Enzymolysis extraction method of ginseng polysaccharide and application of ginseng polysaccharide in food

Abstract

The invention belongs to the technical field of purification processes, and particularly relates to an enzymolysis extraction method of ginseng polysaccharide and application of the ginseng polysaccharide in food. The enzymolysis extraction method comprises the following steps: S1, drying a ginseng medicinal material, crushing, sieving with a 30-50-mesh sieve to obtain dry ginseng powder, adding distilled water, decocting for 2-4 hours, extracting for 2-3 times, combining decoction, concentrating, adding ethanol until the alcohol content of a system is greater than or equal to 80%, standing for 24 hours, centrifuging to obtain a precipitate, and drying the precipitate; s2, mixing the dried precipitate with distilled water, and adding enzyme for enzymolysis extraction to obtain an extracting solution; s3, concentrating the extracting solution until the volume of the extracting solution is 20-25% of that of the extracting solution, centrifuging to remove precipitates, dialyzing, and concentrating and drying to obtain crude ginseng polysaccharide; and S4, dissolving the crude ginseng polysaccharide in distilled water for separation and purification. The variety, adding sequence and conditions of the enzymes are optimized, the extraction rate and purity of the ginseng polysaccharide are improved, and meanwhile the biological activity of the ginseng polysaccharide is kept. The extracted ginseng polysaccharide can be widely applied to various foods.

Description

Technical Field

[0001] The invention belongs to the technical field of purification technology, and specifically relates to an enzymatic extraction method of ginseng polysaccharide and its application in food. Background Art

[0002] As a precious traditional Chinese medicine, ginseng has a long history and wide application value. Its main active ingredients include saponins, polysaccharides, volatile oils, amino acids and peptides, which give ginseng a variety of physiological effects, such as improving immunity, anti-aging, promoting learning and memory, detoxification, antibacterial, etc. In particular, ginseng polysaccharides, as an important bioactive ingredient in ginseng, have attracted widespread attention from researchers in recent years due to their unique chemical structure and diverse biological activities.

[0003] Traditional ginseng polysaccharide extraction methods mainly include water extraction and alcohol precipitation. However, these methods have disadvantages such as low extraction efficiency, easy destruction of polysaccharide structure, and low product purity. In order to improve the extraction efficiency and product quality of ginseng polysaccharides, researchers are constantly exploring new extraction methods. Among them, enzymatic hydrolysis, as a method with mild conditions, little effect on polysaccharide structure, and high extraction efficiency, has gradually become a research hotspot for ginseng polysaccharide extraction. However, there are still some problems in the practical application of enzymatic hydrolysis, such as enzyme selectivity, enzyme stability, optimization of extraction conditions, etc. These problems limit the wide application of enzymatic hydrolysis in ginseng polysaccharide extraction.

[0004] The present invention aims to provide an enzymatic extraction method for ginseng polysaccharides, which realizes efficient extraction and purification of ginseng polysaccharides by optimizing the types of enzymes and extraction conditions, and applies the method to food, thereby providing new ideas and technical support for the development of functional foods. Summary of the invention

[0005] The purpose of the present invention is to provide an enzymatic extraction method of ginseng polysaccharide and its application in food, which significantly improves the extraction rate and purity of ginseng polysaccharide by optimizing the type of enzyme, the order of addition and the conditions, while maintaining its biological activity. The extracted high-purity ginseng polysaccharide can be widely used in a variety of foods.

[0006] In order to achieve the above object, the present invention provides the following technical solutions: A method for enzymatic extraction of ginseng polysaccharides comprises the following steps: S1. Dry the ginseng material and grind it through a 30-50 mesh sieve to obtain dry ginseng powder, add distilled water and boil for 2-4 hours, extract 2-3 times and combine the decoctions, add ethanol after concentration until the alcohol content of the system is ≥80%, let it stand for 24 hours and then centrifuge to obtain a precipitate, and dry the precipitate; S2, mixing the dried precipitate with distilled water, adding enzyme for enzymatic extraction, and obtaining an extract; S3, concentrating the extract to 20% to 25% of the volume of the extract, removing the precipitate by centrifugation, placing the supernatant into a dialysis bag, dialyzing it, and concentrating and drying it to obtain crude ginseng polysaccharides; S4. Dissolve the crude ginseng polysaccharide in distilled water for separation and purification.

[0007] Preferably, the enzymes include cellulase, pectinase, papain, saccharifying enzyme, lignin peroxidase, laccase.

