Glycosides Research Articles

Purification and structural characterization of a neutral polysaccharide from Boletus auripes using self-made quaternary chitosan cryogel.

The purification of polysaccharides is an essential preliminary step in determining their chemical structure, although it presents significant challenges. In this research, a macro-porous monolith of quaternary chitosan cryogel was synthesized for the purification of a neutral polysaccharide from Boletus auripes. A homogeneous neutral polysaccharide (BAP-1a1) with a weight-average molecular weight of 4.13×105Da and a polydispersity index of 1.28 was successfully isolated. The structure of BAP-1a1 was elucidated through a comprehensive characterization utilizing size exclusion chromatography (SEC) combined with laser light scattering (LLS), infrared spectroscopy, monosaccharide composition analysis, methylation analysis, and nuclear magnetic resonance (NMR) spectroscopy. The results revealed that the BAP-1a1 was characterized as a glucan with a backbone structure consisting of 1,4-α-D-Glcp and 1,3-β-D-Glcp glycosidic linkages in a molar ratio of 2:1. Additionally, a minority of branched chains of 1-α-D-Glcp are attached to 1,3-β-D-Glcp residues at the C6 position. In vitro antioxidant activity assays demonstrated that BAP-1a1 exhibits a dose-dependent scavenging effect on ABTS and DPPH radicals with EC50 values of 0.58 and 1.04mg/mL, respectively. These findings indicated that Boletus auripes possesses the potential to be utilized as a natural agent in antioxidant functional foods.

Composition-dependent MRM transitions and structure-indicative elution segments (CMTSES)-based LC-MS strategy for disaccharide profiling and isomer differentiation.

Carbohydrates exhibit diverse functions and extensive biological activities and are notable in the field of life sciences. However, their inherent diversity and complexity-steaming from variations in isomeric monomers, glycosidic bonds, configurations, etc.-present considerable challenges in structural analysis. Considering these challenges, the disaccharide building blocks with simpler structures could provide more structural information. Although various approaches have been explored, sufficient standards or specialized equipment are required to differentiate and characterize isomers. Therefore, a strategy that addresses these challenges is urgently needed. A Composition-dependent MRM Transitions and Structure-indicative Elution Segments (CMTSES)-based liquid chromatography-triple quadrupole mass spectrometry (LC-QQQ-MS) strategy was developed to comprehensively profile disaccharide units and differentiate isomers. First, the composition-related precursor and structure-specific product ions of disaccharides were generated by QQQ-MS. Thereout, MRM transitions were proposed to enable the comprehensive profiling of disaccharides and rapid annotation of their compositions and saccharide types at both termini. Next, the linkage, composition, and configuration isomers of disaccharides were effectively differentiated and presented characteristic LC elution. Furthermore, low-cost and available "location references" (mannose, galactose, and isomaltose) were sought to define structure-indicative elution segments for the identification of isomeric hexose disaccharides. Building on this foundation, the novel CMTSES-based LC-MS strategy was designed, and its feasibility was further verified by successfully differentiating and identifying mixed homogenous and/or heterogenous disaccharide isomers in real samples. Sufficient structural information was obtained even for those consisting of diversified monomer types. This strategy comprehensively profiles both major and minor disaccharides and effectively differentiates multiple types of isomers. The use of readily available "location references" facilitated the identification of isomeric hexose disaccharide with reduced dependence on standards, thereby broadening the applicability of this strategy. However, the characterization of disaccharides with other compositions is challenging. Further in-depth investigations into intramolecular hydrogen bond simulation should provide solutions. Additionally, CMTSES-based LC-MS strategy is promising to analyze complex structures and samples.

Effects of in vitro simulated digestion on the hypoglycaemic capacity of wheat bran-soluble dietary fibre.

