Structural characteristics of a bioactive polysaccharide from Sorghum arundinaceum

A polysaccharide, an alpha-D-glucan with an apparent molecular weight of 6.85 x 10(4), called PSa glucan, was isolated from fresh seeds of Sorghum arundinaceum by fractionation on Sephacryl S-300 HR and Sephadex G-25. Chemical and spectroscopic studies indicated that it has a highly branched glucan type structure composed of alpha-(1-->4) linked D-glucopyranose residues with (1-->3), (1-->6) branching points, and a significant amount of alpha-(1-->6) branching to alpha-(1-->3) linked D-glucopyranose residues. The anti-inflammatory activity of the polysaccharide was performed using the capillary permeability assay.     

Structure determination,apoptosis induction,and telomerase inhibition of CFP-2, a novel lichenin from Cladonia furcata

A great deal of experimental evidence has accumulated in the past several decades, suggesting that polysaccharides have wide bioactivities. Cladonia furcata polysaccharide, CFP-2, a water-soluble lichenin with a mean Mr 7.6 x 10(4), was first obtained by 0.25 M NaOH solution extraction, ethanol precipitation, DEAE-cellulose, and Sephadex G-200 column chromatography. Gas chromatography of acid hydrolyzate of CFP-2 suggested that it was composed of D-glucose, D-galactose, and D-mannose in the molar ratios of 8:1:1. Periodate oxidation, Smith degradation, IR, and NMR spectroscopy analysis revealed that CFP-2 had a backbone consisting of alpha-(1-->3) and alpha-(1-->4)-linked D-glucopyranosyl residues substituted at O-6 with beta-(1-->6)-linked D-galactopyranosyl residue and alpha-(1-->6)-linked D-mannopyranosyl residue. CFP-2 was able to reduce viability of cultured HL-60 and K562 cells. The antiproliferative properties of CFP-2 appeared to be attributable to its induction of apoptotic cell death as determined by ultrastructural change, internucleosomal DNA fragmentation, and increased proportion of the subdiploid cell population. To elucidate molecular events in the apoptosis, protein expressions of Bcl-2, Bax, Fas, and FasL were measured by Western blotting using specific antibodies in HL-60 cells. The level of Bcl-2 remained largely unchanged, but the Bax, Fas, and FasL expression showed up-regulation. Moreover, the telomerase activity analyzed by TRAP-ELISA assay in HL-60 cells treated with CFP-2 decreased as compared with the untreated control cells. These results suggest that CFP-2 could have a possible cancer therapeutic potential.     

Experimental study on anticoagulant and antiplatelet aggregation activity of a chemically sulfated marine polysaccharide YCP

A polysaccharide YCP was prepared from a marine filamentous fungus Keissleriella sp. YS4108, which exhibited as a molecular weight (Mw) of 2.4x10(3) kDa and its three sulfated derivatives (YCP-SL, YCP-SM and YCP-SH) were synthesized, the degree of substitution (DS) of which were determined to be 0.13, 0.99 and 1.3, with the average molecular weight 0.64x10(3), 0.57x10(3) and 0.45x10(3) kDa, respectively. Anticoagulant activity and antiplatelet aggregation activity of these sulfated derivates were evaluated by activated partial thromboplastin time (APTT), prothrombin time (PT), thrombin time (TT) and platelet aggregation assay. The results showed that YCP sulfates significantly prolonged APTT, TT and PT. The derivates showed no effects on thrombin in the presence or in the absence of antithrombin III (AT III) or heparin cofactor II (HC II), while the derivates effectively inhibited factor Xa in the presence of AT III. At the same time, YCP-SH also possessed potent antiplatelet aggregation activity in vitro compared with aspirin. YCP sulfates specifically interfered with different stages of the coagulation cascade, and the anticoagulant activity improved with the increasing DS and decreased Mw.     

