raffinose and stachyose

A large number of soybean components have diverse biological activities. These include hormonal, immunological, bacteriological and digestive effects. The presently known allergens are listed. The divergence between chemical evaluation and biological value is highlighted. The following components are discussed: Kunitz inhibitor, Bowman-Birk inhibitor, saponins, soyacystatin, phytoestrogens (daidzein, glycitein, genistein), Maillard products, soybean hydrophobic protein, soy allergens, lecithin allergens, raffinose, stachyose, 2-pentyl pyridine. The studies describing the effects of the isolated components are reviewed.

Topics: Allergens; Animal Feed; Animals; Cystatins; Glycine max; Maillard Reaction; Oligosaccharides; Phosphatidylcholines; Phytoestrogens; Plant Lectins; Raffinose; Saponins; Soybean Proteins; Trypsin Inhibitors

Research was conducted to determine the effects of enzymatically hydrolyzing raffinose and stachyose from soybean meal (SBM) on fecal oligosaccharide concentration and growth performance of chicks fed a corn-SBM diet. The alpha-galactosidase treatment was optimized for oligosaccharide degradation. Enzyme treatment degraded raffinose and stachyose in SBM by 69 and 54%, respectively, compared to untreated soybean meal (USBM). Diets containing enzyme-treated soybean meal (ESBM) resulted in excreta raffinose and stachyose concentrations reduced to below measurable levels (<0.1 mg/ g feces). Enzyme treatment increased (P < 0.05) TME from 2,974 to 3,328 kcal/kg. Three chick growth studies were conducted to determine the effect of feeding ESBM on growth performance. There were no statistical differences (P > 0.05) in growth performance among treatments. Chicks fed the ESBM diet had an increased (P < 0.05) fecal neutral detergent fiber (NDF) content in one of two studies. A fourth experiment was conducted to determine if heating, used to enhance enzyme treatment, would decrease lysine availability. Heating significantly (P < 0.05) reduced lysine availability compared to USBM. These experiments demonstrated that feces could be made void of raffinose and stachyose, but chick growth performance was not significantly (P > 0.05) improved by enzyme treatment.

Topics: alpha-Galactosidase; Animal Feed; Animals; Chickens; Dietary Fiber; Digestion; Feces; Food Handling; Glycine max; Hydrolysis; Male; Oligosaccharides; Raffinose; Solutions

Adhering to a galactose-free diet by strictly avoiding dairy products and known hidden sources of galac-tose does not completely normalize galactose-1-phosphate (gal-1-P) in erythrocytes from patients with galactosemia. Major neurological complications, even in the best treated patients, are threatening a good clinical outcome and dictate a continuous search for leaks in the dietary regimen. Raffinose and stachyose, present in important amounts in various vegetables, contain alpha-1,4 linked galactose which is cleaved only by bacterial alpha-galactosidases, presumably in the lower part of the gut. In order to test the hypothesis whether galactose released from raffinose and stachyose could be a source of absorbed galactose and a cause of elevated gal-1-P six patients with galactosemia (aged 6-24 years), underwent a raffinose- and stachyose-poor dietary regimen for 2 weeks. Before, after, and during the test period, the daily intake of stachyose and raffinose as well of protein, carbohydrate, fat and minerals was calculated from food protocols obtained from the patients. Plasma galactose and erythrocyte gal-1-P were measured at the end of the three test phases. Stachyose and raffinose intake was reduced to 5%-10% during the experimental diet, which was well tolerated, except for constipation in some patients. In five of the six patients gal-1-P in erythrocytes was somewhat lower (statistically not significant) during the test phase than during regular diet while plasma galactose remained unchanged. Galactose released from raffinose and stachyose may be absorbed and contribute to elevated gal-1-P values in erythrocytes of galactosemic patients.

Topics: Adult; Child; Fabaceae; Female; Galactose; Galactosemias; Galactosephosphates; Humans; Oligosaccharides; Plants, Medicinal; Raffinose

Photosynthetic active radiation (PAR) refers to photons between 400 and 700 nm. These photons drive photosynthesis, providing carbohydrates for plant metabolism and development. Far-red radiation (FR, 701-750 nm) is excluded in this definition because no FR is absorbed by the plant photosynthetic pigments. However, including FR in the light spectrum provides substantial benefits for biomass production and resource-use efficiency. We investigated the effects of continuous FR addition and end-of-day additional FR to a broad white light spectrum (BW) on carbohydrate concentrations in the top and bottom leaves of sweet basil (

Topics: Carbohydrates; Ocimum basilicum; Oligosaccharides; Plant Leaves; Plants; Raffinose; Sucrose

A Raffinose family oligosaccharides (RFOs) is one of the major translocated sugars in the vascular bundle of cucumber, but little RFOs can be detected in fruits. Alpha-galactosidases (α-Gals) catalyze the first catabolism step of RFOs. Six α-Gal genes exist in a cucumber genome, but their spatial functions in fruits remain unclear. Here, we found that RFOs were highly accumulated in vascular tissues. In phloem sap, the stachyose and raffinose content was gradually decreased, whereas the content of sucrose, glucose and fructose was increased from pedicel to fruit top. Three alkaline forms instead of acid forms of

Topics: alpha-Galactosidase; Cucumis sativus; Fruit; Gene Expression Regulation, Plant; Oligosaccharides; Phloem; Promoter Regions, Genetic; Raffinose; Substrate Specificity

Alternative polyadenylation (APA) is a pervasive mechanism for gene regulation in eukaryotes. Stachyose is the main assimilate translocated in the cucumber phloem. Stachyose synthase (CsSTS) catalyzes the last step of stachyose biosynthesis in cucumber leaves and plays a key role in the regulation of assimilate partitioning between source and sink. In this study, three CsSTS mRNAs with the same open reading frame and the 5`untranslated region (UTR), but differing in their 3`UTRs, named CsSTS1 (short), CsSTS2 (medium), and CsSTS3 (long), were identified. Southern blot and sequence analysis of the cucumber genome confirmed that these transcripts are regulated through APA from a single gene. No significant difference of in vitro translation efficiency was found among three mRNAs. However, the relative stabilities of three transcripts varied among different tissues and different leaf development stages of cucumber. CsSTS1 expression in cucumber calli was up-regulated by the raffinose (substrate of CsSTS) and down-regulated by stachyose (product of CsSTS), respectively. In cucumber plants, all three isoforms have considerable expression in non-fruit node leaves. However, in fruit-carrying node leaves, the expression of CsSTS2 and CsSTS3 was severely inhibited and only CsSTS1 was highly expressed, indicating fruit setting has a remarkable effect on the relative expression level of three transcripts. This "fruit setting" effect could be observed until at least 36 h after the fruit was removed from the node. Our results suggest that abundant expression of CsSTS1 is beneficial for stachyose loading in source leaves, and APA is a delicate mechanism for CsSTS to regulate cucumber source-sink balance.

Topics: 3' Untranslated Regions; Carbohydrate Metabolism; Cucumis sativus; Fruit; Galactosyltransferases; Gene Expression Regulation, Plant; Oligosaccharides; Phloem; Plant Leaves; Plant Proteins; Polyadenylation; Raffinose; RNA Isoforms

Soybean oligosaccharides have been previously shown to be associated with the production of major odor-causing compounds in broilers, although little is known about the role of stachyose and raffinose, which are key components of soybean oligosaccharide, in broiler cecal microbiota and odor compound production. To this end, soybean oligosaccharide, stachyose, and raffinose were added to the birds' diets to investigate their effects on odor compound production and the microbial community characteristics of the cecum in broilers. A total of 300 one-day-old Arbor Acre broilers with similar initial live weight were randomly allocated into 5 dietary groups with 6 replicates of 10 birds. The diets included soybean meal (positive control), soybean meal-free (negative control), 0.6% soybean oligosaccharide, 0.6% stachyose, or 0.6% raffinose. After a 49-D feeding period, both ceca were aseptically removed postmortem, and the contents were collected and analyzed for skatole, indole, volatile fatty acids, and lactic acid by using high performance liquid chromatography. Bacterial communities were detected by using a high-throughput sequencing platform based on IlluminaMiSeq 2500. Levels of skatole and indole tended to be lower in the dietary supplementation of oligosaccharides. The lowest levels of skatole and indole were observed in the stachyose group (P < 0.05), while the highest levels were found in the negative control group (P < 0.05). Concentrations of acetic acid and propionic acid in the stachyose group were increased (P < 0.05) while those of butyric acid and lactic acid were decreased (P < 0.05) compared with the soybean oligosaccharide and raffinose groups. Firmicutes and Bacteroidetes were prevalent in all groups, the proportion of Bacteroidetes was slightly decreased in the stachyose group, and Verrucomicrobia was abundant in the raffinose group (P > 0.05). Bacterial genera Alistipes and Parabacteroides were comparably abundant in the stachyose group, while Bacteroides, Lactobacillus, and Akkermansia were more abundant in the negative control, stachyose, and raffinose groups, respectively. Collectively, these findings demonstrated that dietary oligosaccharide supplementation significantly reduced odor compound production by modulating the cecal microbial community. Compared with soybean oligosaccharide and raffinose, the addition of stachyose into diets may help improve gut fermentation and minimize odor compound generation in broilers.

Topics: Animal Feed; Animals; Cecum; Chickens; Diet; Dietary Supplements; Dose-Response Relationship, Drug; Gastrointestinal Contents; Gastrointestinal Microbiome; Glycine max; Odorants; Oligosaccharides; Raffinose; Random Allocation

Strains of

Topics: Animals; Galactose; Gastrointestinal Tract; Genome, Bacterial; Germ-Free Life; Lac Operon; Lactose; Limosilactobacillus reuteri; Mice; Mutation; Oligosaccharides; Probiotics; Raffinose; Synbiotics

Soybean seeds produce valuable protein that is a major component of livestock feed. However, soybean seeds also contain the anti-nutritional raffinose family oligosaccharides (RFOs) raffinose and stachyose, which are not digestible by non-ruminant animals. This requires the proportion of soybean meal in the feed to be limited, or risk affecting animal growth rate or overall health. While reducing RFOs in soybean seed has been a goal of soybean breeding, efforts are constrained by low genetic variability for carbohydrate traits and the difficulty in identifying these within the soybean germplasm. We used reverse genetics Targeting Induced Local Lesions in Genomes (TILLING)-by-sequencing approach to identify a damaging polymorphism that results in a missense mutation in a conserved region of the

Topics: Alleles; Galactosyltransferases; Genetics, Population; Glycine max; Oligosaccharides; Plant Breeding; Polymorphism, Genetic; Raffinose; Seeds; Sequence Analysis, DNA; Soybean Proteins

Legumes are widely consumed by humans, being an important source of nutrients; however, they contain non-nutritional factors (NNFs), such as phytic acid (IP

Topics: Cooking; Flavonoids; Food Handling; Germination; Nutritive Value; Oligosaccharides; Phenols; Proanthocyanidins; Raffinose; Seeds; Vicia sativa

The α-galactosidases, which can catalyze the removal of α-1,6-linked terminal galactose residues from galactooligosaccharide materials, have good potential for industrial applications. The high-level and efficient secretion of the α-galactosidases into the extracellular space has greatly simplified the downstream bioengineering process, facilitating their bioapplications. In this study, the effects of gene dosage and endoplasmic reticulum secretion-associated factors (ERSAs) on the secretory expression of an α-galactosidase gene derived from a Aspergillus oryzae strain were investigated by constructing multicopy expression cassettes and coexpressing the α-galactosidase gene with ERSAs. With the increase in the gene copy-number in the host genome, the expression of GalA was improved. However, the secretory expression level was not linearly related to the copy number. When the number was higher than four copies, the expression level of GalA gene declined. The ERSAs factors HAC1, PDI, and Ero1 improved the secretory expression of α-galactosidase, while Hsp40 inhibited its secretion. After methanol-induced expression in a bench-top bioreactor, Pichia recombinants carrying four copies of GalA genes reached 3520 U/mL in the supernatant of the culture. We further optimized the parameters for α-galactosidase to hydrolyze two types of galactooligosaccharides: raffinose and stachyose. This study has fulfilled the scale-up production of α-galactosidase, thus facilitating its industrial applications.

Topics: alpha-Galactosidase; Aspergillus oryzae; Basic-Leucine Zipper Transcription Factors; Cloning, Molecular; Endoplasmic Reticulum; Fungal Proteins; Gene Dosage; Gene Expression; Genetic Vectors; Glycoproteins; Humans; Hydrolysis; Industrial Microbiology; Oligosaccharides; Oxidoreductases Acting on Sulfur Group Donors; Pichia; Protein Disulfide-Isomerases; Raffinose; Recombinant Proteins; Repressor Proteins; Saccharomyces cerevisiae Proteins

Topics: alpha-Galactosidase; Bacillus subtilis; Bacterial Proteins; Galactosides; Melibiose; Oligosaccharides; Operon; Raffinose; Sucrose

A modelling approach was developed to better understand the behavior of the flatulence-causing oligosaccharides in cowpea seeds during isothermal water soaking-cooking process. Concentrations of verbascose, stachyose and raffinose were measured both in the seed and in the soaking water during the process (T=30, 60 and 95°C). A reaction-diffusion model was built for the three considered alpha-galactosides both in the seed and in the soaking water, together with a model of water transport in the seed. The model reproduced coupled reaction-diffusion of alpha-galactosides during the soaking-cooking process with a good fit. Produced, diffused and degraded alpha-galactoside fractions were identified by performing a mass balance. During soaking at 30°C, degradation predominated (maximum found for raffinose degradation rate constant of 3.22×10

Topics: Carbohydrate Metabolism; Cooking; Diffusion; Galactosides; Hot Temperature; Models, Theoretical; Oligosaccharides; Raffinose; Seeds; Transition Temperature; Vigna; Water

The prebiotic effects of GOS (galactooligosaccharides) are known to depend on the glycosidic linkages, degree of polymerization (DP), and the monosaccharide composition. In this study, a novel form of α-GOS with a potentially improved prebiotic effect was synthesized using bifidobacterial α-galactosidase (α-Gal) purified from recombinant Escherichia coli. The carbohydrate produced was identified as α-d-galactopyranosyl-(1→6)-O-α-d-glucopyranosyl-(1→2)-[α-d-galactopyranosyl-(1→6)-O-β-d-fructofuranoside] and was termed stachyobifiose. Among 17 nonprobiotics, 16 nonprobiotics showed lower growth on stachyobifiose than β-GOS. In contrast, among the 16 probiotics, 6 probiotics showed higher growth on stachyobifiose than β-GOS. When compared with raffinose, stachyobifiose was used less by nonprobiotics than raffinose. Moreover, compared with stachyose, stachyobifiose was used less by Escherichia coli, Enterobacter cloacae, and Clostridium butyricum. The average amounts of total short-chain fatty acids (SCFA) produced were in the order of stachyobifiose > stachyose > raffinose > β-GOS. Taken together, stachyobifiose is expected to contribute to beneficial changes of gut microbiota.

