CN112375153A - Method for extracting ginseng flower polysaccharide by utilizing multistage countercurrent-enzymolysis coupling technology - Google Patents
Method for extracting ginseng flower polysaccharide by utilizing multistage countercurrent-enzymolysis coupling technology Download PDFInfo
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- GNPVGFCGXDBREM-UHFFFAOYSA-N germanium atom Chemical compound [Ge] GNPVGFCGXDBREM-UHFFFAOYSA-N 0.000 description 1
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Classifications
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/0003—General processes for their isolation or fractionation, e.g. purification or extraction from biomass
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/163—Sugars; Polysaccharides
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/125—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives containing carbohydrate syrups; containing sugars; containing sugar alcohols; containing starch hydrolysates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/04—Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/14—Preparation of compounds containing saccharide radicals produced by the action of a carbohydrase (EC 3.2.x), e.g. by alpha-amylase, e.g. by cellulase, hemicellulase
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
Abstract
The invention relates to a method for extracting ginseng flower polysaccharide by utilizing a multistage countercurrent-enzymolysis coupling technology, belonging to the field of extraction of plant active ingredients. In order to overcome the technical defects of low extraction rate of ginseng flower polysaccharide and high water consumption in the production process of products in the prior art, the invention provides a method for extracting ginseng flower polysaccharide by taking ginseng flowers as raw materials and utilizing a multistage countercurrent-enzymolysis coupling technology. The method applies a multistage countercurrent-enzymolysis coupling technology to the extraction of the ginseng flower polysaccharide, adopts one or more of cellulase, hemicellulase, ligninase, amylase, protease and naringinase to be matched with a tank-group countercurrent extraction process to extract the ginseng flower polysaccharide, and obtains the ginseng flower polysaccharide by ultrafiltration, impurity removal, concentration and drying of an extracting solution.
Description
Technical Field
The invention relates to a method for extracting ginseng flower polysaccharide by utilizing a multistage countercurrent-enzymolysis coupling technology, in particular to a method for extracting ginseng flower polysaccharide by taking ginseng flower as a raw material and utilizing one or more of cellulase, hemicellulase, ligninase, amylase and protease to match a tank-group countercurrent extraction process, and belongs to the field of extraction of plant active ingredients.
Background
Ginseng flower, also known as Shencao flower, is an unopened umbelliform inflorescence of Panax ginseng C.A.Meyer of Panax of Araliaceae. Collected in bud stage and naturally dried. The ginseng flower is full of body, crisp, easy to twist and break, unique in fragrance and bitter and panne in taste. Each ginseng blooms after growing for 3 years, only one inflorescence exists in each ginseng every year, 35 kilograms of ginseng can only collect one kilogram of ginseng flowers, and the ginseng flower is prepared by manual collection, solarization and drying, belongs to ginseng essence, is a natural tonic, belongs to precious medicinal materials in the world, and is called green gold. The compendium of the herbal records that the ginseng flower is sweet and bitter in taste and mild in nature, is suitable for people who have deficiency of yin and hyperactivity of fire and are not suitable for nourishing ginseng, the ginseng flower has special fragrance, and drinking the ginseng flower tea can quench thirst, detoxify, promote the production of body fluid and consume no gas, and is suitable for hot drink after meals. Its nutrient composition is 3.2 times of ginseng, and the ginseng flower contains germanium element which is called "miraculous element" by medical field, and its content is rich, and is 3 times of ganoderma and 55 times of luhui. Has excellent activity and oxidation resistance. It has effects in eliminating free radicals, improving organism environment, promoting metabolism, preventing cell aging, improving immunity, and preventing and treating tumor. The ginseng flower elements can make eaters have vigorous qi and blood, full energy, brain and intelligence development, and have outstanding health care effects in the aspects of refreshing, resisting cancer, conditioning gastrointestinal functions, relieving climacteric syndrome and the like, and the ginseng flower elements are identified by a plurality of authorities in the world and are called as 'universal health care products for immune health care'.
