An extracellular polysaccharide produced by Zoogloea ramigera 115

A weakly acidic polysaccharide was purified from the extracellular zoogloeal matrix produced by Zoogloeal ramigera 115. The purified polysaccharide was homogeneous as judged by sedimentation analysis, and the average molecular weight was estimated to be about 10(5) by gel permeation chromatography of the fully methylated preparation. The polysaccharide was composed of D-glucose, D-galactose and pyruvic acid in an approximate molar ratio 11:3:1.5. On the basis of methylation, periodate oxidation, Smith degradation and partial hydrolysis, the following highly branched structure was deduced for the polysaccharide: a long chain mainly consisting of beta 1 leads to 4-linked glucose residues branching at the C-3 or C-6 position of galactose residues which are present in beta 1 leads to 4 or beta 1 leads to 3 linkages as the minor component of the long chain; pyruvic acid residues, the sole acidic component, are linked to the nonreducing end and/or 1,3-linked glucose residues through 4,6-ketal linkages. The purified polysaccharide was not readily soluble in water and had a high affinity for several metallic ions (e.g, 0.25 mumol Fe3+/mg, and 0.17 mumol Fe2+ mg). Upon addition of metallic ions (1 mM) to a gelatinous aqueous solution of the polysaccharide (K+ form, 0.125%), more than 80% of it immediately coprecipitated out with them.     

Ester Synthesis by Zoogloea ramigera 115 Grown in the Presence of Ethanol

The ester ethyl butyrate is produced by Zoogloea ramigera 115, a bacterium isolated from an aerobic waste treatment plant, when ethanol is present in culture media. The cells appear to produce butyric acid which is then esterified with residual ethanol in the culture medium.     

Isolation of Zoogloea ramigera I-16-M exopolysaccharide biosynthetic genes and evidence for instability within this region.

The genetics of the biosynthesis of an exocellular polysaccharide (EPS) from Zoogloea ramigera I-16-M is being investigated. Tn5 insertion mutants deficient in EPS production were isolated by screening for the absence of fluorescence on plates containing the dye Cellufluor (Polysciences Chemicals). Complementation of these mutations was achieved with a Z. ramigera I-16-M gene library constructed in a broad-host-range cosmid vector and introduced into the I-16-M mutants by conjugation. Four recombinant plasmids able to restore EPS production to all of these mutants were found to contain at least 14 kilobases of common insert DNA. Subcloning of the common region and restriction mapping the locations of Tn5 insertions have identified two complementation groups contained within a chromosomal segment of DNA that is between 4.6 and 6.5 kilobases in size. We have clearly demonstrated genetic instability in this region which leads to spontaneous deletions and possibly rearrangements resulting in the loss of EPS production.     

Isolation, Identification, and Metabolic Role of the Sudanophilic Granules of Zoogloea ramigera

Organisms isolated from activated sludge and identified as Zoogloea ramigera accumulated large amounts of sudanophilic granules as the cultures flocculated. The granules were extracted by chloroform and precipitated with ether from acid-hydrolyzed cells. Identification of the sudanophilic granules as poly-β-hydroxybutyric acid (PHB) was confirmed by physical, chemical, and infrared spectral analyses. The isolated polymer accounted for 12.0 to 50.5% of the dry weight of the cells. The polymer was not synthesized when the culture was grown in a growth-limiting concentration of organic substrate; it did accumulate when the culture was grown in medium enriched with carbon and energy sources. An increase in concentration of intracellular PHB was directly proportional to optical density and uptake of glucose. Aside from intracellular storage of PHB as endogenous metabolite, the accumulation of PHB is noted as a possible mechanism of flocculation.     

Determination of the biosorption heats of heavy metal ions on Zoogloea ramigera and Rhizopus arrhizus

The biosorption of Fe(III), Cr(VI), Pb(II), Cu(II) and Ni(II) ions on Zoogloea ramigera (activated sludge bacterium) and Rhizopus arrhizus (filamentous fungus) has been studied as a function of initial metal ion concentration and temperature. The applicability of the Langmuir model for each metal-microorganism system has been tested at different temperatures. The enthalpy change for the biosorption process has been evaluated by using the Langmuir constant b, related to the energy of adsorption. Thermodynamic parameters indicate the exothermic nature of Cu(II) and Ni(II) biosorption on both microorganisms. Fe(III), Cr(VI) and Pb(II) biosorption is determined to be an endothermic process since increased binding occurs as the temperature is increased in the range 15-45 degrees C.     

