Determination of Molecular Dimensions of Carbohydrate Polymers (Polysaccharides) by Size Exclusion Chromatography (SEC).

Calibrated size exclusion chromatography (SEC) is a useful tool for the analysis of molecular dimensions of polysaccharides. The calibration takes place with a set of narrow distributed dextran standards and peak position technique. Adapted columns systems and dissolving processes enable for the adequate separation of carbohydrate polymers. Plant-extracted fructan (a homopolymer with low molar mass and excellent water solubility) and mucilage (differently structured, high molar mass heteropolysaccarides that include existing supramolecular structures, and require a long dissolving time) are presented as examples of the versatility of this technique. Since narrow standards similar to the samples (chemically and structurally) are often unavailable, it must be noted that the obtained molar mass values and distributions by this method are only apparent (relative) values, expressed as dextran equivalents.

Phytochemical Characterization of Purple Coneflower Roots (Echinacea purpurea (L.) Moench.) and Their Extracts.

Echinacea purpurea is a perennial plant that belongs to the Asteraceae family. It has a wide range of applications mainly in the treatment and prevention of inflammations in the respiratory system. The current study aimed to perform a phytochemical characterization of purple coneflower (Echinacea purpurea) roots and their extracts (water, 40%, 50%, 60% ethanol, and 60% glycerol). Phytochemical characterization was carried out by gravimetric, spectrophotometric, and chromatographic methods. Echinacea roots were characterized by a low lipid (0.8%) content. In contrast, carbohydrates (45%) and proteins (20%) occupied a large part of the dry matter. Amongst the extracts, the highest yield was obtained using water as a solvent (53%). Water extract was rich in protein and carbohydrates as fructans (inulin) were the most abundant carbohydrate constituent. The most exhaustive recovery of the phenolic components was conducted by extraction with 40% ethanol and 60% glycerol. It was found that water is the most suitable extractant for obtaining a polysaccharide-containing complex (PSC) (8.87%). PSC was composed mainly of fructans (inulin) and proteins with different molecular weight distributions. The yield of PSC decreased with an increasing ethanol concentration (40% > 50% > 60%) but the lowest yield was obtained from 60% glycerol extract. The obtained results showed that Echinacea roots contained a large amount of biologically active substances-phenolic components and polysaccharides and that glycerol was equally efficient to ethanol in extracting caffeic acid derivatives from purple coneflower roots. The data can be used for the preparation of extracts having different compositions and thus easily be incorporated into commercial products.

Physical Properties and Prebiotic Activities (Lactobacillus spp.) of Gelatine-Based Gels Formulated with Agave Fructans and Agave Syrups as Sucrose and Glucose Substitutes.

This research developed model foods of gelatine-based gels, where carbohydrates from Agave tequilana Weber var. Azul (agave syrups or/and agave fructans) were incorporated into gel formulations as healthy sucrose and glucose substitutes. The sugars (sucrose and glucose) were substituted by agave carbohydrates (agave syrups and agave fructans), obtaining the subsequent gel formulations: 100% agave syrup (F2 gel), 100% agave fructan (F3 gel), and 50% agave syrup−50% agave fructan (F4 gel). The unsubstituted gel formulation was used as a control (F1 gel). The prebiotic activities, physical properties, thermal stability (HP-TLC), and texture of gelatine-based gels were evaluated. The gel formulations showed translucent appearances with approximately 36 g/100 g of water and water activities values between 0.823 and 0.929. The HP-TLC analysis validated that agave fructans did not hydrolyse during the thermal process of gels production. Gels produced with agave syrup and agave fructan (F2-F4 gels) provided higher hardness, gumminess, and springiness values (p < 0.05) than those produced with glucose and sucrose (F1 gel). Gelatine-based gel formulations displayed prebiotic activities correlated to the ability of Lactobacillus plantarum, Lactobacillus paracasei, and Lactobacillus rhamnosus to use agave carbohydrates as carbon sources. Based on the prebiotic effect and physical and textural properties, the F2 and F4 gel formulations displayed the best techno-functional properties to produce gel soft candies.

Effect of an inulin-type fructan from Platycodon grandiflorum on the intestinal microbiota in rats exposed to PM2.5.

