Determination of the molecular weight of microbial polysaccharides using high performance exclusion chromatography

An automatic method of determining the molecular weight parameters (Mw, Mn) of microbial polysaccharides such as dextran, pullulan was developed based on the use of high performance size-exclusion chromatography on the two types of columns: Zorbax PSM 60 + 300 + 1000 and SynChropack GPC 100 + 500 + 1000. The Mw and Mn values were determined for a number of domestic and foreign dextran preparations. Changes in the molecular weight of pullulan and hydroxyethylstarch resulted from acid and enzymatic hydrolysis were estimated.     

Fermentation of mucin and plant polysaccharides by strains of Bacteroides from the human colon.

Ten Bacteroides species found in the human colon were surveyed for their ability to ferment mucins and plant polysaccharides ("dietary fiber"). A number of strains fermented mucopolysaccharides (heparin, hyaluronate, and chondroitin sulfate) and ovomucoid. Only 3 of the 188 strains tested fermented beef submaxillary mucin, and none fermented porcine gastric mucin. Many of the Bacteroides strains tested were also able to ferment a variety of plant polysaccharides, including amylose, dextran, pectin, gum tragacanth, gum guar, larch arabinogalactan, alginate, and laminarin. Some plant polysaccharides such as gum arabic, gum karaya, gum ghatti and fucoidan, were not utilized by any of the strains tested. The ability to utilize mucins and plant polysaccharides varied considerably among the Bacteroides species tested.     

The effects of plant polysaccharides and buffer additives on PCR.

A survey of the inhibitory effects of various plant polysaccharides on PCR amplification of a 974-bp section of rbcL in spinach revealed that most of the polysaccharides tested (arabinogalactan, carrageenan, dextran, gum guar, gum karaya, gum locust bean, inulin, mannan, pectin, starch and xylan) were not inhibitory. In contrast, two of the acidic polysaccharides (dextran sulfate and gum ghatti) were inhibitory. The addition of 0.5% Tween 20 reversed the inhibitory effects of gum ghatti (polysaccharide:DNA ratio of 500:1). The inhibitory effect of dextran sulfate (50:1) could be reversed by the addition of Tween 20 (0.25% or 0.5%), DMSO (5%) or polyethylene glycol 400 (5%), but none of these three additives were effective at a 100:1 ratio of dextran sulfate/DNA.     

Interaction of gum arabic, maltodextrin and pullulan with lipids in emulsions

The interaction of gum arabic, maltodextrin and pullulan with lipids in emulsion systems was investigated. Interfacial tension and interfacial viscosity measurements revealed that only gum arabic could adsorb and form a viscoelastic film at the oil-water interface. Good emulsifying activity was demonstrated for gum arabic, whereas fine emulsions could not be produced from the other polysaccharide solutions and oil. Frequency-dependent increases in the storage and loss moduli were observed for all the polysaccharide solutions. Such rheological behavior did not substantially change when maltodextrin and pullulan were mixed with oil to form emulsions. However, the frequency-dependence of the dynamic moduli disappeared in the gum arabic-stabilized emulsion, suggesting the formation of a network structure in which oil droplets could form junctions with gum arabic chains. The results on the inhibition of lipid oxidation by polysaccharides suggest that gum arabic protected lipids from the attack of lipoxygenase and free radicals by adsorbing at the oil droplet surface.     

Inhibition of random amplified polymorphic DNAs (RAPDs) by plant polysaccharides

A survey of the inhibition of the amplification of spinach DNA by various plant polysaccharides revealed that neutral polysaccharides (arabinogalactan, dextran, gum guar, gum locust bean, inulin, mannan, and starch) were not inhibitory. In contrast, the acidic polysaccharides (carrageenan, dextran sulfate, gum ghatti, gum karaya, pectin, and xylan)were inhibitory. In the process of preparing random amplified polymorphic DNAs (RAPDs), the loss of large DNA bands appears to be an indicator that the fingerprint pattern has been affected by polysaccharides. The addition of various concentrations of Tween 20, DMSO, or PEG 400 to the PCR reaction mixture resulted in partial restoration of amplification of RAPDs for the acidic polysaccharides. The most effective way to eliminate the effects of polysaccharide inhibition was by diluting the DNA extracts, and thereby diluting the polysaccharide inhibitors.     

