A high-efficiency extraction method of a large molecular weight astragalus polysaccharide

Abstract

The application discloses a high-efficiency extraction method of macromolecular astragalus polysaccharide, and belongs to the technical field of plant extraction, and comprises the following steps: cleaning and filtering astragalus membranaceus, and crushing the astragalus membranaceus; mixing a beta-1, 3-glucanase aqueous solution and a trehalose-based ethylenediamine, and stirring for 10-30 min to obtain an enzyme protection system; soaking the astragalus membranaceus powder in a citric acid solution, adding the enzyme protection system, applying a bipolar pulse electric field, and reacting at 40-60 DEG C for 30-40 min; adjusting the ethanol concentration to 40%-50%, and collecting crude astragalus polysaccharide by standing and centrifugation; purifying, decolorizing, concentrating, freeze-drying and crushing the crude astragalus polysaccharide to obtain refined astragalus polysaccharide; and the mass ratio of the trehalose-based ethylenediamine and the beta-1, 3-glucanase is (15:1)-(30:1). The enzyme protection system maintains the activity of the enzyme, the enzyme and the pulse electric field are synergistically used, the extraction time is shortened, the macromolecular astragalus polysaccharide is obtained, and the extraction rate is improved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of plant extraction, in particular to a high-efficiency extraction method of macromolecular astragalus polysaccharide. BACKGROUND

[0002] Radix Astragali is the dried root of Astragalus membranaceus (Fisch) Bge. Var. mongholicus (Bge) Hsiao. or A. membranaceus (Fisch) Bge., belonging to the family Leguminosae, and is a tonic medicine. Astragalus polysaccharide is one of the main components of Radix Astragali and has a wide range of pharmacological effects.

[0003] The main methods for extracting and separating astragalus polysaccharide from Radix Astragali are water extraction and alcohol precipitation and enzymatic hydrolysis. The water extraction and alcohol precipitation method can destroy the structure of polysaccharide, reduce the activity of polysaccharide, and reduce the molecular weight and viscosity after long-time high-temperature cooking. Although the combination of the complex enzyme method and the water extraction and alcohol precipitation method can improve the yield, it is difficult to penetrate the cell wall due to the woody structure of Radix Astragali, the enzymatic hydrolysis efficiency is low, and the time is long. Moreover, the above-mentioned extraction methods are not conducive to obtaining macromolecular astragalus polysaccharide.

[0004] Various washing and caring products on the market usually add thickening agents to adjust the viscosity of the emulsion, and mostly rely on synthetic high-molecular polymers (such as xanthan gum, carbomer, etc.). There is a risk of irritation (such as barrier damage caused by pH-dependent carbomer), and it may also cause allergic problems. Therefore, it is crucial to use naturally occurring substances as thickening agents. Macromolecular astragalus polysaccharide, as a naturally occurring substance, has a long molecular chain and can form a dense network structure, which can avoid skin irritation and is a good thickening agent. Therefore, it is particularly important to efficiently extract macromolecular astragalus polysaccharide from Radix Astragali.

[0005] In view of the problems existing in the prior art, the present application combines years of design and use experience in the relevant field to design a high-efficiency extraction method of macromolecular astragalus polysaccharide to overcome the above-mentioned defects. SUMMARY

[0006] To solve the problems existing in the prior art, the present application provides a high-efficiency extraction method of macromolecular astragalus polysaccharide, which can significantly improve the stability of enzymes in a pulsed electric field, shorten the extraction time, reduce the degradation of astragalus polysaccharide, maintain its high-molecular structure, and can be used as a moisturizing agent and a thickening agent in emulsion, reducing irritation and allergy.

[0007] In order to achieve the above-mentioned purposes, the technical scheme adopted by the present application is as follows: a high-efficiency extraction method of macromolecular astragalus polysaccharide, comprising the following steps:

[0008] (1) Astragalus pretreatment: the astragalus is sequentially cleaned with an acidic solution and deionized water, and then filtered, dried and crushed;

[0009] (2) Enzyme protection system construction: 0.1-1.0 mg / mL of a β-1,3-glucanase aqueous solution and trehalose-based ethylenediamine are mixed to obtain an enzyme protection system after stirring for 10-30 min, and the β-1,3-glucanase enzyme activity is 100,000 U / g;

[0010] (3) Polysaccharide extraction: the astragalus powder of step (1) is soaked in a citric acid solution, the enzyme protection system is added, and a bipolar pulsed electric field is applied, and the reaction is carried out at 40-60°C for 30-40 min;

[0011] (4) Alcohol precipitation: 95% ethanol solution is used to adjust the ethanol concentration of the reaction system of step (3) to 40%-50% to precipitate astragalus polysaccharides, and the precipitate is collected by centrifugation;

[0012] (5) Purification: the crude astragalus polysaccharides are purified, decolorized, concentrated, freeze-dried, and crushed to obtain refined astragalus polysaccharides;

[0013] The mass ratio of the trehalose-based ethylenediamine and the β-1,3-glucanase is (15:1)-(30:1).

[0014] Preferably, the trehalose-based ethylenediamine is synthesized according to the following steps, and the following raw materials are by mass fraction:

[0015] S1, under argon, 1-3 parts of trehalose and 4-10 parts of anhydrous pyridine are added and uniformly mixed, the system temperature is controlled to be ≤5°C, 0.5-1.5 parts of p-toluenesulfonyl chloride is slowly added dropwise, and stirring is carried out for 2-3 h;

[0016] S2, 10-30 parts of ice water are slowly added to quench the excess p-toluenesulfonyl chloride, stirring is carried out for 10 min, 3 times of extraction is carried out with 5-15 parts of dichloromethane, the organic phases are combined, dried and water-free, and white solid crude product is obtained by vacuum rotary evaporation at 40°C, and column chromatography purification is carried out to obtain white solid;

[0017] S3, 1-2 parts of the white solid are taken, 3-6 parts of DMF are added to completely dissolve, 0.2-0.3 parts of ethylenediamine is slowly added and uniformly mixed, stirring is carried out at 40-50°C for 4-6 h, the temperature is cooled to room temperature, DMF and excess ethylenediamine are removed by vacuum rotary evaporation, 5 parts of deionized water is added to dissolve the residue, 3 times of extraction is carried out with 5 parts of ethyl acetate, and the lower aqueous phase is retained;

[0018] S4, the aqueous phase is concentrated at 45°C under vacuum to completely volatilize the water, and light yellow solid trehalose-based ethylenediamine is obtained.

[0019] Preferably, the column chromatography purification in step S2 specifically comprises the following steps: dissolving the white solid crude product in dichloromethane / petroleum ether with a volume ratio of 1:2 and adding to a column filled with silica gel with a mesh of 200-300, eluting with petroleum ether and ethyl acetate with a volume ratio of 3:1 as eluent, concentrating and drying by rotary evaporation to obtain white solid.