In this study, an inulin-type fructan (PGPI-1-a) was isolated from the roots of Platycodon grandiflorum. PGPI-1-a was composed of (2 â†’ 1)-linked ß-D-fructofuranose (Fruf) and a terminal α-d-glucopyranose (Glcp) with a molecular weight of 12.1 kDa. PM2.5 exposure has brought a great threat to human health in recent years. Therefore, this study explored the effect of PGPI-1-a on the intestinal microbial community structure of rats exposed to PM2.5 using the animal model of PM2.5 inhalation exposure. The results showed that PGPI-1-a could regulate the intestinal microbiota by partly restoring the perturbed levels of Peptoniphilaceae_[G-2] and Lachnospiraceae_[G-2] caused by PM2.5 exposure. In addition, the relative abundance of Butyrivibrio, a butyric acid-producing genera, significantly increased after PGPI-1-a intervention. These results indicated that PGPI-1-a could improve the imbalance of intestinal microbiota due to PM2.5 exposure to a certain extent.

In vitro and in silico anti-osteoporosis activities and underlying mechanisms of a fructan, ABW90-1, from Achyranthes bidentate.

Achyranthes bidentata is a traditional Chinese medicine used to treat osteoporosis. AB90, a crude saccharide from A. bidentata, showed excellent osteoprotective effects in ovariectomized rats, and ABW90-1, an oligosaccharide purified from AB90, stimulated significant differentiation of osteoblasts. However, the osteogenic effects and underlying mechanisms of ABW90-1 have remained unknown. In the present study, we found that ABW90-1 significantly promoted ALP activity, mineralization, and the expression of osteogenic markers in MC3T3-E1 cells. ABW90-1 showed strong binding with the WNT signaling complex and BMP2 based on number of interactions, hydrogen bond length, and binding energy in silico. ABW90-1 significantly increased the expression of active-ß-catenin, p-GSK-3ß, LEF-1, BMP2, and p-SMAD1. Importantly, the osteogenic effects of ABW90-1 were partially suppressed by DKK-1 and Noggin, which are specific inhibitors of the WNT and BMP signaling pathways, respectively. Collectively, these findings suggest that ABW90-1 has osteogenic effects through crosstalk between WNT/ß-catenin and BMP2/SMAD1 signaling pathways.

Comprehensive Metabolite Profiling in Genetic Resources of Garlic (Allium sativum L.) Collected from Different Geographical Regions.

Garlic (Allium sativum) is the second most important Allium crop that has been used as a vegetable and condiment from ancient times due to its characteristic flavor and taste. Although garlic is a sterile plant that reproduces vegetatively through cloves, garlic shows high biodiversity, as well as phenotypic plasticity and environmental adaptation capacity. To determine the possible mechanism underlying this phenomenon and to provide new genetic materials for the development of a novel garlic cultivar with useful agronomic traits, the metabolic profiles in the leaf tissue of 30 garlic accessions collected from different geographical regions, with a special focus on the Asian region, were investigated using LC/MS. In addition, the total saponin and fructan contents in the roots and cloves of the investigated garlic accessions were also evaluated. Total saponin and fructan contents did not separate the garlic accessions based on their geographical origin, implying that saponin and fructan contents were clone-specific and agroclimatic changes have affected the quantitative and qualitative levels of saponins in garlic over a long history of cultivation. Principal component analysis (PCA) and dendrogram clustering of the LC/MS-based metabolite profiling showed two major clusters. Specifically, many Japanese and Central Asia accessions were grouped in cluster I and showed high accumulations of flavonol glucosides, alliin, and methiin. On the other hand, garlic accessions grouped in cluster II exhibited a high accumulation of anthocyanin glucosides and amino acids. Although most of the accessions were not separated based on country of origin, the Central Asia accessions were clustered in one group, implying that these accessions exhibited distinct metabolic profiles. The present study provides useful information that can be used for germplasm selection and the development of new garlic varieties with beneficial biotic and abiotic stress-adaptive traits.

Reducing FODMAPs and improving bread quality using type II sourdough with selected starter cultures.

