Subcritical water extraction method and application of polygonatum sibiricum homogeneous polysaccharide

Abstract

The invention relates to the technical field of polygonatum sibiricum polysaccharide extraction, in particular to a polygonatum sibiricum homogeneous polysaccharide subcritical water extraction method and application, and the method comprises the following steps: crushing polygonatum sibiricum, adding a degreasing agent for treatment, and filtering; adding an ammonia water-ethanol solution, performing pretreatment, adding cellulase and hemicellulase for dissolving, and performing centrifugal separation to obtain filtrate and rhizoma polygonati residues; carrying out glycerol modified subcritical water extraction on the polygonatum sibiricum residues to obtain extract liquor; mixing the filtrate and the extract liquor, performing alcohol precipitation, centrifugation, freeze drying, dissolving with distilled water, loading a sample to an anionic cellulose chromatographic column, eluting with distilled water, further purifying with a gel chromatographic column, and eluting with a NaCl solution to obtain the polygonatum homogeneous polysaccharide. According to the method, a specific degreasing mode is combined with enzyme dissolution and subcritical water extraction, so that the extraction rate of the polygonatum sibiricum homogeneous polysaccharide is increased, impurities can be reduced, harmful substances are not used in the extraction process, and the prepared polygonatum sibiricum homogeneous polysaccharide is suitable for the fields of high-end health care products and medicines.

Description

Technical Field

[0001] The present application relates to the technical field of extraction of polygonatum polysaccharide, and in particular to a subcritical water extraction method and use of uniform polygonatum polysaccharide. Background Art

[0002] The Chinese medicine Polygonatum sibiricum is the dried rhizome of Polygonatum sibiricum, Polygonatum yunnanensis and Polygonatum multiflorum, which belong to the genus Polygonatum of the Liliaceae family. It has a long history of medicinal use. Polygonatum sibiricum polysaccharide is one of the most important active compounds in Polygonatum sibiricum. It has multiple biological activities such as anti-oxidation, immune regulation, anti-tumor, anti-osteoporosis, hypoglycemic, hypolipidemic and anti-atherosclerotic, and has become a hot spot in the development and application of Polygonatum sibiricum.

[0003] In order to extract active ingredients such as polysaccharides from Polygonatum sibiricum, various extraction methods have been developed, among which traditional methods using organic solvents are widely used. However, most of these traditional methods are time-consuming, energy-inefficient, and involve some organic solvents that are potentially toxic to the environment or human health.

[0004] Subcritical water has attracted widespread attention due to its environmental friendliness, short time consumption, no organic solvent residue and good ability to maintain the biological activity of polysaccharides. However, the existing subcritical water extraction method has a single extraction method and contains many impurities, and the extraction rate of polygonatum polysaccharides is still insufficient. Summary of the invention

[0005] The present application aims to overcome at least one of the defects of the prior art and provide a method and use for subcritical water extraction of uniform Polygonatum sibiricum polysaccharide. By combining a specific degreasing method with enzyme dissolution and subcritical water extraction, the extraction rate of uniform Polygonatum sibiricum polysaccharide can be improved while reducing the generation of impurities. No harmful substances are used in the extraction process. The prepared uniform Polygonatum sibiricum polysaccharide is suitable for use in high-end health products and the field of medicine.

[0006] In the first aspect, the present application provides a method for extracting uniform polysaccharides of polygonatum, which is achieved by the following technical solutions:

[0007] A method for extracting uniform polysaccharide from polygonatum sibiricum, comprising the following steps:

[0008] (1) washing and crushing the polygonatum sibiricum medicinal material, adding a degreasing agent to perform degreasing treatment at a temperature of 60-80° C., filtering, and obtaining defatted polygonatum sibiricum;

[0009] (2) adding the defatted polygonatum to an ammonia-ethanol solution, pre-treating for 2 hours, adding cellulase and hemicellulase to dissolve, and centrifuging to obtain a filtrate and a polygonatum residue;

[0010] (3) subjecting the polygonatum residue to glycerol-modified subcritical water extraction to obtain an extract;

[0011] (4) mixing the filtrate and the extract, and subjecting the mixture to alcohol precipitation, centrifugation, and freeze-drying to obtain crude polygonatum polysaccharide;

[0012] (5) After dissolving the crude polygonatum polysaccharide with distilled water, the mixture is loaded onto an anionic cellulose chromatography column, and the chromatography column is eluted with distilled water to obtain a water-washed component polysaccharide of polygonatum, and the water-washed component polysaccharide is further purified with a gel chromatography column, and the gel chromatography column is eluted with a NaCl solution to obtain a uniform polygonatum polysaccharide;

[0013] Wherein, the amount of glycerol used in step (3) is 40-60% of the weight of the solution;

[0014] The degreasing agent in step (1) comprises petroleum ether, hydrochloric acid and an adsorbent;

[0015] The amounts of the petroleum ether, the hydrochloric acid and the adsorbent are in a weight ratio of (5-8):(2-3):(0.5-2);

[0016] The raw materials for preparing the adsorbent include ferric chloride, volcanic mud and zirconium oxide in a weight ratio of 1:(2-4):(0.5-2).

