Culture conditions for the production of an acidic exopolysaccharide by the nitrogen-fixing bacterium Burkholderia tropica

2006 ◽  
Vol 52 (5) ◽  
pp. 489-493 ◽  
Author(s):  
Rodrigo V Serrato ◽  
Guilherme L Sassaki ◽  
Leonardo M Cruz ◽  
Fábio O Pedrosa ◽  
Philip A.J Gorin ◽  
...  
Keyword(s):  
High Performance ◽  
C:N Ratio ◽  
Great Influence ◽  
Nitrogen Sources ◽  
Monosaccharide Composition ◽  
Culture Conditions ◽  
Size Exclusion ◽  
Molar Ratio ◽  
Isolation And Purification ◽  
Synthetic Media

The endophytic diazotrophic bacterium Burkholderia tropica, strain Ppe8, produced copious amounts of exopolysaccharide (EPS) on batch growth in liquid synthetic media containing mannitol and glutamate as carbon and nitrogen sources. The effect of various aeration regimes and carbon source concentrations on EPS production was determined, as well as the effects of temperature and time of growth. The degree of aeration had a great influence on the yield of EPS, in contrast with the C:N ratio of the medium. Growth temperature also affected the EPS yield after the first 24 h of culture but seemed to be irrelevant after that. After isolation and purification, the EPS was analyzed by high-performance size exclusion chromatography and multiangle laser light scattering (HPSEC–MALLS), revealing a molecular mass of 300 kDa. The acid hydrolyzate of EPS was examined by HPLC and found to contain Glc, Rha, GlcA, and an aldobiouronic acid. The latter was found to have a GlcA and a Rha unit. Carboxy-reduced EPS contained Glc and Rha (3:2). The monosaccharide composition of the native acidic EPS was calculated as GlcA, Glc, and Rha in a molar ratio of 1:2:2.Key words: Burkholderia, endophyte, diazotrophic, exopolysaccharide, EPS.

scholarly journals Physicochemical Characterization and Immunomodulatory Activity of a Novel Acid Polysaccharide from Solanum muricatum

Polymers ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 1972 ◽  
Author(s):  
Heng Yue ◽  
Qianqian Xu ◽  
Xianheng Li ◽  
Jeevithan Elango ◽  
Wenhui Wu ◽  
...  
Keyword(s):  
High Performance ◽  
Average Molecular Weight ◽  
Thermal Studies ◽  
Physicochemical Characterization ◽  
Monosaccharide Composition ◽  
Molar Ratio ◽  
Immunomodulatory Activity ◽  
Composition Analysis ◽  
Solanum Muricatum ◽  
Acid Polysaccharide

To investigate the structure and immunomodulatory activity of polysaccharide from Solanum muricatum, a novel acid polysaccharide named SMP-3a was purified from Solanum muricatum pulp through DEAE-52 cellulose column and Sephadex G-200 chromatography. Monosaccharide composition analysis showed that SMP-3a was mainly composed of rhamnose, arabinose, galactose, and galacturonic acid with the molar ratio of 1.09:2.64:1.54:1. The average molecular weight was found to be 227 kDa by high performance gel permeation chromatography (HPGPC). Thermal studies revealed the SMP-3a was a thermally stable polymer. Based on the results of methylation and NMR analysis, the backbone chain of SMP-3a was composed of →2)-α-l-Rhap-(1→, →4)-α-d-GalpA-(1→ and →4)-α-d-Galp-(1→. The side chain was consisted of α-l-Araf-(1→ and →5)-α-l-Araf-(1→. Immunomodulatory assay indicated that SMP-3a could significantly promote the proliferation of macrophages and stimulate the secretion of cytokines, including TNF-α, IL-1β, and IL-6. Our results suggested that SMP-3a could be used as a novel potential immunomodulatory agent in functional food.