[0008] Preferably, the enzymatic activity of the cellulase is ≥10000 U / g.

[0009] Preferably, the enzymatic activity of the pectinase is ≥25000 U / g.

[0010] Preferably, the enzymatic activity of the papain is ≥90,000 U / g.

[0011] Preferably, the saccharifying enzyme has an activity of ≥140,000 U / g.

[0012] In some preferred embodiments, the cellulase, pectinase, papain and saccharifying enzyme are all purchased from Ningxia Xiasheng Industrial Group Co., Ltd.

[0013] Preferably, the enzyme activity of the lignin peroxidase is ≥0.1 U / mg.

[0014] Preferably, the enzyme activity of the laccase is ≥0.5 U / mg.

[0015] In some preferred embodiments, the lignin peroxidase and laccase are both purchased from Shanghai MacLean Biochemical Technology Co., Ltd.

[0016] In step S2, the specific steps of enzymatic extraction are: A1. Mix the dried precipitate with distilled water, heat it to 100℃ and stir until there are no particles, then cool it to 45~55℃ and adjust the pH of the system to 4.5~5.0, add cellulase and pectinase, enzymolyze for 3~4h and then boil to inactivate; A2. Cool down to 50-55℃, adjust the system pH to 6.0-7.0, add papain for enzymatic hydrolysis for 2-3h, and then boil to inactivate; A3. Cool down to 50-60℃, adjust the system pH to 4.5-5.0, add saccharifying enzyme for enzymatic hydrolysis for 2-3h, and then boil to inactivate; A4. Cool down to 30-40℃ and adjust the system pH to 4.5-5.0. Add lignin peroxidase and laccase for enzymolysis for 1-2h and then boil to inactivate to obtain the extract.

[0017] In step A1, the mass ratio of the dried precipitate to distilled water is 1:10-20.

[0018] Preferably, the amount of cellulase is 10~30U / g dry ginseng powder; the amount of pectinase is 5~15U / g dry ginseng powder; the amount of papain is 5~10U / g dry ginseng powder; the amount of saccharifying enzyme is 5~15U / g dry ginseng powder; the amount of lignin peroxidase is 0.5~5U / g dry ginseng powder; and the amount of laccase is 0.5~5U / g dry ginseng powder.

[0019] The inventors found that by selecting six different enzymes to perform enzymolysis treatment on ginseng in sequence, the extraction rate and purity of ginseng polysaccharides in the prepared product can be improved. This may be because cellulase and pectinase work synergistically. Cellulase first degrades cellulose, making the cell wall loose, increasing the chance of pectinase contacting pectin, and pectinase then degrades pectin, further destroying the cell wall structure. Cellulase and pectinase promote each other and gradually destroy the integrity of the cell wall, making the internal polysaccharides more easily released. This synergistic effect improves the extraction rate and purity of polysaccharides. Papain reduces the encapsulation and binding of proteins to polysaccharides by degrading proteins, making polysaccharides easier to extract. At the same time, glucoamylase decomposes starch into smaller molecules, reducing the interference of starch on polysaccharide extraction, and further improving the purity and extraction rate of polysaccharides. Lignin peroxidase degrades lignin by redox reaction, while laccase further degrades lignin by catalyzing oxidation reaction. The synergistic effect of the two can more thoroughly destroy the lignin in the cell wall, thereby improving the extraction rate of polysaccharides. In addition, laccase can not only enhance the activity of lignin peroxidase and improve its degradation efficiency, but also catalyze the oxidation of certain phenolic substances in ginseng, thereby improving the permeability of ginseng cells. Laccase makes it easier for other enzymes to enter the cells to act, thereby promoting the release of polysaccharides. Through the synergistic action of the above enzymes, not only the cell wall is destroyed, but also impurities such as protein and starch are removed, so that polysaccharides can be extracted more efficiently, thereby improving the purity and extraction rate of polysaccharides. However, the action time and optimal reaction conditions of each enzyme are different, so the enzymatic process needs to be strictly controlled. Cellulase and pectinase need to destroy the cell wall first, otherwise the subsequent enzyme may be difficult to penetrate into the cell for enzymatic reaction, affecting the polysaccharide extraction rate and purity. At the same time, the amount of enzyme should also be controlled to avoid excessive degradation of the cell wall, which will lead to the destruction of the integrity and biological activity of the polysaccharide, and to avoid excessive degradation of protein and starch, which will cause the generated amino acids to bind to the polysaccharide and affect the purity of the polysaccharide. The complete decomposition of starch will lead to a decrease in the polysaccharide extraction rate.