Gordonan, an acidic polysaccharide with cell aggregation-inducing activity in insect BM-N4 cells, produced by Gordonia sp

An acidic polysaccharide, termed gordonan, was isolated from the culture medium of Gordonia sp. as an inducer of cell aggregation in an insect cell line, BM-N4. Gordonan had an average molecular weight of 5 x 10(6) and its structure was identified as -->3)-4-O-(1-carboxyethyl)-beta-D-Manp-(1-->4)-beta-D-GlcAp-(1-->4)-beta-D-Glcp-(1--> mainly by acid hydrolysis experiments and NMR analysis. It induces cell aggregation at the concentration of 4 microg/ml. A partially hydrolyzed polysaccharide derived from gordonan with a molecular weight of 5 x 10(5) showed weak activity, while any fragment molecules with lower molecular weights prepared from gordonan showed no activity.     

Solution properties of antitumor sulfated derivative of alpha-(1-->3)-D-glucan from Ganoderma lucidum

Four fractions of a water-insoluble alpha-(1-->3)-D-glucan GL extracted from fruiting bodies of Ganoderma lucidum were dissolved in 0.25 M LiCl/DMSO, and then reacted with sulfur trioxide-pyridine complex at 80 degrees C to synthesize a series of water-soluble sulfated derivatives S-GL. The degree of substitution of DS was measured by using IR infrared spectra, elemental analysis, and 13C NMR to be 1.2-1.6 in the non-selective sulfation. Weight-average molecular weight Mw and intrinsic viscosity [eta] of the sulfated derivatives S-GL were measured by multi-angle laser light scattering and viscometry. The Mw value (2.4 x 10(4)) of sulfated glucan S-GL-1 was much lower than that (44.5 x 10(4)) of original alpha-(1-->3)-D-glucan GL-1. The Mark-Houwink equation and average value of characteristic ratio C(infinity) for the S-GL in 0.2 M NaCl aqueous solution at 25 degrees C were found to be: [eta] = 1.32 x 10(-3) Mw(1.06) (cm3 g(-1)) and 16, respectively, in the Mw range from 1.1 x 10(4) to 2.4 x 10(4). It indicated that the sulfated derivatives of the alpha-(1-->3)-D-glucan in the aqueous solution behave as an expanded chain, owing to intramolecular hydrogen bonding or interaction between charge groups. Interestingly, two sulfated derivatives synthesized from the alpha-(1-->3)-D-glucan and curdlan, a beta-(1-->3)-D-glucan, all had significant higher antitumor activity against Ehrlich ascites carcinoma (EAC) than the originals. The effect of expanded chains of the sulfated glucan in the aqueous solution on the improvement of the antitumor activity could not be negligible.     

Purification, structure and immunobiological activity of a water-soluble polysaccharide from the fruiting body of Pleurotus ostreatus

The water-soluble polysaccharide (POP), with a molecular mass of 2.4x10(4)Da, was obtained from the fruiting body of Pleurotus ostreatus. Structure features of the purified polysaccharide were investigated by a combination of chemical and instrumental analysis, such as methylation analysis, Smith degradation, GC-MS, (13)C and (1)H NMR and FTIR. The results indicated that the backbone of POP was composed of (1-->6)-linked-alpha-D-galactopyranosyl and (1-->2,6)-linked-alpha-D-galactopyranosyl residues, which were terminated with a single terminal (1-->)-beta-D-glucose residue at the O-2 position of galactosyl along the main chain in the ratio of 1:1:1. Preliminary tests in vitro showed POP is capable of enhancing concanavalin A (ConA)- or lipopolysaccharide (LPS)-induced lymphocyte proliferation, which suggested that POP could be a potential immunostimulating agent for use in functional foods or medicine against both pathogens and cancer.     

