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Application of a marine Halomonas exopolysaccharide in the preparation of immune enhancer

A technology of Halomonas and exopolysaccharide, which is applied in the preparation of marine Halomonas exopolysaccharide and the field of preparation of immune enhancers, can solve problems such as adverse immune response of immunity, and achieve increased expression and improved immune function. Effect

Active Publication Date: 2022-07-19
秦皇岛益尔生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] The main function of the immune system is to identify and eliminate pathogens to maintain the physiological balance and stability of the body, but too strong or too low immunity can cause a variety of adverse immune reactions

Method used

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  • Application of a marine Halomonas exopolysaccharide in the preparation of immune enhancer
  • Application of a marine Halomonas exopolysaccharide in the preparation of immune enhancer
  • Application of a marine Halomonas exopolysaccharide in the preparation of immune enhancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] The preparation method of marine Halomonas exopolysaccharide EPS2E1, the specific steps are as follows:

[0030] 1) Extraction of crude polysaccharide

[0031] Halomonas sp.2E1 was inoculated into 2216E (containing 1% sucrose) medium, placed on a shaker at 28°C, 150rpm for 48h. After the fermentation, centrifuge the Halomonas fermentation broth (4000rpm) for 15min to remove the bacteria, collect the supernatant, add three times the volume of 95% ethanol solution to the supernatant while stirring, stir evenly, and place it in a refrigerator at 4°C for static Set overnight, followed by centrifugation (4000 rpm, 15 min) to collect the precipitate, redissolved in water, dialyzed (molecular weight cut-off is 3.5 kDa), and freeze-dried to obtain crude polysaccharide.

[0032] 2) Purification of exopolysaccharides

[0033] The crude polysaccharide was purified by DEAE Fast Flow ion-exchange column chromatography. After loading, eluted with pure water for 2 column volumes (CV...

Embodiment 2

[0041] Effects of EPS2E1 administration on the proliferation activity of RAW264.7 cells in vitro

[0042] experimental method:

[0043] Take the RAW264.7 cells in the logarithmic phase of growth, pipetting to make a single cell suspension, and after cell counting, the RAW264. 5 Cells / mL were cultured in a 96-well plate. When the cells grew to about 50%, the old medium was replaced with 100 μL of FBS-free medium, and starved for 12 h. Then, the old medium was replaced with 100 μL blank or medium containing EPS2E1 with different concentrations, and the culture was continued for 24 h. Add 20 μL of MTT solution with a concentration of 5 mg / mL to each well, continue to incubate for 4 h, discard the supernatant, add 150 μL of DMSO to each well, shake gently for 10 min at room temperature to fully dissolve the crystals, and measure the absorbance at a wavelength of 490 nm.

[0044] Cell survival rate=(OD value of experimental group / OD value of blank control group)×100%.

[0045] T...

Embodiment 3

[0047] In vitro administration of EPS2E1 inhibited the secretion of NO secreted by RAW264.7 cells.

[0048] experimental method:

[0049] RAW264.7 cells were placed in 5 × 10 5 Cells / mL were placed in a 96-well plate and cultured in a constant temperature incubator (37°C, 5% CO2). When the cells grew to about 50%, the old medium was replaced with 100 μL of FBS-free medium, and starved for 12 hours. . Add samples according to the following groups: (1) blank control group; (2) LPS treatment group (final LPS concentration 1 μg / mL); (3) EPS2E1 treatment group (final concentration 6.25-200 μg / mL); Placed in an incubator for 24h. Collect the supernatant and measure the NO concentration by the Griess method: take 50 μL of the supernatant and add 50 μL of Griess reagent I and II solutions in turn, mix well and place at room temperature for 10 min to fully react, measure the absorbance at a wavelength of 540 nm, and use sodium nitrite The solution was a standard solution to establi...

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Abstract

The invention relates to the field of biotechnology, in particular to a preparation method of a marine Halomonas exopolysaccharide and its application in the preparation of an immune enhancer. An extracellular polysaccharide (EPS2E1) was isolated and purified from the fermentation broth of a marine Halomonas sp.2E1, and its physicochemical analysis and in vitro immune-enhancing activity were determined. The results showed that EPS2E1 is a mannan that can increase inflammatory factors (NO, IL-6, IL-1β and TNF-α) and cyclooxygenase- 2 (COX-2) expression, with significant immune-enhancing activity, and no obvious cytotoxicity. Therefore, the marine Halomonas exopolysaccharide EPS2E1 provided by the present invention can be used for preparing medicines or health care products related to immune enhancement, and has good development and utilization value.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a preparation method of a marine Halomonas exopolysaccharide and its use in preparing an immune enhancer. Background technique [0002] The main function of the immune system is to identify and eliminate pathogens to maintain the body's physiological balance and stability, but too strong or too low immunity can cause a variety of adverse immune responses. Immunodeficiency is a manifestation of immunocompromised, which is one of the main factors leading to infection and malignancy. Currently, the most common treatment for immunodeficiency is the use of immunopotentiators, such as fungal polysaccharides, thymosin, interferon, and BCG. Among these drugs, polysaccharides have been shown to be safe and nontoxic as immune enhancers. Polysaccharides can bind to receptors on the surface of macrophages, activate signal transduction pathways, and promote the production of cytokines such as i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P19/04C08B37/00A61K31/736A61P37/02A23L33/125C12R1/01
CPCC12P19/04C08B37/0003A61K31/736A61P37/02A23L33/125A23V2002/00A23V2200/324A23V2250/51
Inventor 张全斌王清池孙超岷王晶吴宁岳洋耿丽华
Owner 秦皇岛益尔生物科技有限公司
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