Optimization, purification, characterization and antioxidant activity of an extracellular polysaccharide produced by Paenibacillus polymyxa SQR-21

The optimization, purification and characterization of an extracellular polysaccharide (EPS) from a bacterium Paenibacillus polymyxa SQR-21 (SQR-21) were investigated. The results showed that SQR-21 produced one kind of EPS having molecular weight of 8.96 × 10⁵ Da. The EPS was comprised of mannose, galactose and glucose in a ratio of 1.23:1.14:1. The ratio of monosaccharides and glucuronic acid was 7.5:1. The preferable culture conditions for EPS production were pH 6.5, temperature 30 °C for 96 h with yeast extract and galactose as best N and C sources, respectively. The maximum EPS production (3.44 g L⁻¹) was achieved with galactose 48.5 g L⁻¹, Fe³⁺ 242 μM and Ca²⁺ 441 μM. In addition, the EPS showed good superoxide scavenging, flocculating and metal chelating activities while moderate inhibition of lipid peroxidation and reducing activities were determined. These results showed the great potential of EPS produced by SQR-21 to be used in industry in place of synthetic compounds.     

微生物胞外多糖的合成及其在重金属修复中的作用机制与应用

重金属的生物不可降解性使其在环境中长期存在,导致严重的环境污染,对人类健康和生态系统构成威胁。与传统的物化修复技术相比,微生物修复具有成本低廉、环境友好和高效等特点。在面对重金属胁迫或营养不均衡时,微生物会被激发以分泌合成胞外多糖(exopolysaccharides, EPS)。由此可见,EPS的产生是微生物对抗重金属胁迫的重要策略之一。EPS不仅能保护微生物在低温、高温、高盐等极端环境或受毒性化合物胁迫的条件下存活,并且在细胞内外进行信息和物质的交流与传递,既作为保护屏障限制重金属离子进入细胞,又作为介质进行交流。EPS结构中含有多个带负电荷的官能团,能够与重金属离子发生络合、离子交换、氧化还原等反应,从而降低重金属的生物有效性并减轻其毒性。微生物EPS在重金属胁迫环境中的修复具有重要意义。然而,目前缺乏关于微生物EPS合成过程、与重金属互作机制及其在重金属胁迫环境中应用现状的系统综述。本文概述了微生物EPS及其分类,详细阐述了细菌EPS胞内及胞外的生物合成机制,并探讨了微生物EPS与重金属互作机制,以及微生物EPS修复水、土环境中重金属污染方面的研究进展。最后,展望了EPS合成及其在重金属修复中的作用机制研究,可为微生物EPS进一步应用于环境重金属污染修复提供支持。     

Biosynthesis, characterisation, and design of bacterial exopolysaccharides from lactic acid bacteria

Lactic acid bacteria (LAB) are characterised by their conversion of a large proportion of their carbon feed, fermentable sugars, to lactic acid. However, in addition to lactic acid production, the LAB are able to divert a small proportion of fermentable sugars towards the biosynthesis of exopolysaccharides (EPSs) that are independent of the cell surface and cell wall material. These microbial EPSs when suspended or dissolved in aqueous solution provide thickening and gelling properties, and, as such, there is great interest in using EPSs from food grade microorganisms (such as the LAB that are traditionally used for food fermentations) for use as thickening agents. The current review includes a brief summary of the recent literature describing features of the biosynthetic pathways leading to EPS production. Many aspects of EPS biosynthesis in LAB are still not fully understood and a number of inferences are made regarding the similarity of the pathway to those involved in the synthesis of other cell polysaccharides, e.g., cell wall components. The main body of the review will cover practical aspects concerned with the isolation and characterisation of EPS structures. In the last couple of years, a substantial number of structures have been published and a summary of the common elements of these structures is included as is a suggestion for a system for representing structures. A brief highlight of the attempts that are being made to design ‘tailor’-made polysaccharides using genetic modification and control of metabolic flux is presented.     

Integrated hydrogen production process from cellulose by combining dark fermentation, microbial fuel cells, and a microbial electrolysis cell

Hydrogen gas production from cellulose was investigated using an integrated hydrogen production process consisting of a dark fermentation reactor and microbial fuel cells (MFCs) as power sources for a microbial electrolysis cell (MEC). Two MFCs (each 25 mL) connected in series to an MEC (72 mL) produced a maximum of 0.43 V using fermentation effluent as a feed, achieving a hydrogen production rate from the MEC of 0.48 m³ H₂/m³/d (based on the MEC volume), and a yield of 33.2 mmol H₂/g COD removed in the MEC. The overall hydrogen production for the integrated system (fermentation, MFC and MEC) was increased by 41% compared with fermentation alone to 14.3 mmol H₂/g cellulose, with a total hydrogen production rate of 0.24 m³ H₂/m³/d and an overall energy recovery efficiency of 23% (based on cellulose removed) without the need for any external electrical energy input.     