[0020] Preferably, 2-3 parts of anhydrous sodium sulfate is added for drying in step S2.

[0021] Preferably, in step (1), the acidic solution is a citric acid solution with a pH of 5-6.

[0022] Preferably, the pH of the citric acid solution in step (3) is 5-6.

[0023] Preferably, the pulverized astragalus membranaceus in step (1) has a mesh of 80.

[0024] Preferably, the electric field strength of the bipolar pulsed electric field is 21-29 kV / cm, the pulse width is 15-25 μs, the positive / negative pulse ratio is 1:1, and the frequency is 100 Hz.

[0025] Preferably, the purification process in step (5) specifically comprises the following steps:

[0026] A1, adding deionized water to the crude astragalus polysaccharide until it just dissolves to obtain a sample solution, adding the sample solution to a macroporous adsorption resin chromatographic column, the column loading speed is 0.5-2 mL / min, eluting with 30% ethanol solution and collecting the eluate, the elution speed is 0.5-0.8 mL / min;

[0027] A2, evaporating the eluate and dissolving in water, adding activated carbon to decolorize the astragalus polysaccharide solution to be colorless and transparent, then reducing pressure to concentrate, freeze-drying and pulverizing to 200 mesh to obtain refined astragalus polysaccharide;

[0028] The activated carbon accounts for 0.1%-1% of the astragalus polysaccharide solution by weight.

[0029] Preferably, in step (3), the β-1,3-glucanase is added in an amount of 30-40 U / mg based on the mass of the astragalus powder.

[0030] Preferably, in step (3), the solid-liquid ratio of the astragalus powder and the citric acid solution is (1:15)-(1:30).

[0031] The present application has the following advantages:

[0032] 1.The present application utilizes trehalose-based ethylenediamine to bind with beta-1,3-glucanase, forming a "molecular shield" on the surface of beta-1,3-glucanase, effectively maintaining the spatial structure and activity of beta-1,3-glucanase, and synergizing with pulsed electric fields; pulsed electric fields form reversible or irreversible micropores (electroporation effect) on the cell wall of Astragalus membranaceus, causing the membrane structure to vibrate and break, promoting the release of substances such as Astragalus polysaccharide encapsulated in the cell wall; beta-1,3-glucanase (5.2 nm*4.1 nm) penetrates into the cell through the micropores, directly contacts the substrate for enzymatic hydrolysis, and obtains high extraction yield of Astragalus polysaccharide in a short time, significantly reduces the structural degradation of Astragalus polysaccharide, and obtains Astragalus polysaccharide with large molecular weight, with a molecular weight of 50-500 wDa accounting for more than 52.73%, and an extraction yield of more than 16.92%.

[0033] 2.In the present application, the viscosity of 1% Astragalus polysaccharide solution is 5000-9000 mPa.s, and Astragalus polysaccharide can be used as a moisturizing agent and thickening agent in emulsions at the same time. The large molecular structure of Astragalus polysaccharide can significantly improve the moisturizing efficiency, and the skin feels smooth and not greasy, which can improve the skin feel and texture of the formula, and at the same time, it can absorb excess oil on the skin, reducing the greasy feeling caused by oily ingredients in serums, emulsions and creams. BRIEF DESCRIPTION OF DRAWINGS

[0034] Figure 1 The synthesis route of trehalose-based ethylenediamine in the present application. DETAILED DESCRIPTION

[0035] In order to facilitate the understanding of those skilled in the art, the present application will be further described below in conjunction with specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present application and do not limit the present application.

[0036] The present application provides a high-efficiency extraction method of Astragalus polysaccharide with large molecular weight, comprising the following steps:

[0037] (1) Astragalus membranaceus pretreatment: sequentially clean Astragalus membranaceus with an acidic solution with a pH of 5-6 and deionized water, filter, and then dry and crush the Astragalus membranaceus to 80 mesh;

[0038] (2) Enzyme protection system construction: mix 0.1-1.0 mg / mL aqueous solution of beta-1,3-glucanase and trehalose-based ethylenediamine, stir for 10-30 min to obtain an enzyme protection system, the mass ratio of trehalose-based ethylenediamine to beta-1,3-glucanase is (15:1)-(30:1), and the beta-1,3-glucanase enzyme activity is 100,000 U / g;

[0039] (3) polysaccharide extraction: the astragalus powder is soaked in a citric acid solution with pH of 5-6, the solid-liquid ratio is (1:15)-(1:30), an enzyme protection system is added, a bipolar pulse electric field is applied, and the reaction is carried out at 40-60 DEG C for 30-40 min, the beta-1,3-glucanase is added in an amount of 30-40 U / mg based on the mass of the astragalus powder, and the beta-1,3-glucanase is added in an amount of 30-40 U per mg of the astragalus powder;

[0040] (4) alcohol precipitation: 95% ethanol solution is used to adjust the ethanol concentration of the reaction system in step (3) to 40%-50% to precipitate astragalus polysaccharide, and the precipitate is collected by centrifugation;

[0041] (5) purification: the crude astragalus polysaccharide is purified, decolorized, concentrated, freeze-dried, and crushed to obtain refined astragalus polysaccharide, the proportion of the astragalus polysaccharide with a molecular weight of 50-500 wDa is more than 52.73%, the viscosity of a 1% astragalus polysaccharide solution is more than 6189 mPa.s, the transparency of a 5% astragalus polysaccharide solution is more than 92.8%, the purity of the astragalus polysaccharide is more than 91.0%, and the extraction rate is more than 16.92%.

[0042] The citric acid solution is used as a solvent for extracting astragalus polysaccharide, which can avoid the presence of flavonoids in the extraction solution and reduce the influence of flavonoids on subsequent processing.

[0043] The pulse electric field can destroy the structural integrity of the cell wall of the astragalus powder, form reversible or irreversible micropores (electroporation effect) on the cell wall, the pore size is about 80-120 nm, the membrane structure is shaken and broken, the release of the internal wrapped astragalus polysaccharide and other substances is promoted, and the beta-1,3-glucanase (5.2 nm*4.1 nm) can penetrate into the cell through the micropores, directly contact the substrate and react, promote the extraction of the astragalus polysaccharide, and synergistically act.

