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.     

Molecular signature of kappa-carrageenan mimics chondroitin-4-sulfate and dermatan sulfate and enables interaction with arylsulfatase B

The common food additive kappa-carrageenan (κ-CGN) is a sulfated polysaccharide that resembles chondroitin-4-sulfate (C4S) and dermatan sulfate (DS). All have a sulfate group on C4 of a glycoside (galactose for CGN and N-acetylgalactosamine for C4S), and the sulfate-bearing glycoside is linked in a β-1,4-configuration to an unsulfated, six-carbon sugar (galactose for CGN, glucuronate for C4S and iduronate for DS). The enzyme arylsulfatase B (ARSB; N-acetylgalactosamine-4-sulfate) is the highly selective enzyme that removes the four-sulfate group from the nonreducing terminus of C4S and DS, thereby regulating subsequent degradation. In this report, κ-CGN is shown to be a substrate for recombinant human ARSB (rhARSB). Sulfate was generated from both C4S and κ-CGN following incubation with rhARSB. Exposure of human colonic epithelial cells to κ-CGN, but not to C4S, produced reactive oxygen species (ROS) and increased interleukin (IL)-8 secretion. The ROS production from κ-CGN was reduced by exposure to rhARSB, but increased by competition from C4S or DS, but not from chondroitin-6-sulfate. Prior treatment of either lambda- or iota-CGN with rhARSB had no impact on ROS, IL-8 or inorganic sulfate production, demonstrating a specific effect of the molecular configuration of κ-CGN. By mimicry of C4S and DS and by interaction with ARSB, κ-CGN can directly interfere with the normal cellular functions of C4S, DS and ARSB. Since C4S and DS are present in high concentration in tissues, the impact of κ-CGN exposure may be due to some extent to interference with the normal biological functions of ARSB, C4S and DS.     

Low sulfated glycosaminoglycans are excreted in patients with the Lowe syndrome

Glycosaminoglycans (GAGs) were prepared from the urine of three patients and from normal individuals by cetylpyridinium chloride precipitation and Pronase digestion. The GAGs were analyzed by electrophoresis, anion-exchange chromatography, and enzymatic and chemical degradation. Each of the three patients showed a four- to fivefold increase in urinary GAG excretion compared to normal controls and in one patient a tenfold increase was measured during a period of behavioral agitation which included joint swelling. Urinary GAGs from affected individuals were characterized by a high proportion of low sulfated molecules. The predominant low sulfated component was chondroitin-4-sulfate (C4S); however, small amounts of chondroitin-6-sulfate (C6S) were also present. Heparan sulfate (HS) was present in normal proportion (5-10%) and most of it was not low sulfated. Abnormal excretion of chondroitin (Ch), hyaluronic acid (HA), and dermatan sulfate (DS) was not detected. These findings suggest that the clinical manifestations of Lowe syndrome may be caused by a defect in GAG metabolism.     

Structure elucidation and sulfated derivatives preparation of two alpha-D-glucans from Gastrodia elata Bl. and their anti-dengue virus bioactivities

The structures of two glucans, WGEW and AGEW, isolated from Gastrodia elata Bl. were elucidated using monosaccharide composition analysis by gas chromatography (GC), methylation analysis by gas chromatography-mass spectrometry (GC-MS) and nuclear magnetic resonance (NMR) spectroscopy. Their structures were deduced as an alpha-D-(1-->4)-glucan with an alpha-(1-->4) linked branch attached to O-6 branch points with different branch degrees. Their sulfate derivatives with distinct degrees of substitution (DS) were prepared. The substitution position was assigned to O-6 according to the (13)C NMR spectra. All sulfated derivatives showed strong anti-dengue virus bioactivities. The structure-activity relationships (SAR) between the polysaccharides and their sulfated derivatives were also investigated. Results showed that the higher the DS is, the more potent the impact on the dengue virus infection would be.     