Topics: alpha-Galactosidase; Bacteria; Bifidobacterium; Escherichia coli; Galactose; Gastrointestinal Microbiome; Oligosaccharides; Prebiotics; Probiotics; Raffinose; Recombinant Proteins

New eating habits and diversification of tastes of consumers have led to the scientific community and the food industry to expand the range of probiotic foods and novel probiotic ingredients. Scant information is available about the viability and functionality of probiotics during shelf life and its effect on the nutritional characteristics of dairy-free products. The aim of the study was to formulate a fermented dairy-free dessert using a novel food ingredient based on a pumpkin by-product and containing Lactobacillus casei (ATCC®393™) (NFI). The effect of NFI and the soluble solids (SS) of soy milk on the probiotic viability, physical stability, colour, and firmness of dairy-free dessert was studied using a response surface methodology. The different levels of SS and NFI significantly (p < 0.05) affected the response variables. Thereafter, two formulations were selected and the physico-chemical, nutritional and organoleptic characterization were evaluated. The L. casei count reached the desired therapeutic level (>107 UFC mL-1) after gastrointestinal digestion at 21 days of storage. In general, both the fermentation process and storage reduced (p < 0.05) the content of phytic acid, raffinose and stachyose, which implies a nutritional improvement of the final product. Scores above 5.0 on a 9-point scale were obtained for colour, odour, texture and overall acceptability in the consumer acceptance test. Therefore, a dairy-free dessert with good physical properties, suitable nutritional characteristics, and sensorial acceptability could be successfully formulated with the NFI.

Topics: Adult; Chemical Phenomena; Consumer Behavior; Female; Fermentation; Food Analysis; Food Handling; Food Ingredients; Food Microbiology; Humans; Lacticaseibacillus casei; Male; Microbial Viability; Middle Aged; Nutritive Value; Oligosaccharides; Phytic Acid; Probiotics; Raffinose; Soy Milk; Taste; Young Adult

This study investigates the effectiveness of two types of prebiotics-stachyose and raffinose-which are present in staple food crops that are widely consumed in regions where dietary Fe deficiency is a health concern. The hypothesis is that these prebiotics will improve Fe status, intestinal functionality, and increase health-promoting bacterial populations in vivo (

Topics: Animals; Bifidobacterium; Biological Availability; Chickens; Clostridium; Disease Models, Animal; Escherichia coli; Ferritins; Gastrointestinal Microbiome; Intestinal Mucosa; Intestines; Iron; Lactobacillus; Liver; Microvilli; Oligosaccharides; Prebiotics; Probiotics; Raffinose

The putative gene cluster involved in the degradation of the raffinose family oligosaccharides (RFO) was identified in Caldicellulosiruptor bescii. Within the cluster, the gene encoding a putative α-galactosidase (CbAga36) was cloned and expressed in Escherichia coli. Size exclusion chromatography of the purified rCbAga36 indicated that the native form was a tetramer. Its primary sequence was similar to the family of glycoside hydrolase 36. The purified recombinant CbAga36 (rCbAga36) was optimally active at pH 5.0 and 70°C and had a half-life of 15 h and 10 h at 70°C and 80°C, respectively. rCbAga36 showed high activity with the artificial substrate (p-nitrophenyl α-d-galactopyranoside, pNPαGal) exhibiting lower K

Topics: alpha-Galactosidase; Enzyme Stability; Escherichia coli; Galactose; Glycolysis; Glycosides; Gram-Positive Bacteria; Half-Life; Hydrogen-Ion Concentration; Hydrolysis; Kinetics; Melibiose; Oligosaccharides; Raffinose; Substrate Specificity

In many species, Suc en route out of the leaf migrates from photosynthetically active mesophyll cells into the phloem down its concentration gradient via plasmodesmata, i.e. symplastically. In some of these plants, the process is entirely passive, but in others phloem Suc is actively converted into larger sugars, raffinose and stachyose, and segregated (trapped), thus raising total phloem sugar concentration to a level higher than in the mesophyll. Questions remain regarding the mechanisms and selective advantages conferred by both of these symplastic-loading processes. Here, we present an integrated model-including local and global transport and kinetics of polymerization-for passive and active symplastic loading. We also propose a physical model of transport through the plasmodesmata. With these models, we predict that (1) relative to passive loading, polymerization of Suc in the phloem, even in the absence of segregation, lowers the sugar content in the leaf required to achieve a given export rate and accelerates export for a given concentration of Suc in the mesophyll and (2) segregation of oligomers and the inverted gradient of total sugar content can be achieved for physiologically reasonable parameter values, but even higher export rates can be accessed in scenarios in which polymers are allowed to diffuse back into the mesophyll. We discuss these predictions in relation to further studies aimed at the clarification of loading mechanisms, fitness of active and passive symplastic loading, and potential targets for engineering improved rates of export.

Topics: Biological Transport; Biophysics; Cucumis melo; Malus; Mesophyll Cells; Oligosaccharides; Phloem; Plant Leaves; Plasmodesmata; Raffinose; Xylem

An acidic α-galactosidase designated as PDGI (Pleurotus djamor α-galactosidase) was purified to homogeneity with 290-fold purification and a specific activity of 52.18 units/mg by means of ion exchange chromatography and gel filtration chromatography. PDGI is a monomeric protein exhibiting a molecular mass of 60kDa in SDS-PAGE and gel filtration. The optimum pH and temperature of the enzyme with pNPGal as substrate were 5.0 and 53.5°C, respectively. It displayed great pH stability within the pH range 3.0-10.0. Besides, the enzyme harbored remarkable resistance to acid protease and varying degrees of tolerance to other proteases: trypsin>collagenase Type-I>α-chymotrypsin neutral protease>proteinaseK. It was strongly inhibited by K

Topics: alpha-Galactosidase; Amino Acid Sequence; Chymotrypsin; Enzyme Stability; Fungal Proteins; Hydrogen-Ion Concentration; Hydrolysis; Kinetics; Metals; Molecular Weight; Oligosaccharides; Pleurotus; Raffinose; Substrate Specificity

To understand raffinose family oligosaccharides (RFO) metabolism in chickpea (Cicer arietinum L.) seeds, RFO accumulation and corresponding biosynthetic enzymes activities were determined during seed development of chickpea genotypes with contrasting RFO concentrations. RFO concentration in mature seeds was found as a facilitator rather than a regulating step of seed germination. In mature seeds, raffinose concentrations ranged from 0.38 to 0.68 and 0.75 to 0.99 g/100 g, whereas stachyose concentrations varied from 0.79 to 1.26 and 1.70 to 1.87 g/100 g indicating significant differences between low and high RFO genotypes, respectively. Chickpea genotypes with high RFO concentration accumulated higher concentrations of myo-inositol and sucrose during early seed developmental stages suggesting that initial substrate concentrations may influence RFO concentration in mature seeds. High RFO genotypes showed about two to three-fold higher activity for all RFO biosynthetic enzymes compared to those with low RFO concentrations. RFO biosynthetic enzymes activities correspond with accumulation of individual RFO during seed development.

Topics: Cicer; Galactosyltransferases; Genotype; Oligosaccharides; Raffinose; Seeds; Sucrose

Dietary restriction of fermentable oligosaccharides, disaccharides, monosaccharides and polyols (FODMAPs) is an effective management approach for functional bowel disorders; however, its application is limited by the paucity of food composition data available for ethnic minority groups. The aim was to identify and measure the FODMAP content of these commonly consumed foods. According to their perceived importance to clinical practise, the top 20 ranked foods underwent FODMAP analysis using validated analytical techniques (total fructans, Megazyme hexokinase (HK) assay; all others, high-performance liquid chromatography (HPLC) with evaporative light scattering detectors). Of the 20 foods analysed, five were identified as significant sources of at least one FODMAP. Fructans and galacto-oligosaccharides were the major FODMAPs in these foods, including channa dal (0.13 g/100 g; 0.36 g/100 g), fenugreek seeds (1.11 g/100 g; 1.27 g/100 g), guava (0.41 g/100 g; not detected), karela (not detected; 1.12 g/100 g) and tamarind (2.35 g/100 g; 0.02 g/100 g). Broadening the availability of FODMAP composition data will increase the cultural application of low FODMAP dietary advice.

Topics: Asian People; Black People; Caribbean Region; Culturally Competent Care; Diet; Diet Surveys; Diet, Carbohydrate-Restricted; Disaccharides; Fermentation; Focus Groups; Food Analysis; Fructans; Humans; Internet; Minority Groups; Monosaccharides; Nutritionists; Oligosaccharides; Raffinose; Sugar Alcohols; United Kingdom

The impact of extrusion cooking on the chemical composition and functional properties of bean powders from four common bean varieties was investigated. The raw bean powders were extruded under eight different conditions, and the extrudates were then dried and ground (particle size⩽0.5mm). Compared with corresponding non-extruded (raw) bean powders (particle size⩽0.5mm), the extrusion treatments did not substantially change the protein and starch contents of the bean powders and showed inconsistent effects on the sucrose, raffinose and stachyose contents. The extrusion cooking did cause complete starch gelatinization and protein denaturation of the bean powders and thus changed their pasting properties and solvent-retention capacities. The starch digestibilities of the cooked non-extruded and cooked extruded bean powders were comparable. The extruded bean powders displayed functional properties similar to those of two commercial bean powders.

Topics: Cooking; Gels; Oligosaccharides; Phaseolus; Plant Proteins; Powders; Raffinose; Seeds; Starch

Cryopreservation of human spermatozoa is a commonly used technique in assisted reproduction, however freezing low concentrations of sperm while maintaining adequate post-thaw motility remains a challenge. In an effort to optimize post-thaw motility yields, low volumes of human sperm were frozen in polyimide-coated fused silica micro-capillaries using 0.065 M, 0.125 M, 0.25 M, or 0.5 M trehalose as the only cryoprotectant. Micro-capillaries were either initially incubated in liquid nitrogen vapor before plunging into liquid nitrogen, or directly plunged into liquid nitrogen. Post thaw sperm counts and motility were estimated. Spermatozoa that were initially incubated in liquid nitrogen vapor had greater post thaw motility than those plunged immediately into liquid nitrogen independent of trehalose concentration. The protective effect of 0.125 M d-glucose, 3-O-methyl-d-glucopyranose, trehalose, sucrose, raffinose, or stachyose were evaluated individually. Trehalose and sucrose were the most effective cryoprotectants, recovering 69.0% and 68.9% of initial sperm motility, respectively.

Topics: 3-O-Methylglucose; Animals; Cryopreservation; Cryoprotective Agents; Freezing; Glucose; Humans; Male; Oligosaccharides; Raffinose; Semen Preservation; Sperm Motility; Spermatozoa; Sucrose; Trehalose

In Ajuga reptans, raffinose oligosaccharides accumulated during winter. Stachyose, verbascose, and higher RFO oligomers were exclusively found in the vacuole whereas one-fourth of raffinose was localized in the stroma. The evergreen labiate Ajuga reptans L. can grow at low temperature. The carbohydrate metabolism changes during the cold phase, e.g., raffinose family oligosaccharides (RFOs) accumulate. Additionally, A. reptans translocates RFOs in the phloem. In the present study, subcellular concentrations of metabolites were studied in summer and winter leaves of A. reptans to gain further insight into regulatory instances involved in the cold acclimation process and into the function of RFOs. Subcellular metabolite concentrations were determined by non-aqueous fractionation. Volumes of the subcellular compartments of summer and winter leaves were analyzed by morphometric measurements. The metabolite content varied strongly between summer and winter leaves. Soluble metabolites increased up to tenfold during winter whereas the starch content was decreased. In winter leaves, the subcellular distribution showed a shift of carbohydrates from cytoplasm to vacuole and chloroplast. Despite this, the metabolite concentration was higher in all compartments in winter leaves compared to summer leaves because of the much higher total metabolite content in winter leaves. The different oligosaccharides did show different compartmentations. Stachyose, verbascose, and higher RFO oligomers were almost exclusively found in the vacuole whereas one-fourth of raffinose was localized in the stroma. Apparently, the subcellular distribution of the RFOs differs because they fulfill different functions in plant metabolism during winter. Raffinose might function in protecting chloroplast membranes during freezing, whereas higher RFO oligomers may exert protective effects on vacuolar membranes. In addition, the high content of RFOs in winter leaves may also result from reduced consumption of assimilates.

Topics: Adaptation, Physiological; Ajuga; Biological Transport; Carbohydrate Metabolism; Chloroplasts; Cold Temperature; Cytoplasm; Freezing; Microscopy, Electron, Transmission; Oligosaccharides; Plant Leaves; Raffinose; Seasons; Subcellular Fractions

The second α-galactosidase gene (designated as RmgalB) was cloned from the thermophilic fungus Rhizomucor miehei and expressed in Pichia pastoris. The gene belonging to glycoside hydrolase (GH) family 36 has an open reading frame (ORF) of 2241bp encoding 746 amino acids with two introns. The recombinant α-galactosidase (RmgalB) was secreted at high levels of 1953.9Uml(-1) in high cell density fermentor, which is the highest yield obtained for a α-galactosidase. The purified enzyme as a tetramer gave a single band corresponding to a molecular mass of 83.1kDa in SDS-PAGE. The enzyme exhibited a very high specific activity of 505.5Umg(-1). The optimum temperature and pH of RmgalB were determined to be 55°C and pH 5.5, respectively. It was stable within pH 5.5-9.5 and up to 55°C. RmgalB displayed specificity toward raffinose and stachyose, and completely hydrolyzed the anti-nutritive raffinose family oligosaccharides (RFOs). These properties make RmgalB useful in the food and feed industries.

Topics: alpha-Galactosidase; Amino Acid Sequence; Base Sequence; Cloning, Molecular; Enzyme Stability; Fermentation; Fungal Proteins; Gene Expression; Hydrogen-Ion Concentration; Introns; Kinetics; Molecular Sequence Data; Oligosaccharides; Open Reading Frames; Pichia; Plasmids; Protein Multimerization; Raffinose; Recombinant Proteins; Rhizomucor; Substrate Specificity; Temperature

We identified and characterized a mutant of soybean stachyose synthase gene controlling reduced stachyose content which benefit the soybean seed composition breeding program in the future. It has been shown that in soybean, increased sucrose and reduced raffinose family oligosaccharides would have a positive impact on the world's feed industry by improving digestibility and feed efficiency. We searched for new sources of modified oligosaccharide content in a subset of the USDA Soybean Germplasm Collection and then identified plant introduction (PI) 603176A as having ultra-low stachyose content (0.5%). We identified a 33-bp deletion mutant in the putative stachyose synthase gene (STS gene, Glyma19g40550) of PI 603176A. A co-dominate indel marker was successfully developed from this 33-bp deletion area and was genetically mapped into two F 2:3 populations and a F 4:5 population, which associated with low stachyose content in the progeny lines. These observations provided strong evidence that the STS gene is responsible for stachyose biosynthesis in the soybean plant. Expression of the sts gene remained at the normal level, suggesting the loss of function in the gene is due to defective protein function. This gene-based perfect genetic marker for low stachyose content can be useful for marker-assisted selection in soybean molecular breeding programs.

Topics: Chromosome Mapping; DNA, Plant; Galactosyltransferases; Genetic Markers; Glycine max; INDEL Mutation; Oligosaccharides; Plant Breeding; Plant Proteins; Raffinose; Seeds; Sequence Analysis, DNA; Sequence Deletion

This work deals with the development of a new bioprocess for the efficient synthesis of lactosucrose, a potential prebiotic oligosaccharide with a high value-added, from two important and inexpensive agro-industrial by-products such as tofu whey and cheese whey permeate. The bioconversion is driven by the ability of the enzyme levansucrase SacB from Bacillus subtilis CECT 39 to transfructosylate lactose contained in the cheese whey permeate by using not only sucrose but also raffinose and stachyose, which are present in considerable amounts in the tofu whey, as suitable donors of fructosyl moieties. The maximum lactosucrose concentration obtained from both by-products was 80.1 g L-1 after a short reaction time 120 min at 37°C, leading to productivity and specific productivity values of 40.1 g lactosucrose L-1 h-1 and 80.1 mg lactosucrose U enzyme-1 h-1, respectively. Findings contained in this work could provide a new strategy to valorize agro-industrial by-products as cheese whey permeate and, specially, tofu whey by means of their use as renewable resources in the enzymatic synthesis of bioactive oligosaccharides.

