阅读说明：本技术 一种白芨多糖的制备方法及得到的白芨多糖、应用 (Preparation method of bletilla striata polysaccharide, bletilla striata polysaccharide obtained by preparation method and application of bletilla striata polysaccharide ) 是由 金志强 高合意 刘田静 雷登凤 于 2020-12-10 设计创作，主要内容包括：本发明涉及医药化工技术领域,特别是一种白芨多糖的制备方法及得到的白芨多糖、应用。本发明提供了一种白芨多糖的提取方法,包括以下步骤：S1.将白芨粉用乙醇溶液进行预处理,得到滤渣；S2.将得到的滤渣加入到低浓度的乙醇溶液中搅拌分散,离心,得含白芨多糖清液；S3.将白芨多糖清液中加入淀粉酶和/或葡萄糖苷酶,35-85℃搅拌3-7小时,得到酶解液；向酶解液中加乙醇,离心,得酶解液上清液；S4.酶解液上清液过滤,即得白芨多糖。(The invention relates to the technical field of pharmaceutical chemicals, in particular to a preparation method of bletilla striata polysaccharide, the obtained bletilla striata polysaccharide and application. The invention provides a method for extracting bletilla striata polysaccharide, which comprises the following steps: s1, pretreating bletilla striata powder by using an ethanol solution to obtain filter residues; s2, adding the obtained filter residue into a low-concentration ethanol solution, stirring and dispersing, and centrifuging to obtain a clear liquid containing bletilla striata polysaccharide; s3, adding amylase and/or glucosidase into the bletilla polysaccharide clear liquid, and stirring for 3-7 hours at 35-85 ℃ to obtain enzymatic hydrolysate; adding ethanol into the enzymolysis liquid, and centrifuging to obtain enzymolysis liquid supernatant; and S4, filtering the supernatant of the enzymolysis liquid to obtain the bletilla striata polysaccharide.)

1. The extraction method of bletilla striata polysaccharide is characterized by comprising the following steps:

s1, pretreating bletilla striata powder by using an ethanol solution to obtain filter residues;

s2, adding the obtained filter residue into a low-concentration ethanol solution, stirring and dispersing, and centrifuging to obtain a clear liquid containing bletilla striata polysaccharide;

s3, adding amylase and/or glucosidase into the bletilla polysaccharide clear liquid, and stirring for 3-7 hours at 35-85 ℃ to obtain enzymatic hydrolysate; adding ethanol into the enzymolysis liquid, and centrifuging to obtain enzymolysis liquid supernatant;

and S4, filtering the supernatant of the enzymolysis liquid to obtain the bletilla striata polysaccharide.

2. The extraction method of bletilla striata polysaccharide as claimed in claim 1, wherein the specific operation of step S1 is: adding rhizoma bletilla powder into 50-95% ethanol water solution at a material-to-liquid ratio of 1:5-20g/mL, treating at 40-95 deg.C for 20-50min, and filtering to obtain crude product; repeating the steps for 2-4 times, and finally filtering to obtain filter residue.

3. The method for extracting bletilla striata polysaccharide as claimed in claim 1, wherein the low-concentration ethanol solution is 0-20% ethanol water solution, excluding 0.

4. The extraction method of bletilla striata polysaccharide as claimed in claim 1, wherein the specific operation of step S2 is: adding the obtained filter residue into low concentration ethanol solution, stirring at 40-100 deg.C for 20-50min, centrifuging to obtain precipitate and primary supernatant; repeating the above steps for 1-3 times, and collecting all supernatant to obtain clear solution containing rhizoma bletilla polysaccharide.

5. The method for extracting bletilla striata polysaccharide as claimed in claim 1, wherein the amylase is alpha-amylase and/or beta-amylase; the glucosidase is alpha-glucosidase and/or beta-glucosidase.

6. The method for extracting bletilla striata polysaccharide as claimed in claim 1, wherein in step S3, ethanol is added to the enzymolysis solution to a concentration of 20-40%; preserving heat at 40-100 ℃ for 20-50min, then cooling to room temperature, and centrifuging to obtain supernatant of the enzymolysis liquid.

