Preparation method of tremella polysaccharide

Abstract

The invention relates to the technical field of deep processing of edible fungi, and discloses a preparation method of tremella polysaccharide, which comprises the following steps: S1, cleaning and crushing a tremella raw material, and carrying out repeated freezing and thawing treatment; s2, mixing the frozen and thawed tremella fuciformis raw material with a buffer solution, adding a compound enzyme, and putting the mixture into microwave equipment to obtain an extracting solution; s3, performing enzyme deactivation and centrifugation on the extracting solution, and performing primary purification through two-aqueous-phase extraction; the tremella fuciformis is used as a raw material, the natural structural integrity of polysaccharide can be better kept through repeated freezing and thawing pretreatment, polysaccharide activity is maximally released and protected at low temperature through microwave and compound enzyme synergistic extraction, accurate and green purification is achieved through two-aqueous-phase extraction and ultrafiltration classification, and the method is suitable for large-scale industrial production. The moisture retention and oxidation resistance of the tremella polysaccharide are endowed and remarkably enhanced through directional carboxymethylation modification, and finally, perfect shaping is performed through freeze drying, so that the tremella polysaccharide with high activity and high additional value is obtained.

Description

Technical Field

[0001] This invention relates to the field of edible fungi deep processing technology, specifically a method for preparing Tremella polysaccharide. Background Technology

[0002] Tremella, commonly known as white fungus or snow fungus, is a precious medicinal and edible fungus with a long history of cultivation and consumption in my country and East Asia. Modern scientific research reveals that many of Tremella's health benefits are mainly attributed to its core active ingredient—Tremella polysaccharide. Tremella polysaccharide is a class of high-molecular-weight heteropolysaccharides extracted from the fruiting body or fermented mycelium of Tremella. Its main structure is an acidic heteropolysaccharide with α-(1→3)-mannose as the main chain and branches connected to monosaccharides such as glucuronic acid, xylose, and fucose, forming a highly branched and complex spatial structure. This unique structure, especially its acetylation modification... The decorative and helical conformations are considered the material basis for its excellent biological activity. Unlike common polysaccharides such as starch and cellulose, Tremella polysaccharide has good water solubility and viscosity, and can form a stable gel. Tremella polysaccharide can enhance the body's specific and non-specific immune responses through multiple pathways, such as activating macrophages, promoting lymphocyte proliferation, and inducing cytokine production, without the toxic side effects of typical immunosuppressants. Moreover, as an effective free radical scavenger, Tremella polysaccharide can reduce oxidative stress damage and protect cell membranes and biological macromolecules, showing potential in delaying skin and body aging.

[0003] Currently, most methods for preparing polysaccharides from Tremella fuciformis involve hot water extraction. This method relies primarily on heat and the diffusion of water molecules. Heat softens and swells the cell walls, allowing water molecules to penetrate and dissolve the polysaccharides. The polysaccharides then diffuse into the solvent outside the cells. This process is entirely passive, and its efficiency depends on the thermal motion and concentration gradient of the molecules, making it very slow. Tremella fuciformis polysaccharides exist within the solid mycelial cell walls and gelatinous matrix, resulting in a dense structure. Hot water soaking alone is insufficient to completely destroy the cell structure in a short time, leading to a large amount of polysaccharides not being fully released. The yield from a single extraction is usually low. To improve the yield, industrial processes often require 2-3 repeated extractions, followed by merging the extracts. While this increases the overall yield, it significantly prolongs the total extraction time and dilutes the extract. Summary of the Invention

[0004] The purpose of this invention is to provide a method for preparing Tremella polysaccharide to solve the problems mentioned in the background art.

[0005] To achieve the above objectives, the present invention provides the following technical solution: a method for preparing Tremella polysaccharide, comprising the following steps:

[0006] S1. After cleaning and crushing the raw material of white fungus, it is subjected to repeated freeze-thaw treatment;

[0007] S2. Mix the frozen and thawed tremella raw material with buffer solution, add compound enzyme and place in a microwave device to obtain extract;

[0008] S3. After enzyme inactivation and centrifugation, the extract is initially purified by aqueous two-phase extraction.