[0044] When the beta-1,3-glucanase is directly added to the pulsed electric field, due to the action of the electric field force, the beta-1,3-glucanase may be destroyed in structure stability because of the conformational change of the active center, the stretching of the peptide chain and the abnormal folding, and further affect the activity of the beta-1,3-glucanase in the pulsed electric field, so that the enzymatic effect cannot be fully exerted. The present application adopts trehalose-based ethylenediamine to pretreat the beta-1,3-glucanase. Since the amino group of the trehalose-based ethylenediamine preferentially combines with the high negative charge region on the surface of the enzyme, i.e. the vicinity of aspartic acid, and the glutamic acid in the active center of the enzyme is located in the deep hydrophobic pocket, the amino group on the trehalose-based ethylenediamine cannot reach the deep amino acid residues (glutamic acid) of the active site due to the influence of the steric hindrance of the trehalose-based ethylenediamine, so the trehalose-based ethylenediamine can produce a “molecular shield” effect on the beta-1,3-glucanase, form protection for the beta-1,3-glucanase, and block the ion impact induced by the electric field, and at the same time, will not affect the activity of the beta-1,3-glucanase. The size of the enzyme protection system is 5.5nm*4.3nm, which can enter the inside of the cell through micropores. The present application can fully exert the effect of the beta-1,3-glucanase, so that the pulsed electric field and the beta-1,3-glucanase can synergistically act, greatly shorten the processing time of the radix astragali, reduce the damage to the molecular weight and structure of polysaccharides due to long-time extraction, and reduce the types and amount of enzymes used.

[0045] The synthesis of the trehalose-based ethylenediamine includes the following steps, referring to Figure 1 The following raw materials are all calculated by mass parts:

[0046] S1, under the condition of argon, 1-3 parts of trehalose and 4-10 parts of anhydrous pyridine are added and uniformly mixed, the system temperature is controlled to be ≤5℃, 0.5-1.5 parts of p-toluenesulfonyl chloride is slowly added dropwise, and stirring reaction is carried out for 2-3h;

[0047] S2, 10-30 parts of ice water is slowly added to quench the excess p-toluenesulfonyl chloride, stirring is carried out for 10min, 3 times of extraction is carried out with 5-15 parts of dichloromethane, the organic phases are combined, dried and water is removed, dichloromethane and residual pyridine solvents are removed by vacuum rotary evaporation at 40℃, white solid crude product is obtained, and column chromatography purification is carried out to obtain white solid;

[0048] S3, 1-2 parts of the white solid is taken, 3-6 parts of DMF is added to completely dissolve, 0.2-0.3 parts of ethylenediamine is slowly added and uniformly mixed, stirring is carried out at 40-50℃ for 4-6h, the temperature is cooled to room temperature, DMF and excess ethylenediamine are removed by vacuum rotary evaporation, 5 parts of deionized water is added to dissolve the residue, 3 times of extraction is carried out with 5 parts of ethyl acetate, and the lower aqueous phase is reserved;

[0049] S4, the aqueous phase is concentrated to completely volatilize the water by vacuum rotary evaporation at 45℃, and light yellow solid-trehalose-based ethylenediamine is obtained.

[0050] The column chromatography purification step specifically comprises: dissolving the white solid crude product in dichloromethane / petroleum ether mixture and adding to a column filled with silica gel with a particle size of 200-300 mesh, using petroleum ether and ethyl acetate with a volume ratio of 3:1 as eluent, concentrating and drying by rotary evaporation to obtain a white solid, and the volume ratio of dichloromethane to petroleum ether in the dichloromethane / petroleum ether mixture is 1:2.

[0051] As a preferred embodiment, the electric field strength of the bipolar pulse electric field in the present application is 21-29 kV / cm, the pulse width is 15-25 μs, the positive-to-negative pulse ratio is 1:1, and the frequency is 100 Hz. Electric field strength <20 kV / cm is insufficient for penetration, affecting the release of astragalus polysaccharides, and electric field strength >30 kV / cm is prone to electrolysis of astragalus polysaccharides, destroying its structure; pulse width <15 μs is not thorough for wall breaking, reducing the number of micropores, and pulse width >25 μs is prone to local overheating, causing astragalus polysaccharides to degrade and reduce molecular weight. The acid solution in step (1) is a citric acid solution,

[0052] The purification process in step (5) specifically comprises the following steps:

[0053] A1, adding deionized water to the crude astragalus polysaccharides until it just dissolves to obtain a sample solution, adding the sample solution to a macroporous adsorption resin chromatographic column, the column loading flow rate is 0.5-2 mL / min, eluting with 30% ethanol solution and collecting the eluate, the elution flow rate is 0.5-0.8 mL / min, and the ratio of eluent to sample solution is (400-1000) mL:1 g;

[0054] A2, evaporating the eluate to dryness and dissolving in water, adding activated carbon to decolorize the astragalus polysaccharide solution to colorless and transparent, then concentrating under reduced pressure, freeze-drying and crushing to 200 mesh to obtain refined astragalus polysaccharides, the activated carbon accounts for 0.1%-1% of the astragalus polysaccharide solution by weight, and the macroporous adsorption resin in the macroporous adsorption resin chromatographic column is preferably AB-8.

[0055] The specific implementation is shown as follows:

[0056] Transparency test method: add 0.50 g of refined astragalus polysaccharides to 99.50 g of deionized water, stir with a magnetic stirrer to disperse, add 3% citric acid (based on the mass of refined astragalus polysaccharides), and then stir with a magnetic stirrer until the viscosity is released, keep the temperature at 25°C for 2 h, and measure the light transmittance at 550 nm.

[0057] Viscosity measurement method: add 1 g of refined astragalus polysaccharides to 99 g of deionized water, stir with a magnetic stirrer to disperse, add 3% citric acid (based on the mass of refined astragalus polysaccharides), and then stir with a magnetic stirrer until the viscosity is released, keep the temperature at 25°C for 2 h, and measure the viscosity with a Brookfield viscometer.

[0058] Method for testing the proportion of the molecular weight of 50-500wDa in Huangqi polysaccharide: 5g refined Huangqi polysaccharide is dissolved in 100g deionized water, and ultrafiltration is performed through a 50wDa ultrafiltration membrane. The retentate is dried and the proportion in Huangqi polysaccharide is calculated.

