A kind of exopolysaccharide and its application

Abstract

The invention discloses an exopolysaccharide and its application, belonging to the technical field of bioengineering. The exopolysaccharide (EPS) of the present invention is a kind of glucosamine, arabinose, galactosamine, galactose, glucose and Heteropolysaccharide composed of mannose, which has higher ability to scavenge free radicals than ascorbic acid; can provide good DNA protection for oxidative stress induced by AAPH and hydroxyl radicals; at the same time, it can also prevent direct damage to DNA activity by ultraviolet radiation destroy.

Description

technical field

[0001] The invention relates to an exopolysaccharide and its application, belonging to the technical field of bioengineering. Background technique

[0002] Deoxyribose nucleic acid (Deoxyribonucleic acid, DNA), as the storage center of biological genetic information, controls the proliferation and differentiation of cells, and is an important class of active macromolecules in organisms. Reactive oxygen species (Reactive oxygen species, ROS) produced by oxidative stress caused by radiation and chemical mutagens can cause oxidative damage to DNA, leading to various diseases and endangering human health.

[0003] ROS mainly include hydroxyl radicals (OH), superoxide radicals (O 2- ) and hydrogen peroxide (H 2 o 2 ). Under normal circumstances, ROS can be effectively eliminated by the body's antioxidant defense system, and its production and elimination are in balance; however, when exogenous ROS invades or there is radiation oxidative stress, the balance of ...

Examples

Embodiment 1

[0058] Embodiment 1: the identification of bacterial strain of the present invention

[0059] Specific steps are as follows:

[0060] (1) After the contents of chicken cecum were heat-treated at 85°C, they were enriched and cultured with Clostridium-enriched medium, and then 53 isolates were isolated after anaerobic culture on solid medium for bacteria screening;

[0061] (2) After the isolates undergo colony morphology, growth characteristics, Gram staining, and colony drawing experiments, a strain that can grow well on an anaerobic plate, has a smooth surface, is Gram-positive, produces acid, produces gas, and can produce Strains that utilize starch, cannot hydrolyze gelatin, and produce higher polysaccharides;

[0062] (3) The genomic DNA of the strain was extracted, and the 16S rRNA gene of the strain was amplified, recovered and sequenced, and the obtained 16S rRNA nucleotide sequence was submitted to NCBI for BLAST analysis. %above;

[0063] (4) Select a sequence with...

Embodiment 2

[0064] Embodiment 2: Preparation of exopolysaccharide of the present invention

[0065] (1) Clostridium butyricum (Clostridium butyricum) RO-07 in Example 1 is activated and expanded;

[0066] (2) Inoculate the expanded strain obtained in step (1) into the fermentation medium, and perform anaerobic culture at 30° C. for 48 to 72 hours to obtain a fermentation broth containing the exopolysaccharide of Clostridium butyricum;

[0067] (3) centrifuging the liquid fermentation broth prepared in step (2), and collecting the supernatant;

[0068] (4) Concentrate the supernatant obtained in step (3) by ultrafiltration to 1 / 5 of the original volume to obtain a concentrated solution, then add three times the volume of absolute ethanol to the concentrated solution, and after alcohol precipitation, centrifuge, collect sediment;

[0069] (5) Dissolve the precipitate collected in step (4) with water, add 1 / 4 volume of Seveage reagent to remove protein, and prepare a crude sugar solution; ...

Embodiment 3

[0073] Example 3: Identification of exopolysaccharides of the present invention

[0074] 1. Determination of molecular weight

[0075] Specific steps are as follows:

[0076] (1) Filter the EPS sample (5 mg / mL, 200 μL) obtained in Example 2 with a syringe filter (pore size 0.22 μm);

[0077] (2) Inject the filtered EPS sample into two Ultrahydrogel TM linear series columns (ID 7.8mm×300mm), and use NaNO 3 The solution (0.1M) was eluted at a flow rate of 0.9mL / min to obtain the response value data;

[0078] (3) The obtained data was collected and processed with Empower (Waters, USA), and the molecular weight (Mw) was directly calculated according to the molecular weight of each elution volume and the defined Mw of the RI signal value.