This study focussed on lactic acid bacteria (LAB) screening for sourdough type II elaboration and evaluating the effects of sourdough fermentation in bread making, focussing mainly on reducing FODMAPs. After a technological performance screening, six strains (Levilactobacillus brevis, Weissella minor, Lactiplantibacillus plantarum, Leuconostoc citreum, Limosilactobacillus fermentum, and Companilactobacillus farciminis) were selected for sourdough preparation. Total titratable acidity, pH, specific volume, and enumeration of microorganisms were carried out on sourdoughs, doughs, and breads. Breads were subjected to texture profile and colour analysis, moulds and yeast enumeration, and total fructans (main group of FODMAPs) quantification. Breads produced with sourdough showed a significant reduction of fructans, greater acidity, volume, and better performance during storage when compared to fermentation using only baker's yeast. Including specific cultures as starters in sourdough reduced fructans content by >92%, thereby producing a low FODMAP bread suitable for Irritable Bowel Syndrome patients with improved nutritional and technological properties.

Fundamental study on changes in the FODMAP profile of cereals, pseudo-cereals, and pulses during the malting process.

Whole grains and pulses are rich in nutrients but often avoided by individuals with gastrointestinal disorders, due to high levels of fermentable oligo-, di-, monosaccharides and polyols (FODMAPs). This study investigated the impact of malting as delivery-system for endogenous enzymes. Malts from barley and wheat (naturally high in fructans), lentils and chickpeas (high in galactooligosaccharides), oat and buckwheat (low in FODMAPs) were produced. While barley and wheat malts had slightly elevated fructan-levels, in oat malt 0.8 g/100 g DM fructans were de novo synthesized. In lentils and chickpeas galactooligosaccharides diminished by 80-90%. Buckwheat did not contain any FODMAPs commonly investigated, but fagopyritols which may have a similar physiological effect. Also fagopyritols were degraded. While malted pulses and buckwheat are directly suitable for low FODMAP applications, using the combined approach of malting and fermentation, malted cereals could contribute to high nutritional values of such products.

Effects of fructans and probiotics on the inhibition of Klebsiella oxytoca and the production of short-chain fatty acids assessed by NMR spectroscopy.

Generally, the selection of fructans prebiotics and probiotics for the formulation of a symbiotic has been based on arbitrary considerations and in vitro tests that fail to take into account competitiveness and other interactions with autochthonous members of the intestinal microbiota. However, such analyzes may be a valuable step in the development of the symbiotic. The present study, therefore, aims to investigate the effect of lactobacilli strains and fructans (prebiotic compounds) on the growth of the intestinal competitor Klebsiella oxytoca, and to assess the correlation with short-chain fatty acids production. The short-chain fatty acids formed in the fermentation of the probiotic/prebiotic combination were investigated using NMR spectroscopy, and the inhibitory activities were assessed by agar diffusion and co-culture methods. The results showed that Lactobacillus strains can inhibit K. oxytoca, and that this antagonism is influenced by the fructans source and probably associated with organic acid production.

Arctium lappa root extract containing L-arginine prevents TNF-α-induced early atherosclerosis in vitro and in vivo.

Atherosclerosis is a chronic inflammatory disease affecting the aorta and is a major cause of cardiovascular disease. Arctium lappa root is a plant widely used in traditional Chinese medicine (TCM), and Arctium lappa root extract (ALE) has been reported to exhibit anti-inflammatory capacity and to ameliorate endothelial dysfunction. Thus, we hypothesized that ALE would inhibit the early atherosclerotic stage. In this study, we evaluated the inhibitory effect of ALE on early arteriosclerosis and its mechanisms of action. ALE suppressed TNF-α-induced monocyte adhesion to the vascular endothelium by suppressing NF-κB signaling in HUVECs. In an acute mouse model of atherosclerosis, ALE suppressed TNF-α-induced monocyte infiltration of the vascular endothelium and the expression of genes encoding inflammatory cytokines including IL-1ß, IL-6, TNF-α, and MCP-1 in the mouse aorta. Moreover, inulin-type fructan and amino acids, especially L-aspartate and L-arginine (60.27 and 42.17 mg/g, respectively) were detected by NMR, MALDI-TOF MS, and HPLC analysis as the main components of ALE. Notably, L-arginine suppressed TNF-α-induced monocyte adhesion to HUVECs. Therefore, these results suggest that ALE may be a functional food for the suppression or prevention of early stages of atherosclerosis.

Raman spectroscopy for the analysis of different exo-polysaccharides produced by bacteria.