[0059] The purity of Huangqi polysaccharide in the extract is determined according to the phenol-sulfuric acid method. Example 1

[0060] Trehalose-based ethylenediamine is synthesized according to the following steps, and the following raw materials are by mass fraction:

[0061] S1, under argon, add 1 part of trehalose and 5 parts of anhydrous pyridine and mix evenly, control the system temperature ≤5℃, slowly drop 0.5 parts of p-toluenesulfonyl chloride, stir for 2h;

[0062] S2, slowly add 10 parts of ice water to quench the excess p-toluenesulfonyl chloride, stir for 10min, extract 3 times with 5 parts of dichloromethane, combine the organic phase, add 2 parts of anhydrous sodium sulfate to dry, filter out the drying agent, evaporate the dichloromethane and residual pyridine solvent at 40℃ under vacuum rotary evaporation, to obtain white solid crude product;

[0063] Column chromatography purification is performed on the crude product, the white solid crude product is dissolved with dichloromethane / petroleum ether (volume ratio 1:2) and added to a column filled with 200 mesh silica gel, petroleum ether and ethyl acetate (volume ratio 3:1) are used as eluent, rotary evaporation is performed to concentrate and dry, to obtain white solid pure product;

[0064] S3, take 1 part of white solid, add 3 parts of DMF to completely dissolve, slowly add 0.2 parts of ethylenediamine and mix, stir at 40℃ for 4h, cool to room temperature, remove DMF and excess ethylenediamine by vacuum rotary evaporation, add 5 parts of deionized water to dissolve the residue, extract 3 times with 5 parts of ethyl acetate, and reserve the lower aqueous phase;

[0065] S4, the aqueous phase is concentrated by vacuum rotary evaporation at 45℃ until the water is completely evaporated, to obtain light yellow solid - trehalose-based ethylenediamine.

[0066] A high-efficiency extraction method of macromolecular Huangqi polysaccharide

[0067] (1) Huangqi pretreatment: the Huangqi is sequentially cleaned with a citric acid solution with pH 5 and deionized water, filtered, and then dried and crushed to 80 mesh;

[0068] (2) Enzyme protection system construction: mix 0.5mg / mL aqueous solution of β-1,3-glucanase and trehalose-based ethylenediamine, stir for 30min to obtain the enzyme protection system, the mass ratio of β-1,3-glucanase and trehalose-based ethylenediamine is 1:20, and the enzyme activity of β-1,3-glucanase is 100,000 U / g;

[0069] (3) polysaccharide extraction: soak the astragalus powder in a citric acid solution with pH 5, the solid-liquid ratio is 1:15, add an enzyme protection system, apply a bipolar pulsed electric field, react at 40℃ for 30 min, the β-1, 3-glucanase is added in an amount of 30 U / mg based on the mass of the astragalus powder, the electric field intensity is 25 V / cm, the pulse width is 20 μs, the positive-to-negative pulse ratio is 1:1, and the frequency is 100 Hz;

[0070] (4) alcohol precipitation: adjust the ethanol concentration of the reaction system of step (3) to 45% using a 95% ethanol solution to precipitate the astragalus polysaccharide, stand still, and collect the precipitated crude astragalus polysaccharide by centrifugation;

[0071] (5) purification: add deionized water to the crude astragalus polysaccharide until it just dissolves to obtain a sample solution, and then add the sample solution to a macroporous adsorption resin chromatographic column, the column loading speed is 0.5 mL / min, elute with a 30% ethanol solution, and collect the eluate, the elution speed is 0.6 mL / min;

[0072] After the eluate is evaporated, it is dissolved in water, decolorized with activated carbon until the astragalus polysaccharide solution is colorless and transparent, the activated carbon accounts for 1% of the weight of the astragalus polysaccharide solution, then it is concentrated under reduced pressure, freeze-dried, and pulverized to 200 mesh to obtain refined astragalus polysaccharide, wherein the macroporous adsorption resin is AB-8, and the ratio of the eluent to the sample solution is 400 mL:1 g.

[0073] In this embodiment, the extraction rate of astragalus polysaccharide is 16.92%, the content of astragalus polysaccharide is 92.4%, the viscosity of 1% astragalus polysaccharide solution is 6530 mPa.s, the transparency of 5% astragalus polysaccharide solution reaches 94.5%, and the proportion of molecular weight in the range of 50-500 wDa is 59.61%. Example 2

[0074] S1, under argon, add 2 parts of trehalose and 7 parts of anhydrous pyridine and mix uniformly, control the system temperature ≤5℃, slowly add 0.7 parts of p-toluenesulfonyl chloride, stir for 2h;

[0075] S2, slowly add 15 parts of ice water to quench the excess p-toluenesulfonyl chloride, stir for 10 min, extract 3 times with 10 parts of dichloromethane, combine the organic phases, add 3 parts of anhydrous sodium sulfate for drying, filter out the drying agent, and evaporate the dichloromethane and residual pyridine solvents under vacuum at 40℃ to obtain a white solid crude product;

[0076] Column chromatography purification is performed on the crude product, the white solid crude product is dissolved in dichloromethane / petroleum ether (volume ratio 1:2) and added to a column filled with 200 mesh silica gel, petroleum ether and ethyl acetate with a volume ratio of 3:1 are used as eluent, rotary evaporation is performed for concentration and drying to obtain a white solid;

[0077] S3, take white solid 1 part, add 3 parts of DMF to completely dissolve, slowly add ethylenediamine 0.3 parts, mix, stir at 45 DEG C for 5h, cool to room temperature, remove DMF and excess ethylenediamine by vacuum rotary evaporation, add deionized water 5 parts to dissolve the residue, extract with 5 parts of ethyl acetate 3 times, retain the lower aqueous phase;

[0078] S4, the aqueous phase is concentrated to completely evaporate the water at 45 DEG C by vacuum rotary evaporation, and a light yellow solid-trehalose-based ethylenediamine is obtained.

[0079] A high-efficiency extraction method of large molecular weight astragalus polysaccharide

[0080] (1) Astragalus pretreatment: Astragalus is sequentially cleaned with citric acid solution with pH 6 and deionized water, filtered, dried and crushed to 80 mesh after filtration;

[0081] (2) Enzyme protection system construction: 1.0 mg / mL aqueous solution of β-1, 3-glucanase and trehalose-based ethylenediamine are mixed to obtain an enzyme protection system after stirring for 20 min, the mass ratio of β-1, 3-glucanase and trehalose-based ethylenediamine is 1:15, and the β-1, 3-glucanase enzyme activity is 100,000 U / g;

[0082] (3) Polysaccharide extraction: the astragalus powder is soaked in citric acid solution with pH 6, the solid-liquid ratio is 1:30, the enzyme protection system is added, and a bipolar pulse electric field is applied, the β-1, 3-glucanase addition amount is 40 U / mg based on the mass of astragalus powder, the electric field strength is 21 V / cm, the pulse width is 15 μs, the positive-negative pulse ratio is 1:1, and the frequency is 100 Hz at 60 DEG C for 35 min;

[0083] (4) Alcohol precipitation: 95% ethanol solution is used to adjust the ethanol concentration of the reaction system of step (3) to 50% to precipitate astragalus polysaccharide, and the precipitate crude astragalus polysaccharide is collected by centrifugation after standing;

[0084] (5) Purification: deionized water is added to the crude astragalus polysaccharide to just dissolve to obtain a sample solution, the sample solution is added to a macroporous adsorption resin chromatographic column, the column flow rate is 2 mL / min, 30% ethanol solution is used for elution and the eluate is collected, the elution flow rate is 0.5 mL / min, and the ratio of eluent to sample solution is 800 mL:1 g;

[0085] After the eluate is evaporated, it is dissolved in water, activated carbon is added for decolorization to make the astragalus polysaccharide solution colorless and transparent, the activated carbon accounts for 0.1% of the weight of the astragalus polysaccharide solution, then it is concentrated under reduced pressure, freeze-dried and crushed to 200 mesh to obtain fine astragalus polysaccharide, and the macroporous adsorption resin is AB-8.