Structure of the high-molecular weight exopolysaccharide isolated from Lactobacillus pentosus LPS26

The strain Lactobacillus pentosus LPS26 produces a capsular polymer composed of a high- (2.0 × 10⁶ Da) (EPS A) and a low-molecular mass (2.4 × 10⁴ Da) (EPS B) polysaccharide when grown on semi-defined medium containing glucose as the carbon source. The structure of EPS A and its deacetylated form has been determined by monosaccharide and methylation analysis as well as by 1D/2D NMR studies (¹H and ¹³C). We conclude that EPS A is a charged heteropolymer, with a composition of d-glucose, d-glucuronic acid and l-rhamnose in a molar ratio 1:2:2. The repeating unit is a pentasaccharide with two O-acetyl groups at O-4 of the 3-substituted α-d-glucuronic acid and at O-2 of the 3-substituted β-l-rhamnose, respectively.→4)-α-d-Glcp-(1→3)-α-d-GlcpA4Ac-(1→3)-α-l-Rhap-(1→4)-α-d-GlcpA-(1→3)-β-l-Rhap2Ac-(1→This unbranched structure is not common in EPSs produced by Lactobacilli. Moreover, the presence of acetyl groups in the structure is an unusual feature which has only been reported in L. sake 0-1 [Robijn et al. Carbohydr. Res., 1995, 276, 117-136].     

A novel exopolysaccharide from deep-sea bacterium Zunongwangia profunda SM-A87: low-cost fermentation,moisture retention,and antioxidant activities

Many marine microorganisms can secrete exopolysaccharides (EPSs) which have important applications in biotechnology. We have purified a novel EPS from deep-sea bacterium Zunongwangia profunda SM-A87, identified its glycosyl composition and linkage, and optimized its production to 8.9 g/l in previous studies. To reduce the fermentation cost, an economical fermentation medium containing 60.9 % whey, 10 g/l soybean meal, and 2.9 % NaCl was developed. The EPS yield of batch fermentation in this medium reached 12.1?±?0.3 g/l. Fed-batch fermentation was conducted and led to an EPS yield of 17.2?±?0.4 g/l, which represents the highest EPS yield ever reported for a marine bacterium. The EPS was extracted and it displayed good rheological properties, moisture-retention ability, and antioxidant activity. Particularly, its moisture-retention ability is superior to that of other marine bacterial EPSs reported to date. SM-A87 EPS also showed high antioxidant activity. These results suggest that SM-A87 EPS has promising potentials in biotechnology.     

Factors affecting current production in microbial fuel cells using different industrial wastewaters

This study evaluated how different types of industrial wastewaters (bakery, brewery, paper and dairy) affect the performance of identical microbial fuel cells (MFCs); and the microbial composition and electrochemistry of MFC anodes. MFCs fed with paper wastewater produced the highest current density (125 ± 2 mA/m²) at least five times higher than dairy (25 ± 1 mA/m²), brewery and bakery wastewaters (10 ± 1 mA/m²). Such high current production was independent of substrate degradability. A comprehensive study was conducted to determine the factor driving current production when using the paper effluent. The microbial composition of anodic biofilms differed according to the type of wastewater used, and only MFC anodes fed with paper wastewater showed redox activity at −134 ± 5 mV vs NHE. Electrochemical analysis of this redox activity indicated that anodic bacteria produced a putative electron shuttling compound that increased the electron transfer rate through diffusion, and as a result the overall MFC performance.     

Production of extracellular polysaccharide by Bacillus megaterium RB-05 using jute as substrate

Bacillus megaterium RB-05 was grown on glucose and on “tossa-daisee” (Corchorus olitorius)-derived jute, and production and composition of extracellular polysaccharide (EPS) were monitored. An EPS yield of 0.065 ± 0.013 and of 0.297 g ± 0.054 g⁻¹ substrate after 72 h was obtained for glucose and jute, respectively. EPS production in the presence of jute paralleled bacterial cellulase activity. High performance liquid chromatography (HPLC), matrix assisted LASER desorption/ionization-time of flight (MALDI-ToF) mass spectroscopy, and fourier transform infrared (FT-IR) spectroscopy demonstrated that the EPS synthesized in jute culture (JC) differed from that synthesized in glucose mineral salts medium (GMSM). While fucose was only a minor constituent (4.9 wt.%) of EPS from GMSM, it a major component (41.9 wt.%) of EPS synthesized in JC. This study establishes jute as an effective fermentation substrate for EPS production by a cellulase-producing bacterium.     