Protein--protein docking of electron transfer complexes: cytochrome c oxidase and cytochrome c

Electron transferring protein complexes form only transiently and the crystal structures of electron transfer protein--protein complexes involving cytochrome c could so far be determined only for the pairs of yeast cytochrome c peroxidase (CcP) with iso-1-cytochrome c (iso-1-cyt c) and with horse heart cytochrome c (cyt c). This article presents models from computational docking for complexes of cytochrome c oxidase (COX) from Paracoccus denitrificans with horse heart cytochrome c, and with its physiological counterpart cytochrome c552 (c552). Initial docking is performed with the FTDOCK program, which permits an exhaustive search of translational and rotational space. A filtering procedure is then applied to reduce the number of complexes to a manageable number. In a final step of structural and energetic refinement, the complexes are optimized by rigid-body energy minimization with the molecular mechanics package CHARMM. This methodology was first tested on the CcP:iso-1-cyt c complex, in which the complex with the lowest CHARMM energy has an RMSD from the crystal structure of only 1.8 A (C(alpha) carbon atoms). Notably, the crystal conformation has an even lower energy. The same procedure was then applied to COX:cyt c and COX:c552. The lowest-energy COX:cyt c complex is very similar to a docking model previously described for the complex of bovine cytochrome c oxidase with horse heart cytochrome c. For the COX:c552 complex, cytochrome c552 is found in two different orientations, depending on whether it is docked against COX from a two-subunit or from a four-subunit crystal structure, respectively. Both conformations are discussed critically in the light of the available experimental data.     

Purified curdlan and its hydroxyalkyl derivatives: preparation, properties and applications

Curdlan, a β-1,3-glucan produced by fermentation of Alkaligenes faecalis, is a non-ionic gel-forming polysaccharide that is, along with its hydroxalkyl derivatives, a potentially important matrix for life science applications. The commercially available material contains residual nucleic acids, cellular debris and other contaminants that can interfere with electrophoretic separations and visualization procedures. Simple procedures have been developed for purification of the curdlan and coherent gel formation. Curdlan gels can be formed in a variety of chaotrope-containing, or chaotropic, solvents, including 40% formamide, 7 M urea. The chaotropes can be retained or subsequently removed by leaching. Heat treatment before or after leaching enables thermostable gel formation. Hydroxyethyl and glyceryl derivatives of curdlan have been prepared. Depending on the degree of substitution (DS), a whole spectrum of derivatives with a range of unique properties can be obtained. Lower DS hydroxyethyl derivatives form clear, elastic gels in 6 M urea, while the higher DS hydroxyethyl derivatives are soluble in hot water and gel on cooling to form clear, elastic, thermoreversible gels. Partial depolymerization of curdlan by γ-irradiation reduces the viscosity of subsequent preparations, enabling the preparation of higher concentration, more sieving gels. Use of selected preparations to form unique matrices for electrophoretic separations has been demonstrated.     

Solution Properties of Antitumor Sulfated Derivative of α-(1→3)-D-Glucan from Ganoderma lucidum

Four fractions of a water-insoluble α-(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°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 M_w and intrinsic viscosity [η] of the sulfated derivatives S-GL were measured by multi-angle laser light scattering and viscometry. The M_w value (2.4×10⁴) of sulfated glucan S-GL-1 was much lower than that (44.5×10⁴) of original α-(1→3)-D-glucan GL-1. The Mark-Houwink equation and average value of characteristic ratio C_∞ for the S-GL in 0.2 M NaCl aqueous solution at 25°C were found to be: [η]=1.32×10^-3M_w^1.06 (cm³ g^-1) and 16, respectively, in the M_w range from 1.1×10⁴ to 2.4×10⁴. It indicated that the sulfated derivatives of the α-(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 α-(1→3)-D-glucan and curdlan, a β-(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.     

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.     