Flocculant and Chemical Properties of a Polysaccharide from Pullularia pullulans

An extracellular polysaccharide, PP-floc, was synthesized from glucose by Pullularia pullulans (or Aureobasidium pullulans) in a pilot plant batch fermentor containing 175 liters of culture medium. At 58 h of fermentation, the concentration of PP-floc was 1.03 g/100 ml, giving a 25.8% conversion of initial glucose to polysaccharide. The flocculant activity of the culture medium increased during the fermentation process and reached its maximum at 50 h of culture age. Less PP-floc (0.33 lb/ton of slimes [approximately 149.7 g/0.907 t]) was required to give the same flocculant activity as a synthetic polymer of acrylamide, Separan NP-10 (0.5 lb/ton of slimes [approximately 226.8 g/0.907 t]), at all temperatures from 25 to 100 C. The degree of inactivation of PP-floc and Separan NP-10 at elevated temperatures was almost identical, and they were completely inactivated at about the same temperature (80 C). PP-floc also gave better compaction of slimes than Separan NP-10 at all temperatures tested. PP-floc was soluble in water and its specific optical rotation was [alpha](D) (25) + 194 degrees in water (c, 0.4). PP-floc contained 83.3% carbohydrate, 3.2% protein, and 8.1% water. Glucose was found to be the principal sugar monomer with traces (>5%) of galactose and mannose present. Structural studies on the fractions of purified polysaccharide by methylation and by periodate oxidation techniques prove that PP-floc is linear and composed of alpha-(1 --> 4) and alpha-(1 --> 6) glucopyranosyl units in the approximate ratio of 2:1. The action of pullulanase on crude PP-floc suggested the ordered arrangement of two consecutive alpha-(1 --> 4)-linked glucopyranosyl units flanked by alpha-(1 --> 6)-linked glucopyranose residues.     

Purification, structure and immunobiological activity of a new water-soluble polysaccharide from the mycelium of Polyporus albicans (Imaz.) Teng

A new water-soluble intracellular polysaccharide named as PTP, with a molecular mass of 3.7x10(4) Da, was obtained from the mycelium of Polyporus albicans (Imaz.) Teng. Structure features of the purified polysaccharide were investigated by a combination of chemical and instrumental analysis. The results indicated that PTP consisted of a backbone composed of (1-->3)-linked-beta-d-mannopyranosyl, (1-->3,6)-linked-beta-d-mannopyranosyl and (1-->6)-linked-alpha-d-galactopyranosyl residues in the ratio of 3:1:1, and terminated with a single non-reducing terminal (1-->)-beta-d-mannopyranosyl residues at the C-6 position of (1-->3,6)-linked-beta-d-mannopyranosyl, along the main chain. This is the first report describing the isolation and structure elucidation of a new intracellular polysaccharide produced from the mycelium of P. albicans (Imaz.) Teng. Preliminary tests in vitro showed PTP have potent stimulating effects on murine lymphocyte proliferation induced by concanavalin A or lipopolysaccharide and its branches are extremely important for the expression of the enhancement of the immunological activity.     

Characterization of two cell-wall polysaccharides from Fusicoccum amygdali

1. The nature of two polysaccharides (s(0) (20) values 6S and 2S respectively in 1m-sodium hydroxide), comprising a fragment (fraction BB, [alpha](D) +236 degrees in 1m-sodium hydroxide), previously isolated from cell walls of Fusicoccum amygdali, has been investigated. 2. Both the major (2S) and minor (6S) components were affected by incubation with alpha-amylase. The 6S polysaccharide was also attacked by exo-beta-(1-->3)-glucanase, which is evidence that it contained both alpha-(1-->4)- and beta-(1-->3)-glucopyranose linkages. By fractionation of the products of alpha-amylase-treated fraction BB it was possible to obtain a water-insoluble polysaccharide, fraction P ([alpha](D) +290 degrees in 1m-sodium hydroxide, 67% of fraction BB) and a water-soluble polysaccharide, fraction Q ([alpha](D) +16 degrees in 1m-sodium hydroxide, 11% of fraction BB), both of which sedimented as single boundaries with s(0) (20) values (in 1m-sodium hydroxide) of 1.7S and 4.6S respectively. 3. Evidence from periodate oxidation, methylation analysis, i.r. spectroscopy and partial acid hydrolysis showed that fraction P consisted of linear chains of alpha-(1-->3)-glucopyranose units with blocks of one or two alpha-(1-->4)-glucopyranose units interspersed at intervals along the main chain. The 2S polysaccharide, from which fraction P is derived, evidently also contains longer blocks of alpha-(1-->4)-glucopyranose units, that are susceptible to alpha-amylase action. 4. Fraction Q consisted of glucose (88%) with small amounts of galactose, mannose and rhamnose. Evidence from digestion with exo- and endo-beta-(1-->3)-glucanases, periodate oxidation and methylation analysis suggests that fraction Q consists of a branched galactomannorhamnan core, to which is attached a beta-(1-->3)-, beta-(1-->6)-glucan. In the cell wall, chains of alpha-(1-->4)-linked glucopyranose units are linked to fraction Q to form the 6S component of fraction BB.     