Influence of pH and Metal Cations on Aggregative Growth of Non-Slime-forming Strains of Zoogloea ramigera

Aggregative growth of non-slime-forming strains of Zoogloea ramigera was induced by growing the organisms at a depressed pH. Calcium and magnesium ion was found to reverse aggregative growth of the organisms. Conversely, aggregation was stimulated when the available inorganic cation concentration of the growth medium was lowered by the use of a chelating agent. The aggregative effects of pH depression or cation depletion and the dispersal effects of cation supplementation were observed only during cellular growth. The data suggest that aggregate formation of non-slime-forming strains of Z. ramigera may be related to the calcium or magnesium metabolism of the organisms, or both.     

Effects of a sulfated exopolysaccharide produced by Altermonas infernus on bone biology

The growth and differentiation of bone cells is controlled by various factors, which can be modulated by heparan sulfates. Here, we investigated the effects of an oversulfated exopolysaccharide (OS-EPS) on the bone. We compared the effect of this compound with that of a native EPS. Long-term administration of OS-EPS causes cancellous bone loss in mice due, in part, to an increase in the number of osteoclasts lining the trabecular bone surface. No significant difference in cancellous bone volume was found between EPS-treated mice and age-matched control mice, underlying the importance of sulfation in trabecular bone loss. However, the mechanism sustaining this osteoporosis was unclear. To clarify OS-EPS activities, we investigated the effect of OS-EPS on osteogenesis. Our results demonstrated that OS-EPS inhibited osteoclastogenesis in two cell models. Using the surface plasmon resonance technique, we revealed that OS-EPS can form a hetero-molecular complex OS-EPS/receptor activator of NF-κB ligand (RANKL)/RANK and that RANK had a higher affinity for RANKL pre-incubated with OS-EPS than for RANKL alone, which would be in favor of an increase in bone resorption. However, in vitro, OS-EPS inhibited the early steps of osteoclast precursor adhesion and therefore inhibited the cell fusion step. In addition, we showed that OS-EPS reduced proliferation and accelerated osteoblastic differentiation, leading to strong inhibition of mineralized nodule formation, which would be in favor of an increase in bone resorption. Taken together, these data show different levels of bone resorption regulation by EPSs, most of them leading to proresorptive effects.     

Structure of the exopolysaccharide produced by Enterobacter amnigenus

The bacterial species Enterobacter amnigenus was isolated from sugar beets harvested in Finland. It produced an exopolysaccharide rich in l-fucose, which gave viscous water solutions. Its primary structure was determined mainly by NMR spectroscopy and ESIMS of oligosaccharides and a polysaccharide with decreased molecular weight, obtained by Smith degradation of the O-deacetylated native polymer [carbohydrate structure: see text]     

Pectin lyase production by Penicillium griseoroseum: Effect of tea extract, caffeine, yeast extract, and pectin

Summary Mycelial growth and production of extracellular pectin lyase by Penicillium griseoroseum at different concentrations of inducers were investigated. The fungus was cultured in mineral medium using sucrose as a carbon source and caffeine, yeast extract, tea extract or pectin as inducers. Caffeine, yeast extract and tea extract in the presence of sucrose, and tea extract alone were capable of inducing pectin lyase in P. griseoroseum, even at low concentrations.     

Effects of the oral administration of the exopolysaccharide produced by Lactobacillus kefiranofaciens on the gut mucosal immunity

The probiotic effects ascribed to lactic acid bacteria (LAB) and their fermented dairy products arise not only from whole microorganisms and cell wall components but also from peptides and extracellular polysaccharides (exopolysaccharides) produced during the fermentation of milk. There is a lack of knowledge concerning the immune mechanisms induced by exopolysaccharides produced by lactic acid bacteria, which would allow a better understanding of the functional effects described to them. The aim of this study was to investigate the in vivo immunomodulating capacity of the exopolysaccharide produced by Lactobacillus kefiranofaciens by analyzing the profile of cytokines and immunoglobulins induced at the intestinal mucosa level, in the intestinal fluid and blood serum. BALB/c mice received the exopolysaccharide produced by L. kefiranofaciens for 2, 5 or 7 consecutive days. At the end of each period of administration, control and treated mice were sacrificed and the numbers of IgA+ and IgG+ cells were determined on histological slices of the small and large intestine by immunofluorescence. Cytokines (IL-4, IL-6, IL-10, IL-12, IFNγ and TNFα) were also determined in the gut lamina propria as well as in the intestinal fluid and blood serum. There was an increase of IgA+ cells in the small and large intestine lamina propria, without change in the number of IgG+ cells in the small intestine. This study reports the effects of the oral administration of the exopolysaccharide produced by L. kefiranofaciens in the number of IgA+ cells in the small and large intestine, comparing simultaneously the production of cytokines by cells of the lamina propria and in the intestinal fluid and blood serum. The increase in the number of IgA+ cells was not simultaneously accompanied by an enhance of the number of IL-4+ cells in the small intestine. This finding would be in accordance with the fact that, in general, polysaccharide antigens elicit a T-independent immune response. For IL-10+, IL-6+ and IL-12+ cells, the values found were slightly increased compared to control values, while IFNγ+ and TNFα+ cells did not change compared to control values. The effects observed on immunoglobulins and in all the cytokines assayed in the large intestine after kefiran administration were of greater magnitude than the ones observed in the small intestine lamina propria, which may be due to the saccharolytic action of the colonic microflora. In the intestinal fluid, only IL-4 and IL-12 increased compared to control values. In blood serum, all the cytokines assayed followed a pattern of production quite similar to the one found for them in the small intestine lamina propria. We observed that the exopolysaccharide induced a gut mucosal response and it was able to up and down regulate it for protective immunity, maintaining intestinal homeostasis, enhancing the IgA production at both the small and large intestine level and influencing the systemic immunity through the cytokines released to the circulating blood.     