[0009] S4. Collect the polysaccharide phase after aqueous two-phase extraction and separate it through an ultrafiltration membrane;

[0010] S5. Dissolve the fractionated polysaccharide in NaOH solution, react with chloroacetic acid, neutralize with acid, and then perform ultrafiltration.

[0011] S6. The modified polysaccharide solution was rapidly frozen at ultra-low temperature, and then the ice was directly sublimated in a vacuum environment to obtain Tremella polysaccharide.

[0012] Preferably, the repeated freeze-thaw process in step S1 includes:

[0013] S1.1 Select dried white fungus without sulfur fumigation as raw material;

[0014] S1.2 Rinse the surface with 5℃ deionized water for 30 seconds, with a material-to-water ratio of 1:6, and immediately blow with 0.3MPa clean air for 90 seconds to remove free water from the surface;

[0015] S1.3. The raw material of white fungus is coarsely ground by liquid nitrogen continuous feeding low temperature grinder and passed through a 40-mesh sieve to obtain fine white fungus powder;

[0016] S1.4 Add the fine powder of tremella to 4℃ deionized water at a ratio of 1:2, stir quickly to form a slurry, dispense into 500mL sterile PET bottles, and let stand in a -80℃ refrigerator for 2 hours.

[0017] S1.5 After freezing, remove the PET bottle and thaw it by shaking in a 25°C water bath for 15 minutes until the core temperature is ≥15°C. Then, put it back into the refrigerator to freeze and repeat the cycle 2-3 times.

[0018] S1.6. The freeze-thawed slurry is centrifuged and dehydrated using a 200-mesh nylon filter cloth to obtain a wet material with a moisture content of 65–70%.

[0019] Preferably, in step S1, when the raw material of Tremella is repeatedly frozen and thawed, the freeze-thaw process can be stopped when the mycelial breakage rate is ≥90% as determined by microscopic examination.

[0020] Preferably, the collaborative extraction process in step S2 includes:

[0021] S2.1 First, prepare a 0.05M phosphate buffer solution with a pH between 4.5 and 6.0, and pre-cool the buffer solution to 10°C.

[0022] S2.2. Mix the pretreated tremella raw material with a sufficient amount of buffer solution to ensure that the raw material is completely submerged and forms a homogenate;

[0023] S2.3 Add the prepared compound enzyme to the mixture and stir until homogeneous;

[0024] S2.4 Place the mixture in a constant temperature water bath and stir slowly at the optimal temperature of the enzyme for 20-40 minutes for pretreatment.

[0025] S2.5 Place the pre-enzymatically hydrolyzed mixture container into the microwave reactor, add the magnetic stir bar, turn on the stirring, set the temperature to the optimal range for the enzyme, and operate the microwave in pulse mode to quickly reach the set temperature for synergistic extraction.

[0026] Preferably, the formulation of the complex enzyme in S2 is: 0.59% cellulase, 0.43% pectinase, and 0.98% papain.

[0027] Preferably, the process of performing aqueous two-phase extraction in step S3 includes:

[0028] S3.1 After microwave co-processing, immediately transfer the container to a boiling water bath and heat for 5-10 minutes to completely inactivate the enzyme and terminate the reaction. While the mixture is hot, centrifuge and collect the supernatant, which is the crude extract containing Tremella polysaccharide.

[0029] S3.2. Prepare an aqueous two-phase system with 24.68% ethanol and 28% ammonium sulfate. Mix the crude polysaccharide extract with the prepared aqueous two-phase system in a container at a volume ratio of 1:1 to 1:5.