[0086] The extraction rate of astragalus polysaccharide in the embodiment is 18.15%, the content of astragalus polysaccharide is 91.9%, the viscosity of 1% astragalus polysaccharide solution is 6189 mPa.s, the transparency of 5% astragalus polysaccharide solution reaches 93.6%, and the molecular weight of more than 50-500 wDa accounts for 52.73%. Example 3

[0087] Trehalose-based ethylenediamine is synthesized according to the following steps, and the following raw materials are calculated by mass parts:

[0088] S1, under the condition of argon, 3 parts of trehalose and 10 parts of anhydrous pyridine are added and uniformly mixed, the temperature of the system is controlled to be ≤5℃, 1 part of p-toluenesulfonyl chloride is slowly added dropwise, and stirring reaction is carried out for 2 hours;

[0089] S2, 10 parts of ice water are slowly added to quench the excess p-toluenesulfonyl chloride, stirring is carried out for 10 minutes, 5 parts of dichloromethane are extracted for 3 times, the organic phase is combined, 3 parts of anhydrous sodium sulfate is added for drying, the drying agent is filtered out, and dichloromethane and residual pyridine solvents are evaporated by vacuum rotary evaporation at 40℃, to obtain white solid crude product;

[0090] The crude product is purified by column chromatography, the white solid crude product is dissolved with dichloromethane / petroleum ether (volume ratio 1:2) and added to a column filled with 200 mesh silica gel, petroleum ether and ethyl acetate with a volume ratio of 3:1 are used as eluent, rotary evaporation is carried out for concentration and drying, and white solid is obtained;

[0091] S3, 1.5 parts of white solid are taken, 5 parts of DMF are added to completely dissolve, 0.3 parts of ethylenediamine are slowly added and uniformly mixed, stirring is carried out at 45℃ for 6 hours, the temperature is cooled to room temperature, DMF and excess ethylenediamine are removed by vacuum rotary evaporation, 5 parts of deionized water are added to dissolve the residue, 5 parts of ethyl acetate are extracted for 3 times, and the lower aqueous phase is reserved;

[0092] S4, the aqueous phase is concentrated by vacuum rotary evaporation at 45℃ until the water is completely evaporated, to obtain light yellow solid-trehalose-based ethylenediamine.

[0093] A high-efficiency extraction method of macromolecular astragalus polysaccharide

[0094] (1) Astragalus pretreatment: the astragalus is sequentially cleaned with a citric acid solution with a pH of 5 and deionized water, filtered, dried and crushed to 80 mesh after filtration;

[0095] (2) Construction of enzyme protection system: 1.0 mg / mL of β-1,3-glucanase aqueous solution and trehalose-based ethylenediamine are mixed to obtain an enzyme protection system after stirring for 10 minutes, the mass ratio of β-1,3-glucanase and trehalose-based ethylenediamine is 1:30, and the enzyme activity of β-1,3-glucanase is 100,000 U / g;

[0096] (3) polysaccharide extraction: soak the astragalus powder in a citric acid solution with pH of 5, the solid-liquid ratio is 1:30, add the enzyme protection system, apply a bipolar pulsed electric field, react at 50℃ for 40min, the β-1, 3-glucanase is added at 35U / mg of the astragalus powder, the electric field intensity is 29V / cm, the pulse width is 25μs, the positive-negative pulse ratio is 1:1, and the frequency is 100Hz;

[0097] (4) alcohol precipitation: adjust the ethanol concentration of the reaction system of step (3) to 50% by using 95% ethanol solution to precipitate the astragalus polysaccharide, stand still, and centrifuge to collect the precipitated crude astragalus polysaccharide;

[0098] (5) purification: add deionized water to the crude astragalus polysaccharide until it just dissolves to obtain a sample solution, add the sample solution to a macroporous adsorption resin chromatographic column, the column loading speed is 0.6mL / min, elute with 30% ethanol solution and collect the eluate, the elution speed is 0.8mL / min, and the ratio of eluent to sample solution is 1000mL:1g;

[0099] After the eluate is evaporated, it is dissolved in water, activated carbon is added for decolorization to make the astragalus polysaccharide solution colorless and transparent, the activated carbon accounts for 0.5% of the weight of the astragalus polysaccharide solution, then it is concentrated under reduced pressure, freeze-dried and pulverized to 200 mesh to obtain fine astragalus polysaccharide, and the macroporous adsorption resin is AB-8.

[0100] In this embodiment, the extraction rate of astragalus polysaccharide is 17.98%, the content of astragalus polysaccharide is 91.0%, the viscosity of 1% astragalus polysaccharide solution is 6291mPa.s, the transparency of 5% astragalus polysaccharide solution reaches 93.4%, and the molecular weight of 50-500wDa accounts for 53.5%. Example 4

[0101] Trehalose-based ethylenediamine is synthesized according to the following steps, and the following raw materials are calculated by mass fraction:

[0102] S1, under the condition of argon, add trehalose 1 part and anhydrous pyridine 4 parts and mix uniformly, control the system temperature ≤5℃, slowly drop p-toluenesulfonyl chloride 1.5 parts, and stir for 3h;

[0103] S2, slowly add 30 parts of ice water to quench the excess p-toluenesulfonyl chloride, stir for 10min, extract with 10 parts of dichloromethane for 3 times, combine the organic phase, add 3 parts of anhydrous sodium sulfate for drying, filter out the drying agent, and evaporate the dichloromethane and residual pyridine solvent at 40℃ under vacuum rotary evaporation to obtain white solid crude product;

[0104] The white solid crude product was dissolved in dichloromethane / petroleum ether (1:2 by volume) and added to a column filled with 300-mesh silica gel, eluted with petroleum ether and ethyl acetate (3:1 by volume), concentrated by rotary evaporation and dried to obtain a white solid;

[0105] S3, 2 parts of white solid were taken, 6 parts of DMF were added to completely dissolve, 0.25 parts of ethylenediamine were slowly added and mixed, stirred at 50°C for 6h, cooled to room temperature, and the DMF and excess ethylenediamine were removed by vacuum rotary evaporation, 5 parts of deionized water were added to dissolve the residue, and the lower aqueous phase was retained by extracting 3 times with 5 parts of ethyl acetate;

[0106] S4, the aqueous phase was concentrated by vacuum rotary evaporation at 45°C until the water was completely evaporated to obtain a light yellow solid-trehalose-based ethylenediamine.