Wheat bran-soluble dietary fibre (WB-SDF) is known for its hypoglycaemic properties and its potential to control postprandial blood glucose levels in individuals with hyperglycaemia. However, the digestive process may alter its glucose-lowering potential. This study investigated the effects of in vitro simulated digestion on the hypoglycaemic efficacy of WB-SDF. The hypoglycaemic effects of WB-SDF were determined by examining its glucose-binding capacity, glucose dialysis retardation index and ability to inhibit glucose uptake and transport in Caco-2cells. Structural changes after digestion were analysed via polysaccharide conformation analysis, microstructure observation and particle size measurements to evaluate their impacts on hypoglycaemic efficacy. Results indicate that WB-SDF and digested wheat bran-dietary fibre significantly decreased glucose adsorption and α-glucosidase activity compared with the control group in Caco-2cells. However, simulated digestion resulted in a relatively smaller reduction in α-glucosidase activity compared with the WB-SDF treatment group. The massive loss of surface laminar structure, reduction of -OH groups and partial glycosidic bond breakage in digested wheat bran-dietary fibre after digestion led to reduced glucose adsorption capacity and glucose dialysis retardation index. Moreover, the reduction in particle size after digestion enhanced the inhibition of glucose transport-related gene expression in Caco-2cells. Thus, although digestion weakens the glucose adsorption of WB-SDF, it improves its ability to inhibit glucose transport, highlighting the intricate interplay between structural modifications and hypoglycaemic efficacy.

Microbe-assisted fabrication of circularly polarized luminescent bacterial cellulosic hybrids

The fabrications of circularly polarized luminescent (CPL) material are mainly based on the chemical and physical strategies. Controlled biosynthesis of CPL-active materials is beset with difficulties due to the lack of bioactive luminescent precursors and bio-reactors. Enlighted by microbe-assisted asymmetric biosynthesis, herein, we show the in situ bacterial fermentation of Komagataeibacter sucrofermentants to fabricate a series of bacterial cellulosic biofilms with CPL of green, orange, red, and near-infrared colors. This process can trigger CPL emission for CPL-silent glycosylated luminophores and amplify the glum of weak CPL-active luminophores up to a 10−2 scale. To confirm glycosidic bonds formation during the bacterial copolymerization process, we develop an assay utilizing the cellulase-catalyzed biodegradation of BC hybrids. More importantly, we achieve the information encryption and Fe3+ dual-channel detection based on hybrid bacterial cellulosic biofilms. Therefore, this study not only provides another vision for CPL materials preparation but also broadens the application of bacterial cellulosic hybrids.

Identification of Oligosaccharide Isomers Using Electrostatically Asymmetric OmpF Nanopore.

Glycans, unlike uniformly charged DNA and compositionally diverse peptides, are typically uncharged and exhibit rich stereoisomeric diversity in the glycosidic bonds between two monosaccharide units. This heterogeneity of charge and the structural complexity present significant challenges for accurate analysis. Herein, we developed a novel single-molecule oligosaccharide sensor, OmpF nanopore. The natural electroosmotic flow within OmpF provides robust driving force for unlabeled neutral oligosaccharides, accomplishing a detection concentration as low as 6.4 μM. Furthermore, the asymmetric constriction zone of OmpF was employed to construct a stereoselective recognition site, enabling sensitive identification of glycosidic bond differences in cell lysate samples. Assisted by machine learning algorithms, a recognition accuracy of 99.9% for tetrasaccharides differing in only one glycosidic bond was achieved. This nanopore sensor offers a highly sensitive analytical tool with a broad dynamic range for the chiral recognition of oligosaccharides at low concentrations, applicable to both low-abundance and practical samples.

Identification of Oligosaccharide Isomers Using Electrostatically Asymmetric OmpF Nanopore

Glycans, unlike uniformly charged DNA and compositionally diverse peptides, are typically uncharged and exhibit rich stereoisomeric diversity in the glycosidic bonds between two monosaccharide units. This heterogeneity of charge and the structural complexity present significant challenges for accurate analysis. Herein, we developed a novel single‐molecule oligosaccharide sensor, OmpF nanopore. The natural electroosmotic flow within OmpF provides robust driving force for unlabeled neutral oligosaccharides, accomplishing a detection concentration as low as 6.4 μM. Furthermore, the asymmetric constriction zone of OmpF was employed to construct a stereoselective recognition site, enabling sensitive identification of glycosidic bond differences in cell lysate samples. Assisted by machine learning algorithms, a recognition accuracy of 99.9% for tetrasaccharides differing in only one glycosidic bond was achieved. This nanopore sensor offers a highly sensitive analytical tool with a broad dynamic range for the chiral recognition of oligosaccharides at low concentrations, applicable to both low‐abundance and practical samples.