Topics: Cheese; Chromatography, High Pressure Liquid; Gas Chromatography-Mass Spectrometry; Hexosyltransferases; Lactose; Melibiose; Oligosaccharides; Raffinose; Refractometry; Soy Foods; Sucrose; Trisaccharides; Whey

The objective of this study was to evaluate the changes in oligosaccharides and isoflavone aglycone content in soymilk during fermentation with commercial kefir culture. Soymilk was fermented with kefir culture at 25 °C for 30 h. The counts of lactic acid bacteria, Lactococcus lactis, Leuconostoc sp and yeasts; measurements of pH, acidity, α-galactosidase and β-glucosidase activity, sugar and isoflavone contents were performed at the intervals of time. In the fermented soymilk, the lactic acid bacteria counts increased from 7.6 log to 9.1 CFU g(-1), pH reached to 4.9 and lactic acid reached 0.34 g 100  g(- 1). The α-galactosidase was produced (0.016 AU g(-1)) with 100% raffinose and 92% stachyose hydrolysis being observed after the depletion of galactose, glucose and sucrose. Kefir culture produced β-glucosidase (0.0164 AU g(-1)), resulting in 100% bioconversion of glycitin and daidzin and 89% bioconversion of genistin into the corresponding aglycones. The fermented soymilk presented 1.67 μmol g(-1) of daidzein, 0.28 μmol g(-1) of glicitein and 1.67 μmol g (-1) of genistein.

Topics: alpha-Galactosidase; beta-Glucans; beta-Glucosidase; Cell Survival; Colony Count, Microbial; Cultured Milk Products; Fermentation; Food Handling; Food Microbiology; Genistein; Hydrogen-Ion Concentration; Hydrolysis; Isoflavones; Lactococcus lactis; Leuconostoc; Levilactobacillus brevis; Oligosaccharides; Raffinose; Saccharomyces cerevisiae; Soy Milk

Immature seeds of five bean cultivars (flageolet-type and those intended for dry-seed production) were assessed for changes in water-soluble carbohydrates including raffinose family oligosaccharides (RFOs) due to boiling, sterilization, and storage of the sterilized product. About 100 g fresh weight of edible portion of fresh bean seeds contained 2449.3-3182.6 mg total soluble sugars, of which RFOs comprised 44-49%. The highest amounts of these compounds were found in the seeds of the cultivars Laponia and Mona. The dominant oligosaccharide was stachyose. Boiling fresh seeds to consumption consistency reduced total soluble sugars and RFOs: average values were 57% and 55%, respectively. Sterilization in cans resulted in 65% reductions of both total soluble sugars and RFOs. In general, there were no changes in the content of soluble sugars in canned and sterilized products stored for 12 months.

Topics: Carbohydrates; Cooking; Crops, Agricultural; Dietary Carbohydrates; Dietary Sucrose; Food Preservation; Food Storage; Molecular Weight; Oligosaccharides; Phaseolus; Poland; Raffinose; Seeds; Solubility; Sterilization

To develop genetic improvement strategies to modulate raffinose family oligosaccharides (RFO) concentration in chickpea ( Cicer arietinum L.) seeds, RFO and their precursor concentrations were analyzed in 171 chickpea genotypes from diverse geographical origins. The genotypes were grown in replicated trials over two years in the field (Patancheru, India) and in the greenhouse (Saskatoon, Canada). Analysis of variance revealed a significant impact of genotype, environment, and their interaction on RFO concentration in chickpea seeds. Total RFO concentration ranged from 1.58 to 5.31 mmol/100 g and from 2.11 to 5.83 mmol/100 g in desi and kabuli genotypes, respectively. Sucrose (0.60-3.59 g/100 g) and stachyose (0.18-2.38 g/100 g) were distinguished as the major soluble sugar and RFO, respectively. Correlation analysis revealed a significant positive correlation between substrate and product concentration in RFO biosynthesis. In chickpea seeds, raffinose, stachyose, and verbascose showed a moderate broad sense heritability (0.25-0.56), suggesting the use of a multilocation trials based approach in chickpea seed quality improvement programs.

Topics: Africa; Asia; Cicer; Environment; Genotype; Oligosaccharides; Raffinose; Seeds; South America; Sucrose

Members of the genus Bifidobacterium are common inhabitants of the gastrointestinal tracts of humans and other mammals, where they ferment many diet-derived carbohydrates that cannot be digested by their hosts. To extend our understanding of bifidobacterial carbohydrate utilization, we investigated the molecular mechanisms by which 11 strains of Bifidobacterium breve metabolize four distinct α-glucose- and/or α-galactose-containing oligosaccharides, namely, raffinose, stachyose, melibiose, and melezitose. Here we demonstrate that all B. breve strains examined possess the ability to utilize raffinose, stachyose, and melibiose. However, the ability to metabolize melezitose was not common to all B. breve strains tested. Transcriptomic and functional genomic approaches identified a gene cluster dedicated to the metabolism of α-galactose-containing carbohydrates, while an adjacent gene cluster, dedicated to the metabolism of α-glucose-containing melezitose, was identified in strains that are able to use this carbohydrate.

Topics: Bifidobacterium; Gene Expression Profiling; Humans; Melibiose; Metabolic Networks and Pathways; Multigene Family; Oligosaccharides; Raffinose

Dietary fiber (DF) has important health benefits in the human diet. Developing dry edible bean (Phaseolus vulgaris L.) cultivars with improved DF and reduced nondigestible oligosaccharide content is an important goal for dry bean breeders to increase consumer acceptance. To determine if genetic variation exists among dry bean cultivars for DF, two populations of diverse dry bean cultivars/lines that represent two centers of dry bean domestication were evaluated for dietary fiber using the Integrated Total Dietary Fiber Assay (AOAC 2011.25). This assay was adapted to measure water insoluble dietary fiber, water soluble dietary fiber, oligosaccharides raffinose and stachyose, and the calculated total dietary fiber (TDF) content of cooked dry bean seed. The AOAC 2011.25 protocol was modified by using a quick, simple, and sensitive high-performance liquid chromatography method paired with an electrochemical detection method to separate and quantify specific oligosaccharides, and using duplicate samples as replicates to generate statistical information. The TDF of dry bean entries ranged from 20.0 to 27.0% in population I and from 20.6 to 25.7% in population II. Total oligosaccharides ranged from 2.56 to 4.65% in population I and from 2.36 to 3.84% in population II. The results suggest that significant genetic variation exists among dry bean cultivars/lines to allow for genetic selection for improved DF content in dry beans and that the modifications to the AOAC 2011.25 method were suitable for estimating DF in cooked dry edible beans.

Topics: alpha-Amylases; Cooking; Dietary Fiber; Fabaceae; Oligosaccharides; Raffinose; Seeds

Galactinol synthase (GolS, EC 2.4.1.123) catalyzes formation of galactinol and the subsequent synthesis of raffinose family oligosaccharides. The relationship of GolS to drought and salt tolerance has been well documented, however, little information is available about the role of GolS gene in cold tolerance. A coding sequence of MfGolS1 cDNA was cloned from Medicago sativa spp falcata (i.e. M. falcata), a species that exhibits greater cold tolerance than alfalfa (M. sativa). MfGolS1 transcript was not detected in untreated vegetative tissues using RNA blot hybridization; however, it was greatly induced in leaves, but not in stem and petiole, after cold treatment. Higher levels of MfGolS1 transcript were induced and maintained in M. falcata than in M. sativa during cold acclimation. Accordingly, more sugars including sucrose, galactinol, raffinose and stachyose were accumulated in M. falcata than in M. sativa. The data indicated that MfGolS1 transcript and its resultant sugar accumulation were associated with the differential cold tolerance between M. falcata and M. sativa. MfGolS1 transcript was weakly induced by dehydration and salt stresses, but not responsive to abscisic acid. MfGolS1 could be induced by myo-inositol, which is proposed to participate in cold-induced MfGolS1 expression. Overexpression of MfGolS1 in tobacco resulted in elevated tolerance to freezing and chilling in transgenic plants as a result of enhanced levels of galactinol, raffinose and stachyose. Tolerance to drought and salt stresses was also increased in the transgenic tobacco plants. It is suggested that MfGolS1 plays an important role in plant tolerance to abiotic stresses.

Topics: Abscisic Acid; Acclimatization; Cold Temperature; Disaccharides; Freezing; Galactosyltransferases; Gene Expression Regulation, Plant; Inositol; Medicago; Medicago sativa; Molecular Sequence Data; Nicotiana; Oligosaccharides; Plant Leaves; Plant Proteins; Plants, Genetically Modified; Raffinose; Salt Tolerance; Stress, Physiological

There is a great need to search for natural compounds with superior prebiotic, antioxidant and immunostimulatory properties for use in (food) applications. Raffinose family oligosaccharides (RFOs) show such properties. Moreover, they contribute to stress tolerance in plants, acting as putative membrane stabilizers, antioxidants and signalling agents.. A large-scale soluble carbohydrate screening was performed within the plant kingdom. An unknown compound accumulated to a high extent in early-spring red deadnettle (Lamium purpureum) but not in other RFO plants. The compound was purified and its structure was unravelled with NMR. Organs and organ parts of red deadnettle were carefully dissected and analysed for soluble sugars. Phloem sap content was analysed by a common EDTA-based method.. Early-spring red deadnettle stems and roots accumulate high concentrations of the reducing trisaccharide manninotriose (Galα1,6Galα1,6Glc), a derivative of the non-reducing RFO stachyose (Galα1,6Galα1,6Glcα1,2βFru). Detailed soluble carbohydrate analyses on dissected stem and leaf sections, together with phloem sap analyses, strongly suggest that stachyose is the main transport compound, but extensive hydrolysis of stachyose to manninotriose seems to occur along the transport path. Based on the specificities of the observed carbohydrate dynamics, the putative physiological roles of manninotriose in red deadnettle are discussed.. It is demonstrated for the first time that manninotriose is a novel and important player in the RFO metabolism of red dead deadnettle. It is proposed that manninotriose represents a temporary storage carbohydrate in early-spring deadnettle, at the same time perhaps functioning as a membrane protector and/or as an antioxidant in the vicinity of membranes, as recently suggested for other RFOs and fructans. This novel finding urges further research on this peculiar carbohydrate on a broader array of RFO accumulators.

Topics: Biological Transport; Edetic Acid; Lamiaceae; Magnetic Resonance Spectroscopy; Oligosaccharides; Phloem; Plant Exudates; Plant Leaves; Plant Roots; Plant Stems; Raffinose; Seasons; Solubility; Species Specificity; Trisaccharides

Rice bean, a less known and underutilized legume, has emerged as a potential legume because of its nutritional potential. The nutritional quality of rice bean is higher as compared to many other legumes of Vigna family. In the present study, 16 diverse rice bean genotypes were evaluated for major nutritional constituents viz; protein content, total lipids, dietary fiber, total carbohydrates, vitamins, minerals, protein fractions, amino acid, and fatty acid profile. The protein content to the extent of 25.57% was observed in the genotype BRS-2 with in vitro digestibility of 54.23%. The fatty acid profile revealed the higher percentage of unsaturated fatty viz., linoleic and linolenic acid, which are nutritionally desirable in the diet. Albumins (6.13% to 7.47%) and globulins (13.11% to 15.56%) constituted the major portion of proteins. Anti-nutritional factors were in the range of: total phenolics (1.63% to 1.82%), total tannins (1.37% to 1.55%), condensed tannins (0.75% to 0.80%), hydrolysable tannins (0.56% to 0.79%), trypsin inhibitor (24.55 to 37.23 mg/g), phytic acid (7.32 to 8.17 mg/g), lipoxygenase activity (703 to 950 units/mg), and saponin content (1.2 to 3.1 mg/100 g). The oligosaccharides associated with the production of flatulence viz., raffinose, stachyose, and verbascose were in the limits of 1.66% to 2.58%, 0.94% to 1.88%, and 0.85% to 1.23%, respectively. In vitro protein digestibility up to 55.57% was observed in rice bean genotypes. The present study has revealed that rice bean is a nutritionally rich legume as compared to many other legumes of the category. Among different genotypes BRS-2 was observed superior and could be advocated for consumption as well as for inclusion in crop improvement programs.. Rice bean is nutritionally rich legume, but despite its nutritional excellence, it has been put in underutilized category. Because of this and several other reasons the people are not aware of its nutritional benefits. Moreover, the complete nutritional details are also not available on this pulse. The present study gives the vivid description of nutritional attributes of this legume for making people aware of its nutritional excellence and provoking improved work in rice bean.

Topics: Amino Acids; Ascorbic Acid; Dietary Carbohydrates; Dietary Fiber; Dietary Proteins; Fabaceae; Fatty Acids; Genotype; Lipoxygenase; Niacin; Nutritive Value; Oligosaccharides; Phenols; Phytic Acid; Raffinose; Saponins; Tannins; Trace Elements; Trypsin Inhibitors; Vitamins

Soybeans are an important source of protein-rich meal for livestock feed formulations. Recent changes in the cost of commodity-based sources of metabolizable energy (ME) inputs has put pressure on soybean meal to deliver both protein and ME in feed formulations. The non-oil fraction of soybean contains approximately 12% soluble carbohydrates, principally sucrose, raffinose, and stachyose. Of these carbohydrates, only sucrose is positive for ME. Both raffinose and stachyose, belonging to the raffinose family of oligosaccharides (RFOs), are considered antinutritional because of the negative consequences of their fermentation in the gut of monogastric animals when RFOs are consumed in the diet. Therefore, there is an interest in improving soybean seed composition so that it contains higher ME and fewer antinutritional components by increasing the sucrose content while lowering the RFOs. Several soybean lines have been discovered that contain altered levels of RFOs, and recent molecular genetic investigations have shown the phenotype to be caused by mutations in a raffinose synthase 2 (RS2) gene encoding the enzyme that is the committed step for RFO biosynthesis. The objective of this research was to determine the variation in carbohydrate profile for different soybean lines grown in a single location containing one of three different alleles of the RS2 gene. The results indicate that, although there is variation in the carbohydrate profiles for each line, different lines with the same RS2 genotype tend to produce a characteristic carbohydrate profile. Although the carbohydrate profile for each RS2 genotype class was consistent in different genetic backgrounds under two conditions grown at one location, more research will be necessary to determine the environmental stability of the carbohydrate profiles in multiple locations over different years.