The ginseng flower contains similar components in ginseng root, such as ginsenoside, polysaccharide, protein, volatile oil and other active components. Ginsenoside has effects of invigorating qi, strengthening body constitution, and delaying aging; the ginseng flower polysaccharide has various biological activities of oxidation resistance, immunoregulation, tumor resistance, radiation resistance, blood sugar reduction and the like, wherein the extraction of the ginseng flower polysaccharide is the key point of resource utilization of ginseng flowers. The reflux method is a traditional polysaccharide extraction method and has the defects of long extraction time, high energy consumption, large solvent consumption, low yield, relatively low polysaccharide activity and the like. In recent years, methods such as flash extraction (HE), Ultrasonic Extraction (UE), Microwave Extraction (ME) and the like are widely applied to extraction of plant polysaccharides, and the methods have the characteristics of short time, small solvent consumption, high extraction yield and the like, but different polysaccharide extraction methods not only influence the extraction efficiency of polysaccharides, but also influence the structure and activity of the polysaccharides, thereby influencing the nutritional characteristics of the ginseng flower extract. In addition, the development of plant polysaccharide industry is directly restricted by a large amount of waste water generated in the process of extracting plant polysaccharide. Therefore, establishing an extraction method of ginseng flowers with high oxidation resistance and low wastewater discharge is the most important technical disadvantage in the field.
Disclosure of Invention
In order to solve the problems of low extraction efficiency, high wastewater discharge amount and high energy consumption of the ginseng flower polysaccharide in the prior art, the invention aims to provide a method for extracting the ginseng flower polysaccharide by taking ginseng flowers as raw materials and utilizing a multistage countercurrent-enzymolysis coupling technology. The method applies a multi-stage countercurrent-enzymolysis coupling technology to a preparation method of ginseng flower polysaccharide for the first time, enzymolysis extracting solution is obtained by adopting an enzymolysis matched tank-group type multi-stage countercurrent extraction process, then ultrafiltration concentrated solution is obtained by an ultrafiltration method, impurities are removed, and the ginseng flower polysaccharide can be obtained by drying the concentrated solution.
The invention realizes the technical effects through the following technical scheme:
the invention relates to a method for extracting ginseng flower polysaccharide by taking ginseng flower as a raw material and utilizing a multistage countercurrent-enzymolysis coupling technology, which comprises four steps of ginseng flower pretreatment, multistage countercurrent-enzymolysis coupling extraction, purification and spray drying, and the method comprises the following specific steps of:
1) pretreatment: pulping fresh flos Ginseng with water content of 30-50% (if the water content is too low, adding appropriate amount of purified water) to obtain paste, and performing multi-stage countercurrent-enzymolysis coupled extraction in a multi-stage countercurrent tank; or pulverizing dried flos Ginseng to below 20 mesh, adding 6-10 times of purified water, and performing multi-stage countercurrent-enzymolysis coupled extraction in a multi-stage countercurrent tank set;
2) multistage countercurrent-enzymolysis coupling extraction: 3-5 soaking extraction tanks are arranged for continuous countercurrent operation, in the extraction process, the reverse operation of an extraction phase and a raffinate phase is realized by reverse transfer of an extraction aqueous solution, and the dissolved polysaccharide is removed in time, so that a polysaccharide concentration gradient is formed among the tank groups. Keeping the temperature in the tank at 40-60 ℃ and the pH value at 6.0-8.0 in the reverse extraction process, reacting for 0.5-2h, adding one or more of cellulase, hemicellulase, ligninase, protease and amylase, improving the polysaccharide dissolution rate through enzymolysis, discharging the final extract from the first-stage feeding tank, and discharging the residual phase, namely the extracted residue from the last-stage adding tank of extraction water to obtain an enzymolysis extract;
3) and (3) purification: filtering or centrifuging the enzymolysis extracting solution, and passing the obtained filtrate or centrifugate through an ultrafiltration membrane with the molecular weight cutoff of 2000-3500Da to achieve the purposes of desalting and deproteinizing to obtain an ultrafiltration concentrated solution;
4) and (3) drying: concentrating the ultrafiltered concentrated solution to obtain solid content of 25-35%, and spray drying to obtain flos Ginseng oligosaccharide.
Preferably, the adding amount of the cellulase, the hemicellulase and the ligninase in the step 2) is 500 enzyme activity units per kilogram of dry ginseng flowers, and the optimal enzymolysis conditions of the cellulase, the hemicellulase and the ligninase are as follows: the pH value in the extraction tank is 6.0, the enzymolysis temperature is 50 ℃, and the heat preservation time is 1 hour.
Preferably, the adding amount of the amylase in the step 2) is 450 enzyme activity units per kilogram of the dry ginseng flowers, and the optimal enzymolysis conditions of the amylase are as follows: the pH value in the extraction tank is 7.0, the enzymolysis temperature is 55 ℃, and the heat preservation time is 2 hours.
