Compound Mutation Breeding of Streptococcus equi JC-63 for Producing High Molecular Weight Hyaluronic Acid and Optimization of Culture Conditions

2016 ◽  
Vol 703 ◽  
pp. 337-342
Author(s):  
Rong Hu Zhang ◽  
Jian Cheng Feng
Keyword(s):  
Molecular Weight ◽  
Hyaluronic Acid ◽  
High Molecular Weight ◽  
Corn Steep Liquor ◽  
Fermentation Medium ◽  
Mutation Breeding ◽  
Culture Conditions ◽  
Compound Mutation ◽  
Steep Liquor ◽  
Orthogonal Tests

A strain of mutant Streptococcus equi JC-63, which can produce high molecular weight hyaluronic acid, was obtained from compound mutation breeding of Streptococcus equi JC-0 using 5-Bromouracil, UV rays and nitrosoguanidine as mutagenic agents. After inoculation for 25 generations, Streptococcus equi JC-63 showed inheritance stability, but not hemolysis. The culture conditions for producing HMW-HA were optimized in a 1-L shake flask through multi-factor orthogonal tests. Results showed the optimum ratio of fermentation medium comprised 50 g/L sucrose, 80 g/L corn steep liquor, 10 g/L glucose, 2 g/L MgSO4, 2 g/L NaHCO3, 2 g/L Na2HPO4, and 0.1 g/L UTP. Under these fermentation conditions, the molecular weight of the produced HA was 2.76 × 106 Da, and HA yield reached 4.12 g/L.

Selection of a Mutant Strain Streptococcus equi SH-109 and Optimization of Culture Conditions for the Production of High Molecular Weight Hyaluronic Acid

2014 ◽  
Vol 915-916 ◽  
pp. 909-912 ◽  
Author(s):  
Jian Cheng Feng ◽  
Rong Hu Zhang
Keyword(s):  
Molecular Weight ◽  
Hyaluronic Acid ◽  
High Molecular Weight ◽  
Culture Conditions ◽  
Culture Broth ◽  
Agitation Speed ◽  
Wild Type ◽  
Streptococcus Equi ◽  
Optimal Ph ◽  
Selection Of

A high molecular weight hyaluronic acid-producing mutant, designated asStreptococcus equiSH-109 that did not produce streptolysin or hyaluronidase was selected from wild-typeStreptococcus equiSH-0 by NTG treatment. The selected mutant could produce high molecular weight HA and had perfect genetic stability after reproduction. Several culture conditions were optimized in a 5-l fermentor for the production of hyaluronic acid. It was found that the optimal pH and agitation speed were 7.6 and 550rpm, respectively. Under the optimized culture condition, hyaluronic acid with a molecular weight of 4.0 x 106was accumulated in the culture broth.

scholarly journals Addition of High Molecular Weight Hyaluronic Acid to Fibroblast-Like Stromal Cells Modulates Endogenous Hyaluronic Acid Metabolism and Enhances Proteolytic Processing and Secretion of Versican

Cells ◽  
2020 ◽  
Vol 9 (7) ◽  
pp. 1681
Author(s):  
Jiapeng Xue ◽  
Jinnan Chen ◽  
Quan Shen ◽  
Deva Chan ◽  
Jun Li ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Hyaluronic Acid ◽  
High Molecular Weight ◽  
Stromal Cells ◽  
De Novo ◽  
Linear Chain ◽  
Primary Cultures ◽  
Culture Conditions ◽  
Proteolytic Processing

We have examined the effect of exogenous linear chain high molecular weight hyaluronic acid (HMW HA) on endogenously synthesized hyaluronic acid (HA) and associated binding proteins in primary cultures of fibroblast-like stromal cells that were obtained by collagenase digestion of the murine peripatellar fat pad. The cultures were expanded in DMEM that was supplemented with fetal bovine serum and basic fibroblast growth factor (bFGF) then exposed to macrophage-colony-stimulating factor (MCSF) to induce macrophage properties, before activation of inflammatory pathways using E. coli lipopolysaccharide (LPS). Under all culture conditions, a significant amount of endogenously synthesized HA localized in LAMP1-positive lysosomal vesicles. However, this intracellular pool was depleted after the addition of exogenous HMW HA and was accompanied by enhanced proteolytic processing and secretion of de novo synthesized versican, much of which was associated with endosomal compartments. No changes were detected in synthesis, secretion, or proteolytic processing of aggrecan or lubricin (PRG4). The addition of HMW HA also modulated a range of LPS-affected genes in the TLR signaling and phagocytosis pathways, as well as endogenous HA metabolism genes, such as Has1, Hyal1, Hyal2, and Tmem2. However, there was no evidence for association of endogenous or exogenous HMW HA with cell surface CD44, TLR2 or TLR4 protein, suggesting that its physiochemical effects on pericelluar pH and/or ionic strength might be the primary modulators of signal transduction and vesicular trafficking by this cell type. We discuss the implications of these findings in terms of a potential in vivo effect of therapeutically applied HMW HA on the modification of osteoarthritis-related joint pathologies, such as pro-inflammatory and degradative responses of multipotent mesenchymal cells residing in the synovial membrane, the underlying adipose tissue, and the articular cartilage surface.