7. The extraction method of bletilla striata polysaccharide as claimed in claim 1, wherein the specific steps of step S4 are: concentrating the supernatant of the enzymolysis liquid until the ethanol content is less than 1%, and then treating the concentrated solution by a membrane core.

8. The method for extracting bletilla striata polysaccharide as claimed in claim 7, wherein the molecular weight cut-off of the membrane core is 5000 Da.

9. A bletilla striata polysaccharide obtained by the extraction method of bletilla striata polysaccharide as claimed in any one of claims 1-8.

10. Use of bletilla striata polysaccharide as defined in claim 9 in cosmetics.

Technical Field

The invention relates to the technical field of pharmaceutical chemicals, in particular to a preparation method of bletilla striata polysaccharide, the obtained bletilla striata polysaccharide and application.

Background

Bletilla striata (Thunb.) reichb. f) is a plant of bletilla belonging to the family Orchidaceae, also called bletilla striata, snowy tail, etc., and has the main functions of healing sore, stopping bleeding, tonifying lung, reducing swelling, promoting granulation, etc. Bletilla striata contains a large amount of water-soluble polysaccharide, and the chemical component of bletilla striata is glucomannan which is the main functional component of bletilla striata. The bletilla striata polysaccharide is composed of D-glucose and D-mannose, the ratio of glucose to mannose is about 1:4, and a small amount of metal ions are combined in the molecule. The relative molecular mass of bletilla striata polysaccharides is generally considered to be about 10 to 20 ten thousand, and in addition, the viscosity average molecular weight of bletilla striata crude polysaccharides is also reported to be more than 250 ten thousand.

The infrared spectrum measurement of Zhuxiaping et al shows that the glucosidic bonds in the bletilla striata polysaccharide are beta-type, no alpha-type glucosidic bonds and no acyl modifying groups exist. Researches on Chenjing and Ranunculus japonicus prove that three residues of beta-1, 4-mannose, beta-1, 4-glucose and alpha-1, 6-glucose exist in bletilla polysaccharide at the same time. In view of the fact that the type, amount and distribution characteristics of the side chains on the main chain have great influence on the characteristics of the polysaccharide gum, and starch grains and pigment components contained in the bletilla striata medicinal material have great influence on the stability of the product. At present, the bletilla striata polysaccharide is usually extracted by water or ethanol, new extraction solvent needs to be supplemented or extraction batches need to be added in due to the viscosity and the water swelling property of the bletilla striata polysaccharide, the production period is long, the solvent amount is large, the energy consumption is high, and the purity is low.

Disclosure of Invention

In order to solve the above technical problems, a first aspect of the present invention provides a method for extracting bletilla striata polysaccharide, comprising the following steps:

s1, pretreating bletilla striata powder by using an ethanol solution to obtain filter residues;

s2, adding the obtained filter residue into a low-concentration ethanol solution, stirring and dispersing, and centrifuging to obtain a clear liquid containing bletilla striata polysaccharide;

s3, adding amylase and/or glucosidase into the bletilla polysaccharide clear liquid, and stirring for 3-7 hours at 35-85 ℃ to obtain enzymatic hydrolysate; adding ethanol into the enzymolysis liquid, and centrifuging to obtain enzymolysis liquid supernatant;

and S4, filtering the supernatant of the enzymolysis liquid to obtain the bletilla striata polysaccharide.

As a preferred technical solution, the specific operation of step S1 is: adding rhizoma bletilla powder into 50-95% ethanol water solution at a material-to-liquid ratio of 1:5-20g/mL, treating at 40-95 deg.C for 20-50min, and filtering to obtain crude product; repeating the steps for 2-4 times, and finally filtering to obtain filter residue.

As a preferable technical scheme, the low-concentration ethanol solution is an ethanol water solution with the mass concentration of 0-20%, and 0 is not included.

As a preferred technical solution, the specific operation of step S2 is: adding the obtained filter residue into low-concentration ethanol solution with a material-liquid ratio of 1:20-100g/mL, stirring at 40-100 deg.C for 20-50min, and centrifuging to obtain precipitate and primary supernatant; repeating the above steps for 1-3 times, and collecting all supernatant to obtain clear solution containing rhizoma bletilla polysaccharide;

as a preferred technical scheme, the amylase is alpha-amylase and/or beta-amylase; the glucosidase is alpha-glucosidase and/or beta-glucosidase.