Isolation and characterization of the exopolysaccharide produced by Daedalea quercina

The production of exopolysaccharide (EPS) by a strain of the basidiomycete Daedalea quercina was investigated. Of seven different carbon sources, glucose and dextrins gave the highest crude polysaccharide yield (4.7–5 g l^–1, 55–60% carbohydrate content) in shake-flask cultures, at 14 days of fermentation. Experiments carried out in a 10 l fermenter, at two different agitation speeds, gave the best results at 300 rpm, resulting in 12–14 g l^–1 of crude exopolysaccharide in 9–11 days. Fractionation of the EPS samples, carried out by tangential flow ultrafiltration, evidenced a single EPS fraction (MW >30 000 Da) in samples from glucose, while two fractions (MW > 30 000 Da and 30 000 > MW > 10 000 Da) were present in samples from dextrins. Fractions characterization by HPLC and proton NMR spectroscopy revealed diversity in composition and structure in the obtained EPS: from glucose mainly an -linked mannan, and from dextrins mainly an - and -linked glucan.     

Effects of the oral administration of the exopolysaccharide produced by Lactobacillus kefiranofaciens on the gut mucosal immunity

The probiotic effects ascribed to lactic acid bacteria (LAB) and their fermented dairy products arise not only from whole microorganisms and cell wall components but also from peptides and extracellular polysaccharides (exopolysaccharides) produced during the fermentation of milk. There is a lack of knowledge concerning the immune mechanisms induced by exopolysaccharides produced by lactic acid bacteria, which would allow a better understanding of the functional effects described to them. The aim of this study was to investigate the in vivo immunomodulating capacity of the exopolysaccharide produced by Lactobacillus kefiranofaciens by analyzing the profile of cytokines and immunoglobulins induced at the intestinal mucosa level, in the intestinal fluid and blood serum. BALB/c mice received the exopolysaccharide produced by L. kefiranofaciens for 2, 5 or 7 consecutive days. At the end of each period of administration, control and treated mice were sacrificed and the numbers of IgA+ and IgG+ cells were determined on histological slices of the small and large intestine by immunofluorescence. Cytokines (IL-4, IL-6, IL-10, IL-12, IFNgamma and TNFalpha) were also determined in the gut lamina propria as well as in the intestinal fluid and blood serum. There was an increase of IgA+ cells in the small and large intestine lamina propria, without change in the number of IgG+ cells in the small intestine. This study reports the effects of the oral administration of the exopolysaccharide produced by L. kefiranofaciens in the number of IgA+ cells in the small and large intestine, comparing simultaneously the production of cytokines by cells of the lamina propria and in the intestinal fluid and blood serum. The increase in the number of IgA+ cells was not simultaneously accompanied by an enhance of the number of IL-4+ cells in the small intestine. This finding would be in accordance with the fact that, in general, polysaccharide antigens elicit a T-independent immune response. For IL-10+, IL-6+ and IL-12+ cells, the values found were slightly increased compared to control values, while IFNgamma+ and TNFalpha+ cells did not change compared to control values. The effects observed on immunoglobulins and in all the cytokines assayed in the large intestine after kefiran administration were of greater magnitude than the ones observed in the small intestine lamina propria, which may be due to the saccharolytic action of the colonic microflora. In the intestinal fluid, only IL-4 and IL-12 increased compared to control values. In blood serum, all the cytokines assayed followed a pattern of production quite similar to the one found for them in the small intestine lamina propria. We observed that the exopolysaccharide induced a gut mucosal response and it was able to up and down regulate it for protective immunity, maintaining intestinal homeostasis, enhancing the IgA production at both the small and large intestine level and influencing the systemic immunity through the cytokines released to the circulating blood.     