[0107] A high-efficiency extraction method of a large-molecular-weight astragalus polysaccharide

[0108] (1) Astragalus pretreatment: The astragalus was sequentially cleaned with an acidic solution with a pH of 6 and deionized water, filtered, and then dried and crushed to 80 mesh;

[0109] (2) Enzyme protection system construction: 1 mg / mL of a β-1,3-glucanase aqueous solution and trehalose-based ethylenediamine were mixed to obtain an enzyme protection system after stirring for 30 min, the mass ratio of β-1,3-glucanase to trehalose-based ethylenediamine was 1:30, and the β-1,3-glucanase enzyme activity was 100,000 U / g;

[0110] (3) Polysaccharide extraction: The astragalus powder was soaked in a citric acid solution with a pH of 5, the solid-liquid ratio was 1:20, the enzyme protection system was added, a bipolar pulse electric field was applied, the reaction was carried out at 40°C for 40 min, the β-1,3-glucanase addition amount was 40 U / mg based on the mass of the astragalus powder, the electric field strength was 25 V / cm, the pulse width was 20 μs, the positive-to-negative pulse ratio was 1:1, and the frequency was 100 Hz;

[0111] (4) Alcohol precipitation: 95% ethanol solution was used to adjust the ethanol concentration of the reaction system of step (3) to 50% to precipitate astragalus polysaccharide, and the precipitate was collected by centrifugation;

[0112] (5) Purification: Deionized water was added to the crude astragalus polysaccharide to just dissolve to obtain a sample solution, the sample solution was added to a macroporous adsorption resin chromatographic column, the column loading flow rate was 0.8 mL / min, 30% ethanol solution was used for elution, and the eluate was collected, the elution flow rate was 0.8 mL / min, and the ratio of eluent to sample solution was 600 mL:1 g;

[0113] The eluent was evaporated to dryness, dissolved in water, and decolorized with activated carbon to make the atractylodes rhizome polysaccharide solution colorless and transparent, the activated carbon accounting for 0.5% of the weight of the atractylodes rhizome polysaccharide solution, then concentrated under reduced pressure, freeze-dried, and pulverized to 200 meshes to obtain refined atractylodes rhizome polysaccharide, wherein the macroporous adsorption resin is AB-8.

[0114] In this embodiment, the extraction rate of atractylodes rhizome polysaccharide is 17.45%, the content of atractylodes rhizome polysaccharide is 92.1%, the viscosity of 1% atractylodes rhizome polysaccharide solution is 6217 mPa.s, the transparency of 5% atractylodes rhizome polysaccharide solution reaches 92.8%, and the molecular weight is 50-500 wDa, accounting for 54.96%.

[0115] Comparative Example 1

[0116] This comparative example extracts atractylodes rhizome polysaccharide according to the method of Example 1, except that the β-1, 3-glucanase is not treated, step (2) is omitted, and β-1, 3-glucanase is used instead of the enzyme protection system in step (3), and the amount of β-1, 3-glucanase in the system is kept unchanged.

[0117] Comparative Example 2

[0118] Commercial atractylodes rhizome polysaccharide

[0119] Comparative Example 3

[0120] This comparative example extracts atractylodes rhizome polysaccharide according to the steps of Example 1, except that the electric field parameters of the pulsed electric field are different, the electric field strength is 15 V / cm, the pulse width is 70 μs, the positive-to-negative pulse ratio is 1:1, and the frequency is 100 Hz.

[0121] Comparative Example 4

[0122] (1) Atractylodes rhizome pretreatment: The atractylodes rhizome is sequentially cleaned with an acidic solution with a pH of 5 and deionized water, filtered, and then dried and pulverized to 80 mesh;

[0123] (2) Polysaccharide extraction: The atractylodes rhizome powder is soaked in a citric acid solution with a pH of 5, the solid-liquid ratio is 1:15, a bipolar pulsed electric field is applied, the reaction is carried out at 40°C for 30 min, the electric field strength is 25 V / cm, the pulse width is 20 μs, the positive-to-negative pulse ratio is 1:1, and the frequency is 100 Hz;

[0124] (3) Alcohol precipitation: 95% ethanol solution is used to adjust the ethanol concentration of the reaction system to 45% to precipitate atractylodes rhizome polysaccharide, and the precipitate is collected by centrifugation;

[0125] (4) Purification: Deionized water is added to the crude atractylodes rhizome polysaccharide to just dissolve it to obtain the sample solution, and the sample solution is added to a macroporous adsorption resin chromatographic column, the flow rate is 0.5 mL / min, 30% ethanol solution is used for elution, and the eluate is collected, the flow rate is 0.6 mL / min;

[0126] The eluent was evaporated to dryness, dissolved in water, and activated carbon was added to decolorize the astragalus polysaccharide solution to colorless and transparent. The activated carbon accounted for 1% of the weight of the astragalus polysaccharide solution. Then, the solution was concentrated under reduced pressure, freeze-dried, and ground to 200 mesh to obtain fine astragalus polysaccharide. The macroporous adsorption resin was AB-8, and the ratio of eluent to sample solution was 400 mL: 1 g.

[0127] Comparative Example 5

[0128] (1) Astragalus pretreatment: The astragalus was sequentially cleaned with an acidic solution with a pH of 5 and deionized water, and then filtered. The astragalus was dried and ground to 80 mesh;

[0129] (2) Polysaccharide extraction: The astragalus powder was soaked in a citric acid solution with a pH of 5, with a solid-liquid ratio of 1:15. β-1, 3-glucanase without trehalose-based ethylenediamine treatment was added, with an enzyme activity of 100,000 U / g. The β-1, 3-glucanase was added at a dosage of 30 U / mg based on the mass of the astragalus powder, and the reaction was carried out at 40°C for 30 min.

[0130] (3) Alcohol precipitation: 95% ethanol solution was used to adjust the ethanol concentration of the reaction system to 40% to precipitate astragalus polysaccharide. After standing and centrifugation, the precipitated crude astragalus polysaccharide was collected.

[0131] (4) Purification: Deionized water was added to the crude astragalus polysaccharide to just dissolve the sample solution. The sample solution was added to a macroporous adsorption resin chromatographic column, with an upper column flow rate of 0.5 mL / min. The eluent was eluted with 30% ethanol solution and collected, with an elution flow rate of 0.6 mL / min.