Linking microbial community structure to membrane biofouling associated with varying dissolved oxygen concentrations

In order to elucidate how dissolved oxygen (DO) concentration influenced the generation of extracellular polymeric substance (EPS) and soluble microbial products (SMP) in mixed liquor and biocake, 16S rDNA fingerprinting analyses were performed to investigate the variation of the microbial community in an aerobic membrane bioreactor (MBR). The function of microbial community structure was proved to be ultimately responsible for biofouling. Obvious microbial community succession from the subphylum of Betaproteobacteria to Deltaproteobacteria was observed in biocake. High concentration of EPS in biocake under the low DO concentration (0.5 mg L⁻¹) caused severe biofouling. The correlation coefficient of membrane fouling rate with EPS content in biocake (0.9941-0.9964) was much higher than that in mixed liquor (0.6689-0.8004).     

Genetics and engineering of microbial exopolysaccharides for food: approaches for the production of existing and novel polysaccharides.

Microbial exopolysaccharides (EPSs) are used in the food industry for their unique properties as viscosifiers, stabilisers, emulsifiers or gelling agents. In recent years, significant progress in the understanding of the genetics and biochemistry of microbial EPS synthesis by both Gram-negative and Gram-positive bacteria has been made. Biosynthesis pathways have been elucidated, and several of the genes involved have been characterised. This knowledge can now be applied to start EPS engineering or to improve EPS production.     

Small-scale analysis of exopolysaccharides from Streptococcus thermophilus grown in a semi-defined medium

Background  

Exopolysaccharides (EPSs) produced by lactic acid bacteria are important for the texture of fermented foods and have received a great deal of interest recently. However, the low production levels of EPSs in combination with the complex media used for growth of the bacteria have caused problems in the accurate analysis of the EPS. The purpose of this study was to find a growth medium for physiological studies of the lactic acid bacterium Streptococcus thermophilus, and to develop a simple method for qualitative and quantitative analysis of EPSs produced in this medium.     

Production and partial characterization of the exopolysaccharide from Pleurotus sajor caju

Microbial exopolysaccharides (EPSs) are very important because they are used in biotechnological applications in different industrial areas. The aim of the study was to determine the best EPS producer Pleurotus sp., to optimize EPS production and to perform partial purification and characterization of the produced EPS. After the production conditions were optimized, the EPS was isolated and partially purified. EPS was characterized by HP-TLC, 1H-NMR, FT-IR, and TGA. Hydroxyl, superoxide, and DPPH radical scavenging activities of the EPS were also investigated spectrophotometrically. The best EPS producer and its incubation period in submerged fermentation were determined as Pleurotus sajor caju and on 5 days, respectively. Culture conditions to increase EPS production were optimized as follows (in per liter): 90 g of glucose, 10 g of yeast extract, 10 g of peptone, and 100 mM of Mg2+. The optimal initial pH, temperature, and an agitation rate of culture were determined as 5.0, 25 °C, and 150 rate min−1, respectively. The highest EPS production was determined as 33.32 ± 1.6 g L−1. After isolation of EPS, one active fraction was obtained by gel filtration chromatography. EPS is composed mainly of glucose according to HP-TLC analysis. To the results, EPS had a complex structure by having carbohydrate and protein contents. The produced EPS had high degradation temperature as well as high antioxidant activity.     

ENVIRONMENTAL EFFECTS ON EXOPOLYMER PRODUCTION BY MARINE BENTHIC DIATOMS: DYNAMICS,CHANGES IN COMPOSITION,AND PATHWAYS OF PRODUCTION1