That with Glycosidic Visage: Undergraduate Students’ Interpretation of Glycosidic Bonds

That with Glycosidic Visage: Undergraduate Students’ Interpretation of Glycosidic Bonds

Development of a comprehensive food glycomic database and its application: Associations between dietary carbohydrates and insulin resistance.

Carbohydrates are an integral part of a healthy diet. The molecular compositions of carbohydrates encompass a very broad range of unique structures with many being ill-defined. This vast structural complexity is distilled into vague categories such as total carbohydrates, sugars, starches, and soluble/insoluble fibers. Structural elucidation of the food glycome is until recently extremely slow and immensely challenging. Dietary carbohydrates, including monosaccharides, oligosaccharides, glycosidic linkages, and polysaccharides were determined for the most consumed foods in the US consisting of 250 common foods using a multiglycomic platform. The food glycome was then correlated with clinical data from the National Health and Nutrition Examination Survey (NHANES) consisting of dietary recalls from 13,550 adults to determine associations between dietary carbohydrates, their structural features and insulin resistance. Several features were more powerful predictors compared to traditional measures indicating the need for molecular fine-scale food carbohydrate data in guiding precision nutrition initiatives and clinical studies.

Enzymatic β-Mannosylation of Phenylethanoid Alcohols.

Phenylethanoid glycosides (PhGs) are widely occurring secondary metabolites of medicinal plants with interesting biological activities such as antioxidant, anti-inflammatory, neuroprotective, antiviral, hepatoprotective, immunomodulatory, etc. They are characterized by a structural core formed by a phenethyl alcohol, usually tyrosol or hydroxytyrosol, attached to β-D-glucopyranose via a glycosidic bond. This core is usually further decorated by attached phenolic acids or another saccharide. Several studies suggest an important role of the saccharidic fragment in the biological activities of PhGs, provoking demand for new glycovariants of natural PhGs. This study presents the preparation of β-mannosylated analogs of tyrosol β-D-glucopyranoside (salidroside) and hydroxytyrosol β-D-glucopyranoside (hydroxysalidroside). While the chemical synthesis of β-D-mannopyranosides is rather challenging, they can be prepared by enzymatic catalysis. We found that Novozym 188, an industrial β-glucosidase, also contains β-mannosidase and used this enzyme in the preparation of tyrosol β-D-mannopyranoside and hydroxytyrosol β-D-mannopyranoside in 12 and 16% chemical yields, respectively, by transglycosylation from β-D-mannopyranosyl-(1→4)-D-mannose. The mannosylation was chemoselective and occurred exclusively on the primary hydroxyls of tyrosol and hydroxytyrosol, and the glycosylation of phenolic moieties of the aglycons was observed.

Structural characterization and anti-weightless bone loss activity of an anionic polysaccharide from Dictyophora indusiata.

This study explores the extraction and purification of polysaccharides from Dictyophora indusiata (D. indusiata) with its antioxidant and anti-weightlessness bone loss properties. An anionic polysaccharide (SADIP) with a molecular weight of 13.42 kDa was isolated and purified from D. indusiata. SADIP consists of a glucose-based sugar chain backbone. Its backbone consists of glycosidic linkages of →4)-α-D-Glcp-(1 → and →4)-β-D-Glcp-(1→, with sulfate groups attached at the O-6 position of →4)-β-D-Glcp-(1 → residues. Side chains were α-D-Glcp-(1 → 6)-α-D-Glcp-(1 → linked at C-6 position of →4,6)-α-D-Glcp-(1 → through O-6 bonds. In vitro, SADIP scavenged up to 99.32 ± 0.24 % of 2,2'-Azinobis-(3-ethylbenzthiazoline-6-sulphonate) (ABTS) radicals and 76.72 ± 0.34 % of hydroxyl radicals. In vivo, SADIP reduced malondialdehyde (MDA) levels induced by weightlessness and increased low levels of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) caused by weightlessness. Additionally, three-point mechanical bending tests and Micro-CT analysis demonstrated that SADIP has ability to alleviate bone loss induced by weightlessness. This study provides a theoretical foundation for the rational developmation of D. indusiata resources.