[0020] Preferably, the dialysis bag has a molecular weight cutoff of 3500-5000Da and is purchased from Jiangsu Meg Biotechnology Co., Ltd.

[0021] In step S4, the specific steps of separation and purification are: B1. Dissolve the crude ginseng polysaccharide in distilled water, stir evenly and let stand for 24 hours, filter to remove insoluble matter, pass through a macroporous resin at a flow rate of 2.5-3.5 BV / h, and elute with 3-5 BV of water and 2-4 BV of 20%-28% ethanol solution in sequence, collect the eluate, concentrate and dry under reduced pressure to obtain substance 1; B2. Dissolve the substance in distilled water, stir evenly and let it stand for 24 hours, filter to remove insoluble matter, use an anion exchange column for secondary separation and purification, collect the sample after elution, concentrate and dry to obtain ginseng polysaccharide.

[0022] Preferably, the solid-liquid ratio of crude ginseng polysaccharide to distilled water in step B1 is 1 g: 5-20 mL; the solid-liquid ratio of substance 1 to distilled water in step B2 is 1 g: 5-20 mL.

[0023] Preferably, the macroporous resin is a weakly polar macroporous resin, the particles with a diameter between 0.3 and 1.25 mm account for 95% to 100%, the water content is 60% to 70%, and the specific surface area is 480 to 520 m 2 / g, average pore diameter 12~16nm, pore volume 0.73~0.77mL / g.

[0024] In some preferred embodiments, the macroporous resin is purchased from Shanghai MacLean Biochemical Technology Co., Ltd., AB-8.

[0025] The inventors found that the selection of a specific macroporous resin to enrich and purify ginseng crude polysaccharides and elution under specific conditions can further increase the content of ginseng polysaccharides in the product. This may be because on the one hand, AB-8 macroporous resin has a very large specific surface area and a suitable pore structure, which makes a large number of adsorption sites on its surface. During the adsorption process, ginseng polysaccharide molecules can interact with the resin surface in two ways: physical adsorption and chemical adsorption. The interaction between the van der Waals force on the resin surface and the polysaccharide molecules and the interaction between the polar groups on the resin surface and certain functional groups in the polysaccharide molecules make AB-8 resin have special selectivity for ginseng polysaccharides, so it can effectively adsorb polysaccharide molecules, thereby achieving separation and purification of polysaccharides. At the same time, AB-8 macroporous resin has a weak adsorption capacity for impurities such as proteins, inorganic salts and small molecule hydrophilic organic matter, so the resin can selectively adsorb polysaccharide molecules and exclude impurities. On the other hand, by selecting a specific concentration of ethanol as the eluent, most of the polysaccharides can be washed out while minimizing the elution of impurities, thereby achieving efficient recovery of polysaccharides, avoiding the loss and waste of polysaccharides, and improving the purity of ginseng polysaccharides in the product while also improving the recovery rate of ginseng polysaccharides. It should be noted that the ethanol concentration during elution should not be too high. High concentrations of ethanol may cause problems such as destruction of the polysaccharide structure or poor elution effect of low concentrations of ethanol, affecting the content of ginseng polysaccharides in the product.

[0026] Preferably, the ligand of the anion exchange column is diethylaminoethyl, the ligand density is 35-45 μmol / mL, the filler particle size is 45-55 μm, and the adsorption capacity is 170-190 mg HSA / mL.

[0027] In some preferred embodiments, the anion exchange column is purchased from Shanghai Chenqiao Biotechnology Co., Ltd., DE-52.

[0028] Preferably, the flow rate during elution in step B2 is 11-14 mL / min, and distilled water, 0.1 mol / L sodium chloride aqueous solution, 0.3 mol / L sodium chloride aqueous solution, and 0.5 mol / L sodium chloride aqueous solution are used for elution in sequence.