Marine benthic diatoms excrete large quantities of extracellular polymeric substances (EPS), both as a function of their motility system and as a response to environmental conditions. Diatom EPS consists predominantly of carbohydrate‐rich polymers and is important in the ecology of cells living on marine sediments. Production rates, production pathways, and monosaccharide composition of water‐soluble (colloidal) carbohydrates, EPS, and intracellular storage carbohydrate (glucans) were investigated in the epipelic (mud‐inhabiting) diatoms Cylindrotheca closterium (Ehrenburg), Navicula perminta (Grün.) in Van Heurck, and Amphora exigua Greg. under a range of experimental conditions simulating aspects of the natural environment. Cellular rates of colloidal carbohydrate, EPS, and glucan production were significantly higher during nutrient‐replete compared with nutrient‐limited growth for all three species. The proportion of EPS in the extracellular carbohydrate pool increased significantly (to 44%–69%) as cells became nutrient limited. Cylindrotheca closterium produced two types of EPS differing in sugar composition and production patterns. Nutrient‐replete cells produced a complex EPS containing rhamnose, fucose, xylose, mannose, galactose, glucose, and uronic acids. Nutrient‐limited cells produced an additional EPS containing mannose, galactose, glucose, and uronic acids. Both EPS types were produced under illuminated and darkened conditions. ¹⁴C‐labeling revealed immediate production of ¹⁴C‐glucan and significant increases in ¹⁴C‐EPS between 3 and 4 h after addition of label. The glucan synthesis inhibitor 2,6‐dichlorobenzonitrile significantly reduced ¹⁴C‐colloidal carbohydrate and ¹⁴C‐EPS. The glucanase inhibitor P‐nitrophenyl β‐d ‐glucopyranoside resulted in accumulation of glucan within cells and lowered rates of ¹⁴C‐colloidal and ¹⁴C‐EPS production. Cycloheximide prevented glucan catabolism, but glucan production and EPS synthesis were unaffected.     

Exopolysaccharides production in Lactobacillus bulgaricus and Lactobacillus casei exploiting microfiltration

The physiology of Lactobacillus delbrueckii ssp. bulgaricus and Lactobacillus casei, extensively used in the dairy industry, was studied in order to evaluate key parameters in the synthesis of exopolysaccharides and to improve their production through novel fermentation processes. Selected strains were studied in shake flasks and in fermentor experiments using glucose and lactose as main carbon sources and bacto casitone as the only complex component, in a temperature range between 35 and 42°C. The production of exopolysaccharides was monitored and correlated to the growth conditions using both a colorimetric assay and chromatographic methods. Fermentor experiments in batch mode yielded 100 mg l⁻¹ of EPS from L. bulgaricus and 350 mg l⁻¹ from L. casei. Moreover, the use of a microfiltration (MF) bioreactor resulted in exopolysaccharides (EPS) concentrations threefold and sixfold those of batch experiments, respectively. The monosaccharidic composition of the two analyzed polymers differed from those previously reported. The optimization of the production of EPSs using the MF fermentation strategy could permit the use of these molecules produced by generally recognised as safe (GRAS) microorganisms in the place of other polysaccharides in the food industry.     

Bacterial Exopolysaccharides from Extreme Marine Environments with Special Consideration of the Southern Ocean, Sea Ice, and Deep-Sea Hydrothermal Vents: A Review

Exopolysaccharides (EPSs) are high molecular weight carbohydrate polymers that make up a substantial component of the extracellular polymers surrounding most microbial cells in the marine environment. EPSs constitute a large fraction of the reduced carbon reservoir in the ocean and enhance the survival of marine bacteria by influencing the physicochemical environment around the bacterial cell. Microbial EPSs are abundant in the Antarctic marine environment, for example, in sea ice and ocean particles, where they may assist microbial communities to endure extremes of temperature, salinity, and nutrient availability. The microbial biodiversity of Antarctic ecosystems is relatively unexplored. Deep-sea hydrothermal vent environments are characterized by high pressure, extreme temperature, and heavy metals. The commercial value of microbial EPSs from these habitats has been established recently. Extreme environments offer novel microbial biodiversity that produces varied and promising EPSs. The biotechnological potential of these biopolymers from hydrothermal vent environments as well as from Antarctic marine ecosystems remains largely untapped.     

Composition of extracellular polymeric substances influences the autoaggregation capability of hydrogen-producing bacterium Ethanoligenens harbinense

Non-spore-forming Ethanoligenens, a novel genus of hydrogen-producing bacteria, is endowed with great application potential in biohydrogen production due to acidophilic and autoaggregating growth. In order to elucidate the mechanism of autoaggregation of Ethanoligenens harbinense, extracellular polymeric substances (EPS) from YUAN-3 had been extracted and analyzed. The EPS was mainly produced during the exponential phase and with protein, carbohydrate and DNA as its main components, with yields of 21.0 ± 0.8 mg/g-cell dry weight (CDW), 16.9 ± 0.8 mg/g-CDW and 3.5 ± 0.5 mg/g-CDW, respectively. Compared with the EPS composition of semi-autoaggregating hydrogen-producing bacteria W1 and non-autoaggregating hydrogen-producing bacteria B49, carbohydrate and protein played an important part in the autoaggregation of YUAN-3.     