As a preferred technical scheme, adding ethanol into the enzymolysis liquid to the concentration of 20-40%; preserving heat at 40-100 ℃ for 20-50min, then cooling to room temperature, and centrifuging to obtain supernatant of the enzymolysis liquid.

As a preferred technical solution, the specific steps of step S4 are: concentrating the supernatant of the enzymolysis liquid until the ethanol content is less than 1%, and then treating the concentrated solution by a membrane core.

As a preferred technical scheme, the molecular weight cut-off of the membrane core is 5000 Da.

The second aspect of the invention provides bletilla striata polysaccharide obtained by the method for extracting bletilla striata polysaccharide.

The third aspect of the invention provides the application of the bletilla striata polysaccharide in cosmetics.

Has the advantages that: the extraction method adopted by the application can effectively remove coloring components such as flavonoids, bibenzyls (phenanthrenes) and the like and high-ionic components; flavones, bibenzyl (phenanthrenes) and the like are easy to oxidize and have poor light stability, and the stability of the product is influenced when the flavones, the bibenzyl (phenanthrenes) and the like are added into common cosmetics, so that the shelf life of the cosmetics is shortened; the high ionic components in plant, such as mineral salts and low molecular organic acids, can destroy the thickening system in cosmetic, thereby limiting the application of rhizoma bletilla extract.

Detailed Description

In order to solve the above problems, the invention provides a method for extracting bletilla striata polysaccharide, which comprises the following steps:

s1, pretreating bletilla striata powder by using an ethanol solution to obtain filter residues;

s2, adding the obtained filter residue into a low-concentration ethanol solution, stirring and dispersing, and centrifuging to obtain a clear liquid containing bletilla striata polysaccharide;

s3, adding amylase and/or glucosidase into the bletilla polysaccharide clear liquid, and stirring for 3-7 hours at 35-85 ℃ to obtain enzymatic hydrolysate; adding ethanol into the enzymolysis liquid, and centrifuging to obtain enzymolysis liquid supernatant;

and S4, filtering the supernatant of the enzymolysis liquid to obtain the bletilla striata polysaccharide.

Step S1

The specific operation of step S1 is: adding rhizoma bletilla powder into 50-95% ethanol water solution at a material-to-liquid ratio of 1:5-20g/mL, treating at 40-95 deg.C for 20-50min, and filtering to obtain crude product; repeating the steps for 2-4 times, and finally filtering to obtain filter residue.

The mesh number of the bletilla striata powder is not particularly limited, and for example, the mesh number of the bletilla striata powder is 60-400 meshes.

Preferably, the bletilla striata powder is added into an ethanol water solution with the mass concentration of 70%, the material-liquid ratio is 1:7g/mL, the treatment is carried out for 30min at the temperature of 70 ℃, and a crude product is obtained by filtration; repeating the steps for 2 times, and finally filtering to obtain filter residue.

The applicant finds that the bletilla striata powder can be well decolored and desalted when ethanol water solution with the mass concentration of 50-95% is adopted and the material-liquid ratio is 1:5-20 g/mL; the reason is presumed to be: yellow parts in bletilla are mostly flavonoid and bibenzyl components, and the yellow parts have certain lipid solubility and are better in solubility only under ethanol with a corresponding proportion; but the solubility of the bletilla striata polysaccharide becomes lower under high-concentration ethanol; along with the increase of the concentration of the ethanol, the solubility of the mineral salt is obviously reduced, so that the conductivity of the filter residue is improved.

Step S2

The low-concentration ethanol solution is an ethanol water solution with the mass concentration of 0-20%, and 0 is not included; preferably, the low-concentration ethanol solution is an ethanol aqueous solution with the mass concentration of 6%.