Production and Rheological Properties of Welan Gum Produced by Sphingomonas sp. ATCC 31555 with Different Nitrogen Sources

2017 ◽  
Vol 27 (1) ◽  
pp. 55-63 ◽  
Author(s):  
Xiaopeng Xu ◽  
Zuoming Nie ◽  
Zhiyong Zheng ◽  
Li Zhu ◽  
Xiaobei Zhan
Keyword(s):  
Molecular Weight ◽  
Rheological Properties ◽  
Biomass Concentration ◽  
Bacterial Biomass ◽  
Nitrogen Sources ◽  
Monosaccharide Composition ◽  
Solution Concentration ◽  
Molar Ratio ◽  
Welan Gum ◽  
Beef Extract

This study aimed to investigate the effect of nitrogen sources on the production and rheological properties of welan gum produced by <i>Sphingomonas</i> sp. ATCC 31555. Six different nitrogen sources were used for ATCC 31555 fermentation, and 2 of these were further analyzed due to their more positive influence on welan gum production and bacterial biomass. Bacterial biomass, welan gum yield, welan viscosity, molecular weight, monosaccharide composition, acyl content, and welan structure were analyzed. Welan gum production and the biomass concentration of ATCC 31555 were higher in media containing NaNO<sub>3</sub> and beef extract. Welan viscosity decreased at higher temperatures of 30-90°C, and it increased with a higher welan concentration. In the media containing NaNO<sub>3</sub> (3 g·L<sup>-1</sup>), welan viscosity was higher at 30-70°C and a welan solution concentration of 6-10 g·L<sup>-1</sup>. With a reduced NaNO<sub>3</sub> concentration, the molecular weight of welan gum and the molar ratio of mannose decreased, but the molar ratio of glucuronic acid increased. With different nitrogen sources, the acetyl content of welan gum differed but its structure was similar. NaNO<sub>3</sub> and beef extract facilitated welan production. A reduced NaNO<sub>3</sub> concentration promoted welan viscosity.

scholarly journals A Sensitive and Rapid UPLC-MS/MS Method for Determination of Monosaccharides and Anti-Allergic Effect of the Polysaccharides Extracted from Saposhnikoviae Radix

Molecules ◽  
2018 ◽  
Vol 23 (8) ◽  
pp. 1924 ◽  
Author(s):  
Yan-Yan Gao ◽  
Yue Jiang ◽  
Guo-Chao Chen ◽  
Shuang-Shuang Li ◽  
Fei Yang ◽  
...  
Keyword(s):  
High Performance ◽  
Monosaccharide Composition ◽  
Water Soluble ◽  
Delayed Type Hypersensitivity ◽  
Molar Ratio ◽  
Serum Ige ◽  
Organ Index ◽  
Wide Range ◽  
Liquid Chromatography Tandem Mass ◽  
Saposhnikoviae Radix

Background: Allergic disease is a common clinical disease. Natural products provide an important source for a wide range of potential anti-allergic agents. This study was designed to evaluate the anti-allergic activities of the water-soluble polysaccharides extracted and purified from Saposhnikoviae Radix (SRPS). The composition and content of monosaccharides were determined to provide a material basis. Methods: An ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was established to determine the composition and content of SRPS. 2,4-dinitrofluorobenzene (DNFB) induced a delayed-type hypersensitivity (DTH) mouse model orally administrated SRPS for seven consecutive days. Ear swelling, organ index, and serum IgE levels were observed to evaluate the anti-allergic activities. Results: The UPLC-MS/MS analysis showed that SRPS was consisted of eight monosaccharides including galacturonic acid, mannose, glucose, galactose, rhamnose, fucose, ribose, and arabinose with a relative molar ratio of 4.42%, 7.86%, 23.69%, 12.06%, 3.10%, 0.45%, 0.71%, and 47.70%, respectively. SRPS could effectively reduce ear swelling, a thymus index, and a serum IgE levels. Conclusions: The method was simple, rapid, sensitive, and reproducible, which could be used to analyze and determine the monosaccharide composition of SRPS. The vivo experiments demonstrated that SRPS may effectively inhibit development of DNFB-induced DTH. SRPS is a novel potential resource for natural anti-allergic drugs.

scholarly journals Homologous Over-Expression of Chain Length Determination Protein EpsC Increases the Molecular Weight of Exopolysaccharide in Streptococcus thermophilus 05-34