In this study, Raman spectroscopy is employed for the characterization and comparison of two different classes of exo-polysaccharides including glucans and fructans which are produced by different bacteria. For this purpose, nine samples are used including five samples of glucans and four of fructans. Raman spectral results of all these polysaccharides show clear differences among various glucans as well as fructans showing the potential of this technique to identify the differences within the same class of the compounds. Moreover, these two classes are also compared on the basis of their Raman spectral data and can be differentiated on the basis of their unique Raman features. Multivariate data analysis techniques, Principle Component Analysis (PCA) is found very helpful for the comparison of the Raman spectral data of these classes of the carbohydrates.

Variability in yeast invertase activity determines the extent of fructan hydrolysis during wheat dough fermentation and final FODMAP levels in bread.

Consumption of fructan-containing cereal products is considered beneficial for most people, but not for those suffering from irritable bowel syndrome (IBS), as they should avoid the consumption of fermentable oligosaccharides, disaccharides, monosaccharides and polyols (acronym: FODMAP). Controlling fructan levels in cereal products is not trivial. However, controlling yeast invertase-mediated hydrolysis of fructan during dough fermentation might offer a handle to modulate fructan concentrations. In this work, the variability in invertase activity and substrate specificity in an extensive set of industrial Saccharomyces cerevisiae strains is investigated. Analysis showed a high variability in the capacity of these strains to hydrolyse sucrose and fructo-oligosaccharides. Industrial yeast strains with a high activity towards fructo-oligosaccharides efficiently reduced wheat grain fructans during dough fermentation to a final fructan level of 0.3% dm, whereas strains with a low invertase activity yielded fructan levels around 0.6% dm. The non-bakery strains produced lower levels of CO2 in fermenting dough resulting in lower loaf volumes. However, CO2 production and loaf volume could be increased by the addition of 3% glucose. In conclusion, this study shows that variation in yeast invertase activity and specificity can be used to modulate the fructan content in bread, allowing the production of low FODMAP breads, or alternatively, breads with a higher soluble dietary fibre content.

Comparative studies on characterization, saccharide mapping and antiglycation activity of polysaccharides from different Polygonatum ssp.

Multiple species of Polygonatum are used as "Huangjing" in China and polysaccharides are the main effective constituents of "Huangjing". However, there is a lack of systematic comparison of polysaccharides from Polygonatum spp., and little work concerning on the polysaccharides from unofficial species. This paper conducted a comparison study on the characterization, high-performance gel permeation chromatography (HPGPC) saccharide mapping and antiglycation assay of polysaccharides and their enzymatic hydrolysates from nine species of Polygonatum. The results showed that polysaccharides from P. macropodium were mainly composed of fructans (3.3 × 103 Da); while polysaccharides from other eight Polygonatum spp. contained both fructans (2.5 × 103-4.1 × 103 Da) and pectins (more than 4.1 × 105 Da). Antiglycation assay showed that polysaccharides from Polygonatum spp. could significantly inhibit the formation of AGEs, and degradation of pectins and fructans could result in loss of activity. The results are helpful to the rational usage and quality control of Polygonatum spp.

Characterisation and saccharide mapping of polysaccharides from four common Polygonatum spp.

Polysaccharides from 14 batches of Polygonatum sibiricum (PS), P. cyrtonema (PC), P. kingianum (PK) and P. odoratum (PO) were compared based on high-performance gel permeation chromatography (HPGPC) saccharide mapping, monosaccharide composition, molecular weight distribution and Fourier transform infrared (FTIR) spectra. Results showed that polysaccharides from PS, PC and PK exhibited two different molecular weight fractions and that one was more than 4.1 × 105 Da (P1) and the other was 2.8-5.4 × 103 Da (P2); while the polysaccharides from PO displayed only one main peak (P2). The analysis of monosaccharide composition and HPGPC saccharide mapping proved that P1 and P2 were composed of pectins and fructans, respectively. The FTIR spectra indicated that these polysaccharides had different degrees of esterification. This study provided a systematic profiling of polysaccharides of Polygonatum spp. and was helpful in understanding the varied functions of different Polygonatum spp., based on chemical composition.

Fermentation capacities of fructan- and pectin-rich by-products and purified fractions via an in vitro piglet faecal model.