[0132] The eluent was evaporated to dryness, dissolved in water, and activated carbon was added to decolorize the astragalus polysaccharide solution to colorless and transparent. The activated carbon accounted for 1% of the weight of the astragalus polysaccharide solution. Then, the solution was concentrated under reduced pressure, freeze-dried, and ground to 200 mesh to obtain fine astragalus polysaccharide. The macroporous adsorption resin was AB-8, and the ratio of eluent to sample solution was 400 mL: 1 g.

[0133] Performance test of astragalus polysaccharide:

[0134] 1. Physical properties

[0135] The physical properties of the astragalus polysaccharide of Examples 1-4 and Comparative Examples 1-5 were determined, as shown in Table 1:

[0136] Table 1 Physical property data of astragalus polysaccharide obtained by different extraction methods

[0137]

[0138] From the data in Table 1, it can be seen that the extraction method of the present application can obtain a higher extraction rate of astragalus polysaccharide in a short time, and the prepared astragalus polysaccharide has a relatively large molecular weight, reduces the pyrolysis degree, and to some extent, retains the molecular structure of the astragalus polysaccharide, and the proportion of the molecular weight in the range of 50-500 wDa is more than 52.73%. Compared with Example 1 and Comparative Example 1, it is shown that the present application successfully protects the β-1,3-glucanase by trehalose-based ethylenediamine, greatly retains the activity of the enzyme, significantly improves the extraction rate of the astragalus polysaccharide, and the extraction rate is improved by 36.2%. Compared with Example 1 and Comparative Example 2, the astragalus polysaccharide extracted by the present application has a higher purity while maintaining a large molecular weight. It can be known from the comparison between Example 1 and Comparative Example 3 that changing the electric field parameters not only reduces the extraction rate of the astragalus polysaccharide, but also reduces the molecular weight, which shows that the electric field parameters in the present application promote the extraction rate and high molecular weight of the astragalus polysaccharide. It can be known from the comparison between Comparative Example 4 and Comparative Example 5 that the β-1,3-glucanase and the pulse electric field play a synergistic role, which together improves the extraction rate of the astragalus polysaccharide, shortens the extraction time, and thus reduces the degradation of the structure of the astragalus polysaccharide. It can be known from the comparison between Comparative Example 1 and Comparative Example 4 that the β-1,3-glucanase in Comparative Example 1 has a small degree of degradation of the structure of the astragalus polysaccharide, and thus the viscosity and the proportion of the molecular weight in the range of 50-500 wDa of Comparative Example 1 and Comparative Example 4 are close. It can be known from the comparison between Comparative Example 3 and Comparative Example 5 that the large pulse width of Comparative Example 3 leads to the degradation of the structure of the astragalus polysaccharide, thereby producing the results that the viscosity and the proportion of the molecular weight in the range of 50-500 wDa of Comparative Example 5 are better than those of Comparative Example 3.

[0139] 2. Hydroxyl radical scavenging rate

[0140] Test steps: The astragalus polysaccharides of Example 1-Example 4 and Comparative Example 1-Comparative Example 4 were respectively prepared into a sample solution of 5 mg / mL with deionized water, 1 mL of the sample solution, 100 μL of ferrous sulfate solution, 100 μL of ethanol-salicylic acid solution, and an appropriate amount of isopropyl alcohol were sequentially added into a centrifugal tube, 100 μL of hydrogen peroxide was finally added, the total volume of the liquid in the centrifugal tube was 1.3 mL, it was shaken, and the absorbance at 510 nm was measured after being placed in a 37℃ water bath for 15 min, and the data was recorded, and the results are shown in Table 2.

[0141] Hydroxyl radical scavenging rate (%) = [A0-(A1-A2)] / A0;

[0142] Wherein A0 is the absorbance of the blank control without adding the sample solution; A1 is the absorbance of the sample; A2 is the hydrogen peroxide solution without adding the chromogenic agent;

[0143] Table 2 Hydroxyl radical scavenging rate (%)

[0144]

[0145] From the data in Table 2, it can be seen that the astragalus polysaccharide obtained by the present application has excellent hydroxyl radical scavenging activity at the same concentration.

[0146] 3. Moisturizing enhancement rate

[0147] Test method:

[0148] Instrument and standard: use a skin moisture content test probe in accordance with ISO 24442:2011 specification;

[0149] Test area: 3 areas of 3 cm x 3 cm on the inner side of the forearm, with a distance of at least 1 cm between adjacent areas on the same arm;

[0150] Test conditions: 80 healthy adults, aged 18-45 years old, female, randomly divided into 8 groups, 10 people in each group, sit quietly for 30 minutes in a constant temperature and humidity environment before testing, clean and dry the test area on the inner side of the forearm (avoid damaged parts);

[0151] Test sample: astragalus polysaccharides of Example 1-Example 4, Comparative Example 1-Comparative Example 4 are dissolved in water to prepare a sample solution of 5 mg / mL;

[0152] Test steps:

[0153] Measure the base moisture value (M0) of the untreated test area;

[0154] Take the test sample and evenly apply it to the test area, with a dosage of 2.0±0.1 mg / cm 2 , massage for 1 minute, and dry with a soft towel;

[0155] The subject remains seated, and the moisture value (M4) of the test area is measured after 4 hours;

[0156] Calculation formula: 4-hour stratum corneum moisture enhancement rate (%) = ;

[0157] The test results are shown in Table 3.

[0158] Table 3 Moisture retention performance

[0159]

[0160] From the data in Table 3, it can be seen that the astragalus polysaccharide extracted by the present application has good moisturizing performance and low water molecule loss rate, indicating that the astragalus polysaccharide molecules with large molecular weight in the present application have long molecular chains and complex structures, can form a dense protective film on the skin to reduce water evaporation, and can be used as a good moisturizing agent in emulsions.

[0161] 4. Thickening performance test

[0162] The Astragalus polysaccharide of Example 1, xanthan gum, carbomer and the Astragalus polysaccharide of Comparative Example 2 were taken as raw materials, and emulsions were prepared according to the raw material addition amount in Table 4 to form emulsion 1-emulsion 4, and the thickening performance was evaluated.

[0163] Table 4 Raw material ratio in emulsion

[0164]

[0165] Sensitivity test: the emulsion 1-emulsion 4 prepared in Table 4 were respectively tested for sensitivity behind the ears of the testers, and after determining that there was no adverse reaction, the evaluation was carried out;

[0166] Test conditions: 10 healthy adults, female volunteers aged 18-45 years old, formed an evaluation group for blind evaluation, and the feeling description and scores were recorded, with a full score of 10;

[0167] Test area: face;

[0168] Test procedure: the test sample was evenly applied to the test area, and the amount was 2.0±0.1 mg / cm 2 , and was massaged for 1 minute;

[0169] The test results are shown in Table 5.

