Beta-1, 3-glucan and preparation method and application thereof

文档序号:1068077 发布日期:2020-10-16 浏览:24次 中文

阅读说明:本技术 一种β-1,3-葡聚糖及制备方法和应用 (Beta-1, 3-glucan and preparation method and application thereof ) 是由 杜彦军 杜玲玲 于 2020-07-31 设计创作,主要内容包括:本发明公开了一种β-1,3-葡聚糖及制备方法和应用,本发明的目的在于提供一种β-1,3-葡聚糖及制备方法,并将β-1,3-葡聚糖引入到修复口腔粘膜损伤的治疗中,公开了β-1,3-葡聚糖的新用途,公开了β-1,3-葡聚糖在制备修复口腔粘膜损伤药物中的应用;本发明通过实验证实了β-1,3-葡聚糖具有明显的抗炎、促进组织再生和促进伤口愈合的作用,可以表明β-1,3-葡聚糖对口腔粘膜具有修复作用,β-1,3-葡聚糖可以应用在制备修复口腔粘膜损伤的药物和日化产品中,为同类药物和日化产品的开发奠定了一定的基础。(The invention discloses beta-1, 3-glucan and a preparation method and application thereof, aims to provide beta-1, 3-glucan and a preparation method, introduces the beta-1, 3-glucan into treatment for repairing oral mucosa injury, discloses new application of the beta-1, 3-glucan, and discloses application of the beta-1, 3-glucan in preparation of a medicament for repairing oral mucosa injury; experiments prove that the beta-1, 3-glucan has obvious effects of resisting inflammation, promoting tissue regeneration and promoting wound healing, and can show that the beta-1, 3-glucan has a repairing effect on oral mucosa, and the beta-1, 3-glucan can be applied to the preparation of medicaments and daily chemical products for repairing the oral mucosa injury, so that a certain foundation is laid for the development of similar medicaments and daily chemical products.)

1. A beta-1, 3-glucan, characterized by: the preparation comprises the following components in parts by weight: 15-25 parts of sodium dihydrogen phosphate, 35-45 parts of potassium nitrate, 3-5 parts of magnesium sulfate, 0.1-0.2 part of calcium chloride, 0.2-0.3 part of ferrous sulfate, 0.04-0.08 part of manganese sulfate, 35-45 parts of rapeseed oil and 620 parts of cane sugar 580-doped materials; water 169900 and 17100 shares;

the preparation also comprises an agrobacterium ZX09 seed solution, and the volume ratio of the agrobacterium ZX09 seed solution to the total volume of the components is 1: 95-105.

2. The beta-1, 3-glucan according to claim 1, characterized in that: the weight parts of each component are respectively as follows: 18-22 parts of sodium dihydrogen phosphate, 38-42 parts of potassium nitrate, 3.5-4.5 parts of magnesium sulfate, 0.12-0.18 part of calcium chloride, 0.22-0.28 part of ferrous sulfate, 0.05-0.07 part of manganese sulfate, 38-42 parts of rapeseed oil and 610 parts of sucrose 590-; 17050 parts of 16952 parts of water;

the volume ratio of the agrobacterium ZX09 seed solution to the total volume of the components is 1: 98-102.

3. The beta-1, 3-glucan according to claim 2, characterized in that: the weight parts of each component are respectively as follows: 20 parts of sodium dihydrogen phosphate, 40 parts of potassium nitrate, 4 parts of magnesium sulfate, 0.14 part of calcium chloride, 0.25 part of ferrous sulfate, 0.06 part of manganese sulfate, 40 parts of rapeseed oil and 600 parts of sucrose; 17000 parts of water;

the volume ratio of the agrobacterium ZX09 seed solution to the total volume of the components is 1: 100.

4. The beta-1, 3-glucan according to any one of claims 1 to 3, wherein: the agrobacterium ZX09 seed solution is a mixture formed after agrobacterium ZX09 grows in a culture medium, and the culture medium comprises the following components in parts by weight: 210 portions of water 190-plus, 1.5-2.5 portions of peptone, 0.8-1.2 portions of yeast powder and 1.5-2.5 portions of sodium chloride.