Characterization and protection on acute liver injury of a polysaccharide MP-I from Mytilus coruscus

In this study, we analyzed a water-soluble polysaccharide MP-I isolated from Mytilus coruscus. MP-I was obtained by hot-water extraction, anion-exchange and gel-permeation chromatography. Complete hydrolysis, periodate oxidation, methylation analysis, as well as Fourier transform infrared spectroscopy (FTIR) and nuclear magnetic resonance (NMR) spectroscopy were conducted to elucidate its structure. MP-I was subjected to investigate the protective effect on carbon tetrachloride (CCl(4)) induced liver damage in male Kunming mice. Based on the data obtained, MP-I was found to be an alpha-(1-->4)-D-glucan, branched with a single alpha-D-glucose at the C-6 position every eight residue, on average, along the main chain. Based on the calibration with Dextran, the glucan had a molecular weight of about 1.35 x 10(6) Da. Pharmacological studies revealed that MP-I could decrease serum alanine aminotransferase (ALT), serum aspartate aminotransferase (AST), and hepatic malondialdehyde aldehydes (MDA) levels, increase the hepatic total superoxide dismutase (T-SOD) activity, and improve hepatic damage in the CCl(4) induced liver injury in mice in a dose-dependent manner. The results suggest that the possible mechanism is due to its antioxidant activity of MP-I.     

Structural characterization of the O-antigenic polysaccharide of the lipopolysaccharide from Rhizobium etli strain CE3. A unique O-acetylated glycan of discrete size, containing 3-O-methyl-6-deoxy-L-talose and 2,3,4-tri-O-,methyl-l fucose

The O-antigenic polysaccharide of the Rhizobium etli CE3 lipopolysaccharide (LPS) was structurally characterized using chemical degradations (Smith degradation and beta-elimination of uronosyl residues) in combination with alkylation analysis, electrospray, and matrix-assisted laser desorption ionization-time of flight mass spectrometry, tandem mass spectrometry, and (1)H COSY and TOCSY nuclear magnetic resonance spectroscopy analyses of the native polysaccharide and the derived oligosaccharides. The polysaccharide was found to be a unique, relatively low molecular weight glycan having a fairly discrete size, with surprisingly little variation in the number of repeating units (degree of polymerization = 5). The polysaccharide is O-acetylated and contains a variety of O-methylated glycosyl residues, rendering the native glycan somewhat hydrophobic. The molecular mass of the major de-O-acetylated species, including the reducing end 3-deoxy-d-manno-2-octulosonic acid (Kdo) residue, is 3330 Da. The polysaccharide is comprised of a trisaccharide repeating unit having the structure -->4)-alpha-d-GlcpA-(1-->4)-[alpha-3-O-Me-6-deoxy-Talp-(1--> 3)]-alpha -l-Fucp-(1-->. The nonreducing end of the glycan is terminated with the capping sequence alpha-2,3, 4-tri-O-Me-Fucp-(1-->4)-alpha-d-GlcpA-(1-->, and the reducing end of the molecule consists of the non-repeating sequence -->3)-alpha-l-Fucp-(1-->3)-beta-d-Manp-(1-->3)-beta-QuiNA cp-(1-->4)-a lpha-Kdop-(2-->, where QuiNAc is N-acetylquinovosamine (2-N-acetamido-2,6-dideoxyglucose). The reducing end Kdo residue links the O-chain polysaccharide to the core region oligosaccharide, resulting in a unique location for a Kdo residue in LPS, removed four residues distally from the lipid A moiety. Structural heterogeneity in the O-chain arises mainly from the O-acetyl and O-methyl substitution. Methylation analysis using trideuteriomethyl iodide indicates that a portion of the 2,3,4-tri-O-methylfucosyl capping residues, typically 15%, are replaced with 2-O-methyl- and/or 2,3-di-O-methylfucosyl residues. In addition, approximately 25% of the 3,4-linked branching fucosyl residues and 10% of the 3-linked fucosyl residues are 2-O-methylated. A majority of the glucuronosyl residues are methyl-esterified at C-6. These unique structural features may be significant in the infection process.     