[0170] Table 5 Thickening experiment data of emulsion

[0171]

[0172] From the use feeling comparison of emulsion 1 and emulsion 4, it can be seen that the skin feeling of the emulsion prepared from the Astragalus polysaccharide of the application is better than that of the emulsion prepared from the commercially available Astragalus polysaccharide, and is also better than that of the emulsion prepared from xanthan gum and carbomer. When the Astragalus polysaccharide of the application is used as a thickening agent, it can improve the skin feeling of the emulsion, the quality is smooth and not greasy, the greasy feeling caused by the oily ingredients in the emulsion is reduced, and the Astragalus polysaccharide is a natural extract, which can avoid skin irritation and allergic reactions caused by carbomer and the like.

[0173] In summary, the extraction method of the application can obtain Astragalus polysaccharide with high extraction rate in a short time, reduce the thermal degradation of Astragalus polysaccharide, and effectively maintain the macromolecular structure of Astragalus polysaccharide.

[0174] It should be understood that the use of these examples is only for the purpose of illustrating the present application and is not intended to limit the scope of protection of the present application. In addition, it should also be understood that after reading the technical content of the present application, those skilled in the art can make various modifications, modifications and / or variations to the present application, and all these equivalent forms also fall within the protection scope defined by the claims attached to the present application.

Claims

1. A highly efficient extraction method for high molecular weight Astragalus polysaccharides, characterized in that, Includes the following steps: (1) Pretreatment of Astragalus membranaceus: Astragalus membranaceus was washed with acidic solution and deionized water in sequence, filtered, dried in the shade and pulverized. (2) Construction of enzyme protection system: Mix 0.1-1.0 mg / mL of β-1,3-glucanase aqueous solution with trehalose ethylenediamine and stir for 10-30 min to obtain enzyme protection system. The enzyme activity of β-1,3-glucanase is 100,000 U / g. (3) Polysaccharide extraction: The Astragalus powder from step (1) was soaked in citric acid solution, an enzyme protection system was added, a bipolar pulsed electric field was applied, and the reaction was carried out at 40-60℃ for 30-40 min. (4) Alcohol precipitation: Use 95% ethanol solution to adjust the ethanol concentration of the reaction system in step (3) to 40%-50% to precipitate Astragalus polysaccharide, let it stand, and centrifuge to collect the precipitated crude Astragalus polysaccharide. (5) Purification: The crude Astragalus polysaccharide is purified, decolorized, concentrated, freeze-dried and pulverized to obtain refined Astragalus polysaccharide; The mass ratio of trehalose-sylethylenediamine to β-1,3-glucanase is (15:1) to (30:1). The electric field strength of the bipolar pulsed electric field is 21-29 kV / cm, the pulse width is 15-25 μs, the positive-to-negative pulse ratio is 1:1, and the frequency is 100 Hz.

2. The efficient extraction method for high molecular weight Astragalus polysaccharides according to claim 1, characterized in that, The trehalose-saccharidized ethylenediamine was synthesized according to the following steps, where all raw materials are by mass parts: S1, under argon atmosphere, add 1-3 parts of trehalose and 4-10 parts of anhydrous pyridine and mix well. Control the system temperature ≤5℃, slowly add 0.5-1.5 parts of p-toluenesulfonyl chloride, and react for 2-3 hours with stirring. S2, slowly add 10-30 parts of ice water to quench excess p-toluenesulfonyl chloride, stir for 10 min, extract three times with 5-15 parts of dichloromethane, combine the organic phases, dry to remove water, and vacuum rotary evaporate at 40℃ to obtain a white solid crude product, which is purified by column chromatography to obtain a white solid. S3, take 1-2 parts of white solid, add 3-6 parts of DMF until completely dissolved, slowly add 0.2-0.3 parts of ethylenediamine and mix well, stir at 40-50℃ for 4-6 hours, cool to room temperature, remove DMF and excess ethylenediamine by vacuum rotary evaporation, add 5 parts of deionized water to dissolve the residue, extract 3 times with 5 parts of ethyl acetate, and retain the lower aqueous phase; S4, the aqueous phase was concentrated under vacuum at 45°C until all water evaporated, yielding a pale yellow solid trehalose ethylenediamine.

3. The efficient extraction method for high molecular weight Astragalus polysaccharides according to claim 2, characterized in that, The column chromatography purification in step S2 specifically includes the following steps: the white solid crude product is dissolved in dichloromethane / petroleum ether at a volume ratio of 1:2 and added to a column packed with 200-300 mesh silica gel. It is eluted with petroleum ether and ethyl acetate at a volume ratio of 3:

1. The product is concentrated by rotary evaporation and dried to obtain a white solid.

4. The efficient extraction method for high molecular weight Astragalus polysaccharides according to claim 2, characterized in that, By weight, add 2-3 parts of anhydrous sodium sulfate in step S2 to dry and remove water, and filter to remove the desiccant.

5. The efficient extraction method for high molecular weight Astragalus polysaccharides according to claim 1, characterized in that, In step (1), the acidic solution is a citric acid solution with a pH of 5-6; And / or, the pH of the citric acid solution in step (3) is 5-6.

6. The efficient extraction method for high molecular weight Astragalus polysaccharides according to claim 1, characterized in that, The Astragalus membranaceus powder in step (1) is pulverized to a mesh size of 80.

7. The efficient extraction method for high molecular weight Astragalus polysaccharides according to claim 1, characterized in that, The purification process in step (5) is detailed below. Includes the following steps: A1. Add deionized water to the crude Astragalus polysaccharide until it is just dissolved to obtain the loading solution. Add the loading solution to the macroporous adsorption resin chromatography column at a flow rate of 0.5-2 mL / min. Elute with 30% ethanol solution and collect the eluent at a flow rate of 0.5-0.8 mL / min. A2. After evaporating the eluent to dryness, add water to dissolve it, add activated carbon to decolorize it until the Astragalus polysaccharide solution is colorless and transparent, then concentrate under reduced pressure, freeze dry and pulverize to 200 mesh to obtain refined Astragalus polysaccharide. The activated carbon accounts for 0.1%-1% of the Astragalus polysaccharide solution by weight.

8. The efficient extraction method for high molecular weight Astragalus polysaccharides according to claim 1, characterized in that, In step (3), the amount of β-1,3-glucanase added is 30-40 U / mg based on the mass of Astragalus powder.

9. The efficient extraction method for high molecular weight Astragalus polysaccharides according to claim 1, characterized in that, In step (3), the ratio of Astragalus powder to citric acid solution is (1:15) to (1:30).

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