[0017] According to a method for extracting uniform polysaccharide of polygonatum according to an embodiment of the present application, there are at least the following beneficial effects:

[0018] The present application uses glycerol-modified subcritical water extraction. Glycerol, as an organic solvent, can increase the solubility of Polygonatum sibiricum polysaccharide in subcritical water, so that the Polygonatum sibiricum polysaccharide, which originally has a low solubility in pure water, can be more easily extracted, thereby improving the extraction efficiency. At the same time, the addition of glycerol can effectively reduce the temperature sensitivity of the extraction system, reduce the occurrence of thermal degradation, and protect the structure and activity of the target compound. The glycerol-modified subcritical water extraction technology can also achieve selective extraction of Polygonatum sibiricum polysaccharide by adjusting the concentration of glycerol and the extraction conditions, which helps to reduce the formation of impurities and by-products and improve the purity and quality of the extract.

[0019] The method of the present application first uses a degreasing agent to treat the Polygonatum sibiricum, which can make the Polygonatum sibiricum fiber swell. The degreasing agent is used to destroy the bonds on the fiber surface, remove the lipid substances on the Polygonatum sibiricum surface, and then improve the hydrophilicity and chemical reactivity of the fiber surface, which is beneficial for the alkali solution and enzyme in the subsequent steps to directly act on the internal structure of the Polygonatum sibiricum, thereby improving the extraction efficiency of the uniform polysaccharide of the Polygonatum sibiricum; the use of alkaline treatment can, on the one hand, destroy the internal cellulose and hemicellulose structure of the Polygonatum sibiricum, which is beneficial to the hydrolysis of the Polygonatum sibiricum, and on the other hand, it can further loosen the molecular structure of the Polygonatum sibiricum, improve the extraction rate of enzyme extraction, and reduce the amount of enzyme added.

[0020] The dosage ratio of petroleum ether, hydrochloric acid and adsorbent in the degreasing agent raw materials of the present application can adjust the balance between the degreasing and oil removal effect and the impurity removal effect of the degreasing agent, enhance the degreasing and oil removal ability of the degreasing agent, and thus enhance the extraction effect of Polygonatum sibiricum uniform polysaccharide. If the dosage of the adsorbent is too small, the enhanced impurity removal effect is not obvious. If the dosage of the adsorbent is too large, it will affect the effect of removing lipid substances on the surface of Polygonatum sibiricum fiber. If the dosage of hydrochloric acid is too small, it will also affect the effect of removing lipid substances on the surface of Polygonatum sibiricum fiber. If the dosage of hydrochloric acid is too large, it will cause the loss of part of Polygonatum sibiricum polysaccharide, affecting the yield of Polygonatum sibiricum uniform polysaccharide.

[0021] The volcanic mud of the present application is combined with ferric chloride and zirconium oxide to fully utilize the electrostatic attraction effect of volcanic mud, the cohesion effect of ferric chloride, and the porous adsorption effect of zirconium oxide to effectively absorb odor, grease, and stains on the surface of Polygonatum sibiricum fiber, thereby enhancing the deep cleaning effect of the degreasing agent on Polygonatum sibiricum fiber.

[0022] The present application adds an ammonia-ethanol solution to make the residual acid react with the ammonia, neutralize the residual acid in the degreasing process, avoid the residual acid from destroying the structure of the polygonatum polysaccharide, and ensure the efficient enzymatic hydrolysis process; utilize the volatility of ammonia to control the alkalinity of the enzymatic hydrolysis solution and avoid a large excess of residual alkali.

[0023] According to some embodiments of the present application, the amount of the degreasing agent added in step (1) is 30%-50% of the weight of the polygonatum sibiricum medicinal material.

[0024] According to some embodiments of the present application, the degreasing treatment time in step (1) is 3-5 hours.

[0025] According to some embodiments of the present application, the method for preparing the adsorbent comprises the following steps:

[0026] A1. Mix volcanic mud, ferric chloride and zirconium oxide according to a weight ratio, and then dry and grind to obtain a powder;

[0027] A2. The powder prepared in step A1 was calcined at 110-150 ℃ for 30-40min;

[0028] A3. Then heat to 650-700℃ and calcine for 20-30min;

[0029] A4. After cooling and magnetization, grinding to a size of 100 mesh or more is performed to obtain the adsorbent.

[0030] Step A1: First, volcanic mud, ferric chloride and zirconium oxide are mixed evenly, so that the ferric chloride and zirconium oxide are fully mixed, and are not easily separated in the subsequent granulation process, so that the agglomeration effect of ferric chloride is evenly distributed in the zirconium oxide, thereby making the adsorption performance of the prepared adsorbent more stable.