Topics: Alleles; Disaccharides; Galactosyltransferases; Genotype; Glycine max; Oligosaccharides; Raffinose; Seeds; Sucrose

Ultrastructural and molecular studies have provided experimental evidence for the classification of cucurbits as symplastic loaders, mainly translocating the raffinose family oligosaccharides (RFOs) raffinose and stachyose. Earlier studies established that cucumber mosaic virus (CMV) infection causes a significant increase in the sucrose-to-RFO ratio in the phloem sap of melon plants. The alteration in phloem sap sugar composition was associated with upregulation of CmSUT1 transcript within the vascular bundles. The current research aimed to explore the effect of CMV infection on the enzymes involved in symplastic phloem loading and RFO biosynthesis. Viral infection did not affect the activity of either raffinose or stachyose synthases in source leaves, but caused upregulation of the respective transcripts. Interestingly, activity of galactinol synthase was higher in CMV-infected leaves, associated with upregulation of CmGAS2. A significant increase in CmGAS2 expression in source leaves of melon plants exposed to high temperatures indicated that this response is common for both biotic and abiotic stresses. However, the effect of CMV or heat stress on phloem sap sugar composition is not due to alteration in RFO biosynthesis.

Topics: Biosynthetic Pathways; Chromatography, Liquid; Cucumovirus; Cucurbitaceae; Galactosyltransferases; Gene Expression Regulation, Plant; Hot Temperature; Mass Spectrometry; Oligosaccharides; Plant Diseases; Plant Leaves; Raffinose; Real-Time Polymerase Chain Reaction; Stress, Physiological; Sucrose; Up-Regulation

Raffinose family oligosaccharides (RFOs) in food are considered anti-nutritional factors. This study elucidated the role of α-galactosidase (α-Gal), levansucrase, and sucrose phosphorylase for conversion of RFOs by lactobacilli. Quantification of gene expression by reverse-transcriptase quantitative PCR revealed that expression of levansucrase and sucrose phosphorylase by Lactobacillus reuteri is increased more than 100 fold when sucrose or raffinose are available. Fava bean (Vicia faba) or field pea (Pisum sativum) flours were fermented with α-Gal positive L. reuteri or α-Gal negative Lactobacillus sanfranciscensis. Isogenic strains lacking levansucrase activity, a L. reuteri ftfA mutant and a L. sanfranciscensis levS mutant, were used for comparison. During growth in pulse flours, L. sanfranciscensis accumulated melibiose and α-galactooligosaccharides (α-GOSs); the levansucrase-negative strain did not grow. L. reuteri metabolized raffinose, stachyose, and verbascose by levansucrase activity and accumulated α-GOSs as metabolic intermediates. Oligosaccharide metabolism in the levansucrase-negative mutant was slower, and accumulation of α-GOSs was not observed. The use of sorghum sourdough fermented with L. reuteri LTH5448 and bean flour in gluten-free baking converted RFOs to α-GOSs by levansucrase and invertase activities. In conclusion, the elucidation of the role levansucrase in RFO metabolism by lactobacilli allowed the conversion or hydrolysis of RFOs in food fermentations.

Topics: Bacterial Proteins; Fermentation; Glucosyltransferases; Hexosyltransferases; Lactobacillus; Oligosaccharides; Raffinose; Sucrose

Ruminococcus gnavus belongs to the 57 most common species present in 90% of individuals. Previously, we identified an α-galactosidase (Aga1) belonging to glycoside hydrolase (GH) family 36 from R. gnavus E1 (M. Aguilera, H. Rakotoarivonina, A. Brutus, T. Giardina, G. Simon, and M. Fons, Res. Microbiol. 163:14-21, 2012). Here, we identified a novel GH36-encoding gene from the same strain and termed it aga2. Although aga1 showed a very simple genetic organization, aga2 is part of an operon of unique structure, including genes putatively encoding a regulator, a GH13, two phosphotransferase system (PTS) sequences, and a GH32, probably involved in extracellular and intracellular sucrose assimilation. The 727-amino-acid (aa) deduced Aga2 protein shares approximately 45% identity with Aga1. Both Aga1 and Aga2 expressed in Escherichia coli showed strict specificity for α-linked galactose. Both enzymes were active on natural substrates such as melibiose, raffinose, and stachyose. Aga1 and Aga2 occurred as homotetramers in solution, as shown by analytical ultracentrifugation. Modeling of Aga1 and Aga2 identified key amino acids which may be involved in substrate specificity and stabilization of the α-linked galactoside substrates within the active site. Furthermore, Aga1 and Aga2 were both able to perform transglycosylation reactions with α-(1,6) regioselectivity, leading to the formation of product structures up to [Hex](12) and [Hex](8), respectively. We suggest that Aga1 and Aga2 play essential roles in the metabolism of dietary oligosaccharides and could be used for the design of galacto-oligosaccharide (GOS) prebiotics, known to selectively modulate the beneficial gut microbiota.

Topics: alpha-Galactosidase; Amino Acid Sequence; Animals; Gastrointestinal Tract; Glycosylation; Melibiose; Molecular Sequence Data; Oligosaccharides; Protein Structure, Tertiary; Raffinose; Rats; Ruminococcus; Sequence Alignment; Sequence Analysis, Protein; Substrate Specificity

Among various environmental factors, temperature is a major regulator affecting plant growth, development and fruit composition. Grapevine is the most cultivated fruit plant throughout the world, and grapes are used for wine production and human consumption. The molecular mechanisms involved in grapevine tolerance to high temperature, especially at the fruit level, are poorly understood. To better characterize the sensitivity of berries to the microenvironment, high temperature conditions were locally applied to Vitis vinifera Cabernet Sauvignon clusters. Two genes, VvGOLS1 and VvHsfA2, up-regulated by this treatment, were identified and further characterized. The expression profile of VvGOLS1 correlated positively with galactinol accumulation in heat-stressed berries. However, no galactinol derivatives, such as raffinose and stachyose, accumulated upon heat stress. Heterologous expression of VvGOLS1 in Escherichia coli showed that it encodes a functional galactinol synthase. Transient expression assays showed that the heat stress factor VvHsfA2 transactivates the promoter of VvGOLS1 in a heat stress-dependent manner. Taken together, our results highlight the intrinsic capacity of grape berries to perceive heat stress and to initiate adaptive responses, suggesting that galactinol may play a signaling role in these responses.

Topics: Amino Acid Sequence; Disaccharides; Escherichia coli; Fruit; Galactosyltransferases; Gene Expression Regulation, Plant; Heat-Shock Response; Molecular Sequence Data; Oligosaccharides; Phylogeny; Plant Proteins; Promoter Regions, Genetic; Raffinose; Sequence Homology, Amino Acid; Vitis

Soybean meal (SBM) is an important protein source in animal feed. However, the levels of SBM inclusion are restricted in some animal species by the presence of antinutritional factors (ANFs), including non-starch polysaccharides (NSPs) and α-galactosides (GOSs). The aim of this study was to reduce the soybean meal NSPs and GOSs by solid-state fermentation (SSF) using a combination of cellulolytic bacteria isolated from different environments (termites, earthworms, corn silage and bovine ruminal content). To analyse the key enzymatic activities, the isolates were grown in minimal media containing NSPs extracted from SBM. The selected bacterial strains belonged to the genera Streptomyces, Cohnella and Cellulosimicrobium. SSF resulted in a reduction of nearly 24% in the total NSPs, 83% of stachyose and 69% of raffinose and an increase in the protein content. These results suggest that cellulolytic bacteria-based SSF processing facilitates SBM nutritional improvement. In addition, the use of fermented SBM in animal diets can be recommended.

Topics: Animal Feed; Animals; Bacteria; Cellulase; Cellulose; DNA Primers; Fermentation; Galactosides; Glycine max; Oligosaccharides; Polymorphism, Restriction Fragment Length; Polysaccharides, Bacterial; Raffinose; RNA, Ribosomal; RNA, Ribosomal, 16S; Rumen; Starch; Streptomyces

The human gastrointestinal tract can be positively modulated by dietary supplementation of probiotic bacteria in combination with prebiotic carbohydrates. Here differential transcriptomics and functional genomics were used to identify genes in Lactobacillus acidophilus NCFM involved in the uptake and catabolism of 11 potential prebiotic compounds consisting of α- and β-linked galactosides and glucosides. These oligosaccharides induced genes encoding phosphoenolpyruvate-dependent sugar phosphotransferase systems (PTS), galactoside pentose hexuronide (GPH) permease, and ATP-binding cassette (ABC) transporters. PTS systems were upregulated primarily by di- and tri-saccharides such as cellobiose, isomaltose, isomaltulose, panose and gentiobiose, while ABC transporters were upregulated by raffinose, Polydextrose, and stachyose. A single GPH transporter was induced by lactitol and galactooligosaccharides (GOS). The various transporters were associated with a number of glycoside hydrolases from families 1, 2, 4, 13, 32, 36, 42, and 65, involved in the catabolism of various α- and β-linked glucosides and galactosides. Further subfamily specialization was also observed for different PTS-associated GH1 6-phospho-β-glucosidases implicated in the catabolism of gentiobiose and cellobiose. These findings highlight the broad oligosaccharide metabolic repertoire of L. acidophilus NCFM and establish a platform for selection and screening of both probiotic bacteria and prebiotic compounds that may positively influence the gastrointestinal microbiota.

Topics: ATP-Binding Cassette Transporters; Cellobiose; Gene Expression Regulation, Bacterial; Glucans; Isomaltose; Lactobacillus acidophilus; Oligosaccharides; Phosphoenolpyruvate Sugar Phosphotransferase System; Prebiotics; Raffinose; Sugar Alcohols

Developing garden pea embryos are able to take up exogenously applied cyclitols: myo-inositol, which naturally occurs in pea, and two cyclitols absent in pea plants: d-chiro-inositol and d-pinitol. The competition in the uptake of cyclitols by pea embryo, insensitivity to glucose and sucrose, and susceptibility to inhibitor(s) of H(+)-symporters (e.g. CCCP and antimycin A) suggest that a common cyclitol transporter is involved. Both d-chiro-inositol and d-pinitol can be translocated through the pea plant to developing embryos. During seed maturation drying, they are used for synthesis of mainly mono-galactosides, such as fagopyritol B1 and galactosyl pinitol A. Accumulation of d-chiro-inositol (and formation of fagopyritols), but not d-pinitol, strongly reduces accumulation of verbascose, the main raffinose oligosaccharide in pea seeds. The reasons for the observed changes are discussed.

Topics: Anti-Bacterial Agents; Antimycin A; Biological Transport; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Disaccharides; Inositol; Oligosaccharides; Pisum sativum; Raffinose; Seeds; Time Factors; Uncoupling Agents

Phloem loading is the process by which photoassimilates synthesized in the mesophyll cells of leaves enter the sieve elements and companion cells of minor veins in preparation for long distance transport to sink organs. Three loading strategies have been described: active loading from the apoplast, passive loading via the symplast, and passive symplastic transfer followed by polymer trapping of raffinose and stachyose. We studied phloem loading in Amborella trichopoda, a premontane shrub that may be sister to all other flowering plants. The minor veins of A. trichopoda contain intermediary cells, indicative of the polymer trap mechanism, forming an arc on the abaxial side and subtending a cluster of ordinary companion cells in the interior of the veins. Intermediary cells are linked to bundle sheath cells by highly abundant plasmodesmata whereas ordinary companion cells have few plasmodesmata, characteristic of phloem that loads from the apoplast. Intermediary cells, ordinary companion cells, and sieve elements form symplastically connected complexes. Leaves provided with (14)CO(2) translocate radiolabeled sucrose, raffinose, and stachyose. Therefore, structural and physiological evidence suggests that both apoplastic and polymer trapping mechanisms of phloem loading operate in A. trichopoda. The evolution of phloem loading strategies is complex and may be difficult to resolve.

Topics: Biological Transport; Magnoliaceae; Microscopy, Electron, Transmission; Oligosaccharides; Phloem; Plant Vascular Bundle; Plasmodesmata; Raffinose; Sucrose

Galactinol synthase (EC 2.4.1.123; GolS) catalyzes the first step in the synthesis of raffinose family oligosaccharides (RFOs). Their accumulation in response to abiotic stresses implies a role for RFOs in stress adaptation. In this study, the expression patterns of three isoforms of galactinol synthase (CaGolS1-2-3) from Coffea arabica were evaluated in response to water deficit, salinity and heat stress. All CaGolS isoforms were highly expressed in leaves while little to no expression were detected in flower buds, flowers, plagiotropic shoots, roots, endosperm and pericarp of mature fruits. Transcriptional analysis indicated that the genes were differentially regulated under water deficit, high salt and heat stress. CaGolS1 isoform is constitutively expressed in plants under normal growth conditions and was the most responsive during all stress treatments. CaGolS2 is unique among the three isoforms in that it was detected only under severe water deficit and salt stresses. CaGolS3 was primarily expressed under moderate and severe drought. This isoform was induced only at the third day of heat and under high salt stress. The increase in GolS transcription was not reflected into the amount of galactinol in coffee leaves, as specific glycosyltransferases most likely used galactinol to transfer galactose units to higher homologous oligosaccharides, as suggested by the increase of raffinose and stachyose during the stresses.

Topics: Adaptation, Physiological; Coffea; Desiccation; Galactosyltransferases; Gene Expression; Glycosyltransferases; Hot Temperature; Oligosaccharides; Plant Leaves; Plant Proteins; Protein Isoforms; Raffinose; Salinity; Salt Tolerance; Sodium Chloride; Stress, Physiological; Water

α-Galactosidases has the potential to hydrolyze α-1-6 linkages in raffinose family oligosaccharides (RFO). Aspergillus terreus cells cultivated on wheat bran produced three extracellular forms of α-galactosidases (E1, E2, and E3). E1 and E2 α-galactosidases presented maximal activities at pH 5, while E3 α-galactosidase was more active at pH 5.5. The E1 and E2 enzymes showed stability for 6 h at pH 4-7. Maximal activities were determined at 60, 55, and 50 °C, for E1, E2, and E3 α-galactosidase, respectively. E2 α-galactosidase retained 90% of its initial activity after 70 h at 50 °C. The enzymes hydrolyzed ρNPGal, melibiose, raffinose and stachyose, and E1 and E2 enzymes were able to hydrolyze guar gum and locust bean gum substrates. E1 and E3 α-galactosidases were completely inhibited by Hg²⁺, Ag⁺, and Cu²⁺. The treatment of RFO present in soy milk with the enzymes showed that E1 α-galactosidase reduced the stachyose content to zero after 12 h of reaction, while E2 promoted total hydrolysis of raffinose. The complete removal of the oligosaccharides in soy milk could be reached by synergistic action of both enzymes.

Topics: alpha-Galactosidase; Aspergillus; Dietary Fiber; Dyspepsia; Enzyme Stability; Food Handling; Galactans; Glycine max; Humans; Hydrogen-Ion Concentration; Hydrolysis; Kinetics; Mannans; Melibiose; Metals, Heavy; Oligosaccharides; Plant Gums; Raffinose; Soy Milk; Substrate Specificity; Temperature

Wholegrain grains and cereals contain a wide range of potentially protective factors that are relevant to gastrointestinal health. The prebiotics best studied are fructans [fructooligosaccharides (FOS), inulin] and galactooligosaccharides (GOS). These and other short-chain carbohydrates can also be poorly absorbed in the small intestine (named fermentable oligo-, di- and monosaccharides and polyols; FODMAPs) and may have important implications for the health of the gut.. In the present study, FODMAPs, including fructose in excess of glucose, FOS (nystose, kestose), GOS (raffinose, stachyose) and sugar polyols (sorbitol, mannitol), were quantified using high-performance liquid chromatography with an evaporative light scattering detector. Total fructan was quantified using an enzymic hydrolysis method.. Fifty-five commonly consumed grains, breakfast cereals, breads, pulses and biscuits were analysed. Total fructan were the most common short-chain carbohydrate present in cereal grain products and ranged (g per portion as eaten) from 1.12 g in couscous to 0 g in rice; 0.6 g in dark rye bread to 0.07 g in spelt bread; 0.96 g in wheat-free muesli to 0.11 g in oats; and 0.81 g in muesli fruit bar to 0.05 g in potato chips. Raffinose and stachyose were most common in pulses..   Composition tables including FODMAPs and prebiotics (FOS and GOS) that are naturally present in food will greatly assist research aimed at understanding their physiological role in the gut.