Water-soluble polymers in agriculture: xanthan gum as eco-friendly alternative to synthetics

Water-soluble polymers (WSPs) are a versatile group of chemicals used across industries for different purposes such as thickening, stabilizing, adhesion and gelation. Synthetic polymers have tailored characteristics and are chemically homogeneous, whereas plant-derived biopolymers vary more widely in their specifications and are chemically heterogeneous. Between both sources, microbial polysaccharides are an advantageous compromise. They combine naturalness with defined material properties, precisely controlled by optimizing strain selection, fermentation operational parameters and downstream processes. The relevance of such bio-based and biodegradable materials is rising due to increasing environmental awareness of consumers and a tightening regulatory framework, causing both solid and water-soluble synthetic polymers, also termed ‘microplastics’, to have come under scrutiny. Xanthan gum is the most important microbial polysaccharide in terms of production volume and diversity of applications, and available as different grades with specific properties. In this review, we will focus on the applicability of xanthan gum in agriculture (drift control, encapsulation and soil improvement), considering its potential to replace traditionally used synthetic WSPs. As a spray adjuvant, xanthan gum prevents the formation of driftable fine droplets and shows particular resistance to mechanical shear. Xanthan gum as a component in encapsulated formulations modifies release properties or provides additional protection to encapsulated agents. In geotechnical engineering, soil amended with xanthan gum has proven to increase water retention, reduce water evaporation, percolation and soil erosion – topics of high relevance in the agriculture of the 21st century. Finally, hands-on formulation tips are provided to facilitate exploiting the full potential of xanthan gum in diverse agricultural applications and thus providing sustainable solutions.     

Partition of polysaccharide-coated liposomes in aqueous two-phase systems

Hydrophobized polysaccharides such as cholesterol-bearing pullulan (CHP), dextran (CHD) and mannan (CHM) effectively coat the liposomal surface. Partition of the hydrophobized polysaccharide-coated liposomes in an aqueous two-phase system (PEO (top)/pullulan (bottom) or PEO (top)/dextran (bottom)) was investigated (PEO = poly(ethylene oxide)). Conventional liposomes without a polysaccharide coat mostly locate at the interface between the two polymer phases. The polysaccharide-coated liposomes, on the other hand, were partly partitioned to the bottom polysaccharide phase depending on the structure of the hydrophobized polysaccharide on the liposomal surface. The affinity between the polysaccharide on the liposomal surface and that in the bulk bottom phase controls the efficiency of partition. The sequence of interaction strength between the two carbohydrates as the following: for the PEO/dextran two-phase system, dextran_(liposome)-dextran_(bulk) > mannan_(liposome)-dextran_(bulk) > pullulan_(liposome)-dextran_(bulk); while for the PEO/pullulan system, the sequence of interaction strength was pullulan_(liposome)-pullulan_(bulk) > dextran_(liposome)-pullulan_(bulk) ≈ mannan_(liposome)-pullulan_(bulk).     

Determination of structural peculiarities of dexran,pullulan and gamma-irradiated pullulan by Fourier-transform IR spectroscopy

Deconvoluted IR-absorbance spectra of dextran, pullulan and gamma-irradiated pullulan were analyzed in order to find the most specific spectral peculiarities that allow one to obtain information about the structure and conformation of these macromolecules in solvents that exhibit different influences on the system of intra- and intermolecular interactions. The changes in intensity and width of the IR bands at about 1040, 1020 and, in the case of pullulan, also at 996 cm(-1), were related to changes in conformation and short-range interactions of the polysaccharides. Furthermore, certain bands within the 1200-900 cm(-1) region were considered as a characteristic for the type of glycosidic linkage. The results of the FTIR spectroscopy study allowed one to suggest a predominant cleavage of the alpha-(1-->4) linkages upon the radiation-chemical destruction of pullulan.     

Polysaccharides as labels for antibodies in electron microscopy.

Two polysaccharides, dextran 250 and dextran 70, were covalently linked to antibody molecules, antihuman immunoglobulin G and antihuman type O red blood cells. In electron microscope preparations exposed to lead citrate, polysaccharides, because they chelate lead, were quite dense. Polysaccharides served as a tag for the antibody molecules. Also, bacterial dextran 1355 was used to demonstrate antibody molecules on the surface of ascites tumor cells which are known to be producing a specific antibody to bacterial dextran 1355. The varying sized polysaccharide molecules that are readily available commercially, the high electron density of the polysaccharides after lead staining and a mild procedure for covalently linking polysaccharide to antibody make polysaccharides attractive as particulate labels for antibody in electronmicroscopy.     