[0029] The inventors found that the use of a specific anion exchange column for secondary separation and purification can further increase the content of ginseng polysaccharides in the product while minimizing the impact on the recovery rate. This may be because the diethylaminoethyl group carries a positive charge under neutral or slightly acidic conditions, and can adsorb negatively charged polysaccharides through electrostatic interactions. For uncharged or positively charged impurities (such as proteins, pigments, etc.), the adsorption force of the diethylaminoethyl group is weak, so these impurities are not easily adsorbed, thereby further improving the purity of the polysaccharide. By using salt solutions of different concentrations for gradient elution and gradually increasing the salt concentration, polysaccharide components with different charge densities can be eluted step by step, ensuring efficient desorption of polysaccharides from the resin, while avoiding excessive desorption and loss of polysaccharides, thereby maintaining a high recovery rate. In addition, an appropriate ligand density can also ensure that polysaccharides are not lost in large quantities during adsorption and elution.

[0030] The ginseng polysaccharide is prepared by the enzymatic extraction method of ginseng polysaccharide.

[0031] The application of the ginseng polysaccharide in food can be applied to functional food, such as nutritional supplements, beverages, cakes, etc.

[0032] Compared with the prior art, the advantages and beneficial effects of the present invention are: 1. The present invention provides an enzymatic extraction method for ginseng polysaccharides, which significantly improves the extraction rate and purity of ginseng polysaccharides by optimizing the type, addition order and conditions of enzymes, while maintaining its biological activity. The extracted high-purity ginseng polysaccharides can be widely used in a variety of foods.

[0033] 2. The present invention can improve the extraction rate and purity of ginseng polysaccharides in the prepared product by selecting six different enzymes to perform enzymatic hydrolysis on ginseng in sequence.

[0034] 3. The present invention uses a specific macroporous resin to enrich and purify the crude ginseng polysaccharide, and elutes it under specific conditions, which can further increase the content of ginseng polysaccharide in the product.

[0035] 4. The present invention uses a specific anion exchange column for secondary separation and purification, which can further increase the content of ginseng polysaccharides in the product while minimizing the impact on the recovery rate. DETAILED DESCRIPTION

[0036] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0037] The raw materials used in the present invention are all commercially available, specifically: Cellulase, enzyme activity of about 11000 U / g; pectinase, enzyme activity of about 30000 U / g; papain, enzyme activity of about 100000 U / g; saccharifying enzyme, enzyme activity of about 150000 U / g; all were purchased from Ningxia Xiasheng Industrial Group Co., Ltd.

[0038] Lignin peroxidase, enzyme activity of about 0.1 U / mg; laccase, enzyme activity of about 0.5 U / mg; both were purchased from Shanghai MacLean Biochemical Technology Co., Ltd.

[0039] Dialysis bags, with a molecular weight cutoff of approximately 4000 Da, were purchased from Jiangsu Mag Biotechnology Co., Ltd.

[0040] Macroporous resin, weak polar macroporous resin, the particle size between 0.3 and 1.25 mm in diameter accounts for 95% to 100%, the water content is 60% to 70%, and the specific surface area is 480 to 520 m 2 / g, average pore size 12~16nm, pore volume 0.73~0.77mL / g; purchased from Shanghai MacLean Biochemical Technology Co., Ltd., AB-8.

[0041] Anion exchange column, ligand is diethylaminoethyl, ligand density is 35~45μmol / mL, filler particle size is 45~55μm, adsorption capacity is 170~190mg HSA / mL; purchased from Shanghai Chenqiao Biotechnology Co., Ltd., DE-52.

[0042] Example 1 This embodiment provides a method for enzymatic extraction of ginseng polysaccharides, the steps of which are: S1. Dry the ginseng and grind it through a 40-mesh sieve to obtain dry ginseng powder, add distilled water and boil for 3 hours, extract 3 times and combine the decoctions, concentrate and add ethanol until the alcohol content of the system is about 80%, let it stand for 24 hours and then centrifuge to obtain a precipitate, and dry the precipitate; S2, mixing the dried precipitate with distilled water, adding enzyme for enzymatic extraction, and obtaining an extract; S3, concentrating the extract to 22% of the volume of the extract, removing the precipitate by centrifugation, placing the supernatant into a dialysis bag, dialyzing it, and concentrating and drying it to obtain crude ginseng polysaccharides; S4. Dissolve the crude ginseng polysaccharide in distilled water for separation and purification.

[0043] In step S2, the specific steps of enzymatic extraction are: A1. Mix the dried precipitate with distilled water, heat to 100°C and stir until there are no particles, then cool to 50°C and adjust the pH of the system to 5.0, add cellulase and pectinase, enzymolyze for 3.5 hours and then boil to inactivate; A2, cool down to 52℃ and adjust the system pH to 6.5, add papain for enzymatic hydrolysis for 2.5h and then boil to inactivate; A3, cool down to 55℃ and adjust the system pH to 5.0, add saccharifying enzyme for enzymatic hydrolysis for 2.5h and then boil to inactivate; A4. Cool down to 35°C and adjust the system pH to 4.5. Add lignin peroxidase and laccase for enzymolysis for 1.5 hours and then boil and inactivate to obtain an extract.