Topics: Chromatography, High Pressure Liquid; Dietary Carbohydrates; Edible Grain; Fermentation; Food Handling; Fructans; Fructose; Gastrointestinal Tract; Humans; Intestinal Absorption; Monosaccharides; Oligosaccharides; Poaceae; Prebiotics; Raffinose; Seeds; Sugar Alcohols

Red gram (Cajanus cajan L.) is an important crop for human and animal nutrition. However, raffinose family oligosaccharides present in red gram seed hinder its consumption as it is not digested by normal human carbohydrases and is further fermented by intestinal microflora, which induces flatulence. In order to make the grain legume more amenable for human consumption, we have tried to shed some light on the effect of germination followed by heat treatment methods such as autoclaving, cooking and pressure cooking on the raffinose family of sugars. These techniques, however, are primary prerequisites before consumption of the gram.. The percent removal of raffinose, stachyose and verbascose after germinating red gram seeds for 8 h followed by autoclaving was 65.6%, 58.9% and 65.3% respectively; and after cooking was 61.6%, 69.2% and 72.5%. Germinating for 16 h followed by autoclaving led to a mean decrease of 53.3% for raffinose, 60.3% for stachyose and 62.3% for verbascose. Germination of red gram seeds for 16 h followed by cooking led to a mean decrease of 71.7% for raffinose, 76.2% for stachyose and 74.0% for verbascose, respectively. The results for the percent removal of raffinose, stachyose and verbascose after germination of red gram seeds for 16 h followed by pressure cooking was 68.3%, 73.3% and 68.2% respectively.. This study demonstrates that local methods of processing reduce raffinose family oligosaccharides in red gram. The technique of germinating the seeds for 16 h followed by autoclaving, cooking and pressure cooking for the reduction of raffinose family oligosaccharides is a promising solution to overcome flatulence and increase the overall acceptance of red gram among general populace.

Topics: Cajanus; Dietary Carbohydrates; Digestion; Fermentation; Flatulence; Food Handling; Galactosides; Germination; Hot Temperature; Humans; Oligosaccharides; Raffinose

Desiccation tolerance of seeds is positively correlated with raffinose family oligosaccharides (RFOs). However, RFOs' role in desiccation tolerance is still a matter of controversy. The aim of this work was to monitor the accumulation of RFO during acquisition of desiccation tolerance in rapeseed (Brassica napus L.). Rapeseeds become desiccation tolerant at 21-24d after flowering (DAF), and the time was coincident with an accumulation of raffinose and stachyose. A gene encoding galactinol synthase (GolS; EC2.4.1.123), involved in RFO biosynthesis, was cloned and functionally characterized. Enzymatic properties of recombinant galactinol synthase were also determined. Accumulation of BnGOLS-1 mRNA in developing rapeseeds was concomitant with dry weight deposition and the acquisition of desiccation tolerance, and was concurrent with the formation of raffinose and stachyose. The physiological implications of BnGOLS-1 expression patterns in developing seeds are discussed in light of the hypothesized role of RFOs in seed desiccation tolerance.

Topics: Adaptation, Physiological; Amino Acid Sequence; Base Sequence; Brassica napus; Desiccation; DNA, Complementary; DNA, Plant; Escherichia coli; Galactosyltransferases; Germination; Hydrogen-Ion Concentration; Models, Molecular; Molecular Sequence Data; Oligosaccharides; Phylogeny; Raffinose; Recombinant Proteins; RNA, Messenger; RNA, Plant; Seeds; Sequence Alignment; Stress, Physiological; Water

Raffinose family oligosaccharides (RFOs), which include raffinose and stachyose, are thought to be an important source of energy during seed germination. In contrast to their potential for promoting germination, RFOs represent anti-nutritional units for monogastric animals when consumed as a component of feed. The exact role for RFOs during soybean seed development and germination has not been experimentally determined; but it has been hypothesized that RFOs are required for successful germination. Previously, inhibition of RFO breakdown during imbibition and germination was shown to significantly delay germination in pea seeds. The objective of this study was to compare the germination potential for soybean seeds with either wild-type (WT) or low RFO levels and to examine the role of RFO breakdown in germination of soybean seeds. There was no significant difference in germination between normal and low RFO soybean seeds when imbibed/germinated in water. Similar to the situation in pea, soybean seeds of wild-type carbohydrate composition experienced a delay in germination when treated with a chemical inhibitor of alpha-galactosidase activity (1-deoxygalactonojirimycin or DGJ) during imbibition. However, low RFO soybean seed germination was not significantly delayed or reduced when treated with DGJ. In contrast to the situation in pea, the inhibitor-induced germination delay in wild-type soybean seeds was not partially overcome by the addition of galactose or sucrose. We conclude that RFOs are not an essential source of energy during soybean seed germination.

Topics: 1-Deoxynojirimycin; Germination; Glycine max; Oligosaccharides; Raffinose; Seeds; Water

Levels of sucrose and raffinose family oligosaccharides (RFOs) (raffinose and stachyose) were determined in beech (Fagus sylvatica L.) seeds during development, maturation, desiccation and storage. An increase in RFOs and a marked decrease in the S:(R+St) ratio (i.e. mass ratio of sucrose to the sum of RFOs) were observed at the time of desiccation tolerance (DT) acquisition by seeds. In seeds stored at -10 degrees C through 1, 4, 7, and 12 years, changes in sucrose, raffinose and stachyose levels and in alpha-galactosidase activity were noted. The S/R+St ratio and alpha-galactosidase activity significantly increased in seeds after 7 and 12 years of storage, when a marked decrease in viability, measured as germination capacity, was recorded. Germination capacity was found to be strongly correlated with sucrose content, the S:(R+St) ratio, and alpha-galactosidase activity. A strong positive correlation was found between germination capacity and stachyose content. The results clearly indicated that the composition of RFOs in beech seeds is closely related to DT acquisition and seed viability during storage.

Topics: alpha-Galactosidase; Carbohydrate Metabolism; Cotyledon; Dehydration; Desiccation; Fagus; Germination; Oligosaccharides; Oxidation-Reduction; Raffinose; Seeds; Sucrose; Water

While classic raffinose family oligosaccharides (RFOs) such as raffinose and stachyose are common in plants, stachyose is absent in the Caryophyllaceae. Instead the tetrasaccharide lychnose α-d-Gal-(1→6)α-d-Glc-(1→2)β-d-Fru-(1→1)α-d-Gal can accumulate. Stellaria media, a representative member of this family, was used to isolate α-d-Gal-(1→6)-[α-d-Gal-(1→4)]α-d-Glc-(1→2)β-d-Fru-(1→1)α-d-Gal, a novel pentasaccharide with a lychnose backbone. Complete NMR characterization using COSY, HSQC, HSQC-TOCSY, HMBC, and NOESY experiments was performed to unequivocally resolve its structure. This is the first report of a natural compound containing a Gal α(1→4)Glc linkage. The trivial name stellariose is proposed for this new pentasaccharide.

Topics: Carbohydrate Sequence; Molecular Structure; Oligosaccharides; Raffinose; Stellaria

Stage-specific analyses of starch and 18 sugars, including pentoses, hexoses, disaccharides, trisaccharides, oligosaccharides and sugar alcohols, were made throughout seed development for zygotic embryo and female gametophyte (FG) tissues of loblolly pine (Pinus taeda L.). Tissue was most often analyzed in triplicate from two open-pollinated families grown in different locations and sampled in different years. Carbohydrates were analyzed by enzymatic assay, high performance liquid chromatography or gas chromatography/mass spectrometry. For all carbohydrates quantified, peak concentrations were higher in embryo tissue than in FG tissue. Significant changes in starch and sugar concentrations occurred over time, with both seed collections showing similar trends in temporal changes. Although concentrations were not always similar, embryo and FG tissues generally showed similar patterns of change in starch and sugar concentrations over time. Total starch concentration was highest during early seed development and decreased as development progressed. The major sugars contributing to osmotic potential during early seed development were D-pinitol, sucrose, fructose and glucose. During mid-seed development, D-pinitol, sucrose, fructose, glucose, melibiose and raffinose provided major contributions to the osmotic environment. During late seed development, sucrose, raffinose, melibiose, stachyose and fructose were the major contributors to osmotic potential. These data suggest stage-specific media composition for each step in the somatic embryogenesis protocol.

Topics: Carbohydrates; Chromatography, High Pressure Liquid; Fructose; Gas Chromatography-Mass Spectrometry; Glucose; Inositol; Melibiose; Oligosaccharides; Pinus taeda; Raffinose; Seeds; Starch; Sucrose

Oligosaccharides have been credited with many health-promoting functions, which had been identified in many clinical studies, such as promoting the growth of Bifidobacterium in human intestine and balance of intestinal bacteria, modulating the immune response, inhibition of cancer and tumor, stimulation of mineral absorption. In this study the effect of processing unit operations on the levels of soybean oligosaccharides during production of soybean sheet were investigated. The concentrations of oligosaccharide in initial raw soybean were: sucrose 43.05 g/kg, raffinose 7.52 g/kg and stachyose 41.32 g/kg (in dry matter). Oligosaccharide losses in the soaking water, in the first filtrating stage, in the second filtrating stage and finally in the sheet formation stage were 0.68, 10.3, 8.15 and 47.22 g/kg (initial dry soybean) respectively, representing 0.74, 11.21, 8.87 and 51.39% of the total oligosaccharides present in the initial soybeans. The recovery of oligosaccharides in the final soybean sheet from the initial soybean was 27.92%. The loss of soybean oligosaccharides in different processing stages, especially in the by-product, the sweet slurry, was considerable. The loss of oligosaccharides was mainly associated with water/matter removal in production process. The analysis of loss profile implied possible ways to improve the technology for production of oligosaccharides-enriched soy-sheets.

Topics: Food Handling; Glycine max; Humans; Nutritional Physiological Phenomena; Nutritive Value; Oligosaccharides; Raffinose

The transport of carbohydrates by Streptococcus mutans is accomplished by the phosphoenolpyruvate-phosphotransferase system (PTS) and ATP-binding cassette (ABC) transporters. To undertake a global transcriptional analysis of all S. mutans sugar transporters simultaneously, we used a whole-genome expression microarray. Global transcription profiles of S. mutans UA159 were determined for several monosaccharides (glucose, fructose, galactose, and mannose), disaccharides (sucrose, lactose, maltose, and trehalose), a beta-glucoside (cellobiose), oligosaccharides (raffinose, stachyose, and maltotriose), and a sugar alcohol (mannitol). The results revealed that PTSs were responsible for transport of monosaccharides, disaccharides, beta-glucosides, and sugar alcohol. Six PTSs were transcribed only if a specific sugar was present in the growth medium; thus, they were regulated at the transcriptional level. These included transporters for fructose, lactose, cellobiose, and trehalose and two transporters for mannitol. Three PTSs were repressed under all conditions tested. Interestingly, five PTSs were always highly expressed regardless of the sugar source used, presumably suggesting their availability for immediate uptake of most common dietary sugars (glucose, fructose, maltose, and sucrose). The ABC transporters were found to be specific for oligosaccharides, raffinose, stachyose, and isomaltosaccharides. Compared to the PTSs, the ABC transporters showed higher transcription under several tested conditions, suggesting that they might be transporting multiple substrates.

Topics: ATP-Binding Cassette Transporters; Biological Transport; Carbohydrates; Cellobiose; Fructose; Galactose; Gene Expression Profiling; Gene Expression Regulation, Bacterial; Glucose; Lactose; Maltose; Mannose; Oligonucleotide Array Sequence Analysis; Oligosaccharides; Phosphoenolpyruvate Sugar Phosphotransferase System; Raffinose; Reverse Transcriptase Polymerase Chain Reaction; Streptococcus mutans; Sucrose; Transcription, Genetic; Trisaccharides

Raffinose oligosaccharides (RO) are the factors primarily responsible for flatulence upon ingestion of soybean-derived products. ROs are hydrolyzed by alpha-galactosidases that cleave alpha-1,6-linkages of alpha-galactoside residues. The objectives of this study were the purification and characterization of extracellular alpha-galactosidase from Debaryomyces hansenii UFV-1. The enzyme purified by gel filtration and anion exchange chromatographies presented an Mr value of 60 kDa and the N-terminal amino acid sequence YENGLNLVPQMGWN. The Km values for hydrolysis of pNP alphaGal, melibiose, stachyose, and raffinose were 0.30, 2.01, 9.66, and 16 mM, respectively. The alpha-galactosidase presented absolute specificity for galactose in the alpha-position, hydrolyzing pNPGal, stachyose, raffinose, melibiose, and polymers. The enzyme was noncompetitively inhibited by galactose (Ki = 2.7 mM) and melibiose (Ki = 1.2 mM). Enzyme treatments of soy milk for 4 h at 60 degrees C reduced the amounts of stachyose and raffinose by 100%.

Topics: alpha-Galactosidase; Amino Acid Sequence; Ascomycota; Flatulence; Hydrolysis; Oligosaccharides; Raffinose; Soy Foods; Soy Milk

Conjugating bovine trypsin with oligosaccharides maltotriose, raffinose and stachyose increased its thermostability and suppressed autolysis, without affecting its cleavage specificity. These conjugates accelerated the digestion of protein substrates both in solution and in gel, compared to commonly used unmodified and methylated trypsins.

Topics: Animals; Cattle; Cytochromes c; Enzyme Stability; Fructose-Bisphosphate Aldolase; Gels; Heating; Myoglobin; Oligosaccharides; Proteomics; Raffinose; Serum Albumin, Bovine; Solutions; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Trisaccharides; Trypsin

The contents of raffinose family oligosaccharides (RFO) and sucrose in Brassica, Lupinus, Pisum, and Hordeum species were investigated by chemometric principal component analysis (PCA). Hordeum samples contained sucrose and raffinose, and Brassica samples all contained sucrose, raffinose, and stachyose. In addition to these, the Pisum samples contained verbascose and the Lupinus samples also contained ajugose. High stachyose and low ajugose contents were found in Lupinus albus in contrast to Lupinus angustifolius, having low stachyose and high ajugose contents. Lupinus luteus had average stachyose and ajugose contents, whereas large amounts of verbascose were accumulated in these seeds. Lupinus mutabilis had high stachyose and low ajugose contents, similar to the composition in L. albus but showing higher raffinose content. The Brassica samples also showed compositional RFO variations within the species, and subgroup formations were discovered within the investigated Brassica napus varieties. PCA results indicated compositional variations between the investigated genera and within the various species of value as chemotaxonomic defined parameters and as tools in evaluations of authenticity/falsifications when RFO-containing plants are used as, for example, feed and food additives.