Preferably, the adding amount of the protease in the step 2) is 200 enzyme activity units per kilogram of the dried ginseng flower, and the optimal enzymolysis conditions of the protease are as follows: the pH value in the extraction tank is 8.0, the enzymolysis temperature is 45 ℃, and the heat preservation time is 1 hour.
Preferably, the mesh number of the filter cloth in the filtering operation in the step 3) is 600-1000 meshes; the centrifugal speed in the centrifugal operation in the step 3) is 3500 rpm, and the centrifugal time is 10 min.
Further preferably, a certain amount of naringinase is added in the step 2), and the addition amount is 200 enzyme activity units per kilogram of dry ginseng flower.
The applicant has conducted intensive studies on the preparation method of the present invention based on the combination of the above conditions, and found that, in a most preferred embodiment of the present invention, the specific steps are as follows:
1) pretreatment: crushing dried ginseng flowers, sieving the crushed ginseng flowers with a 20-mesh sieve, putting the crushed ginseng flowers into a multistage countercurrent soaking tank, adding 6 times of water by weight, and performing multistage countercurrent-enzymolysis coupling extraction;
2) multistage countercurrent-enzymolysis coupling extraction: setting 3-stage countercurrent extraction tanks for continuous countercurrent operation, keeping the temperature of the first-stage tank at 40 ℃, keeping the temperature of the second-stage tank at 50 ℃, keeping the temperature of the third-stage tank at 60 ℃, adjusting the pH of each tank to 7.0, adding cellulase, hemicellulase, ligninase, amylase, protease and naringinase according to 200-plus-500 enzyme activity units per kilogram of dry ginseng flowers, carrying out countercurrent extraction for 1 hour, heating to over 90 ℃, keeping the temperature for 15 minutes, inactivating enzyme, finally discharging an extracting solution from a first-stage feeding tank, and discharging residues from the third-stage tank, namely a feeding tank of extracting water to obtain a ginseng flower polysaccharide crude extracting solution;
3) and (3) purification: filtering or centrifuging the crude extractive solution, and passing the obtained filtrate or centrifugate through ultrafiltration membrane with cut-off molecular weight of 2500Da to achieve the purpose of desalting, deproteinizing and removing other impurities to obtain ultrafiltration concentrated solution;
4) and (3) drying: further concentrating the hyperconcentrated solution until the solid content reaches 30%, and spray drying to obtain flos Ginseng polysaccharide.
The effect example of the invention shows that the ginseng flower polysaccharide prepared according to the invention has high extraction efficiency, less water consumption, low energy consumption and high purity of the ginseng flower polysaccharide product.
The invention also claims the ginseng flower polysaccharide prepared by the method.
The ginseng flower polysaccharide can be further prepared into health food, food additives, feed additives and functional agricultural products, and can remarkably enhance the immunity of organisms when being used for the health food, the food additives or the feed additives.
Compared with the prior art, the invention has the technical advantages that:
1) the ginseng flower polysaccharide has high yield, thereby greatly reducing the production cost. The effect embodiment of the invention shows that the ginseng flower polysaccharide prepared according to the invention has high yield, the yield is between 44% and 81%, and is far higher than the ginseng flower polysaccharide prepared by the prior art, and the purity of the ginseng flower polysaccharide prepared by the invention is high and is not lower than the ginseng flower polysaccharide obtained by the prior art.
2) The water consumption in the production process is low, the discharge of waste water is reduced, and the energy consumption is reduced. The effect example of the invention shows that the water consumption for preparing 1kg of ginseng flower polysaccharide according to the invention is between 12 and 20kg, which is far lower than the water consumption for preparing ginseng flower polysaccharide in the prior art.
3) The ginseng flower polysaccharide can be further prepared into health food, food additives, feed additives and functional agricultural products, has high nutrient content, and can obviously enhance the immunity of organisms when being used for the health food, the food additives or the feed additives;
4) the invention adopts a multistage countercurrent-enzymolysis coupling technology to extract the ginseng flower polysaccharide, and adds naringinase on the basis of extracting enzyme preparations such as common cellulase, hemicellulase, ligninase, amylase, protease and the like from the ginseng flower polysaccharide, thereby not only improving the taste of the ginseng flower polysaccharide, but also obviously improving the extraction rate of the ginseng flower polysaccharide. Experimental results show that the multi-stage countercurrent-enzymolysis coupling technology for extracting the ginseng flower polysaccharide has no obvious difference from direct water extraction when a naringinase single enzyme preparation is adopted.