[0044] In step A1, the mass ratio of the dried precipitate to distilled water is 1:15.

[0045] The amount of cellulase used is 20U / g dry ginseng powder; the amount of pectinase used is 10U / g dry ginseng powder; the amount of papain used is 8U / g dry ginseng powder; the amount of saccharifying enzyme used is 10U / g dry ginseng powder; the amount of lignin peroxidase used is 3U / g dry ginseng powder; and the amount of laccase used is 3 / g dry ginseng powder.

[0046] In step S4, the specific steps of separation and purification are: B1. Dissolve the crude ginseng polysaccharide in distilled water, stir evenly and let stand for 24 hours, filter to remove insoluble matter, pass through a macroporous resin at a flow rate of 3BV / h, elute with 4BV water and 3BV 25% ethanol solution in sequence, collect the eluate, concentrate and dry under reduced pressure to obtain substance 1; B2. Dissolve the substance in distilled water, stir evenly and let it stand for 24 hours, filter to remove insoluble matter, use an anion exchange column for secondary separation and purification, collect the sample after elution, concentrate and dry to obtain ginseng polysaccharide.

[0047] The solid-liquid ratio of crude ginseng polysaccharide to distilled water in step B1 is 1 g:10 mL; the solid-liquid ratio of substance 1 to distilled water in step B2 is 1 g:10 mL.

[0048] The flow rate during elution in step B2 is 12 mL / min, and distilled water, 0.1 mol / L sodium chloride aqueous solution, 0.3 mol / L sodium chloride aqueous solution, and 0.5 mol / L sodium chloride aqueous solution are used for elution in sequence.

[0049] Example 2 The difference between this embodiment and embodiment 1 is that the amount of cellulase used is 30 U / g dry ginseng powder; the amount of pectinase used is 5 U / g dry ginseng powder.

[0050] Example 3 The difference between this embodiment and embodiment 1 is that the dosage of the lignin peroxidase is 2 U / g dry ginseng powder; the dosage of the laccase is 4 U / g dry ginseng powder.

[0051] Comparative Example 1 The difference between this ratio and Example 1 is that in step S2, the specific steps of enzymatic extraction are: mixing the dried precipitate and distilled water, heating to 100° C. and stirring until there are no particles, cooling to 45-55° C. and adjusting the pH of the system to 4.5-5.0, adding enzyme, enzymolysis for 10 hours and then boiling and inactivating to obtain an extract.

[0052] Comparative Example 2 The difference between this ratio and Example 1 is that in step S2, the specific steps of enzymatic extraction are: mixing the dried precipitate and distilled water, heating to 100°C and stirring until there are no particles, cooling to 50°C and adjusting the pH of the system to 5.0, adding cellulase and pectinase, enzymolyzing for 3.5 hours and then boiling and inactivating to obtain an extract.

[0053] Comparative Example 3 The difference between this ratio and Example 3 is that in step S2, the specific steps of enzymatic extraction are: A1. Mix the dried precipitate with distilled water, heat it to 100°C and stir until there are no particles, cool it to 35°C and adjust the pH of the system to 4.5, add lignin peroxidase and laccase for enzymatic hydrolysis for 1.5 hours, and then boil and inactivate; A2, cool down to 52℃ and adjust the system pH to 6.5, add papain for enzymatic hydrolysis for 2.5h and then boil to inactivate; A3, cool down to 55℃ and adjust the system pH to 5.0, add saccharifying enzyme for enzymatic hydrolysis for 2.5h and then boil to inactivate; A4. Cool down to 50°C and adjust the pH of the system to 5.0, add cellulase and pectinase, and inactivate by boiling after enzymolysis for 3.5 hours to obtain an extract.

[0054] Comparative Example 4 The difference between this ratio and Example 1 is that the amount of papain used is 15 U / g dry ginseng powder; the amount of saccharifying enzyme used is 20 U / g dry ginseng powder.

[0055] Comparative Example 5 The difference between this ratio and Example 6 is that the amount of cellulase used is 40 U / g dry ginseng powder; the amount of pectinase used is 20 U / g dry ginseng powder.