[0030] S3.3 Stir thoroughly for 10-30 minutes to ensure that the substances are fully distributed between the two phases. Then stop stirring and let stand for 5-30 minutes. The solution will spontaneously form two clear phases. Polysaccharides will be enriched in the upper ethanol phase, while a large amount of proteins, cell debris and other impurities will be distributed in the lower salt phase.

[0031] S3.4 After the phase separation is clear, use a pipette to collect the target phase rich in polysaccharides, which is the upper alcohol phase.

[0032] Preferably, before separation by ultrafiltration membrane in step S4, the polysaccharide enriched phase collected by aqueous two-phase extraction can be centrifuged at low speed to remove tiny particles and prevent particulate matter from clogging the ultrafiltration membrane pores.

[0033] Preferably, the process of modifying Tremella polysaccharide in step S5 includes:

[0034] S5.1 Prepare a NaOH solution and cool it for later use. Weigh out chloroacetic acid and dissolve it in ethanol.

[0035] S5.2 Dissolve the target polysaccharide fraction after ultrafiltration fractionation in a pre-cooled NaOH solution;

[0036] S5.3 Place the mixture in an ice-water bath and react for 30-60 minutes with vigorous stirring to generate active sodium alkoxide anions;

[0037] S5.4 Under continuous stirring and ice bath conditions, slowly and in batches add chloroacetic acid solution to the alkalization mixture. After the addition is complete, remove the ice bath and gradually heat the reaction system to the set temperature of 50-70℃. Stir the reaction at this temperature for 2-6 hours.

[0038] S5.5 After the reaction is complete, cool the mixture to room temperature. While stirring, slowly adjust the pH to about 7.0 with dilute hydrochloric acid to neutralize the excess alkali and terminate the reaction. Add 2-3 times the volume of anhydrous ethanol and let stand to precipitate the modified polysaccharide. Centrifuge to collect the precipitate.

[0039] S5.6. The product of alcohol precipitation is redissolved in water and subjected to ultrafiltration to concentrate and completely desalinate it.

[0040] Preferably, in step S6, after concentrating the ultrafiltration purified carboxymethyl polysaccharide solution to a solid content of 2%-10%, the sample solution is dispensed into a freeze-drying container. The dispensed sample is then quickly placed in an ultra-low temperature freezer below -50°C to ensure that the sample is completely frozen, opaque, and in a hard solid state. The freeze dryer is started in advance, and the condenser temperature is lowered to below -50°C. The completely frozen sample is then removed from the freezing equipment and quickly transferred into the pre-cooled freeze dryer drying chamber. The chamber door is closed, the vacuum pump is started, and sublimation begins when the system vacuum level stabilizes below 10-30 Pa.

[0041] Preferably, during sublimation in step S6, high vacuum and low temperature must be maintained, and the sample temperature must always be lower than its eutectic point temperature. After the first drying is completed, the sample temperature is gradually and slowly increased and maintained at this temperature for 4-10 hours. After drying is completed, the vacuum valve is closed first, and then dry inert gas is slowly introduced into the drying chamber until the pressure returns to normal. Then the sample is immediately taken out and quickly sealed and packaged in a dry environment to obtain the Tremella polysaccharide product.

[0042] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0043] This invention uses Tremella fuciformis as raw material and employs repeated freeze-thaw pretreatment to better maintain the natural structural integrity of polysaccharides. Microwave and compound enzyme synergistic extraction maximizes the release of polysaccharides at low temperatures while protecting their activity. Furthermore, aqueous two-phase extraction combined with ultrafiltration fractionation achieves precise and green purification. Targeted carboxymethylation modification further endows and significantly enhances the moisturizing and antioxidant properties of Tremella fuciformis polysaccharides. Finally, freeze-drying is used for perfect shaping to obtain highly active and high-value-added Tremella fuciformis polysaccharides. Attached Figure Description

[0044] Figure 1 This is a flowchart of the method of the present invention. Detailed Implementation

[0045] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0046] A method for preparing Tremella polysaccharide includes the following steps:

[0047] S1. After cleaning and crushing the raw material of white fungus, it is subjected to repeated freeze-thaw treatment;

[0048] S2. Mix the frozen and thawed tremella raw material with buffer solution, add compound enzyme and place in a microwave device to obtain extract;

[0049] S3. After enzyme inactivation and centrifugation, the extract is initially purified by aqueous two-phase extraction.