The specific operation of step S2 is: adding the obtained filter residue into low-concentration ethanol solution with a material-liquid ratio of 1:20-100g/mL, stirring at 40-100 deg.C for 20-50min, and centrifuging to obtain precipitate and primary supernatant; repeating the above steps for 1-3 times, and collecting all supernatant to obtain clear solution containing rhizoma bletilla polysaccharide;

the rotation speed and time of the centrifugation are not particularly limited, such as 8000-; in the application, the rotating speed is 10000r/min and 10 min.

Preferably, the obtained filter residue is added into a low-concentration ethanol water solution with the material-liquid ratio of 1:40g/mL, stirred for 30min at the temperature of 70 ℃, and centrifuged to obtain a precipitate and primary supernatant; repeating the above steps for 1 time, and collecting supernatant for 2 times to obtain supernatant containing rhizoma bletilla polysaccharide.

The applicant finds that when a low-concentration ethanol aqueous solution is adopted and the ratio of the material to the liquid is 1:20-100g/mL, the extraction rate of the bletilla striata polysaccharide can be improved under a proper temperature condition; the over-high concentration of the ethanol causes the reduction of the solubility of the bletilla polysaccharide and increases the production cost; however, the absence of ethanol is not favorable for the dispersion of macromolecular bletilla striata polysaccharide, and the bletilla striata polysaccharide is easy to agglomerate.

Step S3

The amylase is alpha-amylase and/or beta-amylase.

Such alpha-amylases include, but are not limited to, moderate temperature alpha-amylases, fungal alpha-amylases.

The glucosidase is alpha-glucosidase and/or beta-glucosidase.

In the step S3, the addition of ethanol to the enzymatic hydrolysate is not particularly required, and the specific steps are as follows: adding ethanol into the enzymolysis solution to reach a concentration of 20-40%; preserving heat at 40-100 ℃ for 20-50min, then cooling to room temperature, and centrifuging to obtain supernatant of the enzymolysis liquid. The method removes the influence of protease residues on the product; different from the traditional high-temperature enzyme deactivation mode, the energy consumption is greatly saved.

The enzyme activity of the alpha-amylase is 1.4 x 10⁵IU/g; the beta-glucosidase enzyme activity is 5 x 10⁴IU/g。

The rotation speed and time of the centrifugation are not particularly limited, such as 8000-; in the application, the rotating speed is 10000r/min and 10 min.

The turbidity and the viscosity of the product are reduced by adding the alpha-amylase and/or the beta-glucosidase; the alpha-amylase mainly carries out enzymolysis on a small amount of alpha-type glycosidic bonds of starch and bletilla polysaccharide in the clear liquid; the spatial structure of the components or functional groups is changed to a certain extent at low temperature, so that the solution becomes mixed and turbid, and the stability of the cosmetic is influenced; the beta-glucosidase is mainly used for carrying out enzymolysis on the bletilla striata polysaccharide, so that a unique molecular weight is formed, and the interaction of the alpha-amylase and the beta-glucosidase ensures that the bletilla striata polysaccharide after enzymolysis has more excellent stability and is clearer and brighter.

Step S4

The specific steps of step S4 are: concentrating the supernatant of the enzymolysis liquid until the ethanol content is less than 1%, and then treating the concentrated solution by a membrane core; the trapped liquid is viscous liquid, namely bletilla striata polysaccharide-1; the permeate is oligosaccharide, namely bletilla striata polysaccharide-2.

The molecular weight cut-off of the membrane core is 5000 Da.

The molecular weight of the bletilla striata polysaccharide-1 is 10-20 ten thousand Da; the molecular weight of the bletilla striata polysaccharide-2 is less than 5000 Da.

The present invention will be specifically described below by way of examples. It should be noted that the following examples are only for illustrating the present invention and should not be construed as limiting the scope of the present invention, and that the insubstantial modifications and adaptations of the present invention by those skilled in the art based on the above disclosure are still within the scope of the present invention.

In addition, the starting materials used are all commercially available, unless otherwise specified.

Examples

Example 1

A method for extracting clear liquid containing bletilla striata polysaccharide comprises the following steps:

s1, adding 2g of bletilla striata powder into 95% ethanol water solution with the mass concentration of 1:20g/mL, treating at 80 ℃ for 30min, and filtering to obtain a crude product; repeating the steps for 2 times, and finally filtering to obtain filter residue.