[0031] The preparation method of the adsorbent includes a two-step roasting process. The first stage of the two-stage roasting is pre-activation, and the second stage of roasting is activation. The two-stage roasting gradually destroys the lattice, making it easier to form an amorphous active material, thereby improving the adsorption effect on impurities.

[0032] Grinding to more than 100 meshes can make the adsorbent particles smaller, enhance the dispersibility of the adsorbent, and facilitate rapid and uniform mixing with other components in the subsequent preparation of the degreasing agent. It also increases the fineness of the prepared degreasing agent when used and is not easy to damage the polygonatum polysaccharide.

[0033] Furthermore, the step A3 further includes adding a pore-forming agent and mixing before heating. Adding a pore-forming agent can increase the porosity of the granulation.

[0034] Furthermore, the pore forming agent includes carboxymethyl cellulose and polyvinyl alcohol. Polyvinyl alcohol has good bonding strength with the powder, is more likely to improve dispersibility, and forms micropores after sintering to improve porosity. Carboxymethyl cellulose further improves porosity to avoid affecting adsorption effect due to blockage of adsorbent pores.

[0035] According to some embodiments of the present application, the amount of the ammonia-ethanol solution added in step (2) is 20%-30% of the weight of the polygonatum sibiricum medicinal material.

[0036] According to some embodiments of the present application, the ammonia-ethanol solution in step (2) is prepared by mixing ammonia and ethanol in a volume ratio of 1:1.

[0037] According to some embodiments of the present application, the amount of cellulase added in step (2) is 1%-3% of the weight of the polygonatum sibiricum medicinal material.

[0038] According to some embodiments of the present application, the amount of hemicellulase added in step (2) is 1%-2% of the weight of the polygonatum sibiricum medicinal material.

[0039] According to some embodiments of the present application, the dissolution time of the cellulase and hemicellulase in step (2) is 2-3 hours.

[0040] According to some embodiments of the present application, the centrifugal separation in step (2) has a centrifugal speed of 4000-6000 r / min.

[0041] Furthermore, the centrifugation time of the centrifugal separation in step (2) is 20 min-30 min.

[0042] According to some embodiments of the present application, the extraction temperature of the glycerol-modified subcritical water extraction in step (3) is 100-140°C.

[0043] According to some embodiments of the present application, the extraction pressure of the glycerol-modified subcritical water extraction in step (3) is 1.5-2.5 MPa.

[0044] According to some embodiments of the present application, the solution flow rate of the glycerol-modified subcritical water extraction in step (3) is 2.5-4.5 mL / min.

[0045] According to some embodiments of the present application, the extraction time of the glycerol-modified subcritical water extraction in step (3) is 90-150 min.

[0046] According to some embodiments of the present application, the pH value of the glycerol-modified subcritical water extraction in step (3) is 7.

[0047] In a second aspect, the embodiments of the present application provide a homogeneous polygonatum polysaccharide obtained by using the above-mentioned method for extracting homogeneous polygonatum polysaccharide.

[0048] The homogeneous Polygonatum sibiricum polysaccharide according to the embodiments of the present application has at least the following beneficial effects:

[0049] The uniform Polygonatum sibiricum polysaccharide obtained in the present application is safe and pollution-free, has high purity and uniform molecular weight distribution, has more significant and stable biological activity, and may show stronger effects in immune regulation, anti-tumor and other aspects.

[0050] On the third aspect, the embodiments of the present application provide the application of the above-mentioned uniform polysaccharide of polygonatum in the field of high-end health products and medicine. DETAILED DESCRIPTION

[0051] To make the purpose, technical solution and advantages of the present application clearer, the following will be further described in detail in conjunction with specific embodiments. The embodiments described here are only part of the embodiments of the present application and cannot be understood as limiting the scope of protection of the present application.

[0052] Example 1

[0053] Extraction of Polygonatum sibiricum Homogeneous Polysaccharide:

[0054] (1) Washing and crushing the polygonatum sibiricum medicinal material, adding a degreasing agent of 40% of the weight of the polygonatum sibiricum medicinal material, performing a degreasing treatment at a temperature of 70° C. for 4 hours, filtering, and obtaining a defatted polygonatum sibiricum;

[0055] (2) adding the defatted polygonatum to an ammonia-ethanol solution in an amount of 25% by weight of the polygonatum medicinal material, pre-treating for 2 hours, adding cellulase in an amount of 2% by weight of the polygonatum medicinal material and hemicellulase in an amount of 1.5% by weight of the polygonatum medicinal material to dissolve for 2.5 hours, and centrifuging at a speed of 5000 r / min for 25 minutes to obtain a filtrate and a polygonatum residue;

[0056] (3) subjecting the polygonatum residue to glycerol-modified subcritical water extraction, wherein the extraction temperature is 120° C., the extraction pressure is 2 MPa, the solution flow rate is 3.5 mL / min, the extraction time is 120 min, the extraction pH value is 7, and the amount of glycerol is 50% of the solution weight, to obtain an extract;