[0050] S4. Collect the polysaccharide phase after aqueous two-phase extraction and separate it through an ultrafiltration membrane;

[0051] S5. Dissolve the fractionated polysaccharide in NaOH solution, react with chloroacetic acid, neutralize with acid, and then perform ultrafiltration.

[0052] S6. The modified polysaccharide solution was rapidly frozen at ultra-low temperature, and then the ice was directly sublimated in a vacuum environment to obtain Tremella polysaccharide.

[0053] The repeated freeze-thaw process in step S1 includes:

[0054] S1.1 Select dried white fungus without sulfur fumigation as raw material;

[0055] S1.2 Rinse the surface with 5℃ deionized water for 30 seconds, with a material-to-water ratio of 1:6, and immediately blow with 0.3MPa clean air for 90 seconds to remove free water from the surface;

[0056] S1.3. The raw material of white fungus is coarsely ground by liquid nitrogen continuous feeding low temperature grinder and passed through a 40-mesh sieve to obtain fine white fungus powder;

[0057] S1.4 Add the fine powder of tremella to 4℃ deionized water at a ratio of 1:2, stir quickly to form a slurry, dispense into 500mL sterile PET bottles, and let stand in a -80℃ refrigerator for 2 hours.

[0058] S1.5 After freezing, remove the PET bottle and thaw it by shaking in a 25°C water bath for 15 minutes until the core temperature is ≥15°C. Then, put it back into the refrigerator to freeze and repeat the cycle 2-3 times.

[0059] S1.6. The freeze-thawed slurry is centrifuged and dehydrated using a 200-mesh nylon filter cloth to obtain a wet material with a moisture content of 65-70% (water crystallizes inside the cells to form ice crystals, which expand in volume. Repeated freeze-thaw cycles cause the ice crystals to mechanically pierce and tear the cell walls and membrane structures, greatly increasing their permeability and making it easier for subsequent extraction solvents and polysaccharides to enter and exit).

[0060] When repeatedly freezing and thawing the raw material of Tremella fuciformis in step S1, the freezing and thawing process can be stopped when the mycelial breakage rate is ≥90% as determined by microscopic examination.

[0061] The collaborative extraction process in step S2 includes:

[0062] S2.1 First, prepare a 0.05M phosphate buffer solution with a pH between 4.5 and 6.0, and pre-cool the buffer solution to 10°C (as a surfactant to improve the efficiency of enzymatic hydrolysis).

[0063] S2.2. Mix the pretreated tremella raw material with a sufficient amount of buffer solution to ensure that the raw material is completely submerged and forms a homogenate;

[0064] S2.3 Add the prepared compound enzyme to the mixture and stir until homogeneous;

[0065] S2.4 Place the mixture in a constant temperature water bath and stir slowly at the optimal temperature of the enzyme for 20-40 minutes for pretreatment.

[0066] S2.5 Place the pre-enzymatically hydrolyzed mixture container into the microwave reactor, add the magnetic stir bar, turn on the stirrer, set the temperature to the optimal range for the enzyme, and operate the microwave in pulse mode (the instantaneous volumetric heating effect of microwaves can greatly accelerate cell rupture and mass transfer processes). The set temperature is quickly reached with microwave assistance for synergistic extraction.

[0067] The formula for the complex enzyme in S2 is: 0.59% cellulase, 0.43% pectinase, and 0.98% papain.