2021 ◽  
Vol 12 ◽  
Author(s):  
Zhengyuan Zhai ◽  
Shuxin Xie ◽  
Hongxing Zhang ◽  
Huaxi Yi ◽  
Yanling Hao
Keyword(s):  
Molecular Weight ◽  
Chain Length ◽  
High Performance ◽  
Gel Permeation Chromatography ◽  
Streptococcus Thermophilus ◽  
Monosaccharide Composition ◽  
Molar Ratio ◽  
Gas Chromatography Mass Spectrometry ◽  
Over Expression ◽  
Length Determination

In Streptococcus thermophilus, EpsC is a polysaccharide co-polymerase which is involved in determining the chain length of EPS synthesized by the Wzx/Wzy-dependent pathway. Our previous study found that there was a positive correlation between transcription level of epsC and molecular weight of EPS in S. thermophilus 05-34. To further investigate the effects of EpsC on EPS biosynthesis, this gene was over-expressed in S. thermophilus 05-34 in this study. Reverse transcription qPCR and Western blotting confirmed the successful transcription and translation of epsC in 05-34, respectively. The yield of EPS was not affected by the over-expression of EpsC. Gas chromatography-mass spectrometry (GC-MS) showed that the monosaccharide composition was still composed of galactose and glucose in a molar ratio of 1.0:0.8, whereas high performance gel permeation chromatography (HPGPC) indicated that the molecular weight of EPS was increased from 4.62 × 105 Da to 9.17 × 105 Da by the over-expression of EpsC. In addition, S. thermophilus 05epsC which could produce higher molecular weight EPS improved the viscoelasticity and water-holding capacity of yogurt, but significantly reduced the level of syneresis in yogurt. In summary, these results indicated that homologous over-expression of EpsC in S. thermophilus could increase the molecular weight of EPS and improve the microrheological or physical properties of yogurt.

Purification and Structure Study on Exopolysaccharides Produced by Lactobacillus paracasei KL1-Liu from Tibetan Kefir

2013 ◽  
Vol 781-784 ◽  
pp. 1513-1518 ◽  
Author(s):  
Hui Liu ◽  
Yuan Hong Xie ◽  
Tao Han ◽  
Hong Xing Zhang
Keyword(s):  
Molecular Weight ◽  
High Performance ◽  
Monosaccharide Composition ◽  
Gradient Elution ◽  
Structure Study ◽  
Lactobacillus Paracasei ◽  
Molar Ratio ◽  
Test Results ◽  
Sample Concentration ◽  
Multi Level

To investigate the pure conditions of exopolysaccharides produced by Lactobacillus paracasei KL1-Liu from Tibetan Kefir, and to analyse the structure, we used Multi-level single-factor test to purify EPS by Sepharose CL-6B. And the purity of EPS was detected by UV scan and high performance liquid chromatogram (HPLC). EPS molecular weight and monosaccharide composition were determined by HPLC. Results: Adoption phosphate buffer gradient elution 0.02-0.10 mol/L, the velocity 0.25 mL/min, the sample concentration 1.0 mg/mL, the sample capacity 1.0 mL. Under this purification conditions, components EPSa and EPSb were obtained. The purities of EPSa and EPSb were 82.82% and 91.74% respectively, which were 1.4 and 1.5 times of the pre-purification. Purity Test results showed that EPSa and EPSb polysaccharide were single components, basically no nucleic acid and protein in them. Structural analysis revealed that the molecular weight of EPSa and EPSb were 4.60×104 Da and 2.12×104 Da detected by HPLC. EPSa monosaccharide components were glucose and rhamnose, and the molar ratio was 1:0.68. EPSb were composed of glucose, xylose and rhamnose, and the molar ratio was 1:0.77:0.69.