[0057] (4) mixing the filtrate and the extract, and subjecting the mixture to alcohol precipitation, centrifugation, and freeze-drying to obtain crude polygonatum polysaccharide;

[0058] (5) After dissolving the crude polygonatum polysaccharide with distilled water, the mixture is loaded onto an anionic cellulose chromatography column, and the chromatography column is eluted with distilled water to obtain a water-washed component polysaccharide of polygonatum, and the water-washed component polysaccharide is further purified with a gel chromatography column, and the gel chromatography column is eluted with a NaCl solution to obtain a uniform polygonatum polysaccharide;

[0059] Wherein, the degreasing agent in step (1) is prepared by mixing petroleum ether, hydrochloric acid and adsorbent in a weight ratio of 7:2.5:1;

[0060] The ammonia-ethanol solution in step (2) is prepared by mixing ammonia and ethanol in a volume ratio of 1:1;

[0061] Preparation of adsorbent:

[0062] A1. Mix ferric chloride, volcanic mud and zirconium oxide in a weight ratio of 1:3:1, and then dry and grind to obtain a powder;

[0063] A2. The powder prepared in step A1 was calcined at 130 ° C for 35 min;

[0064] A3. Then carboxymethyl cellulose and polyvinyl alcohol were added and mixed, and the temperature was raised to 680°C and calcined for 25 minutes;

[0065] A4. After cooling and magnetization, grind to more than 100 mesh to obtain adsorbent.

[0066] Example 2

[0067] Extraction of Polygonatum sibiricum Homogeneous Polysaccharide:

[0068] (1) Washing and crushing the polygonatum sibiricum medicinal material, adding a degreasing agent of 50% of the weight of the polygonatum sibiricum medicinal material, performing a degreasing treatment at a temperature of 60° C. for 5 hours, filtering, and obtaining a defatted polygonatum sibiricum;

[0069] (2) adding the defatted polygonatum to an ammonia-ethanol solution in an amount of 20% by weight of the polygonatum medicinal material, pre-treating for 2 hours, adding cellulase in an amount of 3% by weight of the polygonatum medicinal material and hemicellulase in an amount of 1% by weight of the polygonatum medicinal material to dissolve for 3 hours, and centrifuging at a speed of 4000 r / min for 30 minutes to obtain a filtrate and a polygonatum residue;

[0070] (3) subjecting the polygonatum residue to glycerol-modified subcritical water extraction, wherein the extraction temperature is 100° C., the extraction pressure is 2.5 MPa, the solution flow rate is 2.5 mL / min, the extraction time is 150 min, the extraction pH value is 7, and the amount of glycerol is 40% of the solution weight, to obtain an extract;

[0071] (4) mixing the filtrate and the extract, and subjecting the mixture to alcohol precipitation, centrifugation, and freeze-drying to obtain crude polygonatum polysaccharide;

[0072] (5) After dissolving the crude polygonatum polysaccharide with distilled water, the mixture is loaded onto an anionic cellulose chromatography column, and the chromatography column is eluted with distilled water to obtain a water-washed component polysaccharide of polygonatum, and the water-washed component polysaccharide is further purified with a gel chromatography column, and the gel chromatography column is eluted with a NaCl solution to obtain a uniform polygonatum polysaccharide;

[0073] Wherein, the degreasing agent in step (1) is prepared by mixing petroleum ether, hydrochloric acid and adsorbent in a weight ratio of 8:2:2;

[0074] The ammonia-ethanol solution in step (2) is prepared by mixing ammonia and ethanol in a volume ratio of 1:1;

[0075] Preparation of adsorbent:

[0076] A1. Mix ferric chloride, volcanic mud and zirconium oxide in a weight ratio of 1:2:2, and then dry and grind to obtain a powder;

[0077] A2. The powder prepared in step A1 was calcined at 110 ° C for 40 min;

[0078] A3. Then carboxymethyl cellulose and polyvinyl alcohol were added and mixed, and the temperature was raised to 650°C and calcined for 30 minutes;

[0079] A4. After cooling and magnetization, grind to more than 100 mesh to obtain adsorbent.

[0080] Example 3

[0081] Extraction of Polygonatum sibiricum Homogeneous Polysaccharide:

[0082] (1) Washing and crushing the polygonatum sibiricum medicinal material, adding a degreasing agent of 30% of the weight of the polygonatum sibiricum medicinal material, performing a degreasing treatment at a temperature of 80° C. for 3 hours, filtering, and obtaining a defatted polygonatum sibiricum;

[0083] (2) adding the defatted polygonatum to an ammonia-ethanol solution in an amount of 30% by weight of the polygonatum medicinal material, pre-treating for 2 hours, adding cellulase in an amount of 1% by weight of the polygonatum medicinal material and hemicellulase in an amount of 2% by weight of the polygonatum medicinal material to dissolve for 2 hours, and centrifuging at a speed of 6000 r / min for 30 minutes to obtain a filtrate and a polygonatum residue;