[0056] Comparative Example 6 The difference between this ratio and Example 1 is that the macroporous resin is a non-polar macroporous resin, the particles with a diameter between 0.3 and 1.25 mm account for ≥90%, the water content is 70% to 80%, the specific surface area is 480 to 520 m2 / g, the average pore diameter is 8.5 to 9 nm, and the pore volume is 2.1 to 2.15 mL / g; purchased from Tianjin Haoju Resin Technology Co., Ltd., D3520.

[0057] Comparative Example 7 The difference between this ratio and Example 1 is that in step S4, the specific steps of separation and purification are: B1. Dissolve the crude ginseng polysaccharide in distilled water, stir evenly and let stand for 24 hours, filter to remove insoluble matter, pass through a macroporous resin at a flow rate of 3BV / h, elute with 4BV water and 3BV 35% ethanol solution in sequence, collect the eluate, concentrate and dry under reduced pressure to obtain substance 1; B2. Dissolve the substance in distilled water, stir evenly and let it stand for 24 hours, filter to remove insoluble matter, use an anion exchange column for secondary separation and purification, collect the sample after elution, concentrate and dry to obtain.

[0058] Comparative Example 8 The difference between this ratio and Example 1 is that in step S4, the specific steps of separation and purification are: dissolving the crude ginseng polysaccharide in distilled water, stirring evenly and letting it stand for 24 hours, filtering to remove insoluble matter, passing it through a macroporous resin at a flow rate of 3BV / h, eluting with 4BV of water and 3BV of 25% ethanol solution in turn, collecting the eluate, and concentrating and drying under reduced pressure to obtain the product.

[0059] Comparative Example 9 The difference between this ratio and Example 1 is that the flow rate during elution in step B2 is 18 mL / min, and distilled water, 0.1 mol / L sodium chloride aqueous solution, and 0.3 mol / L sodium chloride aqueous solution are used for elution in sequence.

[0060] Performance Testing Referring to the method in the paper "Study on the Purification and Processing Technology of Mulberry Polysaccharides and Comparison of Anti-fatigue Activity Before and After Polysaccharide Purification" published by Jia Xinhui, the polysaccharide content and polysaccharide recovery rate in the final product were tested. The results are shown in Table 1.

[0061] Table 1 Measurement results Polysaccharide purity / % Polysaccharide recovery rate / % Example 1 95.7 91.5 Example 2 95.6 91.2 Example 3 95.5 91.3 Comparative Example 1 63.2 46.9 Comparative Example 2 78.3 67.2 Comparative Example 3 74.1 62.9 Comparative Example 4 84.5 72.8 Comparative Example 5 87.7 75.2 Comparative Example 6 81.2 68.5 Comparative Example 7 79.7 67.6 Comparative Example 8 85.6 74.2 Comparative Example 9 90.3 83.6 According to statistics, the ginseng polysaccharide prepared by the embodiments of the present invention 1~3, not only the polysaccharide content in the product is high, but also the polysaccharide recovery rate is relatively high, indicating that the extraction efficiency is high. There is no step-by-step enzymolysis during enzymolysis in comparative example 1, and comparative example 2 does not add papain, saccharifying enzyme, lignin peroxidase, laccase, comparative example 3 is not first enzymolyzed with cellulase and pectinase, comparative example 4 papain, saccharifying enzyme dosage is too high, comparative example 5 cellulase and pectinase dosage is too high, comparative example 6 macroporous resin does not meet the requirements, comparative example 7 elution time ethanol concentration is too high, comparative example 8 does not use anion exchange column to carry out secondary separation and purification, comparative example 9 elution time flow rate is high and eluent salt concentration does not meet the requirements, and the polysaccharide content in the prepared ginseng polysaccharide is low, and extraction efficiency is also relatively low. Therefore, the raw material and method described in the present application are used to significantly improve the extraction rate and purity of ginseng polysaccharide, while maintaining its biological activity, in accordance with the relevant provisions in the Food Safety Law of the People's Republic of China, it can therefore be applied to the field of food, with anti-oxidation, anti-inflammatory, immunomodulation and other functional effects.