[0068] The process for aqueous two-phase extraction in step S3 includes:

[0069] S3.1 After microwave co-processing, immediately transfer the container to a boiling water bath and heat for 5-10 minutes to completely inactivate the enzyme and terminate the reaction. While the mixture is hot, centrifuge and collect the supernatant, which is the crude extract containing Tremella polysaccharide.

[0070] S3.2. Prepare an aqueous two-phase system with 24.68% ethanol and 28% ammonium sulfate. Mix the crude polysaccharide extract with the prepared aqueous two-phase system in a container at a volume ratio of 1:1 to 1:5.

[0071] S3.3 Stir thoroughly for 10-30 minutes to ensure that the substances are fully distributed between the two phases. Then stop stirring and let stand for 5-30 minutes. The solution will spontaneously form two clear phases. Polysaccharides will be enriched in the upper ethanol phase, while a large amount of proteins, cell debris and other impurities will be distributed in the lower salt phase.

[0072] S3.4 After the phase separation is clear, use a pipette to collect the target phase rich in polysaccharides, which is the upper alcohol phase.

[0073] Before separation via ultrafiltration membrane in step S4, the polysaccharide-enriched phase collected by aqueous two-phase extraction can be centrifuged at low speed to remove tiny particles and prevent them from clogging the ultrafiltration membrane pores.

[0074] The process of modifying Tremella polysaccharide in step S5 includes:

[0075] S5.1 Prepare a NaOH solution and cool it for later use. Weigh out chloroacetic acid and dissolve it in ethanol.

[0076] S5.2 Dissolve the target polysaccharide component after ultrafiltration fractionation in a pre-cooled NaOH solution (if the polysaccharide is not easily soluble, it can be dispersed in water first, and then the alkali solution can be slowly added to form a uniform paste).

[0077] S5.3 Place the mixture in an ice-water bath and react for 30-60 minutes with vigorous stirring to generate active sodium alkoxide anions (low-temperature alkalization can minimize the oxidative degradation of polysaccharides in strong alkali).

[0078] S5.4 Under continuous stirring and ice bath conditions, slowly and in batches add chloroacetic acid solution to the alkalization mixture. After the addition is complete, remove the ice bath and gradually heat the reaction system to the set temperature of 50-70℃. Stir the reaction at this temperature for 2-6 hours (heating provides the energy required for the reaction, so that chloroacetic acid and sodium polyol can undergo an etherification reaction to generate carboxymethyl polysaccharide).

[0079] S5.5 After the reaction is complete, cool the mixture to room temperature. While stirring, slowly adjust the pH to about 7.0 with dilute hydrochloric acid to neutralize the excess alkali and terminate the reaction. Add 2-3 times the volume of anhydrous ethanol and let stand to precipitate the modified polysaccharide. Centrifuge to collect the precipitate.

[0080] S5.6. The product of alcohol precipitation is redissolved in water and subjected to ultrafiltration to concentrate and completely desalinate (which can efficiently remove all small molecule impurities).

[0081] In step S6, after concentrating the ultrafiltration purified carboxymethyl polysaccharide solution to a solid content of 2%-10%, the sample solution is dispensed into a freeze-drying container. The dispensed sample is then quickly placed in an ultra-low temperature freezer below -50°C (to allow the water in the sample to quickly form tiny ice crystals, preventing the ice crystals from growing and piercing the fine porous structure of the polysaccharide, thus obtaining a more fluffy finished product). This ensures that the sample is completely frozen, opaque, and in a hard solid state. The freeze dryer is started in advance, and the condenser temperature is lowered to below -50°C. The completely frozen sample is then removed from the freezing equipment and quickly transferred into the pre-cooled freeze dryer drying chamber. The chamber door is closed, the vacuum pump is started, and sublimation begins when the system vacuum level stabilizes below 10-30 Pa.