[0084] (3) subjecting the polygonatum residue to glycerol-modified subcritical water extraction, wherein the extraction temperature is 140° C., the extraction pressure is 1.5 MPa, the solution flow rate is 4.5 mL / min, the extraction time is 90 min, the extraction pH value is 7, and the amount of glycerol is 60% of the solution weight, to obtain an extract;

[0085] (4) mixing the filtrate and the extract, and subjecting the mixture to alcohol precipitation, centrifugation, and freeze-drying to obtain crude polygonatum polysaccharide;

[0086] (5) After dissolving the crude polygonatum polysaccharide with distilled water, the mixture is loaded onto an anionic cellulose chromatography column, and the chromatography column is eluted with distilled water to obtain a water-washed component polysaccharide of polygonatum, and the water-washed component polysaccharide is further purified with a gel chromatography column, and the gel chromatography column is eluted with a NaCl solution to obtain a uniform polygonatum polysaccharide;

[0087] Wherein, the degreasing agent in step (1) is prepared by mixing petroleum ether, hydrochloric acid and adsorbent in a weight ratio of 5:3:0.5;

[0088] The ammonia-ethanol solution in step (2) is prepared by mixing ammonia and ethanol in a volume ratio of 1:1;

[0089] Preparation of adsorbent:

[0090] A1. Mix ferric chloride, volcanic mud and zirconium oxide in a weight ratio of 1:4:0.5, and then dry and grind to obtain a powder;

[0091] A2. The powder prepared in step A1 was calcined at 150 ° C for 30 min;

[0092] A3. Then carboxymethyl cellulose and polyvinyl alcohol were added and mixed, and the temperature was raised to 700°C and calcined for 20 minutes;

[0093] A4. After cooling and magnetization, grind to more than 100 mesh to obtain adsorbent.

[0094] Example 4

[0095] Extraction of Polygonatum sibiricum Homogeneous Polysaccharide:

[0096] (1) Washing and crushing the polygonatum sibiricum medicinal material, adding a degreasing agent of 45% of the weight of the polygonatum sibiricum medicinal material, performing a degreasing treatment at a temperature of 65° C. for 4 hours, filtering, and obtaining a defatted polygonatum sibiricum;

[0097] (2) adding the defatted polygonatum to an ammonia-ethanol solution in an amount of 25% by weight of the polygonatum medicinal material, pre-treating for 2 hours, adding cellulase in an amount of 2% by weight of the polygonatum medicinal material and hemicellulase in an amount of 1.6% by weight of the polygonatum medicinal material to dissolve for 2.5 hours, and centrifuging at a speed of 5000 r / min for 25 minutes to obtain a filtrate and a polygonatum residue;

[0098] (3) subjecting the polygonatum residue to glycerol-modified subcritical water extraction, wherein the extraction temperature is 120° C., the extraction pressure is 2 MPa, the solution flow rate is 4 mL / min, the extraction time is 130 min, the extraction pH value is 7, and the amount of glycerol is 45% of the solution weight to obtain an extract;

[0099] (4) mixing the filtrate and the extract, and subjecting the mixture to alcohol precipitation, centrifugation, and freeze-drying to obtain crude polygonatum polysaccharide;

[0100] (5) After dissolving the crude polygonatum polysaccharide with distilled water, the mixture is loaded onto an anionic cellulose chromatography column, and the chromatography column is eluted with distilled water to obtain a water-washed component polysaccharide of polygonatum, and the water-washed component polysaccharide is further purified with a gel chromatography column, and the gel chromatography column is eluted with a NaCl solution to obtain a uniform polygonatum polysaccharide;

[0101] Wherein, the degreasing agent in step (1) is prepared by mixing petroleum ether, hydrochloric acid and adsorbent in a weight ratio of 6:2:1;

[0102] The ammonia-ethanol solution in step (2) is prepared by mixing ammonia and ethanol in a volume ratio of 1:1;

[0103] Preparation of adsorbent:

[0104] A1. Mix ferric chloride, volcanic mud and zirconium oxide in a weight ratio of 1:3:1.4, and then dry and grind to obtain a powder;

[0105] A2. The powder prepared in step A1 was calcined at 120 ° C for 36 min;

[0106] A3. Then add carboxymethyl cellulose and polyvinyl alcohol and mix well, raise the temperature to 670℃ and calcine for 25min;

[0107] A4. After cooling and magnetization, grind to more than 100 mesh to obtain adsorbent.