5. The method for producing β -1, 3-glucan according to any one of claims 1 to 4, wherein: the method comprises the following steps:

(1) adding part of water, sodium dihydrogen phosphate, potassium nitrate, magnesium sulfate, calcium chloride, ferrous sulfate, manganese sulfate, rapeseed oil and sucrose into a fermentation tank according to the proportion, dissolving and uniformly stirring to obtain a mixed solution;

(2) adjusting the pH value of the mixed solution obtained in the step (1) to 6.5-7.5, and then carrying out steam sterilization;

(3) cooling the mixed solution subjected to steam sterilization in the step (2) to room temperature;

(4) adding the seed solution of the agrobacterium ZX09 into the mixed solution cooled in the step (3) according to the proportion, adjusting the ventilation flow to 18-22L/min, setting the stirring speed to 250-270rpm and the fermentation temperature to 28-32 ℃, culturing for 58-62h, and obtaining the fermentation liquid after the fermentation is finished;

(5) putting the fermentation liquor obtained in the step (4) into a barrel, adding 95% ethanol to precipitate the fermentation liquor, taking the precipitate, drying by pressure, dissolving the precipitate in the rest water, adding sodium hydroxide and diatomite, uniformly mixing, heating to 88-92 ℃ to obtain a suspension, and repeatedly filtering the suspension by using a plate-and-frame filter press until the solution is clear and transparent; the precipitate and filtrate are retained;

(6) adding 95% ethanol into the filtrate obtained in the step (5) to precipitate the filtrate, and filtering again; keeping the precipitate;

(7) and (4) mixing the precipitates obtained in the step (5) and the step (6), and sequentially performing press drying, drying and crushing to obtain a finished product of the beta-1, 3-glucan.

6. Use of the beta-1, 3-glucan produced in claim 5 for the manufacture of a medicament and a daily chemical product for repairing oral mucosal lesions.

7. Use of the beta-1, 3-glucan produced in claim 5 for the manufacture of anti-inflammatory drugs and daily use chemicals for the oral mucosa.

8. Use of the beta-1, 3-glucan produced in claim 5 in the manufacture of medicaments and daily use chemicals for promoting regeneration of oral mucosal tissue.

9. Use of the beta-1, 3-glucan produced in claim 5 in the manufacture of medicaments and daily use chemicals for promoting healing of wounds in the oral mucosa.

10. Use according to any one of claims 6-9, characterized in that: the beta-1, 3-glucan can be used alone or in combination with other drugs.

Technical Field

The invention relates to the field of medical technology and daily chemical products, in particular to beta-1, 3-glucan and a preparation method and application thereof.

Background

Beta-glucan is a macromolecular polysaccharide widely existing in cell walls of microorganisms, plants and seaweeds, a main chain structure is connected by beta-1, 3-glycosidic bonds beta and/or beta-1, 4 glycosidic bonds, and usually contains branched chains connected by beta-1, 6 bonds in different proportions, and main cell walls exist in the form of cell structural components. The glucan containing the beta-1, 3-glycosidic bond has stronger induction and activation effects on a foreign host defense system. In the 40 s of the 20 th century, Piller and Ecker, et al found that an immunostimulating active was present in yeast cells, and until the 60 s of the 20 th century, this active was not confirmed to be β -glucan.

Disclosure of Invention

The invention aims to provide beta-1, 3-glucan and a preparation method and application thereof. The technical effects that can be produced by the preferred technical scheme in the technical schemes provided by the invention are described in detail in the following.

The invention provides beta-1, 3-glucan, which is prepared from the following components in parts by weight: 15-25 parts of sodium dihydrogen phosphate, 35-45 parts of potassium nitrate, 3-5 parts of magnesium sulfate, 0.1-0.2 part of calcium chloride, 0.2-0.3 part of ferrous sulfate, 0.04-0.08 part of manganese sulfate, 35-45 parts of rapeseed oil and 620 parts of cane sugar 580-doped materials; water 169900 and 17100 shares;

the preparation also comprises an agrobacterium ZX09 seed solution, and the volume ratio of the agrobacterium ZX09 seed solution to the total volume of the components is 1: 95-105.