Application of a fractionation technique for better understanding of the removal of natural organic matter by alum coagulation

2002 ◽  
Vol 2 (5-6) ◽  
pp. 427-433 ◽  
Author(s):  
J. van Leeuwen ◽  
C. Chow ◽  
R. Fabris ◽  
N. Withers ◽  
D. Page ◽  
...  
Keyword(s):  
Organic Matter ◽  
Natural Organic Matter ◽  
High Performance ◽  
Relative Proportion ◽  
Water Source ◽  
Size Exclusion ◽  
Gas Chromatography Mass Spectrometry ◽  
Pyrolysis Gas ◽  
Alum Treatment ◽  
Hydrophobic Acids

To gain an improved understanding of the types of organic compounds that are recalcitrant to water treatment, natural organic matter (NOM) isolates from two drinking water sources (Mt. Zero and Moorabool reservoirs, Victoria, Australia) were separated into fractions of distinct chemical behaviour using resins. Four fractions were obtained from each water source and were organics absorbed to: (1) XAD-8 (very hydrophobic acids, VHA); (2) DAX-4 (slightly hydrophobic acids, SHA); (3) bound to an anion exchange resin (charged organics, CHAR); and (4) not absorbed or bound to resins (neutrals, NEUT). These fractions were then tested to determine the capacity of alum to remove them from water and to correlate this with the character of each isolate. The fractions were characterised by the application of high performance size exclusion chromatography (HPSEC), bacterial regrowth potential (BRP), trihalomethane formation potential (THMFP), pyrolysis gas-chromatography mass spectrometry (Py-GC-MS) and thermochemolysis. The highest removals of dissolved organic carbon (DOC) by alum treatment were in waters spiked with the CHAR fractions while the NEUT fractions were the most recalcitrant. The number average molecular weights (Mn) of DOC of the CHAR fractions before treatment were the highest, whilst those of the NEUT fractions were the lowest. After alum treatment, the Mn of the NEUT fractions were only slightly reduced. Results from Py-GC-MS and thermochemolysis indicate that the NEUT fractions had the highest relative proportion of saccharide derived organic material. Nonetheless, the BRP of waters spiked with the NEUT fractions differed markedly, indicating that organics recalcitrant to alum treatment can vary substantially in their chemical composition and capacity to support microbial growth.

scholarly journals Periplasmic synthesis and purification of the human prolactin antagonist Δ1-11-G129R-hPRL

AMB Express ◽  
2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Miriam F. Suzuki ◽  
Larissa A. Almeida ◽  
Stephanie A. Pomin ◽  
Felipe D. Silva ◽  
Renan P. Freire ◽  
...  
Keyword(s):  
Size Exclusion Chromatography ◽  
High Performance ◽  
Signal Sequence ◽  
Osmotic Shock ◽  
Size Exclusion ◽  
Specific Expression ◽  
E Coli ◽  
Human Prolactin ◽  
Exclusion Chromatography

AbstractThe human prolactin antagonist Δ1-11-G129R-hPRL is a 21.9 kDa recombinant protein with 188 amino acids that downregulates the proliferation of a variety of cells expressing prolactin receptors. Periplasmic expression of recombinant proteins in E. coli has been considered an option for obtaining a soluble and correctly folded protein, as an alternative to cytoplasmic production. The aim of this work was, therefore, to synthesize for the first time, the Δ1-11-G129R-hPRL antagonist, testing different activation temperatures and purifying it by classical chromatographic techniques. E. coli BL21(DE3) strain was transformed with a plasmid based on the pET25b( +) vector, DsbA signal sequence and the antagonist cDNA sequence. Different doses of IPTG were added, activating under different temperatures, and extracting the periplasmic fluid via osmotic shock. The best conditions were achieved by activating at 35 °C for 5 h using 0.4 mM IPTG, which gave a specific expression of 0.157 ± 0.015 μg/mL/A600 at a final optical density of 3.43 ± 0.13 A600. Purification was carried out by nickel-affinity chromatography followed by size-exclusion chromatography, quantification being performed via high-performance size-exclusion chromatography (HPSEC). The prolactin antagonist was characterized by SDS-PAGE, Western blotting, reversed-phase high-performance liquid chromatography (RP-HPLC) and MALDI-TOF–MS. The final product presented > 95% purity and its antagonistic effects were evaluated in vitro in view of potential clinical applications, including inhibition of the proliferation of cancer cells overexpressing the prolactin receptor and specific antidiabetic properties, taking also advantage of the fact that this antagonist was obtained in a soluble and correctly folded form and without an initial methionine.

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