[0108] Comparative Example 1

[0109] Extraction of Polygonatum sibiricum Homogeneous Polysaccharide:

[0110] (1) Washing and crushing the polygonatum sibiricum medicinal material, adding a degreasing agent of 40% of the weight of the polygonatum sibiricum medicinal material, performing a degreasing treatment at a temperature of 70° C. for 4 hours, filtering, and obtaining a defatted polygonatum sibiricum;

[0111] (2) adding the defatted polygonatum to an ammonia-ethanol solution in an amount of 25% by weight of the polygonatum medicinal material, pre-treating for 2 hours, adding cellulase in an amount of 2% by weight of the polygonatum medicinal material and hemicellulase in an amount of 1.5% by weight of the polygonatum medicinal material to dissolve for 2.5 hours, and centrifuging at a speed of 5000 r / min for 25 minutes to obtain a filtrate and a polygonatum residue;

[0112] (3) subjecting the polygonatum residue to subcritical water extraction, wherein the extraction temperature is 120° C., the extraction pressure is 2 MPa, the solution flow rate is 3.5 mL / min, the extraction time is 120 min, the extraction pH value is 7, and an extract is obtained;

[0113] (4) mixing the filtrate and the extract, and subjecting the mixture to alcohol precipitation, centrifugation, and freeze-drying to obtain crude polygonatum polysaccharide;

[0114] (5) After dissolving the crude polygonatum polysaccharide with distilled water, the mixture is loaded onto an anionic cellulose chromatography column, and the chromatography column is eluted with distilled water to obtain a water-washed component polysaccharide of polygonatum, and the water-washed component polysaccharide is further purified with a gel chromatography column, and the gel chromatography column is eluted with a NaCl solution to obtain a uniform polygonatum polysaccharide;

[0115] Wherein, the degreasing agent in step (1) is prepared by mixing petroleum ether, hydrochloric acid and adsorbent in a weight ratio of 7:2.5:1;

[0116] The ammonia-ethanol solution in step (2) is prepared by mixing ammonia and ethanol in a volume ratio of 1:1;

[0117] Preparation of adsorbent:

[0118] A1. Mix ferric chloride, volcanic mud and zirconium oxide in a weight ratio of 1:3:1, and then dry and grind to obtain a powder;

[0119] A2. The powder prepared in step A1 was calcined at 130 ° C for 35 min;

[0120] A3. Then carboxymethyl cellulose and polyvinyl alcohol were added and mixed, and the temperature was raised to 680°C and calcined for 25 minutes;

[0121] A4. After cooling and magnetization, grind to more than 100 mesh to obtain adsorbent.

[0122] Comparative Example 2

[0123] Extraction of Polygonatum sibiricum Homogeneous Polysaccharide:

[0124] (1) Washing and crushing the polygonatum sibiricum medicinal material, adding a degreasing agent of 40% of the weight of the polygonatum sibiricum medicinal material, performing a degreasing treatment at a temperature of 70° C. for 4 hours, filtering, and obtaining a defatted polygonatum sibiricum;

[0125] (2) adding 2% of cellulase by weight of the polygonatum sibiricum and 1.5% of hemicellulase by weight of the polygonatum sibiricum to the defatted polygonatum sibiricum and dissolving them for 2.5 hours, and centrifuging at a speed of 5000 r / min for 25 minutes to obtain a filtrate and a polygonatum sibiricum residue;

[0126] (3) subjecting the polygonatum residue to glycerol-modified subcritical water extraction, wherein the extraction temperature is 120° C., the extraction pressure is 2 MPa, the solution flow rate is 3.5 mL / min, the extraction time is 120 min, the extraction pH value is 7, and the amount of glycerol is 50% of the solution weight, to obtain an extract;

[0127] (4) mixing the filtrate and the extract, and subjecting the mixture to alcohol precipitation, centrifugation, and freeze-drying to obtain crude polygonatum polysaccharide;

[0128] (5) After dissolving the crude polygonatum polysaccharide with distilled water, the mixture is loaded onto an anionic cellulose chromatography column, and the chromatography column is eluted with distilled water to obtain a water-washed component polysaccharide of polygonatum, and the water-washed component polysaccharide is further purified with a gel chromatography column, and the gel chromatography column is eluted with a NaCl solution to obtain a uniform polygonatum polysaccharide;

[0129] Wherein, the degreasing agent in step (1) is prepared by mixing petroleum ether, hydrochloric acid and adsorbent in a weight ratio of 7:2.5:1;

[0130] Preparation of adsorbent:

[0131] A1. Mix ferric chloride, volcanic mud and zirconium oxide in a weight ratio of 1:3:1, and then dry and grind to obtain a powder;

[0132] A2. The powder prepared in step A1 was calcined at 130 ° C for 35 min;

[0133] A3. Then add carboxymethyl cellulose and polyvinyl alcohol and mix well, raise the temperature to 680℃ and calcine for 25min;

[0134] A4. After cooling and magnetization, grind to more than 100 mesh to obtain adsorbent.