[0079] The measurement result is: the average molecular weight of EPS is 1.23×10 4 The chromatogram of Da; EPS showed a single symmetrical narrow peak, indicating that EPS is a homogeneous material; meanwhile, no peaks were detected from the...

technical field

[0001] The invention relates to an exopolysaccharide and its application, belonging to the technical field of bioengineering. Background technique

[0002] Deoxyribose nucleic acid (Deoxyribonucleic acid, DNA), as the storage center of biological genetic information, controls the proliferation and differentiation of cells, and is an important class of active macromolecules in organisms. Reactive oxygen species (Reactive oxygen species, ROS) produced by oxidative stress caused by radiation and chemical mutagens can cause oxidative damage to DNA, leading to various diseases and endangering human health.

[0003] ROS mainly include hydroxyl radicals (OH), superoxide radicals (O 2- ) and hydrogen peroxide (H 2 o 2 ). Under normal circumstances, ROS can be effectively eliminated by the body's antioxidant defense system, and its production and elimination are in balance; however, when exogenous ROS invades or there is radiation oxidative stress, the balance of ...

Examples

Embodiment 1

[0058] Embodiment 1: the identification of bacterial strain of the present invention

[0059] Specific steps are as follows:

[0060] (1) After the contents of chicken cecum were heat-treated at 85°C, they were enriched and cultured with Clostridium-enriched medium, and then 53 isolates were isolated after anaerobic culture on solid medium for bacteria screening;

[0061] (2) After the isolates undergo colony morphology, growth characteristics, Gram staining, and colony drawing experiments, a strain that can grow well on an anaerobic plate, has a smooth surface, is Gram-positive, produces acid, produces gas, and can produce Strains that utilize starch, cannot hydrolyze gelatin, and produce higher polysaccharides;

[0062] (3) The genomic DNA of the strain was extracted, and the 16S rRNA gene of the strain was amplified, recovered and sequenced, and the obtained 16S rRNA nucleotide sequence was submitted to NCBI for BLAST analysis. %above;

[0063] (4) Select a sequence with...

Embodiment 2

[0064] Embodiment 2: Preparation of exopolysaccharide of the present invention

[0065] (1) Clostridium butyricum (Clostridium butyricum) RO-07 in Example 1 is activated and expanded;

[0066] (2) Inoculate the expanded strain obtained in step (1) into the fermentation medium, and perform anaerobic culture at 30° C. for 48 to 72 hours to obtain a fermentation broth containing the exopolysaccharide of Clostridium butyricum;

[0067] (3) centrifuging the liquid fermentation broth prepared in step (2), and collecting the supernatant;

[0068] (4) Concentrate the supernatant obtained in step (3) by ultrafiltration to 1 / 5 of the original volume to obtain a concentrated solution, then add three times the volume of absolute ethanol to the concentrated solution, and after alcohol precipitation, centrifuge, collect sediment;

[0069] (5) Dissolve the precipitate collected in step (4) with water, add 1 / 4 volume of Seveage reagent to remove protein, and prepare a crude sugar solution; ...

Embodiment 3

[0073] Example 3: Identification of exopolysaccharides of the present invention

[0074] 1. Determination of molecular weight

[0075] Specific steps are as follows:

[0076] (1) Filter the EPS sample (5 mg / mL, 200 μL) obtained in Example 2 with a syringe filter (pore size 0.22 μm);

[0077] (2) Inject the filtered EPS sample into two Ultrahydrogel TM linear series columns (ID 7.8mm×300mm), and use NaNO 3 The solution (0.1M) was eluted at a flow rate of 0.9mL / min to obtain the response value data;

[0078] (3) The obtained data was collected and processed with Empower (Waters, USA), and the molecular weight (Mw) was directly calculated according to the molecular weight of each elution volume and the defined Mw of the RI signal value.

[0079] The measurement result is: the average molecular weight of EPS is 1.23×10 4 The chromatogram of Da; EPS showed a single symmetrical narrow peak, indicating that EPS is a homogeneous material; meanwhile, no peaks were detected from the...

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