Integrated analysis of cellulose structure and properties using solid-state low-field H-NMR and photoacoustic spectroscopy

In this study, we explore the structural intricacies of cellulose, a polymer composed of glucose monomers arranged in a linear chain, primarily investigated through solid-state NMR techniques. Specifically, we employ low-field proton nuclear magnetic resonance (H-NMR) to delve into the diverse hydrogen atom types within the cellulose molecule. The low-field H-NMR technique allows us to discern these hydrogen atoms based on their distinct chemical shifts, providing valuable insights into the various functional groups present in cellulose. Our focus extends to the examination of anomeric protons of glucose units and protons linked to carbon atoms engaged in glycosidic linkages within cellulose chains, which exist in diverse crystalline and amorphous forms. Solid-state low-field H-NMR spectroscopy aids in characterizing the crystallinity degrees and amorphous regions within cellulose, revealing time-dependent changes in free induction decay (FID) signals. Complementing this, we investigate the photo-absorption properties of cellulose fibers under both continuous and modulated irradiation using reversed double-beam photoacoustic spectroscopy (RDB-PAS). This photoacoustic approach allows us to observe ultraviolet- and visible light-induced processes, including electron trap filling and reductive changes on the fiber surface. Our findings suggest that RDB-PAS is a feasible method for estimating the electron trap distribution, serving as a potential measure of the density of crystalline cellulose defects. This integrated approach of combining solid-state low-field H-NMR and RDB-PAS techniques offers a comprehensive understanding of cellulose structure and properties, enhancing our ability to characterize its diverse features.

In Silico Structural Insights into a Glucanase from Clostridium perfringens and Prediction of Structural Stability Improvement Through Hydrophobic Interaction Network and Aromatic Interaction.

Glucanases are widely applied in industrial applications such as brewing, biomass conversion, food, and animal feed. Glucanases catalyze the hydrolysis of glucan to produce the sugar hemiacetal through hydrolytic cleavage of glycosidic bonds. Current study aimed to investigate structural insights of a glucanase from Clostridium perfringens through blind molecular docking, site-specific molecular docking, molecular dynamics (MD) simulation, and binding energy calculation. Furthermore, we aimed to enhance structural stabilization through formation of hydrophobic interaction network. The molecular docking results illustrated that residues Glu222 and Asp187 may act as nucleophile acid/base catalyst. Moreover, the MM/PBSA results illustrated a high binding affinity of 108.71 ± 8.5kJ/mol between glucanase and barely glucan during 100 ns simulation. The RMSF analysis illustrated a high flexible surface loop with the highest mobility at position D130. Therefore, the structural engineering was carried out through introducing a double-mutant S125Y/D130P, and the structural stability was improved by forming the hydrophobic interaction network and one π-π aromatic interaction. The spatial distance between the mutation sites and the catalytic pocket attenuates their direct impact on binding interactions within the catalytic pocket.

4-O-Methylglucuronoxylan from Hygrophila Ringens var. Ringens Seeds: Chemical Composition and Anti-Inflammatory Activity.

Hygrophila ringens var. ringens is a medicinal plant of the Acanthaceae family. A soluble polysaccharide is extracted from H. ringens seeds using warm water, followed by deproteinization and purification using column chromatography. DL1 is characterized comprehensively using spectroscopic and chromatographic techniques and identified as a polymer containing xylose (Xyl; 78.5%) and 4-O-methyl-d-glucuronic acid (4-O-MeGlcA; 21.5 %). The most prominent glycosidic linkages detected are terminal-xylose (T-Xyl); 1,2,3,4-Xylp; 1,2,4-Xylp; and T-4-O-MeGlcA. DL1 belongs to the xylan group and is a 4-O-methylglucuronoxylan. DL1 exhibits inhibition of bovine serum albumin denaturation with IC50 values of 0.35mgmL-1 and a similar activity to diclofenac (non-steroidal anti-inflammatory drug). In a model of lipopolysaccharide-stimulated macrophages, DL1 (20-40µgmL-1) strongly inhibits inflammatory cytokines and reactive oxygen species release without having significant macrophage cytotoxicity. The inhibitory effect of DL1 on inflammatory cytokines is mediated by the activation of mitogen-activated protein kinases by inhibiting the phosphorylation of p38 and extracellular signal-regulated kinase. These results highlight the potential of DL1 for treating inflammation through its cytokine-suppressive activity.