Detailed Description
Example 1:
crushing dehydrated dry ginseng flowers, sieving with a 20-mesh sieve, respectively adding 50kg of crushed ginseng pollen into a 3-grade countercurrent extraction tank, adding 1t of water into the first-grade tank, adjusting the pH to 7.5, adding 75kU cellulase and 30kU papain, heating the first-grade tank to 40 ℃, heating the second-grade tank to 50 ℃, heating the third-grade tank to 60 ℃, sequentially performing countercurrent enzymolysis extraction operation according to the operating procedure of the 3-grade countercurrent extraction, heating to 95 ℃ after 1 hour of extraction, keeping the temperature for 15min, and inactivating enzyme activity. Discharging the extracting solution from the first-stage tank, filtering with a plate-and-frame filter to obtain 780L of filtrate, adding 500kg of tap water for dilution, passing through a nanofiltration membrane with the molecular weight cutoff of 2500Da, concentrating the concentrated solution to 500L until the volume of the concentrated solution is concentrated to 30L, further concentrating the concentrated solution to 30% of solid matters by using a three-way concentrator, and performing spray drying to obtain 77.01kg of ginseng flower polysaccharide.
Example 2 example 10
The ginseng flower polysaccharides were prepared according to the parameters as described in table 1, and examples 2 to 10 were prepared in the same manner as in example 1, except for the following parameters.
Table 1 main parameters of example 2 to example 10
The yield, purity and water consumption of the ginseng flower polysaccharide prepared according to examples 1 to 10 of the present invention were measured. The measurement results are shown in table 2. The yield of the ginseng flower polysaccharide is ginseng flower polysaccharide/dry ginseng flower x 100%, or ginseng flower polysaccharide/(fresh ginseng flower x (1-moisture content)) × 100%. The purity of the ginseng flower polysaccharide is total sugar content/ginseng flower polysaccharide multiplied by 100%. Method for determination of total sugars the phenol-sulfuric acid method disclosed in the prior art was followed. The water consumption is calculated according to the water consumption for producing 1kg of ginseng flower polysaccharide.
Table 2 example 1-example 10 index test results for ginseng flower polysaccharide preparation
As can be seen from Table 2, the ginseng flower polysaccharide prepared according to the present invention has high yield, less water consumption during the production process, and high purity of the product, wherein the product prepared by the method described in example 9 has the highest yield, which is the best example of the present invention.
The above embodiments are only used for illustrating the technical solution of the present invention, and not for limiting the same; although the invention has been described in detail with reference to the foregoing embodiments, it will be apparent to those skilled in the art that various changes may be made and equivalents may be substituted for elements thereof; such modifications and substitutions do not depart from the spirit and scope of the present invention as set forth in the appended claims.
Claims (9)
1. A method for extracting ginseng flower polysaccharide by utilizing a multistage countercurrent-enzymolysis coupling technology is characterized by comprising four steps of ginseng flower pretreatment, multistage countercurrent-enzymolysis coupling extraction, purification and spray drying, and the specific steps are as follows:
1) pretreatment: pulping fresh flos Ginseng with water content of 30-50% (if the water content is too low, adding appropriate amount of purified water) to obtain paste, and performing multi-stage countercurrent-enzymolysis coupled extraction in a multi-stage countercurrent tank; or pulverizing dried flos Ginseng to below 20 mesh, adding 6-10 times of purified water, and performing multi-stage countercurrent-enzymolysis coupled extraction in a multi-stage countercurrent tank set;
2) multistage countercurrent-enzymolysis coupling extraction: arranging 3-5 soaking extraction tanks for continuous countercurrent operation, realizing the reverse operation of an extraction phase and a raffinate phase by reverse transfer of an extraction aqueous solution in the extraction process, timely removing the dissolved polysaccharide to form a polysaccharide concentration gradient among the tank groups, keeping the temperature of the tank at 40-60 ℃ in the reverse extraction process, controlling the pH value at 6.0-8.0, reacting for 0.5-2h, adding one or more of cellulase, hemicellulase, ligninase, protease and amylase, improving the polysaccharide dissolution rate through enzymolysis, finally discharging an extracting solution from a first-stage feeding tank, and discharging raffinate phase, namely extracted residues from a last-stage feeding tank of extraction water to obtain an enzymolysis extracting solution;
3) and (3) purification: filtering or centrifuging the enzymolysis extracting solution, and passing the obtained filtrate or centrifugate through an ultrafiltration membrane with the molecular weight cutoff of 2000-3500Da to achieve the purposes of desalting and deproteinizing to obtain an ultrafiltration concentrated solution;
4) and (3) drying: concentrating the ultrafiltered concentrated solution to obtain solid content of 25-35%, and spray drying to obtain flos Ginseng oligosaccharide.
