ability to inhibit lipid peroxidation was not affected. Unfermented plant material also retained the highest concentration of mangiferin. Overall, extracts of unfermented Cyclopia were either of similar or lower antioxidant activity as compared to the other teas. However, the presence of high levels of mangiferin merits the use of Cyclopia spp. and, in particular, C. genistoides, as an alternative herbal tea and potential dietary supplement.     

Fate of mucilage cell wall polysaccharides during coffee fermentation.

Effects of a 20-h fermentation on cell wall polysaccharides from the mucilage of pulped coffee beans were examined and compared to those of unfermented beans, on alcohol insoluble residues (AIRs), their hot-water-soluble crude pectic substances (PECTs), and their hot-water-insoluble residues (RESs). Yields and compositions were very similar: AIRs, which consisted of approximately 30% highly methylated pectic substances, approximately 9% cellulose, and approximately 15% neutral noncellulosic polysaccharides, exhibited no apparent degradation. However, PECTs from fermented beans were shown to have undergone a slight reduction of their intrinsic viscosity and weight-average molecular weight by capillary viscosimetry and high-performance size-exclusion chromatography. After fermentation, hot-water-insoluble pectic substances of RES exhibited partial de-esterification. Removal of coffee bean mucilage by natural fermentation seems to result from a restricted pectolysis, the mechanism of which remains to be elucidated.     

In vitro fermentation of arabinoxylan oligosaccharides and low molecular mass arabinoxylans with different structural properties from wheat (Triticum aestivum L.) bran and psyllium (Plantago ovata Forsk) seed husk

Ball milling was used for producing complex arabinoxylan oligosaccharides (AXOS) and low molecular mass arabinoxylans (AX) from wheat bran, pericarp-enriched wheat bran, and psyllium seed husk. The arabinose to xylose ratio of the samples produced varied between 0.14 and 0.92, and their average degree of polymerization (avDP) ranged between 42 and 300. Their fermentation for 48 h in an in vitro system using human colon suspensions was compared to enzymatically produced wheat bran AXOS with an arabinose to xylose ratio of 0.22 and 0.34 and an avDP of 4 and 40, respectively. Degrees of AXOS fermentation ranged from 28% to 50% and were lower for the higher arabinose to xylose ratio and/or higher avDP materials. Arabinose to xylose ratios of the unfermented fractions exceeded those of their fermented counterparts, indicating that molecules less substituted with arabinose were preferably fermented. Xylanase, arabinofuranosidase, and xylosidase activities increased with incubation time. Enzyme activities in the samples containing psyllium seed husk AX or psyllium seed husk AXOS were generally higher than those in the wheat bran AXOS preparations. Fermentation gave rise to unbranched short-chain fatty acids. Concentrations of acetic, propionic, and butyric acids increased to 1.9-2.6, 1.9-2.8, and 1.3-2.0 times their initial values, respectively, after 24 h incubation. Results show that the human intestinal microbiota can at least partially use complex AXOS and low molecular mass AX. The tested materials are thus interesting physiologically active carbohydrates.     

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.     

In vitro digestion and fermentation characteristics of temulose molasses, a coproduct of fiberboard production, and select temulose fractions using canine fecal inoculum

It is of interest to discover new fermentable carbohydrate sources that function as prebiotics. This study evaluated the hydrolytic digestibility, fermentative capacity, and microbiota modulating properties of Temulose molasses, four hydrolyzed fractions of Temulose molasses, short-chain fructooligosaccharides (scFOS), and a yeast cell wall preparation (Safmannan). These substrates resisted in vitro hydrolytic digestion. Each substrate was fermented in vitro using dog fecal inoculum, and fermentation characteristics were quantified at 0 and 12 h. All Temulose molasses substrates decreased pH by at least 0.64 unit and resulted in greater (P < 0.05) butyrate and total short-chain fatty acid (SCFA) production compared to scFOS and Safmannan. Temulose molasses substrates resulted in higher (P < 0.01) or equal Bifidobacterium spp. concentrations compared to scFOS. Temulose molasses substrate and its fractions demonstrated prebiotic characteristics as indicated by low hydrolytic digestibility, high fermentability, and enhanced growth of microbiota considered to be beneficial to health.     