Topics: Brassicaceae; Electrophoresis, Capillary; Fabaceae; Galactosides; Glucosides; Hordeum; Oligosaccharides; Pyrans; Raffinose; Sucrose

The antinutrient (raffinose oligosaccharides, tannins, phytic acid and trypsin inhibitors) composition and in vitro protein digestibility of eight improved varieties of Phaseolus vulgaris grown in Ethiopia were determined. Stachyose was the predominant alpha-galactosides in all haricot bean samples. Raffinose was also present in significant quantities but verbascose, glucose and fructose were not detected at all in the samples. The concentrations observed for the protein digestibility and antinutritional factors, varied significantly (P<0.05) between varieties investigated in this study. Mean values for protein digestibility ranged from 80.66% (in Roba variety) to 65.64% (in Beshbesh variety). Mean values for raffinose, stachyose, sucrose, trypsin inhibitors, tannins and phytic acid were 3.14 mg/g, 14.86 mg/g, 24.22 mg/g, 20.68 TUIx10(3)/g, 17.44 mg, catechin equivalents/g and 20.54 mg/g respectively. Statistical analyses of data revealed that antinutritional factors and protein digestibility were influenced by variety (genotype). Relationships between antinutritional factors and protein digestibility were also observed. The possibility of selecting varieties to be used for large-scale cultivation in Ethiopia on the basis of these data is discussed. Among the improved varieties studied, Roba, Redwolaita, Mexican and Awash were found to be the best food and export type of haricot beans in the Ethiopian context, because of their higher protein digestibility, lower antinutrtional factors and other beneficial nutritional parameters. Roba variety can be used by local food processors for the production of value-added bean-based products especially to combat the problem of protein energy malnutrition and related diseases which are very common in developing countries.

Topics: Dietary Proteins; Digestion; Ethiopia; Food Analysis; Nutritional Physiological Phenomena; Oligosaccharides; Phaseolus; Phytic Acid; Plant Proteins; Raffinose; Tannins; Trypsin Inhibitors; Zinc

The oligosaccharide content was determined in 12 different cultivars of black gram. The effect of various treatments such as soaking, cooking, and enzyme treatment on the raffinose family oligosaccharides of dry seeds and flour was studied. Ajugose, a higher oligosaccharide (DP 6) found in trace quantities in seeds, was shown in black gram by HPLC. The percent reduction of raffinose, stachyose, verbascose, and ajugose after soaking for 16 hr was 41.66%, 47.61%, 28.48%, and 26.82%, respectively in Local-I variety and 43.75%, 20.58%, 23.60%, and 15.88%, respectively in Local-II variety. Cooking for 60 min resulted in decrease of 100% for raffinose, 76.19% for stachyose, 36.39% for verbascose, and 60.97% for ajugose in Local-I variety and 100% for raffinose, 55.88% for stachyose, 48.52% for verbascose, and 56.07% for ajugose in Local-II variety. Thin layer chromatographic analysis of 3 hr enzyme-treated samples revealed almost complete hydrolysis of raffinose family of oligosaccharides. Among the different methods employed, enzyme treatment was found to be the most effective for removing alpha-galactosides in black gram.

Topics: alpha-Galactosidase; Chromatography, High Pressure Liquid; Chromatography, Thin Layer; Cooking; Fabaceae; Food Handling; Nutritive Value; Oligosaccharides; Raffinose; Seeds; Time Factors

Consumption of soya-derived products has been hampered by the presence of alpha-galactooligosaccharides (alpha-GOS) because mammals lack pancreatic alpha-galactosidase (alpha-Gal) which is necessary for their hydrolysis. These sugars thus reach the large intestine causing gastrointestinal disorders in sensitive individuals. The use of lactic acid bacteria (LAB) expressing alpha-Gal is a promising solution for the degradation of alpha-GOS in soyamilk.. The capacity of the LAB Lactobacillus fermentum CRL 722 to properly degrade alpha-GOS was studied in vitro using controlled fermentation conditions and in vivo using a rat model. Lactobacillus fermentum CRL 722 was able to grow on commercial soyamilk and completely eliminated stachyose and raffinose during fermentation because of its high alpha-Gal activity. Rats fed soyamilk fermented by this LAB had smaller caecums compared with rats fed unfermented soyamilk.. Soyamilk fermentation by Lact. fermentum CRL 722 results in the reduction of alpha-GOS concentrations in soyamilk, thus eliminating possible undesirable physiological effects normally associated with its consumption.. Fermentation with Lact. fermentum CRL 722 could prevent gastrointestinal disorders in sensitive individuals normally associated with the consumption of soya-based products. This LAB could thus be used in the elaboration of novel fermented vegetable products which better suit the digestive capacities of consumers.

Topics: alpha-Galactosidase; Animals; Cecum; Colony Count, Microbial; Fermentation; Food Microbiology; Galactose; Lactobacillus; Oligosaccharides; Raffinose; Rats; Rats, Wistar; Soy Milk

Human consumption of soy-derived products has been limited by the presence of non-digestible oligosaccharides (NDO), such as the alpha-galactooligosaccharides raffinose and stachyose. Most mammals, including man, lack pancreatic alpha-galactosidase (alpha-Gal), which is necessary for the hydrolysis of these sugars. However, such NDO can be fermented by gas-producing microorganisms present in the cecum and large intestine, which in turn can induce flatulence and other gastrointestinal disorders in sensitive individuals. The use of microorganisms expressing alpha-Gal is a promising solution to the elimination of NDO before they reach the large intestine. In the present study, lactic acid bacteria engineered to degrade NDO have been constructed and are being used as a tool to evaluate this solution. The alpha-Gal structural genes from Lactobacillus plantarum ATCC8014 (previously characterized in our laboratory) and from guar have been cloned and expressed in Lactococcus lactis. The gene products were directed to different bacterial compartments to optimize their possible applications. The alpha-Gal-producing strains are being evaluated for their efficiency in degrading raffinose and stachyose: i) in soymilk fermentation when used as starters and ii) in situ in the upper gastrointestinal tract when administered to animals orally, as probiotic preparations. The expected outcomes and possible complications of this project are discussed.

Topics: alpha-Galactosidase; Animals; Cultured Milk Products; Digestion; Fermentation; Food, Genetically Modified; Lactobacillus plantarum; Lactococcus lactis; Oligosaccharides; Probiotics; Raffinose; Rodentia; Soy Milk

Topics: Adaptation, Physiological; alpha-Galactosidase; Carbon Dioxide; Cold Temperature; Fabaceae; Hydrogen; Methane; Oligosaccharides; Raffinose; Seeds; Sucrose; Water

The objective of this study was to evaluate dietary galactooligosaccharide (Gal OS) addition on swine nutrient digestibility, ileal and fecal bacterial populations, and ileal short-chain fatty acid (SCFA) production, and to determine their impact on ileal fermentative characteristics in vitro. Twelve T-cannulated pigs (BW = 25 kg) were fed a diet free of Gal OS for 21 d. On d 22, ileal digesta samples were collected for an in vitro fermentation experiment (Exp. 1). Substrates included: raffinose/stachyose combination (R + S), soy solubles (SS), and transgalactooligosaccharides (TOS). Also included were the non-OS components of SS and TOS. Nine pigs (three donors per treatment) served as ileal effluent donors. Each substrate was fermented in vitro for 6 h, and pH and SCFA and gas production were determined. Pigs then were allotted to three treatments: a Gal OS-free control diet and the control diet with either 3.5% added Gal OS from SS or TOS. Diets, feces, and digesta samples collected weekly for 6 wk on d 6 (feces) and 7 (digesta) were analyzed for DM, OM, CP, and chromic oxide concentrations. Feces and ileal digesta were analyzed for bifidobacteria and lactobacilli populations. Ileal digesta samples were analyzed for SCFA. On d 64, a second in vitro fermentation experiment (Exp. 2) was conducted using ileal effluent from three pigs per treatment and the same substrates used in Exp. 1. In vivo results showed that ileal and total tract DM and OM digestion were decreased (P < 0.05) by addition of both SS and TOS to the diet. Ileal and total-tract N digestibilities were decreased (P < 0.05) by dietary addition of SS. Fecal bifidobacteria and lactobacilli were increased (P < 0.05) by addition of SS and TOS to the diet. Ileal propionate and butyrate concentrations were greater (P < 0.05) for pigs fed diets containing both sources of Gal OS. In vitro results showed that fermentation data were not affected by donor animal adaptation to treatment. For both in vitro experiments, gas and SCFA production were higher (P < 0.05) for R + S than for SS or TOS. Fermentation of R + S resulted in a higher pH (P < 0.05) than did SS or TOS. Fermentation of non-OS components of SS and TOS resulted in more (P < 0.05) gas and SCFA production, and pH values that did not differ (P > 0.05) compared to SS and TOS. The Gal OS used in this study were prebiotics, increasing beneficial bacteria in vivo and SCFA concentrations both in vivo and in vitro.

Topics: Animal Feed; Animals; Digestion; Fatty Acids, Volatile; Feces; Fermentation; Hydrogen-Ion Concentration; Ileum; Oligosaccharides; Probiotics; Raffinose; Swine; Time Factors

A rapid, easy, and reproducible capillary electrophoresis method for determination of raffinose family oligosaccharides (alpha-galactosides) was developed. Sucrose, raffinose, stachyose, verbascose, and ajugose were determined with indirect UV detection at moderate alkaline pH 9.2, using pyridine-2,6-dicarboxylic acid as background electrolyte in a sodium tetraborate buffer with added cetyltrimethylammonium bromide. The separation efficiency measured by the number of theoretical plates (N) ranged from 1.4 x 10(5) to 2.3 x 10(5). The precision of the method, measured by the relative standard deviation (RSD), was less than 0.53% for the migration times and better than 3.4% for normalized areas (NA), considering all sugars except verbascose (RSD(NA) = 11.8%). Detection limits were about 110 microg/mL, corresponding to 150-320 microM. Relative response factors (RRF) were calculated on the basis of linearity studies and used for quantification of alpha-galactosides in a lupine sample (Lupinus angustifolius).

Topics: Brassicaceae; Electrophoresis, Capillary; Fabaceae; Galactosides; Hydrogen-Ion Concentration; Lupinus; Oligosaccharides; Raffinose; Sensitivity and Specificity; Sucrose; Ultraviolet Rays

Three experiments were conducted to examine the effects of a commercial enzyme preparation on chicks performance and digestibilities of nonstarch polysaccharides (NSP), raffinose (R), stachyose (S), and total oligosaccharides (O) in diets containing whole and dehulled lupin and ethanol-extracted dehulled lupin meal. Ethanol extract was also used to produce a rich oligosaccharide fraction. In the first experiment, the dehulling treatment and the addition of enzyme to the diet improved (P < 0.05) weight gain (24 and 15%), feed-to-gain ratio (13 and 9%), dry matter retention (32 and 8%), apparent protein digestibility (6 and 3%) and ileal digestibilities of raffinose (19 and 119%), stachyose (85 and 204%), and total oligosaccharides (68 and 178%), respectively. In addition, enzyme treatment improved (P < 0.05) excreta digestibility values for NSP (from -1.7 to 5.5%), R (from 64.6 to 92.6%), S (from 48.8 to 82.4%), and O (from 55.8 to 83.5%). In the second experiment, extraction of the ethanol soluble components from dehulled lupin decreased (P < 0.0001) weight gain (51%), and increased (P < 0.0001) feed consumption (34%), feed-to-gain ratio (32%), relative gizzard weight (14%), and the relative ceca length (20%). The addition of the isolated fraction of oligosacharides to the corn-extracted lupin diet (7.5 and 15%) yielded performance values similar to those obtained with the unextracted dehulled lupin. The addition of enzymes to the diets significantly improved the weight gain (11%) and feed to gain ratio (6%), and decreased relative gizzard weight (12%) and relative ceca length (7%). Ileal digestibilities of R, S, and O were considerably lower than corresponding excreta digestibilities. The excreta digestibility of NSP and excreta and ileal digestibilities of R, S, and O were lower (P < 0.05) in chicks fed the lupin diet containing the high concentration of extract compared with those fed the same diet containing the lower concentration of extract. Enzymes, when added to the diets, increased (P < 0.05) the digestibilities of R, S, and O with the relative effects being much greater for ileal than excreta samples. In the third experiment, the ethanol extract (7.5 and 15%) added to a corn-soybean diet improved weight gain, feed consumption, and feed-to-gain ratio by 19, 13, and 6%, respectively. Ileal digestibilities of R, S, and O were low (< 45%), especially in chicks fed the diets containing the high amount of added oligosaccharide fraction. There was no e

Topics: Animal Feed; Animal Nutritional Physiological Phenomena; Animals; Cell Wall; Chickens; Digestion; Enzymes; Ethanol; Lupinus; Male; Oligosaccharides; Plant Extracts; Polysaccharides; Raffinose

Fourteen ileally cannulated pigs (BW = 35 +/- 2 kg) were randomly allotted to a replicated 7 x 7 Latin square design experiment to evaluate the influence of the soybean oligosaccharides (OS), raffinose and stachyose, on ileal nutrient digestibility and fecal consistency. Semipurified diets containing soy protein concentrate (SPC) or soybean meal (SBM) as the sole protein sources were fed. Soy solubles (SS), a by-product of SBM processing containing 3.5% raffinose and 11.5% stachyose, were used to increase dietary raffinose and stachyose concentrations. The seven dietary treatments were SPC, SPC + 9% SS, SBM, SBM + 9% SS, SBM + 18% SS, SBM + 24,000 U alpha-galactosidase enzyme preparation/kg diet, and a low-protein casein (LPC) diet used to calculate true digestibility. Diets, with the exception of the LPC diet, were formulated to contain 17% CP. All diets contained 0.5% chromic oxide as a marker for ileal digestibility determination. The experimental periods were divided into a 5-d diet adaptation followed by 2-d of ileal digesta collection. Diets and digesta were analyzed for DM, N, Cr, amino acids (AA), raffinose, and stachyose. Fecal consistency was determined on d 6 and 7 of each experimental period. The apparent and true ileal AA digestibilities were not different (P < 0.05) for the SPC and SBM control diets. When SS was added to the SPC diet, apparent and true N and AA digestibilities were depressed (P < 0.05) with the exception of Trp and Pro. The apparent and true ileal N and AA digestibilities were not different (P > 0.05) between the SBM control and SBM + 9% SS diets with the exception of Glu. There was a linear decrease (P < 0.05) in apparent and true DM, Val, Gly, and Tyr digestibilities when increasing levels of SS were added to the SBM diet. The addition of alpha-galactosidase did not improve apparent or true ileal N or AA digestibilities except for apparent and true Val and Tyr. Ileal raffinose digestibility was improved (P < 0.05) by addition of a-galactosidase, but was not affected by any other dietary treatment. Ileal stachyose digestibility was not affected (P > 0.58) by treatment. Fecal consistency likewise was not affected (P > 0.36) by dietary treatment. In conclusion, soy OS reduced nutrient digestibilities, but the reductions were small, ranging from approximately 1.1 to 7.4 percentage units. This suggests that other factors may be negatively impacting SBM digestibility.