[0135] Comparative Example 3

[0136] Extraction of Polygonatum sibiricum Homogeneous Polysaccharide:

[0137] (1) Washing and crushing the polygonatum sibiricum medicinal material, adding a degreasing agent of 40% of the weight of the polygonatum sibiricum medicinal material, performing a degreasing treatment at a temperature of 70° C. for 4 hours, filtering, and obtaining a defatted polygonatum sibiricum;

[0138] (2) adding the defatted polygonatum to an ammonia-ethanol solution in an amount of 25% by weight of the polygonatum medicinal material, pre-treating for 2 hours, adding cellulase in an amount of 2% by weight of the polygonatum medicinal material and hemicellulase in an amount of 1.5% by weight of the polygonatum medicinal material to dissolve for 2.5 hours, and centrifuging at a speed of 5000 r / min for 25 minutes to obtain a filtrate and a polygonatum residue;

[0139] (3) subjecting the polygonatum residue to glycerol-modified subcritical water extraction, wherein the extraction temperature is 120° C., the extraction pressure is 2 MPa, the solution flow rate is 3.5 mL / min, the extraction time is 120 min, the extraction pH value is 7, and the amount of glycerol is 50% of the solution weight, to obtain an extract;

[0140] (4) mixing the filtrate and the extract, and subjecting the mixture to alcohol precipitation, centrifugation, and freeze-drying to obtain crude polygonatum polysaccharide;

[0141] (5) After dissolving the crude polygonatum polysaccharide with distilled water, the mixture is loaded onto an anionic cellulose chromatography column, and the chromatography column is eluted with distilled water to obtain a water-washed component polysaccharide of polygonatum, and the water-washed component polysaccharide is further purified with a gel chromatography column, and the gel chromatography column is eluted with a NaCl solution to obtain a uniform polygonatum polysaccharide;

[0142] Wherein, the degreasing agent in step (1) is prepared by mixing petroleum ether, hydrochloric acid and adsorbent in a weight ratio of 7:2.5:1;

[0143] The ammonia-ethanol solution in step (2) is prepared by mixing ammonia and ethanol in a volume ratio of 1:1;

[0144] Preparation of adsorbent:

[0145] A1. Mix volcanic mud and zirconium oxide in a weight ratio of 3:1, then dry and grind to obtain a powder;

[0146] A2. The powder prepared in step A1 was calcined at 130 ° C for 35 min;

[0147] A3. Then carboxymethyl cellulose and polyvinyl alcohol were added and mixed, and the temperature was raised to 680°C and calcined for 25 minutes;

[0148] A4. After cooling and magnetization, grind to more than 100 mesh to obtain adsorbent.

[0149] Experimental example

[0150] Extract Polygonatum sibiricum homogeneous polysaccharide according to the method of Example 1-4 and Comparative Example 1-3, and determine the extraction rate, extraction rate (%) = (Polygonatum sibiricum homogeneous polysaccharide weight / Polygonatum sibiricum medicinal material weight) × 100%;

[0151] The obtained data are shown in Table 1 below:

[0152] Table 1

[0153] Test items Example 1 Example 2 Example 3 Example 4 Comparative Example 1 Comparative Example 2 Comparative Example 3 Extraction rate (%) 33.6 32.8 33.1 33.5 22.4 25.1 27.6

[0154] As can be seen from Table 1, the extraction rates of Polygonatum sibiricum uniform polysaccharides obtained by the extraction methods of Examples 1-4 of the present application are relatively high.

[0155] The subcritical water extraction in the extraction method of Comparative Example 1 is modified by adding glycerol, and the rest is the same as Example 1. The extraction rate of the uniform polysaccharide of Polygonatum sibiricum extracted in Comparative Example 1 is significantly lower than that in Example 1, indicating that the glycerol-modified subcritical water extraction of the present application is very critical for the extraction of the uniform polysaccharide of Polygonatum sibiricum. Glycerol, as an organic solvent, can increase the solubility of Polygonatum sibiricum polysaccharide in subcritical water, so that the Polygonatum sibiricum polysaccharide with a lower solubility in pure water can be more easily extracted, thereby improving the extraction efficiency. At the same time, the addition of glycerol can effectively reduce the temperature sensitivity of the extraction system, reduce the occurrence of thermal degradation, and protect the structure and activity of the target compound. The glycerol-modified subcritical water extraction technology can also achieve selective extraction of Polygonatum sibiricum polysaccharide by adjusting the concentration of glycerol and the extraction conditions, which helps to reduce the formation of impurities and by-products and improve the purity and quality of the extract.

[0156] In step (2) of the extraction method of comparative example 2, no ammonia-ethanol solution pretreatment is performed, and the rest is the same as that of example 1. The extraction rate of the uniform polysaccharide of polygonatum extracted in comparative example 2 is significantly lower than that in example 1, indicating that the ammonia-ethanol solution pretreatment of the present application is very critical for the extraction of the uniform polysaccharide of polygonatum. By adding ammonia-ethanol solution for treatment, the residual acid reacts with ammonia water to neutralize the residual acid in the defatting process, thereby avoiding the residual acid from destroying the structure of the polygonatum polysaccharide and ensuring the efficient enzymatic hydrolysis process. The volatile nature of ammonia water is utilized to control the alkalinity of the enzymatic hydrolysis solution and avoid a large amount of excess residual alkali.