Sulfation of a polysaccharide produced by a marine filamentous fungus Phoma herbarum YS4108 alters its antioxidant properties in vitro

Free radicals and other reactive oxygen species (ROS) are generated by all aerobic cells and are widely believed to play a significant role in aging as well as a number of degenerative or pathological diseases. This study compared the free radical-scavenging properties and antioxidant activity of YCP, a polysaccharide from the mycelium of a marine filamentous fungus Phoma herbarum YS 4108 and its two chemically sulfated derivatives YCP-S1 and YCP-S2. Sulfation, which masks hydroxyl groups of YCP polysaccharide molecule, could introduce new antioxidant activity, such as superoxide and hydroxyl radicals scavenging activity, metal chelating action, lipid peroxidation and linoleic acid oxidation inhibition capability. Furthermore, sulfated YCP was more potent than YCP at protecting erythrocytes against oxidative damage hemolysis. The current data suggest for the first time that sulfation of polysaccharide significantly increases its antioxidant activity and the chemical modification of polysaccharides may allow the preparation of derivatives with new properties and a variety of applications.     

Structural characterization of a water-soluble polysaccharide from the roots of Codonopsis pilosula and its immunity activity

The water-soluble polysaccharide (CPP), with a molecular mass of 1.1x10(4) Da, was obtained from the roots of Codonopsis pilosula. Structure feature investigation by a combination of chemical and instrumental analysis revealed that CPP had a backbone consisting of (1-->3)-linked-beta-D-galactopyranosyl, (1-->2, 3)-linked-beta-D-galactopyranosyl and (1-->3)-linked-alpha-D-rhamnopyranosyl residues, which were branched with two glycosyl residues composed of alpha-L-arabinose-(1-->5)-alpha-L-arabinose(1-->linked residues at the O-2 position of galactosyl along the main chain in the ratio of 1:1:2:1:1. Preliminary immunological tests in vitro showed CPP could stimulate concanavalin A (ConA)- or lipopolysaccharide (LPS)-induced lymphocyte proliferation in a dose-dependent manner.     

Anti-inflammatory and immunologically active polysaccharides of Periandra mediterranea

Three polysaccharides, glucans with mean M(r)'s of 1.5 x 10(5), 3.6 x 10(4) and 2.1 x 10(4), were isolated from dried roots of Periandra mediterranea by fractionation on Sephacryl S-300 HR and Sephadex G-25. Chemical and spectroscopic studies indicated that they have a highly branched glucan type structure composed of alpha-(1-->4) linked D-glucopyranose residues with both (3-->4) and (4-->6) branching points. The polysaccharides enhance phagocytosis in vivo, and exhibit anti-inflammatory activity.     

NMR spectroscopic analysis of exopolysaccharides produced by Leuconostoc citreum and Weissella confusa

Dextrans are the main exopolysaccharides produced by Leuconostoc species. Other dextran-producing lactic acid bacteria include Streptococci, Lactobacilli, and Weissella species. Commercial production and structural analysis has focused mainly on dextrans from Leuconostoc species, particularly on Leuconostoc mesenteroides strains. In this study, we used NMR spectroscopy techniques to analyze the structures of dextrans produced by Leuconostoc citreum E497 and Weissella confusa E392. The dextrans were compared to that of L. mesenteroides B512F produced under the same conditions. Generally, W. confusa E392 showed better growth and produced more EPS than did L. citreum E497 and L. mesenteroides B512F. Both L. citreum E497 and W. confusa E392 produced a class 1 dextran. Dextran from L. citreum E497 contained about 11% alpha-(1-->2) and about 3.5% alpha-(1-->3)-linked branches whereas dextran from W. confusa E392 was linear with only a few (2.7%) alpha-(1-->3)-linked branches. Dextran from W. confusa E392 was found to be more linear than that of L. mesenteroides B512F, which, according to the present study, contained about 4.1% alpha-(1-->3)-linked branches. Functionality, whether physiological or technological, depends on the structure of the polysaccharide. Dextran from L. citreum E497 may be useful as a source of prebiotic gluco-oligosaccharides with alpha-(1-->2)-linked branches, whereas W. confusa E392 could be a suitable alternative to widely used L. mesenteroides B512F in the production of linear dextran.     