Further, the weight parts of the components are respectively as follows: 18-22 parts of sodium dihydrogen phosphate, 38-42 parts of potassium nitrate, 3.5-4.5 parts of magnesium sulfate, 0.12-0.18 part of calcium chloride, 0.22-0.28 part of ferrous sulfate, 0.05-0.07 part of manganese sulfate, 38-42 parts of rapeseed oil and 610 parts of sucrose 590-; 17050 parts of 16952 parts of water;

the volume ratio of the agrobacterium ZX09 seed solution to the total volume of the components is 1: 98-102.

Further, the weight parts of the components are respectively as follows: 20 parts of sodium dihydrogen phosphate, 40 parts of potassium nitrate, 4 parts of magnesium sulfate, 0.14 part of calcium chloride, 0.25 part of ferrous sulfate, 0.06 part of manganese sulfate, 40 parts of rapeseed oil and 600 parts of sucrose; 17000 parts of water;

the volume ratio of the agrobacterium ZX09 seed solution to the total volume of the components is 1: 100.

Further, the agrobacterium ZX09 seed solution is a mixture formed after agrobacterium ZX09 grows in a culture medium, and the culture medium comprises the following components in parts by weight: 210 portions of water 190-plus, 1.5-2.5 portions of peptone, 0.8-1.2 portions of yeast powder and 1.5-2.5 portions of sodium chloride.

The preparation method of the beta-1, 3-glucan provided by the invention comprises the following steps:

(1) adding water, sodium dihydrogen phosphate, potassium nitrate, magnesium sulfate, calcium chloride, ferrous sulfate, manganese sulfate, rapeseed oil and sucrose into a fermentation tank according to the proportion, dissolving and uniformly stirring to obtain a mixed solution;

(2) adjusting the pH value of the mixed solution obtained in the step (1) to 6.5-7.5, and then carrying out steam sterilization;

(3) cooling the mixed solution subjected to steam sterilization in the step (2) to room temperature;

(4) adding the seed solution of the agrobacterium ZX09 into the mixed solution cooled in the step (3) according to the proportion, adjusting the ventilation flow to 18-22L/min, setting the stirring speed to 250-270rpm and the fermentation temperature to 28-32 ℃, culturing for 58-62h, and obtaining the fermentation liquid after the fermentation is finished;

(5) putting the fermentation liquor obtained in the step (4) into a barrel, adding 95% ethanol to precipitate the fermentation liquor, taking the precipitate, drying by pressure, dissolving in water, adding sodium hydroxide and diatomite, uniformly mixing, heating to 88-92 ℃ to obtain a suspension, and repeatedly filtering the suspension by using a plate-and-frame filter press until the solution is clear and transparent; the precipitate and filtrate are retained;

(6) adding 95% ethanol into the filtrate obtained in the step (5) to precipitate the filtrate, and filtering again; keeping the precipitate;

(7) and (4) mixing the precipitates obtained in the step (5) and the step (6), and sequentially performing press drying, drying and crushing to obtain a finished product of the beta-1, 3-glucan.

Further, in the step (2), the steam sterilization is performed at a temperature of 121 ℃ for 25-35 min.

The invention discloses that the beta-1, 3-glucan has the function of repairing oral mucosa for the first time and can be used for treating the injury of the oral mucosa.

The invention particularly discloses application of beta-1, 3-glucan in preparing a medicine for repairing oral mucosa injury.

The invention discloses application of beta-1, 3-glucan in preparation of oral mucosa anti-inflammatory drugs and daily chemical products.

The invention discloses application of beta-1, 3-glucan in preparation of a medicament and a daily chemical product for promoting regeneration of oral mucosa tissues.

The invention discloses application of beta-1, 3-glucan in preparation of a medicine and a daily chemical product for promoting healing of oral mucosa wound.