出芽短梗霉Aureobasidium sp. SRF的菌株特性分析及胞外多糖结构鉴定

菌株SRF是1株从意大利树莓(Rubus corchorifolius)果实表面分离、可产胞外多糖的新菌株。在鉴定其分类归属的基础上,对其产生的胞外多糖进行了结构分析和发酵条件优化,为寻找微生物多糖提供新的菌株,为开发利用资源微生物提供借鉴。通过形态学和ITS序列对比分析进行菌株鉴定;通过薄层层析和红外光谱分析,确定胞外多糖结构;通过单因素检测试验,确定影响产糖量的主要因素;响应面Plackett-Burman和Box-Behnken设计筛选发酵产胞外多糖的最优条件。结果表明,出发菌株SRF隶属于出芽短梗霉属,命名为Aureobasidium sp. SRF;SRF所产胞外多糖为普鲁兰多糖;单因素检测表明,对多糖产量影响最大的因素为碳源浓度、氮源浓度、无机离子浓度,其次是碳源、氮源、无机离子、pH值;根据响应面结果确定最优发酵条件为麦芽糖8%(质量分数)、酵母提取物3%(质量分数)、钙离子0.3 g/L、pH 6,产糖量达5.93 g/L。SRF是1株来源于树莓浆果表面,可产胞外普鲁兰多糖的出芽短梗霉新菌株,是1株产微生物多糖的候选菌株。     

Gelation of Limulus Lysate by Synthetic Dextran Derivatives

The Limulus test has been considered specific for the presence of bacterial endotoxins. To synthesize a simple model of endotoxin, palmitoyldextran phosphate was prepared by modification of dextran by palmitoylation and phosphorylation. The present studies indicated that a variety of polysaccharide derivatives, such as palmitoyldextran phosphate, palmitoyldextran, and dextran phosphate, give a positive Limulus test and show pyrogenic activity, except for low molecular dextran derivatives. On the other hand, polysaccharides, such as dextran, starch (soluble), chitosan, xylan, and lentinan, were negative in these assays. The gelation reaction of Limulus lysate by modified dextran derivatives may depend on the molecular weight or modification of polysaccharides by palmitoylation and/or phosphorylation to a great extent.     

TEMPO-mediated selective oxidation of substituted polysaccharides--an efficient approach for the determination of the degree of substitution at C-6

2,2,6,6-Tetramethyl-1-piperidinyloxy radical (TEMPO)-mediated oxidations of substituted polysaccharides were studied at pH 10.2 and at a temperature of 0 °C with NaOCl as the oxidant. The reaction is highly selective, and it was shown that the oxidation can proceed to a yield of nearly 100%. The oxidation process was investigated for several substituted polysaccharides, especially for a series of hydroxypropyl guar gums with different molar degrees of substitution. It was shown that this oxidation can be used for the determination of the degree of substitution at C-6 of the polysaccharide by comparing the difference in oxidation yield between substituted and natural polysaccharides. Studies on several hydroxypropyl guar gums showed that the degrees of substitution at C-6—for MS of 0.08, 0.34, 0.62, and 1.08—are 0.06, 0.24, 0.40, and 0.44, respectively. The results were extended to other polysaccharides such as carboxymethyl cellulose, cationic guar gum, carboxymethyl pullulan, and methyl cellulose. It can be concluded that the TEMPO-mediated oxidation is a useful method for the determination of the DS at the substituted C-6 position for different kinds of modified polysaccharides.     

Pressate from Peat Dewatering as a Substrate for Bacterial Growth

This study considered the possibility of using water expressed during the drying of fuel-grade peat as a substrate for microbial growth. Highly humified peat pressed for 2.5 min at 1.96 MPa produced water with a chemical oxygen demand of 690 mg/liter. Several biological compounds could be produced by using the organic matter in expressed peat water as a substrate. These included polymers such as chitosan, contained in the cell wall of Rhizopus arrhizus, and two extracellular polysaccharides, xanthan gum and pullulan, produced by Xanthomonas campestris and Aureobasidium pullulans, respectively. A very effective surfactant was produced by Bacillus subtilis grown in the expressed water. Small additions of nutrients to the peat pressate were necessary to obtain substantial yields of products. The addition of peptone, yeast extract, and glucose improved production of the various compounds. Biological treatment improved the quality of the expressed water to the extent that in an industrial process it could be returned to the environment.     