Effects of growth conditions on exopolysaccharide production by Cyanospira capsulata

Batch cultures of Cyanospira capsulata, a heterocystous cyanobacterium possessing a thick polysaccharidic capsule, were characterized by increasing viscosity owing to the continuous release of a soluble polysaccharide (EPS) into the culture medium. Both capsulated trichomes and solubilized EPS contributed to the flow properties of whole cultures. A typical pseudoplastic behaviour, the more marked the more aged were the cultures, was evidenced.The production of EPS was investigated under different growth conditions by changing some nutritional and physical parameters known to affect the synthesis of exopolysaccharides in algae and cyanobacteria. Among the factors tested (Ca²⁺, Mg²⁺ or PO₄⁻⁴ deficiencies, salinity and pH) only Mg²⁺ shortage caused a significant enhancement of the EPS production. Under continuous illumination in open ponds, the EPS productivity of batch cultures on standard mineral medium was about 5·8 g m⁻² day⁻¹, whereas under Mg²⁺ deficiency with a consequent increase of the cultures' viscosity     

营养因子对干酪乳杆菌LC2W胞外多糖合成的影响

针对影响干酪乳杆菌LC2W产胞外多糖的主要营养因子:葡萄糖、酪蛋白胨和酵母提取物进行了单因素及正交试验,确定了合成胞外多糖的最适组成为葡萄糖60 g/L,酪蛋白胨16 g/L ,酵母提取物10 g/L,其中酪蛋白胨影响差异极显著.验证试验表明,以优化后的培养基进行发酵试验,37 ℃培养24 h,所得胞外多糖量为120.37 mg/L,较原来提高67.97 %.     

Influence of the concentrations of d-xylose and yeast extract on ethanol production by Pachysolen tannophilus

To determine the most favorable conditions for the production of ethanol by Pachysolen tannophilus, this yeast was grown in batch cultures with various initial concentrations of two of the constituents of the culture medium: d-xylose (s_o), ranging from 1 g·l⁻¹ to 200 g·l⁻¹, and yeast extract (l_o), ranging from 0 g·l⁻¹ to 8 g·l⁻¹. The most favorable conditions proved to be initial concentrations of S_o=25 g·l⁻¹ and l_o=4 g·l⁻¹, which gave a maximum specific growth rate of 0.26 h⁻¹, biomass productivity of 0.023 g·l⁻¹·h⁻¹, overall biomass yield of 0.094 g·g⁻¹, specific xylose-uptake rate (q_s) of 0.3 g·g⁻¹·h⁻¹ (for t=50 h), specific ethanol-production rate (q_E) of 0.065 g·g⁻¹·h⁻¹ and overall ethanol yield of 0.34 g·g⁻¹; q_s values decreased after the exponential growth phase while q_E remained practically constant.     

Submerged culture process for biomass and exopolysaccharide production by Antarctic yeast: some engineering considerations

Production of biomass and extracellular polysaccharide (EPS) from psychrophilic Sporobolomyces salmonicolor AL₁ in a stirred bioreactor was studied. The aspects of production technical-scale parameters, namely, bioreactor flow field, biomass and EPS production rates, oxygen mass transfer per input power, as well as important product properties, such as rheology and stability of EPS mixtures, were considered. The bioprocess was found to proceed in non-Newtonian flow with consistency coefficient rising typically to 0.03 Pa.sⁿ and flow index declining to 0.7. Flow modeling was carried out and showed good homogenization for substrate delivery at agitation rates exceeding 400 rpm. Agitation rates lower than 400 rpm were considered counterproductive due to flow field non-uniformity. The cell density reached 5 g/l and EPS production yield reached 5.5 g/l at production rate 0.057 g EPS/l per hour (0.01 g EPS/g biomass per hour). Oxygen uptake rate and oxygen transfer rate were in the range of 0.5–1.7 mmolO₂/l per hour and 2–4.7 mmolO₂/l per hour, respectively. The mass transfer coefficient at reaction conditions was found to be in the range $ {K_L}a\tilde{\mkern6mu} 0.004-0.01{{\mathrm{s}}^{-1 }} $ . The bioprocess biological performance was higher at moderate agitation speed and revealed biomass diminution and cell inactivation by increasing impeller revolutions and shear rate. The product EPS was found to introduce shear-thinning behavior in water solutions with apparent viscosity of up to 30 mPa.s and to stabilize 1–2 % oil-in-water emulsions improving their lipophilic properties. The emulsion dispersion index was found to be comparable with the one of Arlacel 165, the emulsifier used in cosmetic. The long-term performance of the complex cream mixtures of the glucomannan prepared in commercial format was found promising for further application.