BACKGROUND: Dietary strategies such as the inclusion of prebiotics have been suggested for modulating intestinal microbiota. In piglets, this strategy could result in a reduction of post-weaning-associated disorders and the use of antibiotics. To date, mainly purified fractions have been tested for their prebiotic effects at weaning while trials of potential health-promoting effects of products and corresponding by-products remain rare. In this study, fructan- and pectin-based ingredients have been tested in a two-step in vitro model for their fermentation kinetics as well as for their short-chain fatty acid production and microbiota profiles in fermentation broth as indicators for their prebiotic activity. RESULTS: Chicory root, in contrast to chicory pulp, exhibited an extensive and rapid fermentation similar to inulin and oligofructose, although butyrate levels of root and pulp did not reach those of the purified fractions. Chicory pulp showed higher relative levels of Lactobacillus spp., Bifidobacterium spp., Clostridium cluster IV and butyryl-CoA:acetate-CoA transferase gene abundance compared to chicory root. Sugar beet pulp, orange and citrus by-products displayed extensive gas fermentation patterns, equivalent to those of purified pectin, and revealed an elevated butyrate production compared to purified pectin. Moreover, several orange and citrus by-products displayed significantly higher relative levels of Bifidobacterium spp. in comparison to purified pectin. CONCLUSIONS: Chicory root and pulp as well as orange and citrus by-products appear to be promising ingredients for piglet diets for modulating intestinal fermentation for health purposes. © 2019 Society of Chemical Industry.

Hydration shells of carbohydrate polymers studied by calorimetry and terahertz spectroscopy.

We present a study of the hydration shells of some carbohydrate polymers of commercial and biological importance, namely, agave fructans, inulin, and maltodextrin, employing terahertz time-domain spectroscopy and differential scanning calorimetry. We observe that the hydration numbers calculated using terahertz spectroscopy are marginally higher than those of the calorimetric values. We attribute this discrepancy to the definition of hydration number, which in a way correlates with the physical process used to quantify it. The aqueous solutions show a non-proportional increase in the absorption coefficient and the hydration number, with a decrease in the carbohydrate concentration. We demonstrate that this behavior is consistent with the "chaotropic" or "structure breaking" model of the hydration shell around the carbohydrates. In addition, the study reveals that agave fructans and inulin have good hydration ability. Given the high glass transition temperature and good hydration ability, these carbohydrates may behave as good bio-protectants and hydrating additives for food and beverages.

Understanding the effect of Pulsed Electric Fields on multilayered solid plant foods: Bunching onions (Allium fistulosum) as a model system.

While it is well known that the nature of the applied electric field and the heterogeneity of the tissue can influence the impact of PEF treatment on the plant tissues found in plant-based foods, few studies have investigated the influence of PEF on plant structures that are made up of multiple structurally similar organs. The aim of this study was to understand the effect of pulsed electric fields (PEF), at different electric field strengths (0, 0.3, 0.7 and 1.2 kV/cm) and specific energy (7, 21 and 52 kJ/kg), on a multilayered plant material, with bunching onion bulb tissues being used as a model system. The present study found that carbohydrates leakage was an appropriate index to assess PEF induced damage and that plasmolysis of epidermal cells was a good indicator of plasma membrane integrity after PEF. In addition, electric field strength had a greater impact on the cell integrity than specific energy applied. While other studies have shown that different cell types have different sensitivities to PEF, using plasmolysis as an indicator of cell damage, this study clearly showed that the same PEF treatment conditions had a greater effect on the epidermal cells of the outer scales compared to the inner scales. Hence, while different plant cell types vary in their sensitivities to PEF the spatial location of the same cell type within a complex plant material made up of multiple similar organs, i.e. an onion bulb, can also influence how cells respond to the PEF treatment. Despite PEF induced disruption at the cellular level being detected by carbohydrate leakage, the epidermal cell plasmolysis test and by cryo-scanning electromicroscopy (cryo-SEM), no gross structural changes at the organ level were observed using cryo-SEM or fluorescence microscopy. This study also reports for the first time that PEF treatment can enhance fructan leakage from onion bulbs, which means that PEF treatments have the potential to manipulate the fructan contents of some plant-based foods.