[0062] The above is a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for enzymatic extraction of ginseng polysaccharides, characterized in that: The following steps are involved: S1. Dry the ginseng material and grind it through a 30-50 mesh sieve to obtain dry ginseng powder, add distilled water and boil for 2-4 hours, extract 2-3 times and combine the decoctions, add ethanol after concentration until the alcohol content of the system is ≥80%, let it stand for 24 hours and then centrifuge to obtain a precipitate, and dry the precipitate; S2, mixing the dried precipitate with distilled water, adding enzyme for enzymatic extraction, and obtaining an extract; S3, concentrating the extract to 20% to 25% of the volume of the extract, removing the precipitate by centrifugation, placing the supernatant into a dialysis bag, dialyzing it, and concentrating and drying it to obtain crude ginseng polysaccharides; S4, dissolving the crude ginseng polysaccharide in distilled water for separation and purification to obtain; The enzymes include cellulase, pectinase, papain, saccharifying enzyme, lignin peroxidase and laccase.

2. The method for enzymatic extraction of ginseng polysaccharides according to claim 1, characterized in that: In step S2, the specific steps of enzymatic extraction are: A1. Mix the dried precipitate with distilled water, heat it to 100℃ and stir until there are no particles, then cool it to 45~55℃ and adjust the pH of the system to 4.5~5.0, add cellulase and pectinase, enzymolyze for 3~4h and then boil to inactivate; A2. Cool down to 50-55℃, adjust the system pH to 6.0-7.0, add papain for enzymatic hydrolysis for 2-3h, and then boil to inactivate; A3. Cool down to 50-60℃, adjust the system pH to 4.5-5.0, add saccharifying enzyme for enzymatic hydrolysis for 2-3h, and then boil to inactivate; A4. Cool down to 30-40℃ and adjust the system pH to 4.5-5.

0. Add lignin peroxidase and laccase for enzymolysis for 1-2h and then boil to inactivate to obtain the extract.

3. The method for enzymatic extraction of ginseng polysaccharides according to claim 2, characterized in that: The amount of cellulase used is 10-30 U / g dry ginseng powder; the amount of pectinase used is 5-15 U / g dry ginseng powder; the amount of papain used is 5-10 U / g dry ginseng powder; the amount of saccharifying enzyme used is 5-15 U / g dry ginseng powder; the amount of lignin peroxidase used is 0.5-5 U / g dry ginseng powder; and the amount of laccase used is 0.5-5 U / g dry ginseng powder.

4. The method for enzymatic extraction of ginseng polysaccharides according to claim 3, characterized in that: The molecular weight cut-off of the dialysis bag is 3500-5000Da.

5. The method for enzymatic extraction of ginseng polysaccharides according to claim 1, characterized in that: In step S4, the specific steps of separation and purification are: B1. Dissolve the crude ginseng polysaccharide in distilled water, stir evenly and let stand for 24 hours, filter to remove insoluble matter, pass through a macroporous resin at a flow rate of 2.5-3.5 BV / h, and elute with 3-5 BV of water and 2-4 BV of 20%-28% ethanol solution in sequence, collect the eluate, concentrate and dry under reduced pressure to obtain substance 1; B2. Dissolve the substance in distilled water, stir evenly and let it stand for 24 hours, filter to remove insoluble matter, use an anion exchange column for secondary separation and purification, collect the sample after elution, concentrate and dry to obtain ginseng polysaccharide.

6. The method for enzymatic extraction of ginseng polysaccharides according to claim 5, characterized in that: The macroporous resin is a weakly polar macroporous resin, with particles having a diameter between 0.3 and 1.25 mm accounting for 95% to 100%, a water content of 60% to 70%, a specific surface area of ​​480 to 520 m2 / g, an average pore diameter of 12 to 16 nm, and a pore volume of 0.73 to 0.77 mL / g.

7. The method for enzymatic extraction of ginseng polysaccharides according to claim 5, characterized in that: The ligand of the anion exchange column is diethylaminoethyl, the ligand density is 35-45 μmol / mL, the filler particle size is 45-55 μm, and the adsorption load is 170-190 mg HSA / mL.

8. The method for enzymatic extraction of ginseng polysaccharides according to claim 5, characterized in that: The flow rate during elution in step B2 is 11-14 mL / min, and distilled water, 0.1 mol / L sodium chloride aqueous solution, 0.3 mol / L sodium chloride aqueous solution, and 0.5 mol / L sodium chloride aqueous solution are used for elution in sequence.

9. Ginseng polysaccharide prepared by the enzymatic extraction method of ginseng polysaccharide according to any one of claims 1 to 8.

10. Use of the ginseng polysaccharide according to claim 9 in the food field.