Topics: alpha-Galactosidase; Amino Acids; Animal Feed; Animals; Chromium Compounds; Digestion; Feces; Glycine max; Ileum; Nutritive Value; Oligosaccharides; Raffinose; Random Allocation; Swine

The objective of this study was to verify the effect of soaking on the factors causing flatulence in the common bean (Phaseolus vulgaris, L.) cv. IAC-Carioca during domestic preparation. A biological assay using recently weaned (21 days) male Wistar rats provided the Food Conversion Efficiency (FCE) and the Net Protein Ratio (NPR). Five treatments were carried out with isocaloric (350.9 +/- 37.9 kcal/100 g) and isoprotein (12.0 +/- 0.5%) experimental diets, with the following protein sources: beans cooked without soaking (BNS), beans soaked and cooked with the soaking water (BSWW), beans soaked and cooked without the residual soaking water (BSNW), control diet (casein) (CC), casein plus the total soluble solids found in the soaking water (CSS) for comparative purposes, and an aproteic diet (AP) for corrective purposes, all diets offered ad libitum. The contents of raffinose-type oligosaccharides were determined in the different domestic preparations of the beans. Significant reductions were observed in the contents of the oligosaccharides raffinose (25.0%), stachyose (24.8%), and verbascose (41.7%), and in the contents of total sugars (80.6%), reducing sugars (58.2%), nonreducing sugars (90.3%), and starch (26.8%) when soaking took place before cooking and elimination of the soaking water not absorbed by the beans (BSNW) was used. No significant difference (p > 0.05) was observed between the values for FCE and NPR of the control diet (casein) and control diet plus soaking water soluble solids. Neither was any significant difference between the values for the different bean treatments found, though the values for FCE and NPR were lower than those obtained for casein treatments. Thus it was verified that although the domestic preparation of the common bean significantly reduced the contents of raffinose-type oligosaccharides, total reducing and nonreducing sugars and starch, it did not interfere with its nutritive value.

Topics: Animals; Carbohydrate Metabolism; Carbohydrates; Cooking; Dietary Proteins; Digestion; Flatulence; Food Handling; Male; Nutritive Value; Oligosaccharides; Phaseolus; Raffinose; Rats; Rats, Wistar; Starch

Raffinose oligosaccharides are major soluble carbohydrates in seeds and other tissues of plants. Their biosynthesis proceeds by stepwise addition of galactose units to sucrose, which are provided by the unusual donor galactinol (O-alpha-d-galactopyranosyl-(1-->1)-l-myo-inositol). Chain elongation may also proceed by transfer of galactose units between raffinose oligosaccharides. We here report on the purification, characterization, and heterologous expression of a multifunctional stachyose synthase (EC ) from developing pea (Pisum sativum L.) seeds. The protein, a member of family 36 of glycoside hydrolases, catalyzes the synthesis of stachyose, the tetrasaccharide of the raffinose series, by galactosyl transfer from galactinol to raffinose. It also mediates the synthesis of the pentasaccharide verbascose by galactosyl transfer from galactinol to stachyose as well as by self-transfer of the terminal galactose residue from one stachyose molecule to another. These activities show optima at pH 7.0. The enzyme also catalyzes hydrolysis of the terminal galactose residue of its substrates, but is unable to initiate the synthesis of raffinose oligosaccharides by galactosyl transfer from galactinol to sucrose. A minimum reaction mechanism which accounts for the broad substrate specificity and the steady-state kinetic properties of the protein is presented.

Topics: Amino Acid Sequence; Catalysis; Cloning, Molecular; Galactosyltransferases; Hydrogen-Ion Concentration; Inositol; Kinetics; Molecular Sequence Data; Oligosaccharides; Pisum sativum; Raffinose; Substrate Specificity

A single, recessive mutation in soybean (Glycine max L. Merr.), which confers a seed phenotype of increased inorganic phosphate, decreased phytic acid, and a decrease in total raffinosaccharides, has been previously disclosed (S.A. Sebastian, P.S. Kerr, R.W. Pearlstein, W.D. Hitz [2000] Soy in Animal Nutrition, pp 56-74). The genetic lesion causing the multiple changes in seed phenotype is a single base change in the third base of the codon for what is amino acid residue 396 of the mature peptide encoding a seed-expressed myo-inositol 1-phospate synthase gene. The base change causes residue 396 to change from lysine to asparagine. That amino acid change decreases the specific activity of the seed-expressed myo-inositol 1-phosphate synthase by about 90%. Radio tracer experiments indicate that the supply of myo-inositol to the reaction, which converts UDP-galactose and myo-inositol to galactinol is a controlling factor in the conversion of total carbohydrate into the raffinosaccharides in both wild-type and mutant lines. That same decrease in myo-inositol 1-phosphate synthetic capacity leads to a decreased capacity for the synthesis of myo-inositol hexaphosphate (phytic acid) and a concomitant increase in inorganic phosphate.

Topics: Amino Acid Sequence; Disaccharides; Galactosyltransferases; Glycine max; Inositol; Mutation; Myo-Inositol-1-Phosphate Synthase; Oligosaccharides; Phenotype; Phosphates; Phytic Acid; Raffinose; Seeds; Sequence Homology, Amino Acid; Sucrose

The effects of cooking treatments (boiling, autoclaving and microwave cooking) and germination on the nutritional composition and antinutritional factors of chickpeas were studied. Cooking treatments and/or germination caused significant (p < 0.05) decreases in fat, total ash, carbohydrate fractions, antinutritional factors, minerals and B-vitamins. Germination was less effective than cooking treatments in reducing trypsin inhibitor, hemagglutinin activity, tannins and saponins; it was more effective in reducing phytic acid, stachyose and raffinose. Cooking treatments and germination decreased the concentrations of lysine, tryptophan, total aromatic and sulfur-containing amino acids. However, cooked and germinated chickpeas were still higher in lysine, isoleucine and total aromatic amino acid contents than the FAO/WHO reference. The losses in B-vitamins and minerals in chickpeas cooked by microwaving were smaller than in those cooked by boiling and autoclaving. Germination resulted in greater retention of all minerals and B-vitamins compared to cooking treatments. In vitro protein digestibility, protein efficiency ratio and essential amino acid index were improved by all treatments. The chemical score and limiting amino acid of chickpeas subjected to the various treatments varied considerably, depending on the type of treatment. Based on these results, microwave cooking appears to be the best alternative for legume preparation in households and restaurants.

Topics: Amino Acids; Cicer; Dietary Carbohydrates; Dietary Fats; Digestion; Germination; Hemagglutinins; Hot Temperature; Minerals; Nutritive Value; Oligosaccharides; Phytic Acid; Raffinose; Tannins; Trypsin Inhibitors; Vitamin B Complex

Raffinose family oligosaccharides (RFOs) are synthesized by a set of galactosyltransferases, which sequentially add galactose units from galactinol to sucrose. The accumulation of RFOs was studied in maturing seeds of two pea (Pisum sativum) lines with contrasting RFO composition. Seeds of the line SD1 accumulated stachyose as the predominant RFO, whereas verbascose, the next higher homolog of stachyose, was almost absent. In seeds of the line RRRbRb, a high level of verbascose was accumulated alongside with stachyose. The increase in verbascose in developing RRRbRb seeds was associated with galactinol-dependent verbascose synthase activity. In addition, a galactinol-independent enzyme activity was detected, which catalyzed transfer of a galactose residue from one stachyose molecule to another. The two enzyme activities synthesizing verbascose showed an optimum at pH 7.0. Both activities were almost undetectable in SD1. Maximum activity of stachyose synthase was about 4-fold higher in RRRbRb compared with SD1, whereas the activities of galactinol synthase and raffinose synthase were only about 1.5-fold higher in RRRbRb. The levels of galactinol synthase and stachyose synthase activity were reflected by steady-state levels of corresponding mRNAs. We suggest that the accumulation of verbascose in RRRbRb was controlled by a coordinated up-regulation of the last steps of verbascose biosynthesis.

Topics: Disaccharides; Galactosyltransferases; Hydrogen-Ion Concentration; Molecular Sequence Data; Oligosaccharides; Pisum sativum; Raffinose; Seeds

The objective of this study was to evaluate the effect of the soaking step and the domestic processing of the common bean, on the chemical composition, the levels of phytate, tannin, starch and flatulence factors by utilizing the follows treatments: raw bean (FC), freeze-dried cooked unsoaked bean (FCSM), freeze-dried cooked bean without the non-absorbed soaking water (FCSAM), freeze-dried cooked bean with the non-absorbed soaking water (FCCAM) and the soaking water (AM). The beans were soaking for a period for 16 hours in the proportion 3:1 (water:beans) at room temperature. The effect of the phytates and tannins on the net protein efficiency ratio (NPR) and protein digestibility using male Wistar rats were studied. A decrease in the phytate content of the beans (85%) with use of soaking was observed. In the case of the tannin content, only the cooking of the beans promoted high decomposition (84%). In the (FCSAM) treatment a decrease in the raffinose (25.0%), stachiose (24.8%), verbascose (41.7%) and starch (26.8%) contents was observed. Diets containing casein (control), casein plus the soluble solids obtain from the soaking water showed no significant difference (p > 0.05) for the NPR, as well as for the different bean treatments, although these showing lower values. The treatment (FCSM) showed the higher digestibility (74.3 +/- 5.8%) of the bean treatments, the casein diets showing 94.6 +/- 0.9%. The reduction of the phytates, tannin, starch contents and flatulence factors in the common bean was most effective when the soaking water not absorbed was discarded (FCSAM).

Topics: Animals; Carbohydrates; Cooking; Digestion; Flatulence; Food Handling; Glucosides; Male; Nutritive Value; Oligosaccharides; Phaseolus; Phytic Acid; Raffinose; Rats; Rats, Wistar; Starch; Tannins

Three conventional soybean meals (CSBM) and five low-oligosaccharide soybean meals (LOSBM) were evaluated. Meals were prepared by solvent-extracting flakes from generic soybean lines and soybean lines genetically modified to contain varying oligosaccharide content. The mean raffinose, stachyose, and galactinol levels in the CSBM were 0.58, 3.23, and 0%, respectively. The mean (range) raffinose, stachyose, and galactinol levels in the LOSBM were 0.08 (0.03 to 0.17%), 0.42 (0.04 to 1.08%), and 0.104 (0 to 0.24%), respectively. The crude fiber, fat, and ash contents of all SBM were similar, whereas the CP and sucrose contents of the LOSBM were slightly higher than the CSBM. The effect of SBM type on digesta passage rate, dry matter digestibility, and TMEN was assessed in conventional and cecectomized roosters using a precision-fed rooster assay (8 x 2 factorial design). Quantitative collection of excreta for 4, 8, 12, 16, 20, 24, and 48 h postfeeding indicated little or no effect of SBM type on digesta passage rate. The dry matter digestibility and TMEN values for the SBM were lower for cecectomized roosters compared with conventional roosters (P < 0.05), and there was no significant SBM x bird type interaction. The mean TMEN values (kcal/kg DM) for the CSBM and LOSBM were 2,739 and 2,931, respectively, which was a difference of 7% (P < 0.05). The two LOSBM with the lowest total raffinose, stachyose, and galactinol levels (0.07% and 0.25%) had average TMEN values that were 9.8% (P < 0.05) higher than their respective genetic controls. Digestibilities of raffinose and stachyose were much higher (P < 0.05) in conventional roosters than in cecectomized roosters. The results of this study indicated that the TMEN of LOSBM was higher than that of CSBM.

Topics: Animal Nutritional Physiological Phenomena; Animals; Chickens; Digestion; Energy Intake; Energy Metabolism; Genetic Engineering; Glycine max; Male; Nutritive Value; Oligosaccharides; Raffinose

The effect of various processing treatments viz., soaking, pressure cooking, open pan cooking, germination followed by pressure cooking and roasting of whole ricebean and pressure cooking, open pan cooking and fermentation and frying of fermented batter of dehulled ricebean flour was assessed for total sugars and oligosaccharide contents. Dehulling caused a significant reduction in the raffinose and stachyose contents. Pre-soaking of ricebean caused appreciable losses in the oligosaccharide content. Germination (48 h) followed by pressure cooking and fermentation (18 h) followed by frying caused the maximum losses in the raffinose and stachyose contents.

Topics: Chromatography, Paper; Cooking; Fabaceae; Fermentation; Flatulence; Food Handling; Germination; Oligosaccharides; Plants, Medicinal; Raffinose

Stachyose synthase (STS) (EC 2.4.1.67) was purified 313-fold from mature seeds of lentil. The final preparation had a specific activity of 9.09 nkat stachyose formed per milligram of protein. The enzyme was a monomeric protein with a molecular mass of 88.6 kDa (SDS-PAGE) and an isoelectric point of 4.8 (chromatofocusing). Western analysis revealed cross-reactivity of polyclonal antibodies raised against STS from adzuki bean with the lentil enzyme. The purified enzyme catalyzed a range of different galactosyl transfer reactions. In addition to the genuine STS reaction (raffinose + galactinol --> stachyose + myo-inositol), the enzyme catalyzed the reversible galactosyl transfer from galactinol to d-pinitol (1d-3-O-methyl-chiro-inositol), yielding galactopinitol A (O-alpha-d-galactopyranosyl-(1 --> 2)-4-O-methyl-d-chiro-inositol) and myo-inositol. Galactopinitol A could be further galactosylated by STS to give ciceritol (O-alpha-d-galactopyranosyl-(1 --> 6)-O-alpha-d-galactopyranosyl-(1 --> 2)-4-O-methyl-d-chiro-inositol). Enzymatic synthesis of galactopinitol A and ciceritol is a new observation. However, STS was not only able to utilize galactopinitol A as galactosyl acceptor, but also as galactosyl donor to form stachyose from raffinose. The role of STS in the metabolism of galactosyl cyclitols and oligosaccharides in plant seeds is discussed.

Topics: Disaccharides; Fabaceae; Galactosyltransferases; Inositol; Kinetics; Oligosaccharides; Plants, Medicinal; Raffinose; Seeds; Substrate Specificity

In order to measure broiler chick hydrogen gas production, a sealed atmosphere chamber was constructed and chicks were intubated with soybean meal (SBM), alpha-galactoside free, ethanol-extracted soybean meal (ESBM), and ESBM with alpha-galactosides added to the levels of SBM (ESBMG). Six male broiler chicks averaging 156 g of weight were deprived of feed for 12 h prior to intubation with 6 g of the test soybean meals. Two chicks were used for each treatment. Following intubation, chicks were placed in the sealed atmosphere chamber for 20 min at 2-h intervals for 28 h. At the end of this 20-min period, a sample of the chamber atmosphere was collected with a gas-tight syringe and analyzed for hydrogen gas by gas-solid chromatography. The hydrogen production of the two chicks intubated with SBM peaked 7 h postintubation at 127 ppm. The ESBM produced a peak at approximately 17 h postintubation at 26 ppm. Intubation with ESBMG resulted in peak hydrogen production at approximately 12 h postintubation at an average of 67 ppm. Results indicate that chicks intubated with SBM produced 3.2 times the amount of total hydrogen gas than those chicks intubated with ESBM. Chicks intubated with ESBMG produced 2.2 times the amount of total hydrogen gas than chicks intubated with ESBM. The research indicates the alpha-galactoside oligosaccharides are a major cause of hydrogen gas production from SBM in poultry.