Sulfated modification and antioxidant activity of exopolysaccahrides produced by Enterobacter cloacae Z0206

Nine modification conditions were designed to sulfate exopolysaccharides (EPS) produced by Enterobacter cloacae Z0206 by chlorosulfonic acid-pyridine (CSA-Pyr) method according to the orthogonal test and focusing on three affecting factors such as the ratio of CSA to Pyr, reaction temperature and reaction time. And nine sulfated derivatives with various degrees of substitution (DS) were obtained. Their antioxidant activities were evaluated in vitro, by scavenging abilities on superoxide radical and hydroxyl radical. The results indicated that sulfated derivatives of EPS showed noticeable effects on scavenging superoxide radical and hydroxyl radical compared with native one, and sulfated derivative with moderate DS of 0.60 showed highest antioxidant activities. The optimum sulfated conditions of EPS were the ratio of CSA to Pyr of 1:2, the reaction time of 2h and reaction temperature of 70 °C.     

Interaction of chondroitin sulfate and dermatan sulfate from various biological sources with heparin-binding growth factors and cytokines

Chondroitin sulfate (CS) and dermatan sulfate (DS) interact with various extracellular molecules such as growth factors, cytokines/chemokines, neurotrophic factors, morphogens, and viral proteins, thereby playing roles in a variety of biological processes including cell adhesion, proliferation, tissue morphogenesis, neurite outgrowth, infections, and inflammation/leukocyte trafficking. CS/DS are modified with sulfate groups at C-2 of uronic acid residues as well as C-4 and/or C-6 of N-acetyl-D-galactosamine residues, yielding enormous structural diversity, which enables the binding with numerous proteins. We have demonstrated that highly sulfated CS-E from squid cartilage, for example, interacts with heparin-binding proteins including midkine, pleiotrophin, and fibroblast growth factors expressed in brain with high affinity (Kd values in the nM range). Here, we analyzed the binding of CS and DS, which have a relatively low degree of sulfation and have been widely used as a nutraceutical and a drug for osteoarthritis etc., with a number of heparin-binding neurotrophic factors/cytokines using surface plasmon resonance (SPR) and structurally characterized the CS/DS chains. SPR showed that relatively low sulfated CS-A, DS, and CS-C also bound with significant affinity to midkine, pleiotrophin, hepatocyte growth factor, monokine-induced by interferon-γ, and stromal cell derived factor-1β, although the binding was less intense than that with highly sulfated CS-D and CS-E. These findings suggest that even low sulfated CS and/or DS chains may contain binding domains, which include fine sugar sequences with specific sulfation patterns, and that sugar sequences, conformations and electrostatic potential are more important than the simple degree of sulfation represented by disaccharide composition.     

Structure and antioxidant activities of sulfated guar gum: homogeneous reaction using DMAP/DCC catalyst

It was essential to understand the chemical structure of polysaccharides for further research and biochemical or medical application of this natural biopolymer. In the present study, sulfated derivatives of guar gum with high degree of sulfation (DS) were synthesized using 4-dimethylaminopyridine (DMAP)/dimethylcyclohexylcarbodiimide (DCC) as catalyst in homogeneous conditions. The effects of the ratio of chlorosulfuric acid to pyridine, the content of catalyst and reaction temperature were investigated. Results of FT-IR, (1)H and (13)C NMR indicated that C-6 substitution was predominant in sulfated polysaccharide. In the sulfation reaction, a sharp decrease in M(W) was observed. The enhanced antioxidant activities of sulfated polysaccharides were not a function of a single factor but a combination of high DS and low molecule weight.     

Synthesis and anticoagulant activity of the quaternary ammonium chitosan sulfates

Quaternary ammonium chitosan sulfates with diverse degrees of substitution (DS) ascribed to sulfate groups between 0.52 and 1.55 were synthesized by reacting quaternary ammonium chitosan with an uncommon sulfating agent (N(SO₃Na)₃) that was prepared from sodium bisulfite (NaHSO₃) through reaction with sodium nitrite (NaNO₂) in the aqueous system homogeneous. The structures of the derivatives were characterized by FTIR, ¹H NMR and ¹³C NMR. The factors affecting DS of quaternary ammonium chitosan sulfates which included the molar ratio of NaNO₂ to quaternary ammonium chitosan, sulfated temperature, sulfated time and pH of sulfated reaction solution were investigated in detail. Its anticoagulation activity in vitro was determined by an activated partial thromboplastin time (APTT) assay, a thrombin time (TT) assay and a prothrombin time (PT) assay. Results of anticoagulation assays showed quaternary ammonium chitosan sulfates significantly prolonged APTT and TT, but not PT, and demonstrated that the introduction of sulfate groups into the quaternary ammonium chitosan structure improved its anticoagulant activity obviously. The study showed its anticoagulant properties strongly depended on its DS, concentration and molecular weight.     