S2, adding 200mL of ethanol water solution with the mass concentration of 6% into the obtained filter residue, stirring at 70 ℃ for 30min at the rotating speed of 10000r/min, and centrifuging for 10min to obtain supernatant, namely the bletilla striata polysaccharide-containing clear liquid.

Example 2

A method for extracting clear liquid containing bletilla striata polysaccharide comprises the following steps:

s1, adding 2g of bletilla striata powder into 50% ethanol water solution with the mass concentration of 1:20g/mL, treating at 95 ℃ for 30min, and filtering to obtain a crude product; repeating the steps for 2 times, and finally filtering to obtain filter residue.

S2, adding 200mL of ethanol water solution with the mass concentration of 6% into the obtained filter residue, stirring at 70 ℃ for 30min at the rotating speed of 10000r/min, and centrifuging for 10min to obtain supernatant, namely the bletilla striata polysaccharide-containing clear liquid.

Example 3

A method for extracting clear liquid containing bletilla striata polysaccharide comprises the following steps:

s1, adding 2g of bletilla striata powder into 70% ethanol water solution with the mass concentration of 1:7g/mL, treating at 70 ℃ for 30min, and filtering to obtain a crude product; repeating the steps for 2 times, and finally filtering to obtain filter residue.

S2, adding 200mL of ethanol water solution with the mass concentration of 6% into the obtained filter residue, stirring at 70 ℃ for 30min at the rotating speed of 10000r/min, and centrifuging for 10min to obtain supernatant, namely the bletilla striata polysaccharide-containing clear liquid.

The bletilla polysaccharide-containing supernatants obtained in examples 1-3 were tested for decolorization (visual inspection), salt rejection (conductivity), and extraction yield, as shown in the following table.

The extraction rate refers to the ratio of solid weight of clear liquid containing rhizoma Bletillae polysaccharide to the weight of rhizoma Bletillae powder.

	Example 1	Example 2	Example 3
				Decolorization ratio	Colorless and colorless	Light yellow	Colorless and colorless
Electrical conductivity of	580μs/cm	50μs/cm	60μs/cm
				Extraction rate	49％	32％	51％

Wherein 2g of the bletilla striata dry powder before treatment is dispersed in 200mL of water, and the electric conductivity is 2100 mus/cm.

Example 4

A method for extracting clear liquid containing bletilla striata polysaccharide comprises the following steps:

s1, adding 2g of bletilla striata powder into 70% ethanol water solution with the mass concentration of 1:7g/mL, treating at 70 ℃ for 30min, and filtering to obtain a crude product; repeating the steps for 2 times, and finally filtering to obtain filter residue.

S2, adding 200mL of 20% ethanol water solution into the obtained filter residue, stirring for 30min at the temperature of 70 ℃, and centrifuging (10000r/min, 10min) to obtain a precipitate and a primary supernatant; repeating the above steps for 1 time (adding 200mL of 20% ethanol water solution into the precipitate, stirring at 70 deg.C for 30min, centrifuging to obtain precipitate and secondary supernatant), and mixing the secondary supernatant to obtain supernatant containing rhizoma bletilla polysaccharide.

Example 5

A method for extracting clear liquid containing bletilla striata polysaccharide comprises the following steps:

s1, adding 2g of bletilla striata powder into 70% ethanol water solution with the mass concentration of 1:7g/mL, treating at 70 ℃ for 30min, and filtering to obtain a crude product; repeating the steps for 2 times, and finally filtering to obtain filter residue.

S2, adding the obtained filter residue into 80mL of ethanol water solution with the mass concentration of 6%, stirring for 30min at the temperature of 70 ℃, and centrifuging (10000r/min, 10min) to obtain a precipitate and a primary supernatant; repeating the above steps for 1 time (adding 80mL ethanol water solution with mass concentration of 6% into the precipitate, stirring at 70 deg.C for 30min, centrifuging to obtain precipitate and secondary supernatant), and mixing the secondary supernatant to obtain clear solution containing rhizoma bletilla polysaccharide.