Role of Lactic Acid Bacteria in the Eating Qualities of Fermented Rice Noodles

To ascertain the role of natural fermentation and its associated microorganisms in the eating quality of fermented rice noodles, we compared changes in main chemical components, microorganisms, and eating qualities of fermented rice noodles compared with unfermented ones. The results indicated that noodles made from rice grains fermented at 40°C have higher tensile strength, hardness, springiness, and chewiness. Statistical analyses show that fermentation significantly changed the content and composition of the starch, lipid, protein, amylose, and ash of the indica rice. High‐throughput sequencing analyses of the supernatant of the fermentation samples revealed the dynamic changes of the bacteria population during fermentation. Lactic acid bacteria are found to be the dominant bacteria (more than 90%) during the entire process. The relatively high abundance of Lactobacillus fermentum and L. delbrueckii at each phase of the fermentation suggests that the two species are likely to be the species that determine eating quality in the rice fermentation process. The results may provide guidance on the maintenance of quality consistency of fermented rice at the industrial scale.     

Faster fermentation of cooked carrot cell clusters compared to cell wall fragments in vitro by porcine feces

Plant cell walls are the major structural component of fruits and vegetables, which break down to cell wall particles during ingestion (oral mastication) or food processing. The major health-promoting effect of cell walls occurs when they reach the colon and are fermented by the gut microbiota. In this study, the fermentation kinetics of carrot cell wall particle dispersions with different particle size and microstructure were investigated in vitro using porcine feces. The cumulative gas production and short-chain fatty acids (SCFAs) produced were measured at time intervals up to 48 h. The results show that larger cell clusters with an average particle size (d(0.5)) of 298 and 137 μm were more rapidly fermented and produced more SCFAs and gas than smaller single cells (75 μm) or cell fragments (50 μm), particularly between 8 and 20 h. Confocal microscopy suggests that the junctions between cells provides an environment that promotes bacterial growth, outweighing the greater specific surface area of smaller particles as a driver for more rapid fermentation. The study demonstrates that it may be possible, by controlling the size of cell wall particles, to design plant-based foods for fiber delivery and promotion of colon fermentation to maximize the potential for human health.     

Dietary fiber from coffee beverage: degradation by human fecal microbiota

Arabinogalactans and galactomannans from coffee beverages are part of the dietary fiber complex. Chemical structures and fermentability of soluble dietary fiber obtained from a standard filter coffee beverage (Coffea arabica, origin Colombia, medium roasted) by human intestinal bacteria were investigated. One cup (150 mL) of filter coffee contained approximately 0.5 g of soluble dietary fiber (enzymatic-gravimetric methodology), 62% of which were polysaccharides. The remainder was composed of Maillard reaction products and other nonidentified substances. Galactomannans and type II arabinogalactans were present in almost equal proportions. Coffee dietary fiber was readily fermented by human fecal slurries, resulting in the production of short-chain fatty acids (SCFA). After 24 h of fermentation, 85% of total carbohydrates were degraded. In general, arabinosyl units from the polysaccharide fraction were degraded at a slower rate than mannosyl and galactosyl units. In the process of depolymerization arabinogalactans were debranched and the ratio of (1-->3)-linked to (1-->6)-linked galactosyl residues decreased. Structural units composed of (1-->5)-linked arabinosyl residues were least degradable, whereas terminally linked arabinosyl residues were easily utilized. The impact of coffee fiber on numerically dominant population groups of the intestinal microbiota was investigated by fluorescence in situ hybridization combined with flow cytometry (FISH-FC). After 24 h of fermentation, an increase of about 60% of species belonging to the Bacteroides-Prevotella group was observed. The growth of bifidobacteria and lactobacilli was not stimulated.     

REVIEW: Rye Arabinoxylans: Molecular Structure,Physicochemical Properties and Physiological Effects in the Gastrointestinal Tract