Functional differences between Arctic seawater and sedimentary microbial communities: contrasts in microbial hydrolysis of complex substrates

The activities and structural specificities of extracellular enzymes that initiate microbial remineralization of high-molecular-weight (MW) organic matter were investigated in surface waters and sediments of an Arctic fjord of Svalbard. Hydrolysis rates of a suite of fluorescently labeled macromolecular substrates, including seven commercially available polysaccharides and three high-carbohydrate-content plankton extracts ranged from rapid to not detectable, and differed markedly between seawater and sediments. Order (fastest to slowest) of hydrolysis in surface water was laminarin, Spirulina extract, xylan>chondroitin, alginic acid, Wakame extract>arabinogalactan, fucoidan>Isochrysis extract>pullulan, while in sediments the order was pullulan, laminarin, alginic acid, Wakame extract>chondroitin, xylan>arabinogalactan, Isochrysis extract>Spirulina extract>fucoidan. These differences cannot be explained by simple scaling factors such as differences in microbial cell numbers between seawater and sediments. Other investigations have shown that microbial community composition of Svalbard sediments and of polar bacterioplankton samples differ markedly. These results demonstrate that sedimentary and seawater microbial communities also differ fundamentally in their abilities to access specific high-MW substrates. Substrate bioavailability depends on the capabilities of a microbial community, as well as the chemical and structural features of the substrate itself.     

Molecular motions of polysaccharides in the solid state: dextran, pullulan and amylose.

Dextran, pullulan and amylose have been investigated by differential scanning calorimetry, thermogravimetric analysis, dynamic mechanical and dielectric spectroscopy over a wide range of temperatures and frequencies. No melting or glass transition is seen below the range of thermal degradation (about 300 degrees C) for either amylose or pullulan; only dextran shows a Tg at 223 degrees C (delta cp = 0.40 J/g deg). The viscoelastic spectrum of the 'dry' polysaccharides is characterized by a low temperature relaxation that occurs at -94, -73 and -59 degrees C, at 1 kHz, (activation energy 32, 39 and 52 kJ/mol) in dextran, pullulan and amylose respectively and is assigned to small entity local motions of the polysaccharide backbone. Absorbed water strongly modifies the relaxation spectrum, inducing a new relaxation below room temperature and dissipation regions associated with water loss above room temperature. The former appears at temperatures higher than the relaxation characteristic of the dry polymer and moves to lower temperature with increasing water content. In normal 'room humidity' conditions (about 10% absorbed water) the water-induced relaxation, attributed to the motion of complex polymer-water relaxing units, is the only observable feature in the dynamic mechanical and dielectric spectrum below room temperature.     

Application of Natural Polysaccharides in Pharmaceutics

Russian Journal of Bioorganic Chemistry - Modern data are presented on the pharmaceutical application of natural polysaccharides, such as pullulan, alginates, hyaluronic acid, dextrans, pectin,...     

Functional Characterization of a Newly Identified Group B Streptococcus Pullulanase Eliciting Antibodies Able to Prevent Alpha-Glucans Degradation

Streptococcal pullulanases have been recently proposed as key components of the metabolic machinery involved in bacterial adaptation to host niches. By sequence analysis of the Group B Streptococcus (GBS) genome we found a novel putative surface exposed protein with pullulanase activity. We named such a protein SAP. The sap gene is highly conserved among GBS strains and homologous genes, such as PulA and SpuA, have been described in other pathogenic streptococci. The SAP protein contains two N-terminal carbohydrate-binding motifs, followed by a catalytic domain and a C-terminal LPXTG cell wall-anchoring domain. In vitro analysis revealed that the recombinant form of SAP is able to degrade α-glucan polysaccharides, such as pullulan, glycogen and starch. Moreover, NMR analysis showed that SAP acts as a type I pullulanase. Studies performed on whole bacteria indicated that the presence of α-glucan polysaccharides in culture medium up-regulated the expression of SAP on bacterial surface as confirmed by FACS analysis and confocal imaging. Deletion of the sap gene resulted in a reduced capacity of bacteria to grow in medium containing pullulan or glycogen, but not glucose or maltose, confirming the pivotal role of SAP in GBS metabolism of α-glucans. As reported for other streptococcal pullulanases, we found specific anti-SAP antibodies in human sera from healthy volunteers. Investigation of the functional role of anti-SAP antibodies revealed that incubation of GBS in the presence of sera from animals immunized with SAP reduced the capacity of the bacterium to degrade pullulan. Of interest, anti-SAP sera, although to a lower extent, also inhibited Group A Streptococcus pullulanase activity. These data open new perspectives on the possibility to use SAP as a potential vaccine component inducing functional cross-reacting antibodies interfering with streptococcal infections.     