Chemical Space of Fluorinated Nucleosides/Nucleotides in Biomedical Research and Anticancer Drug Discovery

Fluorinated nucleos(t)ide drugs have proven to be successful chemotherapeutic agents in treating various cancers. The Food and Drug Administration (FDA) has approved several drugs that fit within the fluorinated nucleoside pharmacophore, and many more are either in preclinical development or clinical trials. The addition of fluorine atoms to nucleos(t)ides improves the metabolic stability of the glycosidic bond and, in certain instances, facilitates additional interactions of nucleons(t)ides with receptors. The insertion of fluorine either on sugar or the base of nucleos(t)ides proved to enhance the lipophilicity, pharmacokinetic, and pharmacodynamic properties. Overall, the fluorine atom feeds diverse advantages to the biological profile of nucleos(t)ide analogs by improving their drug-like properties and therapeutic potential. This review article covers the often-used fluorinating reagents in nucleoside chemistry, the clinical significance of [18F]-labeled nucleosides, the synthesis and anticancer activity of FDA-approved fluoro-nucleos(t)ide drugs, as well as clinical candidates, which are at various stages of clinical development as anticancer agents.

Long, Synthetic Staphylococcus aureus Type 8 Capsular Oligosaccharides Reveal Structural Epitopes for Effective Immune Recognition.

Staphylococcus aureus is a Gram-positive bacterium that is responsible for severe nosocomial infections. The rise of multidrug-resistant strains, which can pose significant health threats, prompts the development of new treatment interventions, and much attention has been directed at the development of prophylactic and therapeutic vaccination strategies. Capsular polysaccharides (CPs) are key protective elements of the S. aureus cell wall and have been proposed as promising candidate antigens. Thirteen different CP serotypes have been identified to date, of which types 5 and 8 are the most prominent. CP8 is composed of trisaccharide repeating units that are built up from an N-acetyl-4-O-acetyl-β-d-mannosaminuronic acid, that carries a C-4-O-acetyl, an N-acetyl-α-d-fucosamine, and an N-acetyl-α-l-fucosamine. Synthetic oligosaccharides are valuable tools to unravel the immunogenicity of bacterial oligosaccharides at the molecular level. However, the rare monosaccharides, cis-glycosidic linkages, and O-acetylation represent significant challenges for the synthesis of CP8 fragments. Here the stereoselective assembly of well-defined CP8 fragments, comprising a trimer, hexamer, nonamer, and dodecamer, is presented. This is the first time that fragments larger than a single repeating trisaccharide, which has been proven to be insufficient for antigenic activity, have been assembled. Structural studies have revealed a linear conformation for the oligosaccharides, with each trisaccharide repeat tilted ∼90° with respect to the flanking repeats, which is stabilized by the acetyl groups that prevent rotation around the glycosidic linkages. The N-acetyl groups in each repeating unit point in the same direction, generating a hydrophobic flank in the trisaccharide repeats. We applied the oligomers to generate model glycoconjugate vaccine modalities, which we then used to raise anti-CP8 antibodies. The antibody interaction and immunization studies have revealed a clear length dependent structure-activity relationship for the oligosaccharides, with an oligosaccharide of at least three repeating units required for an adequate immune response.

Optimized microwave-assisted extraction and characterization of spray dried Luffa aegyptiaca nanomucilage: Physicochemical properties, biological activities, and anticancer efficacy against MCF-7 human breast cancer cells.