[0157] The raw materials of the adsorbent in the extraction method of Comparative Example 3 do not contain ferric chloride, and the rest are the same as Example 1. The extraction rate of the uniform polysaccharide of Polygonatum sibiricum extracted in Comparative Example 3 is significantly lower than that in Example 1, indicating that the combination of volcanic mud, ferric chloride and zirconium oxide in the raw materials of the adsorbent of the present application is very critical for the extraction of uniform polysaccharide of Polygonatum sibiricum. By combining volcanic mud with ferric chloride and zirconium oxide, the electrostatic attraction effect of volcanic mud, the coagulation effect of ferric chloride and the porous adsorption effect of zirconium oxide can be fully utilized to effectively adsorb the odor, grease and stains on the surface of the Polygonatum sibiricum fiber, thereby enhancing the deep cleaning effect of the degreasing agent on the Polygonatum sibiricum fiber.

[0158] Although the embodiments of the present application have been shown and described, those skilled in the art will appreciate that various changes, modifications, substitutions or variations may be made to the embodiments without departing from the principles and purpose of the present application, and that the technical solutions after these changes, modifications, substitutions or variations will fall within the scope of protection of the present application.

Claims

1. A method for extracting uniform polysaccharide from polygonatum, characterized in that: The following steps are involved: (1) washing and crushing the polygonatum sibiricum medicinal material, adding a degreasing agent to perform degreasing treatment at a temperature of 60-80° C., filtering, and obtaining defatted polygonatum sibiricum; (2) adding the defatted polygonatum to an ammonia-ethanol solution, pre-treating for 2 hours, adding cellulase and hemicellulase to dissolve, and centrifuging to obtain a filtrate and a polygonatum residue; (3) subjecting the polygonatum residue to glycerol-modified subcritical water extraction to obtain an extract; (4) mixing the filtrate and the extract, and subjecting the mixture to alcohol precipitation, centrifugation, and freeze-drying to obtain crude polygonatum polysaccharide; (5) After dissolving the crude polygonatum polysaccharide with distilled water, the mixture is loaded onto an anionic cellulose chromatography column, and the chromatography column is eluted with distilled water to obtain a water-washed component polysaccharide of polygonatum, and the water-washed component polysaccharide is further purified with a gel chromatography column, and the gel chromatography column is eluted with a NaCl solution to obtain a uniform polygonatum polysaccharide; Wherein, the amount of glycerol used in step (3) is 40-60% of the weight of the solution; The degreasing agent in step (1) comprises petroleum ether, hydrochloric acid and an adsorbent; The amounts of the petroleum ether, the hydrochloric acid and the adsorbent are in a weight ratio of (5-8):(2-3):(0.5-2); The raw materials for preparing the adsorbent include ferric chloride, volcanic mud and zirconium oxide in a weight ratio of 1:(2-4):(0.5-2).

2. The method for extracting uniform polysaccharide from polygonatum according to claim 1, characterized in that: The amount of the ammonia-ethanol solution added in step (2) is 20%-30% of the weight of the polygonatum sibiricum medicinal material.

3. The method for extracting uniform polysaccharide from polygonatum according to claim 1, characterized in that: The ammonia-ethanol solution in step (2) is prepared by mixing ammonia and ethanol in a volume ratio of 1:

1.

4. The method for extracting uniform polysaccharide from polygonatum according to claim 1, characterized in that: The extraction temperature of the glycerol-modified subcritical water extraction in step (3) is 100-140° C., and / or the extraction pressure of the glycerol-modified subcritical water extraction in step (3) is 1.5-2.5 MPa.

5. The method for extracting uniform polysaccharide from polygonatum according to claim 1, characterized in that: The amount of the degreasing agent added in step (1) is 30%-50% of the weight of the polygonatum sibiricum medicinal material, and / or the time of the degreasing treatment in step (1) is 3-5 hours.

6. The method for extracting uniform polysaccharide from polygonatum according to claim 1, characterized in that: The preparation method of the adsorbent comprises the following steps: A1. Mix volcanic mud, ferric chloride and zirconium oxide according to a weight ratio, and then dry and grind to obtain a powder; A2. The powder prepared in step A1 was calcined at 110-150 ℃ for 30-40min; A3. Then heat to 650-700℃ and calcine for 20-30min; A4. After cooling and magnetization, grinding to a size of 100 mesh or more is performed to obtain the adsorbent.

7. The method for extracting uniform polysaccharide from polygonatum according to claim 1, characterized in that: The amount of the cellulase added in step (2) is 1%-3% of the weight of the polygonatum sibiricum medicinal material, and / or the amount of the hemicellulase added in step (2) is 1%-2% of the weight of the polygonatum sibiricum medicinal material.

8. The method for extracting uniform polysaccharide from polygonatum according to claim 1, characterized in that: The dissolution time of the cellulase and hemicellulase in step (2) is 2-3 hours.

9. A homogeneous polygonatum polysaccharide obtained by the method according to any one of claims 1 to 8.

10. Use of the homogeneous polygonatum polysaccharide as claimed in claim 9 in the fields of high-end health products and medicine.