Example 6

A method for extracting clear liquid containing bletilla striata polysaccharide comprises the following steps:

s1, adding 2g of bletilla striata powder into 70% ethanol water solution with the mass concentration of 1:7g/mL, treating at 70 ℃ for 30min, and filtering to obtain a crude product; repeating the steps for 2 times, and finally filtering to obtain filter residue.

S2, adding 200mL of deionized water into the obtained filter residue, stirring for 30min at the temperature of 100 ℃, and centrifuging (10000r/min, 10min) to obtain a precipitate and a primary supernatant; repeating the above steps for 1 time (adding 200mL deionized water into the precipitate, stirring at 100 deg.C for 30min, centrifuging to obtain precipitate and secondary supernatant), and mixing the secondary supernatant to obtain clear solution containing rhizoma bletilla polysaccharide.

Example 7

A method for extracting clear liquid containing bletilla striata polysaccharide comprises the following steps:

s1, adding 2g of bletilla striata powder into 70% ethanol water solution with the mass concentration of 1:7g/mL, treating at 70 ℃ for 30min, and filtering to obtain a crude product; repeating the steps for 2 times, and finally filtering to obtain filter residue.

S2, adding 40mL of ethanol water solution with the mass concentration of 6% into the obtained filter residue, stirring for 30min at the temperature of 70 ℃, and centrifuging (10000r/min, 10min) to obtain a precipitate and a primary supernatant; repeating the above steps for 1 time (adding 40mL of 6% ethanol water solution into the precipitate, stirring at 70 deg.C for 30min, centrifuging to obtain precipitate and secondary supernatant), and mixing the secondary supernatant to obtain bletilla striata polysaccharide-containing supernatant.

Example 8

A method for extracting clear liquid containing bletilla striata polysaccharide comprises the following steps:

s1, adding 2g of bletilla striata powder into 70% ethanol water solution with the mass concentration of 1:7g/mL, treating at 70 ℃ for 30min, and filtering to obtain a crude product; repeating the steps for 2 times, and finally filtering to obtain filter residue.

S2, adding the obtained filter residue into 80mL of ethanol water solution with the mass concentration of 6%, stirring for 30min at the temperature of 40 ℃, and centrifuging (10000r/min, 10min) to obtain a precipitate and a primary supernatant; repeating the above steps for 1 time (adding 80mL ethanol water solution with mass concentration of 6% into the precipitate, stirring at 40 deg.C for 30min, centrifuging to obtain precipitate and secondary supernatant), and mixing the secondary supernatant to obtain supernatant containing rhizoma bletilla polysaccharide.

The bletilla striata polysaccharide-containing clear solutions obtained in examples 4-8 were tested for extraction yield, as shown in the following table.

The extraction rate refers to the ratio of solid weight of clear liquid containing rhizoma Bletillae polysaccharide to the weight of rhizoma Bletillae powder.

	Example 4	Example 5	Example 6	Example 7	Example 8
						Extraction rate	43％	50％	40％	45％	47％

Example 9

A method for extracting bletilla striata polysaccharide comprises the following steps:

step S1 and step S2 are the same as in example 5;

s3, adding 0.001g of alpha-amylase into the bletilla polysaccharide clear liquid, and stirring for 5 hours at 55 ℃ to obtain enzymatic hydrolysate; concentrating until the solid content of the enzymolysis liquid is 2%, refrigerating at 4 ℃ for 24h, recovering to 25 ℃, and measuring turbidity and viscosity;

example 10

A method for extracting bletilla striata polysaccharide comprises the following steps:

step S1 and step S2 are the same as in example 5;

s3, stirring the bletilla striata polysaccharide clear liquid for 5 hours at 55 ℃; concentrating to solid content of 2%, refrigerating at 4 deg.C for 24 hr, recovering to 25 deg.C, and measuring turbidity and viscosity.

Example 11

A method for extracting bletilla striata polysaccharide comprises the following steps:

step S1 and step S2 are the same as in example 5;

s3, adding 0.001g of protease into the bletilla striata polysaccharide clear liquid, and stirring for 5 hours at 55 ℃ to obtain an enzymolysis liquid; concentrating until the solid content of the enzymolysis liquid is 2%, refrigerating at 4 deg.C for 24h, recovering to 25 deg.C, and measuring turbidity and viscosity.