New poly(A)+RNAs appear coordinately during the differentiation of Naegleria gruberi amebae into flagellates

We have examined the nature of the requirement for RNA synthesis during the differentiation of Naegleria gruberi amebae into flagellates (Fulton, C., and C. Walsh, 1980, J. Cell Biol., 85:346-360) by looking for poly(A)+RNAs that are specific to differentiating cells. A cDNA library prepared from poly(A)+RNA extracted from cells 40 min after initiation of the differentiation (40-min RNA), the time when formation of flagella becomes insensitive to inhibitors of RNA synthesis, was cloned into pBR322. Recombinant clones were screened for sequences that were complementary to 40-min RNA but not to RNA from amebae (0-min RNA). Ten of these differentiation-specific (DS) plasmids were identified. The DS plasmids were found to represent at least four different poly(A)+RNAs based on cross-hybridization, restriction mapping, and Northern blot analysis. Dot blot analysis was used to quantify changes in DS RNA concentration. The four DS RNAs appeared coordinately during the differentiation. They were first detectable at 10-15 min after initiation, reached a peak at 70 min as flagella formed, and then declined to low levels by 120 min when flagella reached full length. The concentration of the DS RNAs was found to be at least 20-fold higher in cells at 70 min than in amebae. The changes in DS RNA concentration closely parallel changes in tubulin mRNA as measured by in vitro translation (Lai, E.Y., C. Walsh, D. Wardell, and C. Fulton, 1979, Cell, 17:867-878).     

Sulfated modification, characterization and antitumor activities of Radix hedysari polysaccharide

Sulfated modification of a polysaccharide obtained from Radix hedysari (RHP) was studied. Four sulfated derivatives (RHPS) with variable degrees of substitution (DS) were obtained by the chlorosulfonic acid method with ionic liquids (ILs) as solvent and 4-dimethylaminopyridine (DMAP) as catalyst. The structures of RHPS were characterized by FT-IR spectra and (13)C NMR spectra, and the results indicated that the sulfated groups were modified mainly at the C-6 position and C-2 position. Four kinds of RHPS showed different DS ranging from 0.63 to 1.45, and different weight-average molecular mass (Mw) ranging from 60.8 to 71.1kDa with a little degradation. Compared with RHP, all of RHPS exhibited obvious antitumor activity on A549 cells and BGC-823 cells in vitro. However, they had no obvious influence on HEK293 cells, which indicated that they had low toxicity to normal cells. Flow cytometric studies indicated that the treatment of RHPS against A549 cells and BGC-823 cells could mediate the cell-cycle arrest in the G1 phase.     

Sulfated hyaluronan derivatives reduce the proliferation rate of primary rat calvarial osteoblasts

Glycosaminoglycans (GAG) and proteoglycans, which are components of the extracellular bone matrix, are also localized in and at the membrane of osteoblasts and in the pericellular matrix. Due to their interaction with several growth factors, water and cations these molecules play an important role in regulating proliferation and differentiation of osteoblasts and bone development. The aim of this study was to assess in vitro the effects of two chemically sulfated hyaluronan (HyaS) derivatives on the proliferation of rat calvarial osteoblasts and to compare with those of native hyaluronan (Hya) and natural sulfated GAG such as chondroitin-4-sulfate (C4S), chondroitin-6-sulfate (C6S), dermatan sulfate (DS) and heparan sulfate (HS). Moderately and highly sulfated HyaS derivatives caused a time-dependent reduction of osteoblast proliferation. The anti-proliferative effect of HyaS was accompanied by a cell cycle arrest in the G1 phase, but was not associated with cell death. Whereas non-sulfated high molecular weight (HMW)- and low molecular weight (LMW)-Hya as well as C4S, C6S, DS and HS showed no effect on the cell proliferation.     