Further, the medicine is an oral medicine or an external medicine; the dosage form of the oral medicine is a tablet or a capsule; the dosage form of the external medicine is gargle, spray, medicine film, aerosol, paste or cataplasm, and of course, special dosage forms such as controlled release agent, sustained release agent and targeting preparation are also included.

Further, the daily chemical product is toothpaste or mouthwash.

Further, the beta-1, 3-glucan can be used alone or in the form of a pharmaceutical composition comprising a therapeutically effective amount of beta-1, 3-glucan.

The structural formula of the beta-1, 3-glucan prepared by the invention is as follows:

the general structural formula is as follows:

poly-3) - β -D-pyran- (l → 3) - [ β -D-pyran- (l → 3) - β -D-pyran-]3- (l → 3) - α -D-pyran- (l → 3) - α -D-pyran- (l → glucose

The action mechanism of the beta-1, 3-glucan in the invention for repairing the oral mucosa is as follows:

on one hand, the beta-1, 3-glucan can promote the secretion of EGF (epidermal growth factor) in the epithelial basal cell layer of the oral mucosa, maintain the regeneration and proliferation of the epithelium of the oral mucosa, reduce the ulcer area, relieve the inflammation of the tissue of the oral mucosa and reduce the infiltration of lymphocytes; on the other hand, beta-1, 3-glucan, as an immunomodulator, can significantly increase lymphocyte activity, promote the production of IgA, IgG and IgM antibodies, which are secretion agents of cytokines such as TNF-alpha, IL-beta and IL-2, and thus exert anti-inflammatory and significantly immune enhancing effects.

Based on the technical scheme, the embodiment of the invention can at least produce the following technical effects:

the invention provides beta-1, 3-glucan and a preparation method thereof, and the prepared beta-1, 3-glucan can be introduced into treatment for repairing oral mucosa injury; experiments prove that the beta-1, 3-glucan has obvious effects of resisting inflammation, promoting tissue regeneration and promoting wound healing, and can show that the beta-1, 3-glucan has a repairing effect on oral mucosa, and the beta-1, 3-glucan can be applied to the preparation of medicaments and daily chemical products for repairing the oral mucosa injury, so that a certain foundation is laid for the development of similar medicaments and daily chemical products.

Drawings

In order to illustrate the embodiments or prior art solutions more clearly, the drawings used in the description of the embodiments or prior art will be briefly described below, it is obvious that the drawings in the following description are only some embodiments of the present description, and it is obvious for a person skilled in the art that other drawings can be obtained from these drawings without inventive effort.

FIG. 1 is a graphical representation of the anti-inflammatory effect of beta-1, 3-glucan on zebrafish inflammation;

FIG. 2 shows the anti-inflammatory effect of beta-1, 3-glucan on zebrafish inflammation (neutrophil count);

FIG. 3 is a graph showing the inflammation resolution rate of beta-1, 3-glucan against inflammation in zebrafish;

FIG. 4 is a typical graph of regeneration (arrow marks) of zebrafish tissue after β -1, 3-glucan treatment;

FIG. 5 shows the regeneration length (in pixels) of zebrafish tissue after beta-1, 3-glucan treatment;

FIG. 6 shows the tissue regeneration promoting effect of zebra fish treated with beta-1, 3-glucan;

FIG. 7 is a phenotype plot of pigment recruitment to the wound site following β -1, 3-glucan treatment;

FIG. 8 is a phenotype graph of angiogenesis after treatment of a test article;

FIG. 9 is the sum of the opacity of the wound site of zebrafish after treatment with β -1, 3-glucan;

FIG. 10 shows the wound healing promoting effect of beta-1, 3-glucan on zebrafish after treatment;

FIG. 11 is a graph of the incidence of neoangiogenesis in zebrafish after treatment with β -1, 3-glucan.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the technical solutions of the present invention will be described in detail below. It is to be understood that the described embodiments are merely exemplary of the invention, and not restrictive of the full scope of the invention. All other embodiments, which can be derived by a person skilled in the art from the examples given herein without any inventive step, are within the scope of the present invention.

First, preparation example:

preparation of beta-1, 3-glucan:

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