Microwave-assisted extraction conditions were optimized using response surface methodology to evaluate the effects of extraction parameters on the yield and carbohydrate content of Luffa aegyptiaca mucilage. Extraction at 540 W for 2 min with a 1:20 (g/mL) was determined as the optimal parameter, resulting in a maximum yield of 5.90 % (w/w) with 63 % carbohydrate content consisting of glucose, galactose, maltose, mannose, and galacturonic acid, with structural linkages of β (1 → 4) and β (1 → 6) glycosidic bonds. The nanomucilage exhibited a monomodal particle size distribution of 145.3 ± 4.60 nm, high thermal stability (-1363.08 J/g), oil and water retention capacity, emulsifying ability (93.06 ± 0.48 %), emulsifying stability (75.02 ± 0.96 %), solubility (95.36 ± 0.89 %), and foaming ability (93.06 ± 0.48 %). Mucilage demonstrated potential in vitro anti-oxidant activity (2.05 ± 0.10 %) against Caco-2 cells, anti-inflammatory activity during membrane stabilization (30.47 ± 0.42 % to 70.46 ± 0.31 %,) and protein denaturation (20.47 ± 0.42 % to 78.39 ± 0.40 %) assays and anticancer activity against human breast cancer cells (MCF-7), with growth inhibition of 100.5 ± 12.45 %. Hence, this evaluation of Luffa aegyptiaca nanomucilage highlights its potential as a multifunctional biomaterial with significant applications in the healthcare industry.

Biopolymer-Levan Characterization in Bacillus Species Isolated from Traditionally Fermented Soybeans (Thua Nao).

Bacterial levans are biopolymers composed of fructose units linked by β-2,6 glycosidic bonds that are degradable, nontoxic and flexible, representing a green technology with significant applications across various industries. Fermented soybeans are a common source of bacteria-producing polysaccharides. In this study, Bacillus siamensis KKSB4, Bacillus velezensis KKSB6 and Bacillus amyloliquefaciens KKSB7 isolated from traditionally fermented soybean (Thua-nao), along with B. velezensis strain 5.18 isolated from Jerusalem artichoke were investigated for their levansucrase activity and subsequent levan production. B. siamensis KKSB4, B. amyloliquefaciens KKSB7 and B. velezensis strain 5.18 exhibited the highest enzyme activity at 30% sucrose, whereas B. velezensis KKSB6 demonstrated optimal activity at 20% sucrose. Characterization of the levans using Fourier transform infrared spectroscopy (FTIR) revealed the peak at 3310.02, 1122.42, 1011.14, and 923.89 cm-1, corresponding to the O-H group in fructose, C-O-H stretching, C-O-C stretching vibrations, and pyranose ring structures, respectively. 13C NMR spectrum of levan aligned with the reference spectrum of B. siamensis, showing proton shifts at H3 (4.19 ppm), H4 (4.10 ppm), H5(3.96 ppm), H6a (3.89 ppm), H1a (3.76 ppm), H1b (3.67 ppm), and H6b (3.56 ppm). All strains produced high molecular weight levans (107-108 Da) with thermal stability, exhibiting melting temperatures exceeding 200 °C. The levans also demonstrated high water and oil holding capacities of over 100%, attributed to their large molecular complexes. Additionally, they exhibited antioxidant activity by scavenging DPPH radicals and oxidizing ABTS, particularly B. siamensis KKSB4 and B. velezensis KKSB6, although low FRAP activity was observed. Furthermore, the levans were produced by B. siamensis KKSB4 and B. velezensis KKSB6 promoted the growth of Streptococcus thermophilus DKT-3, suggesting its potential prebiotic properties. This work highlights B. siamensis KKSB4 and B. velezensis KKSB6 is an outstanding candidate for levan production with diverse functional properties.

Decoding the difference of four species of Cordyceps based on polysaccharides and immunomodulation activity.

Nucleosides and polysaccharides are the main bioactive ingredients of Cordyceps genus. Nucleosides shows significant differences in different Cordyceps species. However, the differences of polysaccharides have not been decoded. Here, the structure characters of polysaccharides including molecular weight (Mw) distribution, compositional monosaccharides and glycosidic linkage types were compared in C. sinensis (CS), C. militaris (CM), silkworm-hosted C. militaris (SCM) and Cordyceps fermented products (CSF). Compositional monosaccharides including mannose, glucose and galactose, and 1,4-Glcp glycosidic linkage were found abundant in Cordyceps species. Chemometric analysis showed that Cordyceps exhibit significant differences in structural information especially glycosidic linkage types. Besides, polysaccharides in CS and CSF-4 had obviously strong capacity of stimulating phagocytic, NO production and cytokines secretion. Gray relational analysis and Pearson correlation analysis were performed to further investigate the relationship between key polysaccharide structure and immunomodulatory activities. The results indicated that polysaccharides with relatively large number of 1, 4-Glcp and Mw in range of 7.16×106Da-7.99×107Da and 1.43×104 D-6.94×105Da probably contributed to its immunomodulatory activities. The chemical and biological evaluation of natural and various cultured cordyceps in this study is useful for understanding and regulating the quality of cultured Cordyceps.