Sulfated modification of the polysaccharides obtained from defatted rice bran and their antitumor activities

Nine sulfated defatted rice bran polysaccharides (sRBPS), with various degrees of sulfation (DS) and carbohydrate content, were prepared by chlorosulfonic acid–pyridine (CSA–Pyr) method according to orthogonal test. Nine sulfated derivatives sRBPS were obtained and their antitumor activities were compared by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The results showed that when DS within the scope of 0.81–1.29, carbohydrate content in the range of 41.41–78.56%, sulfated derivatives exhibit relatively strong antitumor activity in vitro. The optimum modification conditions were reaction temperature of 70 °C, the ratio of chlorosulfonic acid to pyridine of 1:4 and the reaction time of 2 h.     

Catalytic synthesis and antioxidant activity of sulfated polysaccharide from Momordica charantia L.

Sulfated derivatives of polysaccharide from Momordica charantia L. (MCPS) with different degree of sulfation (DS) were synthesized by chlorosulfonic acid method with ionic liquids as solvent. Fourier transform infrared spectra and ¹³C nuclear magnetic resonance spectra indicated that C‐6 substitution was predominant in MCPS compared with the C‐2 position. Compared with the native polysaccharide from Momordica charantia L. (MCP), MCPS exhibited more excellent antioxidant activities in vitro, which indicated that sulfated modification could enhance antioxidant activities of MCP. Furthermore, high DS and moderate molecular weight could improve the antioxidant activities of polysaccharide. © 2013 Wiley Periodicals, Inc. Biopolymers 101: 210–215, 2014.     

Comparison of three S‐β‐CDs with different degrees of substitution for the chiral separation of 12 drugs in capillary electrophoresis

Three kinds of sulfated β‐cyclodextrin (S‐β‐CD), including a single isomer, heptakis‐6‐sulfato‐β‐cyclodextrin (HS‐β‐CD), degree of substitution (DS) of 7, which was synthesized in our laboratory and another two commercialized randomly substituted mixtures, a sulfated β‐cyclodextrin with DS of 7 to 11, as well as a highly sulfated‐β‐cyclodextrin with DS of 12 to 15, were used for the enantioresolution of 12 drugs (the β‐blockers, phenethylamines, and anticholinergic agents) in capillary electrophoresis. The enantioseparation under varying concentrations of S‐β‐CD and background electrolyte pH were systematically investigated and compared. Based on the experimental results, the effect of the nature of S‐β‐CD and analyte structure on the enantioseparation is discussed.     

What initially brought about communications?

This paper reports an intelligent agent equipped with two-layer finite state machines (FSMs) that can communicate by turning lighting on and off, leading to social cooperation that solves the dilemma situation, modeled by a one-shot 2x2 game. This communication between two gaming agents can be observed in hero- and leader-type dilemma games, where alternating reciprocity, repeating cooperation (C)-defeat (D) after D-C, is the equal pareto optimum instead of a sequence of mutual cooperation that is the equal pareto optimum for a prisoner's dilemma (PD) game.     

Chiral recognition ability of curdlan triacetate: solvent and temperature effects

The chiral recognition ability of the chiral stationary phase (CSP) consisting of curdlan (beta-1,3-glucan) triacetate coated on silica gel was clearly changed by the contacting solvents and heat treatment. The chiral recognition ability significantly decreased, particularly at temperatures above 45 degrees C, depending on the racemates. The molecular weight of the curdlan triacetate slightly influenced its ability. The recognition abilities of curdlan tricetate that was lost by heat treatment were partially recovered by contact with methanol. However, when it was contacted with ethanol a different selectivity was observed. The labile chiral recognition ability of curdlan triacetate is in striking contrast to the very stable chiral recognition of cellulose (beta-1,4-glucan) triacetate (Chiralcel OA). This difference may be ascribed to the conformational stability of the acetates consisting of curdlan (beta-1,3-glucan) and cellulose (beta-1,4-glucan) with different sugar linkages.