Optimization and Validation of an HPAEC-PAD Method for the Quantification of FODMAPs in Cereals and Cereal-Based Products.

This study presents an analytical method for the quantification of fermentable oligo-, di-, and monosaccharides and polyols (FODMAPs) in cereals and cereal-based products, considering diverse ingredients, such as different cereals in addition to wheat, pulses, or pseudocereals. All carbohydrates have been separated, identified, and quantified with a high-performance anion-exchange chromatographic system coupled with a pulsed amperometric detection (HPAEC-PAD). The total fructan content and the average degree of polymerization (DPav) have been determined after enzymatic hydrolysis to the monomers glucose and fructose, on the basis of the principle of the official method for fructan quantification in food products, AOAC 997.08. The methods for extraction, separation, and detection as well as fructan determination are based on several other studies and were modified in order to minimize interferences in the analysis. The method has been validated with regard to the limits of detection and quantification, the linearity, the repeatability, and the accuracy as well as the DPav of the fructans.

Determination of Fructan (Inulin, FOS, Levan, and Branched Fructan) in Animal Food (Animal Feed, Pet Food, and Ingredients): Single-Laboratory Validation, First Action 2018.07.

Traditional enzyme-based methods for measurement of fructan were designed to measure just inulin and branched-type (agave) fructans. The enzymes employed, namely exo-inulinase and endo-inulinase, give incompletely hydrolysis of levan. Levan hydrolysis requires a third enzyme, endo-levanase. This paper describes a method and commercial test kit (Megazyme Fructan Assay Kit) for the determination of all types of fructan (inulin, levan, and branched) in a variety of animal feeds and pet foods. The method has been validated in a single laboratory for analysis of pure inulin, agave fructan, levan, and a range of fructan containing samples. Quantification is based on complete hydrolysis of fructan to fructose and glucose by a mixture of exo-inulinase, endo-inulinase, and endo-levanase, followed by measurement of these sugars using the PAHBAH reducing sugar method which gives the same color response with fructose and glucose. Before hydrolysis of fructan, interfering sucrose and starch in the sample are specifically hydrolyzed and removed by borohydride reduction. The single-laboratory validation (SLV) outlined in this document was performed on commercially available inulin (Raftiline) and agave fructan (Frutafit®), levan purified from Timothy grass, two grass samples, a sample of legume hay, two animal feeds and two barley flours, one of which (Barley MAX®) was genetically enriched in fructan through plant breeding. Parameters examined during the validation included working range, target selectivity, recovery, LOD, LOQ, trueness (bias), precision (repeatability and intermediate precision), robustness, and stability. The method is robust, quick, and simple.

Gluten-free and low-FODMAP sourdoughs for patients with coeliac disease and irritable bowel syndrome: A clinical perspective.

Wheat- and gluten-containing products are often blamed for triggering a wide range of gastrointestinal symptoms, and this has fueled demand for gluten-free products worldwide. The best studied 'gluten intolerance' is coeliac disease, an auto-immune disease that affects the small intestine. Coeliac disease occurs in 1% of the population and requires strict, life-long avoidance of gluten-containing foods as the only medical treatment. There is a larger group of individuals (around 10-15% of the population) who report a wide-range of gastrointestinal symptoms that respond well to a 'gluten-free diet', but who do not have coeliac disease - so called 'non-coeliac gluten sensitivity (NCGS)'. The team at Monash University has identified other factors in gluten-containing foods that may be responsible for symptoms in this group of individuals with so-called, NCGS. We have evidence that certain poorly absorbed short chain carbohydrates (called FODMAPs) present in many gluten-containing food products, induce symptoms of abdominal pain, bloating, wind and altered bowel habit (associated with irritable bowel syndrome, IBS). Our research has shown that FODMAPs, and not gluten, triggered symptoms in NCGS. Going forward, there are great opportunities for the food industry to develop low FODMAP products for this group, as choice of grain variety and type of food processing technique can greatly reduce the FODMAP levels in foods. The use of sourdough cultures in bread making has been shown to reduce the quantities of FODMAPs (mostly fructan), resulting in bread products that are well tolerated by patients with IBS. Greater interaction between biomedical- and food-scientists will improve understanding about the clinical problems many consumers face, and lead to the development of food products that are better tolerated by this group.