Arabinoxylans (AX) are the main dietary fiber (DF) polysaccharides in rye where they represent ≈55% of the total polysaccharides. Rye AX consist of a backbone of (1→4)‐β‐d ‐xylopyranosyl residues (X) mainly substituted with α‐l ‐arabinofuranosyl residues (A) to varying degrees at the O‐2 position, the O‐3 position, or both. The A/X ratio of total AX is 0.49–0.82 and extractable AX ratio is 0.34–0.85 in different studies. AX also contain small amounts of ferulate residues bound to arabinose as esters at its O‐5 position. The weight average molecular weight varies from 40,000 to 900,000 with an average of ≈200,000. AX influence physiology in different segments of the gastrointestinal tract. The complex molecular structure of rye AX makes them resistant against microbial modification in the small intestine; consequently, rye AX have a much higher influence on the viscosity in the small intestinal digesta than does β‐glucan from oats and barley. In spite of that, it has not been possible in studies with AX‐rich foods such as bread to demonstrate a significant effect on the postprandial glucose response, however, a significantly reduced insulin response has been seen. Nevertheless, addition of 6 g and 12 g of AX‐rich wheat fiber to a breakfast meal has significantly lowered postprandial glucose and insulin response. Studies with hypercholesterolemic pigs fed rye buns rich in AX have resulted in dramatic reductions in plasma total and LDL cholesterol, whereas a gender difference was seen in studies on the effect of AX on plasma lipids in humans. Only certain species of bacteria from the human gut produce the enzymes needed for the degradation of AX. Nevertheless, wheat AX stimulate prebiotic bacteria presumably brought about by cross feeding of lactobacilli and bifidobacteria with degradation products from versatile carbohydrate‐degrading bacteria. Soluble AX are readily fermented in the large intestine, the majority is broken down between the ileum and the cecum. AX, characterized by a low degree of substitution and virtually no doubly substituted xylose, are slowly degraded at more distal locations. The remaining AX, characterized by a high degree of substitution, are not degraded at all. Although the fermentation pattern of AX may vary in different experimental models, in vitro fermentation studies and in vivo intervention studies with animals and humans point to AX as substrates that enhance the formation of butyrate in the large intestine.     

Fermentation of cottonseed and other feedstuffs in cattle rumen fluid

Bovine rumen fluid was fermented anaerobically over 48 h with cottonseed, corn, alfalfa, or a mixture of these substrates in anaerobic mineral buffer. Samples taken at different incubation times were derivatized with n-butanol and subjected to gas chromatography and mass spectroscopy. No unusual fermentation end-products from the cottonseed substrate were detected. Cottonseed supported rumen fermentation at levels comparable to those of the other substrates. Major components were usually found in the decreasing order of acetate, propionate, butyrate, and valerate, although acetate and propionate concentrations decreased late in the alfalfa and mixed-feed fermentations, eventually allowing butyrate concentrations to exceed those of propionate. As expected, lactate was produced in high concentrations when corn was fermented. The minor components 2-methylpropionate, 2- and 3-methylbutyrate, phenylacetate, phenylpropionate, and caproate also accumulated, with their relative concentrations varying with the substrate. Succinate was produced in substantial amounts only when corn and alfalfa were fermented; it did not accumulate when cottonseed was the substrate. Samples containing cottonseed were derivatized and subjected to reversed-phase high-performance liquid chromatography, revealing that gossypol concentrations did not change during fermentation.     

Polysaccharide profile and content during the vinification and aging of Tempranillo red wines

Passing from must to wine produced a loss of low-molecular-weight grape structural glucosyl polysaccharides, and an important gain in yeast mannoproteins (MP) and grape-derived arabinogalactan proteins (AGP), and rhamnogalacturonans-II (RG-II). AGP were more easily extracted than RG-II, and small quantities of RG-II monomers and galacturonans were detected. Postmaceration produced a reduction in all grape polysaccharide families, particularly acute in AGP. The reduction of polysaccharides during malolactic fermentation only affected grape AGP, and MP were continuously liberated during the entire vinification process. Wine oak and bottle aging was associated with a relative stability of the polysaccharide families. AGP were thus the majority polysaccharides in young wines but, contrary to what may be thought, structural glucosyl oligosaccharides dominated in musts and MP in aged wines. Precipitation of polysaccharides was noticeable during vinification, and it mainly affected high-molecular-weight AGP and MP. Hydrolytic phenomena affected the balance of wine polysaccharides during late maceration-fermentation.     

Effect of fermentation and autoclaving on dietary fiber fractions and antinutritional factors of beans (Phaseolus vulgaris L.)