[0082] During sublimation in step S6, high vacuum and low temperature must be maintained, and the sample temperature must always be lower than its eutectic point temperature. After the first drying, the sample temperature is gradually and slowly increased and maintained at this temperature for 4-10 hours (to break the hydrogen bonds between water and polysaccharide molecules, further reduce the residual moisture of the product to 1%-3%, and improve product stability). After drying, the vacuum valve is closed first, and then dry inert gas is slowly introduced into the drying chamber until the pressure returns to normal. The sample is then immediately removed and quickly sealed and packaged in a dry environment to obtain the Tremella polysaccharide product.

[0083] It should be noted that, in this document, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitations, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0084] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A method for preparing Tremella fuciformis polysaccharide, characterized in that, Includes the following steps: S1. After cleaning and crushing the raw material of white fungus, it is subjected to repeated freeze-thaw treatment; S2. Mix the frozen and thawed tremella raw material with buffer solution, add compound enzyme and place in a microwave device to obtain extract; S3. After enzyme inactivation and centrifugation, the extract is initially purified by aqueous two-phase extraction. S4. Collect the polysaccharide phase after aqueous two-phase extraction and separate it through an ultrafiltration membrane; S5. Dissolve the fractionated polysaccharide in NaOH solution, react with chloroacetic acid, neutralize with acid, and then perform ultrafiltration. S6. The modified polysaccharide solution was rapidly frozen at ultra-low temperature, and then the ice was directly sublimated in a vacuum environment to obtain Tremella polysaccharide.

2. The method for preparing Tremella polysaccharide according to claim 1, characterized in that: The repeated freeze-thaw process in step S1 includes: S1.1 Select dried white fungus without sulfur fumigation as raw material; S1.2 Rinse the surface with 5℃ deionized water for 30 seconds, with a material-to-water ratio of 1:6, and immediately blow with 0.3MPa clean air for 90 seconds to remove free water from the surface; S1.

3. The raw material of white fungus is coarsely ground by liquid nitrogen continuous feeding low temperature grinder and passed through a 40-mesh sieve to obtain fine white fungus powder; S1.4 Add the fine powder of tremella to 4℃ deionized water at a ratio of 1:2, stir quickly to form a slurry, dispense into 500mL sterile PET bottles, and let stand in a -80℃ refrigerator for 2 hours. S1.5 After freezing, remove the PET bottle and thaw it by shaking in a 25°C water bath for 15 minutes until the core temperature is ≥15°C. Then, put it back into the refrigerator to freeze and repeat the cycle 2-3 times. S1.

6. The freeze-thawed slurry is centrifuged and dehydrated using a 200-mesh nylon filter cloth to obtain a wet material with a moisture content of 65–70%.

3. The method for preparing Tremella polysaccharide according to claim 1, characterized in that: In step S1, when repeatedly freezing and thawing the tremella raw material, the freezing and thawing process can be stopped when the mycelial breakage rate is ≥90% as determined by microscopic examination.

4. The method for preparing Tremella polysaccharide according to claim 1, characterized in that: The collaborative extraction process in step S2 includes: S2.1 First, prepare a 0.05M phosphate buffer solution with a pH between 4.5 and 6.0, and pre-cool the buffer solution to 10°C. S2.

2. Mix the pretreated tremella raw material with a sufficient amount of buffer solution to ensure that the raw material is completely submerged and forms a homogenate; S2.3 Add the prepared compound enzyme to the mixture and stir until homogeneous; S2.4 Place the mixture in a constant temperature water bath and stir slowly at the optimal temperature of the enzyme for 20-40 minutes for pretreatment. S2.5 Place the pre-enzymatically hydrolyzed mixture container into the microwave reactor, add the magnetic stir bar, turn on the stirring, set the temperature to the optimal range for the enzyme, and operate the microwave in pulse mode to quickly reach the set temperature for synergistic extraction.

5. The method for preparing Tremella polysaccharide according to claim 1, characterized in that: The formula of the compound enzyme in S2 is: 0.59% cellulase, 0.43% pectinase, and 0.98% papain.