Topics: Animal Feed; Animals; Chickens; Ethanol; Glycine max; Hydrogen; Male; Oligosaccharides; Raffinose

Soybean milk, which serves as a base for a variety of beverages, contains raffinose, stachyose, pentanal and n-hexanal; the former two may be responsible for flatulence after fermentation, whilst the latter two for a beany flavour. Twenty-seven strains of Bifidobacterium were analyzed for their alpha-galactosidase activity and the production of lactic and acetic acids to determine their potential for use in the production of fermented soymilk. The behaviour of three strains in soymilk was studied to determine their ability to reduce alpha-D-galactosyl oligosaccharides and produce lactic and acetic acids. They all were able to reduce stachyose and raffinose. Pentanal and n-hexanal were metabolized by Bifidobacterium breve MB233. These data indicate that bifidobacteria can be used for biotechnological processes that employ soymilk as the substrate. A product with low levels of alpha-D-galactosyl oligosaccharides and alkylic aldehydes may be obtained.

Topics: Acetic Acid; Aldehydes; alpha-Galactosidase; Animals; Bifidobacterium; Chromatography, High Pressure Liquid; Fermentation; Glycine max; Lactic Acid; Milk; Oligosaccharides; Raffinose

Antinutritional factors of anasazi bean were compared to traditional pinto bean (Phaseolus vulgaris L.). Anasazi beans contained less (p<0.001) soluble and bound condensed tannins compared to pinto beans. No differences (p>0.05) in stachyose and raffinose content were found between the two bean types; verbascose was not detected at all. Significant (p<0.05) differences in lectin content were observed between anasazi and pinto bean. The lectins of anasazi beans were classified as non toxic and those of the pinto beans as toxic types. No differences (p>0.05) in inhibitor activity against human and bovine trypsin and chymotrypsin were found between the two bean types.

Topics: Animals; Fabaceae; Galactosides; Humans; Lectins; Nutritive Value; Oligosaccharides; Plant Lectins; Plants, Medicinal; Raffinose; Sucrose; Tannins; Trypsin Inhibitors

A capillary zone electrophoretic (CZE) method is described for the analysis of the raffinose family of oligosaccharides (RFO) in pea seeds. Extraction of RFO was carried out in 80% ethanol and the extract was passed through a Sep-Pak C18 cartridge. This proved to be an improvement on the currently accepted purification technique. High-quality electrophoregrams were obtained which allowed the separation and quantitation of sucrose, raffinose, stachyose and verbascose The CZE results were compared with those obtained by anion-exchange high-performance liquid chromatography coupled to a triple-pulsed amperometric detection (HPAC-PAD). The samples were obtained from four pea strains which were near-isogenic except for genes at the r and rb loci. A high degree of precision and reproducibility was obtained for the RFO compositions of all the pea strains. No statistically significant differences (p < or equal to 0.05) were found between the two analytical techniques using paired Student-t tests.

Topics: Chromatography, High Pressure Liquid; Electrophoresis, Capillary; Fabaceae; Galactosides; Oligosaccharides; Plants, Medicinal; Raffinose; Seeds; Sucrose

One important aspect of the safety assessment of genetically engineered crops destined for food and feed uses is the characterization of the consumed portion of the crop. One crop currently under development, glyphosate-tolerant soybeans (GTS), was modified by the addition of a glyphosate-tolerance gene to a commercial soybean cultivar. The composition of seeds and selected processing fractions from two GTS lines, designated 40-3-2 and 61-67-1, was compared with that of the parental soybean cultivar, A5403. Nutrients measured in the soybean seeds included macronutrients by proximate analyses (protein, fat, fiber, ash, carbohydrates), amino acids and fatty acids. Antinutrients measured in either the seed or toasted meal were trypsin inhibitor, lectins, isoflavones, stachyose, raffinose and phytate. Proximate analyses were also performed on batches of defatted toasted meal, defatted nontoasted meal, protein isolate, and protein concentrate prepared from GTS and control soybean seeds. In addition, refined, bleached, deodorized oil was made, along with crude soybean lecithin, from GTS and control soybeans. The analytical results demonstrated the GTS lines are equivalent to the parental, conventional soybean cultivar.

Topics: Amino Acids; Carbohydrates; Fatty Acids; Food Handling; Glycine; Glycine max; Glyphosate; Herbicides; Isoflavones; Lectins; Oligosaccharides; Phytic Acid; Plant Lectins; Plant Proteins; Raffinose; Seeds; Trypsin Inhibitors; Urease

A nutritionally improved local snack compared to existing kokoro has been developed by extrusion cooking of different formulations of maize, soybean and condiments such as pepper, onion, salt, palm oil, plantain and banana. The improved snack was named as the 'soyabari snack stick'. The chemical composition of representative extruded products indicates a high level of crude protein, fat, energy, available lysine and improved in vitro digestibility compared to the usual maize-based products. The level of stachyose and raffinose were greatly reduced in the extruded products compared to raw soya. Formulations using various additives yielded products suitable for different consumers' preferences such as hot, sweet, bland, gritty or crispy and acceptable to taste assessors. Soyabari snack sticks were equally acceptable as Bombay mix, a product on the market in London. Sensory analysis showed no significant differences in the two products but the crude fibre content of Bombay mix was higher while the protein was slightly lower than for soyabari sticks. Local ingredients can produce acceptable extrudates.

Topics: Dietary Fats; Dietary Proteins; Energy Metabolism; Food Analysis; Food, Fortified; Glycine max; Humans; Lysine; Nigeria; Nutritive Value; Oligosaccharides; Plant Proteins, Dietary; Raffinose; Soybean Proteins; Taste; Zea mays

Topics: Animals; Antibodies, Heterophile; Antibody Affinity; Antibody Specificity; Antigens, Heterophile; Carbohydrate Sequence; Disaccharides; Humans; Immunoglobulin M; Melibiose; Molecular Sequence Data; Oligosaccharides; Raffinose; Swine

Recent literature has suggested that the depression of the phase transition temperature (Tm) in dry phospholipids by sugars may be ascribed to vitrification of the stabilizing solute, rather than by the direct interaction between sugar and phospholipid we have proposed. Koster et al. ((1994) Biochim. Biophys. Acta 1193, 143-150) claim that the only necessity is that the glass transition (Tg) for the sugar exceed Tm for the lipid. Evidence is presented in the present paper that this is not sufficient. Based on the vitrification hypothesis of Koster et al., the predicted order of effectiveness in depressing Tm in dry dipalmitoylphosphatidylcholine (DPPC) is dextran > or = hydroxyethyl starch > stachyose > raffinose > trehalose > sucrose > glucose. In fact, the opposite order was seen. The effect of raffinose, sucrose, or trehalose on Tm in dry DPPC depends on the thermal history of the sample, as we have reported previously. When DPPC dried with trehalose is heated for the first time, Tm is about 55 degrees C, but on the second and subsequent heating scans Tm falls to about 25 degrees C. Koster et al. suggest that this effect is due to heating the sample above Tg rather than to melting the hydrocarbon chains. We present evidence here that all that is required is for the chains to be melted. Further, we show that retention of residual water by DPPC dried with trehalose depends on the drying temperature, but is independent of drying temperature with glucose, a finding that is consistent with direct interaction. We conclude that vitrification is not in itself sufficient to depress Tm in dry phospholipids.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Dextrans; Glucose; Hot Temperature; Hydroxyethyl Starch Derivatives; Membrane Fusion; Oligosaccharides; Raffinose; Sucrose; Trehalose

We investigated digestion responses to conventional and low oligosaccharide soybean meal (SBM) incorporation into diets for dogs. Five female dogs were fitted with T-type cannulas at the terminal ileum and fed five diets in a 5 x 5 Latin square design. Corn grain + poultry meal-based diets containing different levels and types of SBM (0% SBM, 18.55% conventional SBM, 18.55% low oligosaccharide SBM, 37.1% conventional SBM, 37.1% low oligosaccharide SBM) were formulated. Each period consisted of 11 d (7-d diet adaptation; 4-d collection of ileal digesta and feces). Intakes of DM, OM, CP, fat, and GE were not affected (P > .10) by treatment. Higher (P < .01) starch intakes and higher (P < .05) total dietary fiber (TDF) intakes were noted for dogs fed diets with SBM. Digestibilities of CP (P < .04) and starch (P < .002) at the ileum were higher for dogs fed the higher levels of SBM. Ileal digestibilities of most individual amino acids followed the CP response. Total tract digestibility of CP was higher (P < .006) in dogs fed the SBM diets. There were no significant differences in nutrient digestibilities between conventional and low oligosaccharide SBM. Stachyose and raffinose intakes by dogs were decreased dramatically (P < .001) as a result of substituting the low oligosaccharide SBM for conventional SBM at the higher dietary concentration, although sucrose intake by dogs fed low oligosaccharide SBM was higher (P < .001). Galactinol was present in low oligosaccharide SBM but not in conventional SBM. Total tract digestion of all oligosaccharides was near 100%. The low oligosaccharide SBM was digested as extensively, but no better than, conventional SBM.

Topics: Amino Acids; Animals; Diet; Dietary Fiber; Digestion; Dogs; Female; Glycine max; Ileum; Oligosaccharides; Raffinose

The use of intracellular alpha-galactosidase from Gibberella fujikuroi to remove raffinose and stachyose in soymilk was studied. The optimum conditions for the enzymic hydrolysis of raffinose and stachyose was pH 5.5 to 6.0 at 55 degrees C. Alpha-galactosidase showed optimum activity at pH 5.0 and 50 degrees C with the substrate p-nitrophenyl-alpha-D-galacto-pyranoside (PNGP). The enzyme showed no detectable loss of activity when held more than 8 hr at 50 degrees C. Thin layer chromatography (TLC) revealed the following composition of oligosaccharides in local soybean variety: sucrose, 5.53%; raffinose, 1.95%; and stachyose, 6.1%. Investigation by TLC showed complete hydrolysis of raffinose and stachyose in 3 hr. HPLC analysis of hydrolyzate indicated complete hydrolysis of stachyose, and more than 60% hydrolysis of raffinose in 2.5 hr.

Topics: alpha-Galactosidase; beta-Fructofuranosidase; Chromatography, High Pressure Liquid; Chromatography, Thin Layer; Enzyme Stability; Glycine max; Glycoside Hydrolases; Hot Temperature; Hydrogen-Ion Concentration; Kinetics; Oligosaccharides; Plants; Raffinose; Substrate Specificity; Thermodynamics

Forty-six strains of tempe-forming Rhizopus species were screened for their ability to grow on raffinose as the sole carbon source. Six of the strains showed good growth and sporulation. These isolates were one Rhizopus oligosporus, one Rhizopus microsporus var. chinensis, three Rhizopus oryzae and one Rhizopus stolonifer. These six moulds and R. oligosporus strain NRRL 2710 were investigated for their metabolism of the raffinose family of alpha-galactoside carbohydrates. Degradation experiments were performed in submerged culture in a medium containing soybean alpha-protein, sodium phytate and either stachyose, raffinose or melibiose. R. oryzae and R. stolonifer completely consumed the tested carbohydrates as carbon source. R. microsporus var. chinensis failed to hydrolyse the alpha-galactosidic bonds of raffinose, stachyose or melibiose, whereas it was able to use sucrose and the fructose moiety of raffinose or stachyose for growth. R. oligosporus NRRL 2710 was unable to hydrolyse any of the tested carbohydrates. The results of the oligosaccharide degradation experiments could be verified during tempe production from soybeans with the selected fungal species.

Topics: Biodegradation, Environmental; Chromatography, High Pressure Liquid; Fermentation; Melibiose; Oligosaccharides; Raffinose; Rhizopus; Sucrose

Anti-nutritional factors of developing seeds and pod walls of fenugreek were evaluated which affect their nutritional value. Sucrose, raffinose and stachyose decreased with maturity of seeds in all the cultivars. Total, non-reducing sugars increased and reducing sugars decreased at maturity in all the cultivars. The reducing sugars decreased with maturity of pod walls. The flatus producing sugars were maximum in PEB pod walls. Saponin content increased towards maturity of seeds but decreased at maturity of seeds and decreased at maturity in pod walls of all cultivars. Phytate increased with seed maturity up to 95 days after anthesis, while phytate decreased in pod walls of all the cultivars with maturity. Total phenol, catechol and flavonol decreased with advancement of pod wall development. Total phenol decreased except HM 46 a maturity of seeds.

Topics: Catechols; Fabaceae; Flavonoids; Flavonols; Magnoliopsida; Nutritive Value; Oligosaccharides; Phenol; Phenols; Phytic Acid; Plants, Medicinal; Raffinose; Saponins; Seeds; Sucrose

Soybeans contain the enzyme alpha-galactosidase, which hydrolyzes alpha-1, 6 linkages in stachyose and raffinose to give sucrose and galactose. We have found that galactose, a competitive product inhibitor of alpha-galactosidase, strongly promotes the heat stability of the tetrameric form of the enzyme at pH 4.0 and at temperatures of up to 70 degrees C for 60 min. Stachyose and raffinose also protect alpha-galactosidase from denaturation at pH 4.0 although to a lesser extent. Glucose and mannose have little effect. At pH 7.0 the enzyme is a monomer, and galactose has no effect on the heat stability of the enzyme. In the absence of heat protection of the enzyme by added sugars, a series deactivation mechanism was found to describe the deactivation data. In comparison, a unimolecular, non-first order deactivation model applies at pH 4.0, where heat protection effects were observed. At a temperature above 60 degrees C, simple deactivation is a suitable model. The results suggest that alpha-galactosidase conformation and heat stability are directly related.

Topics: alpha-Galactosidase; Enzyme Stability; Galactose; Glycine max; Hot Temperature; Hydrogen-Ion Concentration; Hydrolysis; Models, Biological; Oligosaccharides; Plant Proteins; Raffinose; Sucrose

Galactose oxidase is a fungal enzyme which is known to oxidize the C-6 hydroxymethyl of galactose to an aldehyde group. When the products of a galactose oxidase-catalase treatment of raffinose were examined by gel filtration and ion exchange chromatography, we found that, in addition to the expected 6''-aldehydoraffinose, two other components were present. Of these two components, the major one was retained on a column of AG 1-X8 (formate), gave a positive carbazole reaction for uronic acid, and on paper chromatograms had a mobility identical with that of 6''-carboxyraffinose. The infrared spectrum of the compound showed a carbonyl absorbance at 1725 cm-1 and was distinguishable from the spectra of raffinose and 6''-aldehydoraffinose. These data showed that raffinose was partly converted to 6''-carboxyraffinose when treated with galactose oxidase and catalase. The conversion of [3H]raffinose to [3H]6''-carboxyraffinose increased gradually with time of oxidation from 22% at 6 h to 68% at 96 h. Results of other experiments provided evidence that this was an enzymic conversion and depended on the presence of galactose oxidase. The activities responsible for the formation of aldehyde and uronic acid could not be separated by affinity chromatography, gel electrophoresis, or ion exchange chromatography, indicating that the same enzyme is responsible for both activities. Treatment of galactose, melibiose, and stachyose with galactose oxidase and catalase also resulted in the formation of the corresponding uronic acids. These studies indicate that galactose oxidase not only converts the C-6 hydroxymethyl group of galactose to an aldehyde group, but also catalyzes further oxidation to the carboxyl group.

Topics: Animals; Catalase; Cattle; Chromatography, Affinity; Chromatography, Gel; Chromatography, Ion Exchange; Chromatography, Paper; Galactose; Melibiose; Oligosaccharides; Oxidation-Reduction; Raffinose; Spectrophotometry, Infrared; Time Factors; Uronic Acids

Topics: alpha-Galactosidase; Carbohydrates; Glycoside Hydrolases; Oligosaccharides; Raffinose; Sucrose; Sweetening Agents