Microbial community developments and biomass characteristics in membrane bioreactors under different organic loadings

Microbial community developments and biomass characteristics (concentration, particle size, extracellular polymeric substances (EPS), and membrane fouling propensity) were compared when three MBRs were fed with the synthetic wastewater at different organic loadings. Results showed that the bacterial communities dynamically shifted in different ways and the EPS displayed dissimilar profiles under various organic loadings, which were associated with the ratios of food to microorganism and dissolved oxygen levels in the MBRs. The membrane fouling tendency of biomass in the low-loading MBR (0.57 g COD/L day) was insignificantly different from that in the medium-loading MBR (1.14 g COD/L day), which was apparently lower than that in the high-loading MBR (2.28 g COD/L day). The membrane fouling propensity of biomass was strongly correlated with their bound EPS contents, indicating cake layer fouling (i.e., deposition of microbial flocs) was predominant in membrane fouling at a high flux of 30 L/m² h.     

Microbial behaviors involved in cake fouling in membrane bioreactors under different solids retention times

Biomass characteristics and membrane performances in the MBRs operated at a high flux of 30 L/m² h under different SRTs (10, 30 days, and infinity) were monitored. Results showed that more serious cake-fouling happened in the SRT-infinity MBR, which correlated with the activated sludge characteristics such as smaller floc size and greater EPS amount. DGGE analysis indicated that the microbial community shifted in different ways under various SRTs, which also influenced EPS productions in the MBRs. Different microbial communities were developed on the membrane surfaces at various operating stages and SRTs. Possibly, the activated sludge characteristics (such as MLSS concentration, EPS properties) and hydrodynamic conditions influenced by the SRTs were associated with cake layer development and membrane fouling propensity. Insight into the EPS characteristics and deposition behaviors of bacterial flocs will be crucial to explore appropriate biofouling control strategies in MBRs.     

Immunomodulation and antitumor activities of different-molecular-weight polysaccharides from Porphyridium cruentum

The extracellular polysaccharides (EPSs) isolated from Porphyridium cruentum were degraded by hermetical-microwave and H₂O₂ under ultrasonic waves. Six products were obtained with molecular weights of 6.53, 256, 606, 802.6, 903.3 and 1002 kDa. The antitumor and immunomodulatory activities of different-molecular-weight (MW) polysaccharides were evaluated by the S180-tumor-bearing mouse model in vivo and peritoneal macrophage activation in vitro. The degraded EPSs all showed clear immunomodulation to different extents. The MW of the EPSs had a notable effect on their activity. The 6.53-kDa fragment had the strongest immunoenhancing activity. Different doses of EPS all inhibited the growth of the implanted S180 tumor. The tumor inhibition index at high, middle and low doses was 53.3%, 47.5% and 40.5%, respectively. In addition, three different concentrations of EPS significantly increased lymphocyte proliferation, which indicated the unique mechanism of the antitumor effect of EPS.     

Arabinose content of extracellular polysaccharide plays a role in cell aggregation of Azospirillum brasilense

Extracellular polysaccharides play an important role in aggregation and surface colonization of plant-associated bacteria. In this work, we report the time course production and monomer composition of the exopolysaccharide (EPS) produced by wild type strain and several mutants of the plant growth promoting rhizobacterium (PGPR) Azospirillum brasilense. In a fructose synthetic medium, wild type strain Sp7 produced a glucose-rich EPS during exponential phase growth and an arabinose-rich EPS during stationary and death phase growth. D-glucose or L-arabinose did not support cell growth as sole carbon sources. However, glucose and arabinose-rich EPSs, when used as carbon source, supported bacterial growth. Cell aggregation of Sp7 correlated with the synthesis of arabinose-rich EPS. exoB (UDP-glucose 4'-epimerase), exoC (phosphomannomutase) and phbC (poly-beta-hydroxyburyrate synthase) mutant strains, under tested conditions, produced arabinose-rich EPS and exhibited highly cell aggregation capability. A mutant defective in LPS production (dTDP 4-rhamnose reductase; rmlD) produced glucose-rich EPS and did not aggregate. These results support that arabinose content of EPS plays an important role in cell aggregation. Cell aggregation appears to be a time course phenomenon that takes place during reduced metabolic cell activity. Thus, aggregation could constitute a protected model of growth that allows survival in a hostile environment. The occurrence of exoC and rmlD was detected in several species of Azospirillum.