Example 12

A method for extracting bletilla striata polysaccharide comprises the following steps:

step S1 and step S2 are the same as in example 5;

s3, adding 0.001g of alpha-amylase and 0.001g of beta-glucosidase into the bletilla polysaccharide clear liquid; stirring for 5 hours at 55 ℃ to obtain an enzymolysis liquid; concentrating until the solid content of the enzymolysis liquid is 2%, refrigerating at 4 ℃ for 24h, recovering to 25 ℃, and measuring turbidity and viscosity;

example 13

A method for extracting bletilla striata polysaccharide comprises the following steps:

step S1 and step S2 are the same as in example 5;

s3, adding 0.001g of beta-glucosidase into the bletilla polysaccharide clear liquid; stirring for 5 hours at 55 ℃ to obtain an enzymolysis liquid; concentrating until the solid content of the enzymolysis liquid is 2%, refrigerating at 4 ℃ for 24h, recovering to 25 ℃, and measuring turbidity and viscosity;

example 14

A method for extracting bletilla striata polysaccharide comprises the following steps:

step S1 and step S2 are the same as in example 5;

s3, adding 0.001g of alpha-amylase and 0.001g of beta-glucosidase into the bletilla polysaccharide clear liquid; stirring for 5 hours at 55 ℃ to obtain an enzymolysis liquid; adding ethanol into the enzymolysis solution to reach the concentration of 30%; preserving heat at 70 ℃ for 30min, then cooling to room temperature at 10000r/min, and centrifuging for 10min to obtain supernatant of the enzymolysis solution.

Concentrating the supernatant of the enzymolysis liquid until the solid content of the solution is 2%, refrigerating the solution at 4 ℃ for 24 hours, then recovering the solution to 25 ℃, and measuring turbidity and viscosity;

the supernatants of the enzymatic hydrolysates obtained in examples 9 to 14 were tested for turbidity and viscosity, as specified in the following table.

Example 15

A method for extracting bletilla striata polysaccharide comprises the following steps:

s1, adding 1000g of bletilla striata powder into 70% ethanol water solution with the mass concentration of 1:7g/mL, treating at 70 ℃ for 30min, and filtering to obtain a crude product; repeating the steps for 2 times, and finally filtering to obtain filter residue.

S2, adding the obtained filter residue into 40L of ethanol water solution with the mass concentration of 6%, stirring at 70 ℃ for 30min, 10000r/min, and centrifuging for 10min to obtain a precipitate and a primary supernatant; repeating the above steps for 1 time (adding 40L ethanol water solution with mass concentration of 6% into the precipitate, stirring at 70 deg.C for 30min, 10000r/min, centrifuging for 10min to obtain precipitate and secondary supernatant), and mixing the secondary supernatants to obtain supernatant containing rhizoma bletilla polysaccharide.

S3, adding 0.001g of alpha-amylase and 0.001g of beta-glucosidase into the bletilla polysaccharide clear liquid; stirring for 5 hours at 55 ℃ to obtain an enzymolysis liquid; adding ethanol into the enzymolysis solution to reach the concentration of 30%; preserving heat at 70 ℃ for 30min, then cooling to room temperature at 10000r/min, and centrifuging for 10min to obtain supernatant of the enzymolysis solution.

S4, concentrating the supernatant of the enzymolysis liquid until the ethanol content is less than 1%, and then treating the concentrated liquid by a membrane core (the molecular weight cut-off of the membrane core is 5000 Da); the trapped liquid is viscous liquid, namely bletilla striata polysaccharide-1; the permeate is oligosaccharide, namely bletilla striata polysaccharide-2.

The average molecular weight of bletilla striata polysaccharide-1 is 160000 Da;

the average molecular weight of bletilla striata polysaccharide-2 is 650 Da.

The above description is only a preferred embodiment of the present invention, and is not intended to limit the present invention in other forms, and any person skilled in the art may modify or change the technical content of the above disclosure into equivalent embodiments with equivalent changes, but all those simple modifications, equivalent changes and modifications made to the above embodiments according to the technical spirit of the present invention still belong to the protection scope of the present invention.