Enzymatic degradation products from a marine polysaccharide YCP with different immunological activity and binding affinity to macrophages,hydrolyzed by α-amylases from different origins

YCP is a marine polysaccharide with anti-tumor and immune-modulating effects. This study evaluated the effect of enzymatic degradation of YCP by α-amylases from different origins on its immunological activity and binding ability to the macrophages. YCP was hydrolyzed by α-amylases isolated from Aspergillus oryzae, Bacillus licheniformis, Barley malt, and Porcine pancreas respectively, then four fragments with unique molecular weight (termed: YCP-Ao, YCP-Bl, YCP-Bm, and YCP-Pp, respectively) were obtained. The four fragments showed different immunological activity and the ability to bind to macrophages. Among them, YCP-Ao possessed almost equivalent immunological activity compared to the original YCP, while such properties were not retained in YCP-Bl. Our further study showed that YCP-Ao prevented YCP from binding to macrophages. In conclusion, YCP-Ao and YCP might have similar active regions.     

A bioactive (1-->3)-, (1--4)-beta-D-glucan from Collybia dryophila and other mushrooms

Polysaccharides from higher Basidiomycete mushrooms, mainly beta-D-glucans, are considered to be potent bioactive fungal compounds. In this study a beta-glucan (1.237 x 10(6) Da) consisting of (1-->3) and (1-->4) glucosidic linkages, named Collybia dryophila polysaccharide (CDP), was extracted from the wild mushroom C. dryophila. CDP was shown to strongly inhibit nitric oxide production in activated macrophages suggesting that this polysaccharide displays a potential anti-inflammatory activity. In addition it was shown that polysaccharides similar to CDP (CDP-like) are present in Lentinus edodes and different wild mushrooms collected in northeastern North America.     

A fucoidan fraction from Ascophyllum nodosum.

A fucoidan fraction was purified from the brown alga Ascophyllum nodosum. The polysaccharide contained L-fucose and sulfate as the only constituents. Combination of methylation analysis, Smith degradation, FTIR and NMR spectroscopy on the native and the de-sulfated polymers demonstrated that the fucoidan consisted of a highly branched core region with primarily alpha-(1-->3)-linked fucosyl residues and a few alpha-(1-->4) linkages. Branch points were at position 2 of the -->3-linked internal residues. The side chains consisted of single and multi-unit fucosyl residues. The combined analytical data suggested also a complex sulfation pattern with substitution principally at position 2 and/or position 4. Such diversity in the structural features of this fucoidan may be of importance for its various biological properties.     

Chemical modification,characterization and structure-anticoagulant activity relationships of Chinese lacquer polysaccharides

A natural lacquer polysaccharide with complex branches was separated into two fractions, LPH (MW 16.9x10(4)) and LPL (MW 6.85x10(4)). Results of 13C NMR and FT-IR indicated they had the same structure. The treatment of LPL with sodium periodate led to a partial cut-off of side chains with 4-O-methyl-D-glucuronic acid in the terminal. These polysaccharides were sulfated in the presence of Py*SO3/DMSO. Depending on the reaction conditions, the products showed a different degree of sulfation (DS) ranging from 0.57 to 1.57 and different molecular weights ranging from 1.71x10(4) to 3.49x10(4). FT-IR analysis showed the equatorial primary OH at O-6 and the axial secondary OH at O-4 were sulfated. Activated partial thromboplastin time (APTT), prothrombin time and thrombin time (TT) assays showed the sulfated polysaccharides could prolong APTT and TT, but not TP. These activities strongly depended on the DS, the molecular weights (MW) and the branching structure of polysaccharides. DS of above 0.8 was essential for anticoagulant activity. The anticoagulant activity increased with the DS and the molecular weights. The molecular weights played a more important role. The branching structure of polysaccharides increased the activities. In our studies, the sulfated polysaccharides with the DS of 1.15 and the highest MW of 3.49x10(4) had the best blood anticoagulant activities.