Polysaccharides isolated from Ampelopsis grossedentata and their immunomodulatory activity.

To explore the immunomodulatory activity of polysaccharides from Ampelopsis grossedentata, two polysaccharides named AGP1 and AGP2 were isolated and purified by DEAE-cellulose 52 column and Sephacryl S-300HR chromatography. AGP1 and AGP2 were composed of fucose, arabinose, rhamnose, galactose, glucose, mannose, galacturonic acid, and glucuronic acid, with a ratio of 0.5: 10.2: 0.9: 31.8: 7.4: 3.4: 21.6: 24.2 and 0.4: 6.0: 0.5: 23.3: 3.3: 6.2: 33.5: 26.8, respectively. The average molecular weights of AGP1 and AGP2 were found to be 6.60×105Da and 7.24×105Da, respectively. AGP1 contained →4,6)-Galp-(1→glycosidic linkages, while AGP2 contained →2)-Galp-(1→and →2,3,4)-Glcp-(1→glycosidic linkages. The structures of AGPs were characterized by Fourier transform infrared spectroscopy, nuclear magnetic resonance, and scanning electron microscope. The immunomodulatory activity of AGPs was investigated in RAW264.7 cells, and the results indicated that AGPs significantly activated macrophages, promoted cells differentiation and NO secretion, increased the expression of IL-6 and TNF-α, and induced macrophage M1 polarization. Transcriptomic analysis indicated that AGP1 and AGP2 regulated a total of 1043 and 970 differentially expressed genes respectively, which were identified in different immune related signaling pathways. Moreover, the immunoblot demonstrated that AGPs exerted immune-promoting effects through the TLR4, MAPK and NF-κB signaling pathways in macrophages. Consequently, AGPs have potent immunomodulatory activity and can be considered as immunomodulators in medical and food industries.

Glycosidic linkages of fungus polysaccharides influence the anti-inflammatory activity in mice

Over decades, the source-function relationships of bioactive polysaccharides have been progressively investigated, however, it is still unclear how a defined structure may conduce to the bioactivities of polysaccharides. To explore the structure-function relationship of fungus polysaccharides, we employed a dextran sulfate sodium (DSS)-induced colitis mouse model to compare the anti-inflammatory activity of two fungus polysaccharides from Dictyophora indusiata (DIP) and Tremella fuciformis (TFP), which exhibit distinct glycosidic linkages. The structures of DIP and TFP were characterized through molecular weight detection, molecular morphology analysis, methylation analysis, and NMR analysis. Subsequently, we employed a DSS-induced colitis model to assess the anti-inflammatory efficacy of DIP and TFP. The colitis symptoms, histological morphology, intestinal inflammatory cytokines, and the composition and function of gut microbiota before and after polysaccharides treatment in colitis mice were also investigated. DIP, l,3-β-D-glucan with 1,4-β and 1,6-β-D-Glcp as branched chains, exhibited superior therapeutic effect than that of TFP consisted of a linear 1,3-α-D-mannose backbone with D-xylose and L-fucose in the side chains. Both DIP and TFP relieved DSS-induced colitis in a gut microbiota dependent manner. Furthermore, metagenomics showed that DIP and TFP could partially reverse the bacterial function in colitis mice. Glycoside Hydrolase 1 (GH1) and GH3 were shown to cleave the glucose linkages in DIP, while GH92 and GH29 were likely active on the α-1,3-linked mannose linkages and the glycosidic bonds of L-fucose residues in TFP. Our findings highlight the pivotal role of glycosidic linkages in anti-inflammatory activities of fungus polysaccharides and would promote the design and discovery of polysaccharides with designated activity to be used as functional foods and/or therapeutics.