Preparation and Characterization of the High Molecular Weight [3H]Hyaluronic Acid

1992 ◽  
Vol 57 (10) ◽  
pp. 2151-2156 ◽  
Author(s):  
Peter Chabreček ◽  
Ladislav Šoltés ◽  
Hynek Hradec ◽  
Jiří Filip ◽  
Eduard Orviský
Keyword(s):  
Molecular Weight ◽  
Hyaluronic Acid ◽  
High Molecular Weight ◽  
Methyl Bromide ◽  
High Performance ◽  
Hydrogen Atoms ◽  
Isolation And Characterization ◽  
Labelling Procedure ◽  
Enzymatic Depolymerization

Two methods for the preparation of high molecular weight [3H]hyaluronic acid were investigated. In the first one, hydrogen atoms in the molecule were replaced by tritium. This isotopic substitution was performed in aqueous solution using Pd/CaCO3 as the catalyst. In the second method, the high molecular weight hyaluronic acid was alkylated with [3H]methyl bromide in liquid ammonia at a temperature of -33.5 °C. High-performance gel permeation chromatographic separation method was used for the isolation and characterization of the high molecular weight [3H]hyaluronic acid. Molecular weight parameters for the labelled biopolymers were Mw = 128 kDa, Mw/Mn = 1.88 (first method) and Mw = 268 kDa, Mw/Mn = 1.55 (second method). The high molecular weight [3H]hyaluronic acid having Mw = 268 kDa was degraded further by specific hyaluronidase. Products of the enzymatic depolymerization were observed to be identical for both, labelled and cold biopolymer. This finding indicates that the described labelling procedure using [3H]methyl bromide does not induce any major structural rearrangements in the molecule.

scholarly journals Synthesis of the Microbial Polysaccharide Gellan from Dairy and Plant-Based Processing Coproducts

2021 ◽  
Vol 2 (2) ◽  
pp. 234-244
Author(s):  
Thomas P. West
Keyword(s):  
Corn Steep Liquor ◽  
Nitrogen Sources ◽  
Soluble Starch ◽  
Culture Conditions ◽  
Water Soluble ◽  
Production Costs ◽  
Microbial Polysaccharide ◽  
Steep Liquor ◽  
The Cost ◽  
Condensed Distillers Solubles

This review examines the production of the microbial polysaccharide gellan, synthesized by Sphingomonas elodea, on dairy and plant-based processing coproducts. Gellan is a water-soluble gum that structurally exists as a tetrasaccharide comprised of 20% glucuronic acid, 60% glucose and 20% rhamnose, for which various food, non-food and biomedical applications have been reported. A number of carbon and nitrogen sources have been tested to determine whether they can support bacterial gellan production, with several studies attempting to optimize gellan production by varying the culture conditions. The genetics of the biosynthesis of gellan has been explored in a number of investigations and specific genes have been identified that encode the enzymes responsible for the synthesis of this polysaccharide. Genetic mutants exhibiting overproduction of gellan have also been identified and characterized. Several dairy and plant-based processing coproducts have been screened to learn whether they can support the production of gellan in an attempt to lower the cost of synthesizing the microbial polysaccharide. Of the processing coproducts explored, soluble starch as a carbon source supported the highest gellan production by S. elodea grown at 30 °C. The corn processing coproducts corn steep liquor or condensed distillers solubles appear to be effective nitrogen sources for gellan production. It was concluded that further research on producing gellan using a combination of processing coproducts could be an effective solution in lowering its overall production costs.

Surface Modification of Ultra-High Molecular Weight Polyethylene with Crosslinked Hyaluronic Acid and its Antiwear Performance

2015 ◽  
Vol 642 ◽  
pp. 94-98
Author(s):  
Chau Chang Chou ◽  
Yu Hsiang Hao ◽  
Fu Yin Hsu
Keyword(s):  
Molecular Weight ◽  
Hyaluronic Acid ◽  
High Molecular Weight ◽  
Incident Light ◽  
Contact Angle Measurement ◽  
Angle Measurement ◽  
Coated Sample ◽  
Infrared Spectrometry ◽  
Water Contact ◽  
Ultra High Molecular Weight

The surface of high-pressure crystallized ultra-high molecular weight polyethylene (UHMWPE) was modified for application as an artificial cartilage material. A UHMWPE surface pretreated by a series of processes, including treatment with O2-plasma and ethylenediamine solution, was coated with hyaluronic acid (HA). After that, adipic acid dihydrazide (AAD) was added to partially crosslink the HA coating in order to enhance its durability. The modified samples were verified by water contact angle measurement and Fourier transform infrared spectrometry. Both HA layers, original and crosslinked, were also quantitatively evaluated by carbohydrate chemistry assay according to the absorbance of the incident light. The tribological performance of the samples was evaluated by a pin-on-disk test rig lubricated by normal saline under an average pressure of 18 MPa and at a sliding speed of 0.03 m/s for 45 h. The wear resistance of the HA-coated UHMWPE specimens promoted by the crosslink process was superior to that of the original HA-coated sample, and that resistance was maintained after immersion in saline solution for one month.

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