The effect of fermentation on antinutritional factors and also on total dietary fiber (TDF), insoluble (IDF) and soluble (SDF) dietary fiber fractions was studied in beans (Phaseolus vulgaris L.). The processes studied were two types of fermentation (lactic acid and natural), and a portion of the obtained flours were processed by autoclaving. The dietary fiber (DF) content and its components were determined using the enzymatic-gravimetric and enzymatic-chemical methods. The TDF content ranged from 24.5% dry matter (DM) in the raw to 25.2% DM in the processed beans. All the processing treatments significantly decreased the SDF content, and irrelevant changes were noticed in the IDF content of processed beans. Cellulose content of all samples was reduced by the processing treatments. Correspondingly, higher amounts of resistant starch was observed in the processed beans, except in the lactic acid fermented ones. However, the levels of pectic polysaccharides and Klason lignin were higher in the samples fermented by Lactobacillus plantarum. The action of microorganisms was determinant for the different degradation of the bean cell wall, disrupting the protein-carbohydrate integration, thus reducing the solubility of DF.     

Moderate ferulate and diferulate levels do not impede maize cell wall degradation by human intestinal microbiota

The degradation of plant fiber by human gut microbiota could be restricted by xylan substitution and cross-linking by ferulate and diferulates, for example, by hindering the association of enzymes such as xylanases with their substrates. To test the influence of feruloylation on cell wall degradability by human intestinal microbiota, nonlignified primary cell walls from maize cell suspensions, containing various degrees of ferulate substitution and diferulate cross-linking, were incubated in nylon bags in vitro with human fecal microbiota. Degradation rates were determined gravimetrically, and the cell walls were analyzed for carbohydrates, ferulate monomers, dehydrodiferulates, dehydrotriferulates, and other minor phenolic constituents. Shifting cell wall concentrations of total ferulates from 1.5 to 15.8 mg/g and those of diferulates from 0.8 to 2.6 mg/g did not alter the release of carbohydrates or the overall degradation of cell walls. After 24 h of fermentation, the degradation of xylans and pectins exceeded 90%, whereas cellulose remained undegraded. The results indicate that low to moderate levels of ferulates and diferulates do not interfere with hydrolysis of nonlignified cell walls by human gut microbiota.     

Enzymatic degradation of oligosaccharides in pinto bean flour

The use of dry edible beans is limited due to the presence of flatulence factors, the raffinose oligosaccharides. Our objective was to investigate the process for the removal of oligosaccharides from pinto bean using enzymatic treatment and to compare it to removal by soaking and cooking methods. Crude enzyme preparation was produced by six fungal species on wheat bran- and okara-based substrates with soy tofu whey. The loss of raffinose oligosaccharides after soaking pinto beans for 16 h at the room temperature was 10%, after cooking for 90 min was 52%, and after autoclaving for 30 min was 58%. On the other hand, the treatment using crude alpha-galactosidase (60 U mL(-1)) produced by Aspergillus awamori NRRL 4869 from wheat bran-based substrate with soy tofu whey on pinto bean flour for 2 h completely hydrolyzed raffinose oligosaccharides. These results supported that the enzymatic treatment was the most effective among various processing methods tested for removing the raffinose oligosaccharides, and hence, crude alpha-galactosidases from fungi have potential use in the food industry.     

Digestion residues of typical and high-beta-glucan oat flours provide substrates for in vitro fermentation

In vitro fermentabilities of the oat flour digestion residues (ODR) from two commercial oat lines with 4.7 and 5.3% beta-glucan and from two high-beta-glucan experimental lines with 7.6 and 8.1% beta-glucan were evaluated and compared with fermentations of lactulose, purified oat beta-glucan (POBG), and purified oat starch (POS). Substrates were fermented by using an in vitro batch fermentation system under anaerobic conditions for 24 h. The progress of the fermentation was studied by following the change in pH of the fermentation medium, production of short-chain fatty acids (SCFA) and gases, and consumption of carbohydrates. The substrate from the flour with the greatest amount of beta-glucan tended to have the greatest pH decline and the greatest total SCFA production. A significant correlation occurred between gas production and SCFA formation (R 2 = 0.89-0.99). Acetate was produced in the greatest amounts by all of the substrates except POBG, by which butyrate was produced in the greatest amount. More propionate and butyrate, but less acetate, were produced from high-beta-glucan ODR. With the given fermentation conditions, >80% of the total carbohydrate was depleted by the bacteria after 24 h. Glucose was the most rapidly consumed carbohydrate among other available monosaccharides in the fermentation medium. Overall, the high-beta-glucan experimental lines provided the best conditions for optimal in vitro gut fermentations.     