2. The method for extracting ginseng flower polysaccharide by utilizing the multistage countercurrent-enzymolysis coupling technology according to claim 1, wherein the adding amount of cellulase, hemicellulase and ligninase in the step 2) is 500 enzyme activity units per kilogram of dry ginseng flower, and the optimal enzymolysis conditions of the cellulase, hemicellulase and ligninase are as follows: the pH value in the extraction tank is 6.0, the enzymolysis temperature is 50 ℃, and the heat preservation time is 1 hour.
3. The method for extracting ginseng flower polysaccharide by utilizing the multistage countercurrent-enzymolysis coupling technology according to claim 1, wherein the addition amount of amylase in the step 2) is 450 enzyme activity units per kilogram of dry ginseng flower, and the optimal enzymolysis conditions of the amylase are as follows: the pH value in the extraction tank is 7.0, the enzymolysis temperature is 55 ℃, and the heat preservation time is 2 hours.
4. The method for extracting ginseng flower polysaccharide by utilizing the multistage countercurrent-enzymolysis coupling technology according to claim 1, wherein the adding amount of the protease in the step 2) is 200 enzyme activity units per kilogram of dry ginseng flower, and the optimal enzymolysis conditions of the protease are as follows: the pH value in the extraction tank is 8.0, the enzymolysis temperature is 45 ℃, and the heat preservation time is 1 hour.
5. The method for extracting ginseng flower polysaccharide by utilizing the multistage countercurrent-enzymolysis coupling technology as claimed in claim 1, wherein the mesh number of the filter cloth in the filtering operation in the step 3) is 600-1000 meshes; the centrifugal speed in the centrifugal operation in the step 3) is 3500 rpm, and the centrifugal time is 10 min.
6. The method for extracting ginseng flower polysaccharide by utilizing the multistage countercurrent-enzymolysis coupling technology as claimed in claim 1, wherein a certain amount of naringinase is added in the step 2), and the addition amount is 200 enzyme activity units per kilogram of dry ginseng flower.
7. The method for extracting ginseng flower polysaccharide by utilizing the multistage countercurrent-enzymolysis coupling technology according to claim 1, which is characterized by comprising the following specific steps:
1) pretreatment: crushing dried ginseng flowers, sieving the crushed ginseng flowers with a 20-mesh sieve, putting the crushed ginseng flowers into a multistage countercurrent soaking tank, adding 6 times of water by weight, and performing multistage countercurrent-enzymolysis coupling extraction;
2) multistage countercurrent-enzymolysis coupling extraction: setting 3-stage countercurrent extraction tanks for continuous countercurrent operation, keeping the temperature of the first-stage tank at 40 ℃, keeping the temperature of the second-stage tank at 50 ℃, keeping the temperature of the third-stage tank at 60 ℃, adjusting the pH of each tank to 7.0, adding cellulase, hemicellulase, ligninase, amylase, protease and naringinase according to 200-plus-500 enzyme activity units per kilogram of dry ginseng flowers, carrying out countercurrent extraction for 1 hour, heating to over 90 ℃, keeping the temperature for 15 minutes, inactivating enzyme, finally discharging an extracting solution from a first-stage feeding tank, and discharging residues from the third-stage tank, namely a feeding tank of extracting water to obtain a ginseng flower polysaccharide crude extracting solution;
3) and (3) purification: filtering or centrifuging the crude extractive solution, and passing the obtained filtrate or centrifugate through ultrafiltration membrane with cut-off molecular weight of 2500Da to achieve the purpose of desalting, deproteinizing and removing other impurities to obtain ultrafiltration concentrated solution;
4) and (3) drying: further concentrating the hyperconcentrated solution until the solid content reaches 30%, and spray drying to obtain flos Ginseng polysaccharide.
8. A ginseng flower polysaccharide prepared according to the method of any one of claims 1 to 7.
9. Use of the ginseng flower polysaccharide of claim 8 in the preparation of health food, food additives, feed additives and functional agricultural products.
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| CN115651088A (en) * | 2022-08-23 | 2023-01-31 | 辽宁成大生物股份有限公司 | Preparation method and application of ginseng total polysaccharide, ginseng total polysaccharide vaccine adjuvant and vaccine composition thereof |
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