Film forming microbial biopolymers for commercial applications—A review

Abstract

Microorganisms synthesize intracellular, structural and extracellular polymers also referred to as biopolymers for their function and survival. These biopolymers play specific roles as energy reserve materials, protective agents, aid in cell functioning, the establishment of symbiosis, osmotic adaptation and support the microbial genera to function, adapt, multiply and survive efficiently under changing environmental conditions. Viscosifying, gelling and film forming properties of these have been exploited for specific significant applications in food and allied industries. Intensive research activities and recent achievements in relevant and important research fields of global interest regarding film forming microbial biopolymers is the subject of this review. Microbial polymers such as pullulan, kefiran, bacterial cellulose (BC), gellan and levan are placed under the category of exopolysaccharides (EPS) and have several other functional properties including film formation, which can be used for various applications in food and allied industries. In addition to EPS, innumerable bacterial genera are found to synthesis carbon energy reserves in their cells known as polyhydroxyalkanoates (PHAs), microbial polyesters, which can be extruded into films with excellent moisture and oxygen barrier properties. Blow moldable biopolymers like PHA along with polylactic acid (PLA) synthesized chemically in vitro using lactic acid (LA), which is produced by LA bacteria through fermentation, are projected as biodegradable polymers of the future for packaging applications. Designing and creating of new property based on requirements through controlled synthesis can lead to improvement in properties of existing polysaccharides and create novel biopolymers of great commercial interest and value for wider applications. Incorporation of antimicrobials such as bacteriocins or silver and copper nanoparticles can enhance the functionality of polymer films especially in food packaging applications either in the form of coatings or wrappings. Use of EPS in combinations to obtain desired properties can be evaluated to increase the application range. Controlled release of active compounds, bioactive protection and resistance to water can be investigated while developing new technologies to improve the film properties of active packaging and coatings. An holistic approach may be adopted in developing an economical and biodegradable packaging material with acceptable properties. An interdisciplinary approach with new innovations can lead to the development of new composites of these biopolymers to enhance the application range. This current review focuses on linking and consolidation of recent research activities on the production and applications of film forming microbial polymers like EPS, PHA and PLA for commercial applications.     

Pullulan and dextran: uncommon composition dependent Flory-Huggins interaction parameters of their aqueous solutions

Vapor pressure measurements were performed for aqueous solutions of pullulan ( M w 280 kg/mol) and dextran ( M w 60 and 2100 kg/mol, respectively) at 25, 37.5, and 50 degrees C. The Flory-Huggins interaction parameters obtained from these measurements, plus information on dilute solutions taken from the literature, show that water is a better solvent for pullulan than for dextran. Furthermore, they evince uncommon composition dependencies, including the concurrent appearance of two extrema, a minimum at moderate polymer concentration and a maximum at high polymer concentration. To model these findings, a previously established approach, subdividing the mixing process into two clearly separable steps, was generalized to account for specific interactions between water and polysaccharide segments. Three adjustable parameters suffice to describe the results quantitatively; according to their numerical values, the reasons for the solubility of polysaccharides in water, as compared with that of synthetic polymers in organic solvents, differ in a principal manner. In the former case, the main driving force comes from the first step (contact formation between the components), whereas it is the second step (conformational relaxation) that is advantageous in the latter case.     

Tree gum-based renewable materials: Sustainable applications in nanotechnology,biomedical and environmental fields

The prospective uses of tree gum polysaccharides and their nanostructures in various aspects of food, water, energy, biotechnology, environment and medicine industries, have garnered a great deal of attention recently. In addition to extensive applications of tree gums in food, there are substantial non-food applications of these commercial gums, which have gained widespread attention due to their availability, structural diversity and remarkable properties as ‘green’ bio-based renewable materials. Tree gums are obtainable as natural polysaccharides from various tree genera possessing exceptional properties, including their renewable, biocompatible, biodegradable, and non-toxic nature and their ability to undergo easy chemical modifications. This review focuses on non-food applications of several important commercially available gums (arabic, karaya, tragacanth, ghatti and kondagogu) for the greener synthesis and stabilization of metal/metal oxide NPs, production of electrospun fibers, environmental bioremediation, bio-catalysis, biosensors, coordination complexes of metal–hydrogels, and for antimicrobial and biomedical applications. Furthermore, polysaccharides acquired from botanical, seaweed, animal, and microbial origins are briefly compared with the characteristics of tree gum exudates.