6. The method for preparing Tremella polysaccharide according to claim 1, characterized in that: The process of aqueous two-phase extraction in step S3 includes: S3.1 After microwave co-processing, immediately transfer the container to a boiling water bath and heat for 5-10 minutes to completely inactivate the enzyme and terminate the reaction. While the mixture is hot, centrifuge and collect the supernatant, which is the crude extract containing Tremella polysaccharide. S3.

2. Prepare an aqueous two-phase system with 24.68% ethanol and 28% ammonium sulfate. Mix the crude polysaccharide extract with the prepared aqueous two-phase system in a container at a volume ratio of 1:1 to 1:

5. S3.3 Stir thoroughly for 10-30 minutes to ensure that the substances are fully distributed between the two phases. Then stop stirring and let stand for 5-30 minutes. The solution will spontaneously form two clear phases. Polysaccharides will be enriched in the upper ethanol phase, while a large amount of proteins, cell debris and other impurities will be distributed in the lower salt phase. S3.4 After the phase separation is clear, use a pipette to collect the target phase rich in polysaccharides, which is the upper alcohol phase.

7. The method for preparing Tremella polysaccharide according to claim 1, characterized in that: Before separation via ultrafiltration membrane in step S4, the polysaccharide-enriched phase collected by aqueous two-phase extraction can be centrifuged at low speed to remove tiny particles and prevent particulate matter from clogging the ultrafiltration membrane pores.

8. The method for preparing Tremella polysaccharide according to claim 1, characterized in that: The process of modifying Tremella polysaccharide in step S5 includes: S5.1 Prepare a NaOH solution and cool it for later use. Weigh out chloroacetic acid and dissolve it in ethanol. S5.2 Dissolve the target polysaccharide fraction after ultrafiltration fractionation in a pre-cooled NaOH solution; S5.3 Place the mixture in an ice-water bath and react for 30-60 minutes with vigorous stirring to generate active sodium alkoxide anions; S5.4 Under continuous stirring and ice bath conditions, slowly and in batches add chloroacetic acid solution to the alkalization mixture. After the addition is complete, remove the ice bath and gradually heat the reaction system to the set temperature of 50-70℃. Stir the reaction at this temperature for 2-6 hours. S5.5 After the reaction is complete, cool the mixture to room temperature. While stirring, slowly adjust the pH to about 7.0 with dilute hydrochloric acid to neutralize the excess alkali and terminate the reaction. Add 2-3 times the volume of anhydrous ethanol and let stand to precipitate the modified polysaccharide. Centrifuge to collect the precipitate. S5.

6. The product of alcohol precipitation is redissolved in water and subjected to ultrafiltration to concentrate and completely desalinate it.

9. The method for preparing Tremella polysaccharide according to claim 1, characterized in that: In step S6, after concentrating the ultrafiltration purified carboxymethyl polysaccharide solution to a solid content of 2%-10%, the sample solution is dispensed into a freeze-drying container. The dispensed sample is then quickly placed in an ultra-low temperature freezer below -50°C to ensure that the sample is completely frozen, opaque, and in a hard solid state. The freeze dryer is started in advance, and the condenser temperature is lowered to below -50°C. The completely frozen sample is then removed from the freezing equipment and quickly transferred into the pre-cooled freeze dryer drying chamber. The chamber door is closed, the vacuum pump is started, and sublimation begins when the system vacuum level stabilizes below 10-30 Pa.

10. The method for preparing Tremella polysaccharide according to claim 1, characterized in that: During sublimation in step S6, high vacuum and low temperature must be maintained, and the sample temperature must always be lower than its eutectic point temperature. After the first drying is completed, the sample temperature is gradually and slowly increased and maintained at this temperature for 4-10 hours. After drying is completed, the vacuum valve is closed first, and then dry inert gas is slowly introduced into the drying chamber until the pressure returns to normal. Then the sample is immediately taken out and quickly sealed and packaged in a dry environment to obtain the Tremella polysaccharide product.

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