Metabolism of anthocyanins by human gut microflora and their influence on gut bacterial growth

Consumption of anthocyanins has been related with beneficial health effects. However, bioavailability studies have shown low concentration of anthocyanins in plasma and urine. In this study, we have investigated the bacterial-dependent metabolism of malvidin-3-glucoside, gallic acid and a mixture of anthocyanins using a pH-controlled, stirred, batch-culture fermentation system reflective of the distal human large intestine conditions. Most anthocyanins have disappeared after 5 h incubation while gallic acid remained constant through the first 5 h and was almost completely degraded following 24 h of fermentation. Incubation of malvidin-3-glucoside with fecal bacteria mainly resulted in the formation of syringic acid, while the mixture of anthocyanins resulted in formation of gallic, syringic and p-coumaric acids. All the anthocyanins tested enhanced significantly the growth of Bifidobacterium spp. and Lactobacillus-Enterococcus spp. These results suggest that anthocyanins and their metabolites may exert a positive modulation of the intestinal bacterial population.     

腌制叶用芥菜发酵菌分离鉴定及应用研究

为获得发酵性能优良的菌株,生产优质的发酵叶用芥菜,本研究从自然发酵的芥菜中分离发酵用菌株,通过测定乳酸菌的生长、产酸和亚硝酸盐降解能力以及酵母菌的产气、产酯能力和蛋白酶活性等发酵特性进行筛选,并对筛选获得的优良发酵菌加以应用。结果表明,筛选获得的2株乳酸菌L8短乳杆菌(Lactobacillus brevis)、L9植物乳杆菌(Lactobacillus plantarum)和1株酵母菌Y9酿酒酵母菌(Saccharomyces cerevisiae)在腌制叶用芥菜中的发酵性能优良。与自然发酵的叶用芥菜相比,接菌发酵叶用芥菜的pH值和亚硝酸盐含量分别减少了2.22%和88.13%;总酸含量、氨基酸态氮含量分别增加了9.02%和28.09%。接菌发酵提高了腌制芥菜的营养品质和安全性。本研究丰富了叶用芥菜的发酵菌种,为蔬菜发酵加工产业奠定了技术基础。     

Evaluation of oligosaccharide synthesis from lactose and lactulose using β-galactosidases from Kluyveromyces isolated from artisanal cheeses

The β-galactosidase activity of 15 Kluyveromyces strains isolated from cheese belonging to Kluyveromyces lactis and Kluyveromyces marxianus species was tested for the production of oligosaccharides derived from lactose (GOS) and lactulose (OsLu). All Kluyveromyces crude cell extracts (CEEs) produced GOS, such as 6-galactobiose and 3'-, 4'-, and 6'-galactosyl-lactose. At 4 h of reaction, the main trisaccharide formed was 6'-galactosyl-lactose (20 g/100 g of total carbohydrates). The formation of OsLu was also observed by all CEEs tested, with 6-galactobiose, 6'-galactosyl-lactulose, and 1-galactosyl-lactulose being found in all of the reaction mixtures. The synthesis of trisaccharides predominated over other oligosaccharides. K. marxianus strain O3 produced the highest yields of GOS and OsLu after 4 h of reaction, reaching 42 g/100 g of total carbohydrates (corresponding to 80% lactose hydrolysis) and 45 g/100 g of total carbohydrates (corresponding to 87% lactulose hydrolysis), respectively. Therefore, the present study contributes to a better insight into dairy Kluyveromyces β-galactosidases and shows the feasibility of these enzymes to transglycosylate lactose and lactulose, producing high yields of prebiotic oligosaccharides.     

Rheological Changes in Wheat Sourdough During Controlled and Spontaneous Fermentation

The changing rheological characteristics of wheat doughs during fermentation at 30°C for 72 hr were measured using a controlled stress rheometer. Dynamic oscillation tests were performed at frequencies ranging from 0.01 to 10 Hz. Wheat sourdoughs (dough yield 200) were prepared with a mixed starter culture containing typical hetero- and homofermentative sourdough lactic acid bacteria. Results from the controlled fermentation process were compared to results from spontaneous fermentation. Maximum phase angle values, especially at low frequencies, were closely related to total gas production in the doughs. Complex viscosity decreased during fermentation and reached lower final values for doughs without starter culture. Heating characteristics of doughs after various fermentation times were measured at temperatures ranging from 30 to 80°C. The highest values for complex viscosity were found at ≈65°C. When heated, fermented doughs produced weaker gels than fresh doughs. The temperatures at which these maxima occurred increased significantly with fermentation time for spontaneously fermented dough.