Yeast glucan and preparation method therefor

Abstract

The present invention relates to the technical field of yeast glucans, and specifically relates to a yeast glucan and a preparation method therefor. The preparation method for the yeast glucan comprises: step 1: adding a first solvent to a yeast raw material; step 2: adding an alkali reagent to the solution obtained in step 1 to implement an extraction reaction; step 3: separating the reaction solution obtained in step 2 to obtain solid-phase delipidized cell walls; step 4: adding a second solvent to the solid-phase delipidized cell walls obtained in step 3 to prepare a cell wall emulsion to implement a reaction; step 5: adding an acid reagent to the reaction solution obtained in step 4 to implement an extraction reaction; and step 6: separating the reaction solution obtained in step 5 and collecting a heavy phase to obtain a yeast glucan product. The preparation method for the yeast glucan of the present invention shortens the conventional preparation process, greatly reduces the burden of waste treatment, and effectively reduces production costs, and the prepared yeast glucan has higher purity and has better industrial application prospects.

Description

A yeast glucan and its preparation method

[0001] Related applications

[0002] This application claims priority to the earlier application No. 202411011616.9 filed with the China National Intellectual Property Administration on July 25, 2024, the entire contents of which are incorporated herein by reference. Technical Field

[0003] This invention belongs to the field of yeast glucan technology, specifically relating to a yeast glucan and its preparation method. Background Technology

[0004] Yeast β-glucan, also known as dextran, is a functional polysaccharide found in the inner layer of yeast cell walls. The finished product is a light yellow or yellowish-brown powder. β-glucan has a molecular weight of approximately 20,000-4,000,000 D, and its basic structure is mainly composed of β-1,3-D-glucan as the main chain and 1,6-D-glucan as the branch chain. In recent years, in-depth research has been conducted on the efficacy, molecular mechanism, and applications of yeast β-glucan. It has been found that β-glucan has good biological activity, especially through immunomodulatory effects, exhibiting good anti-upper respiratory tract infection effects. Clinical studies have also confirmed its anti-upper respiratory tract infection effects. Therefore, β-glucan is being developed as a health food with significant functions.

[0005] Currently, the production method of yeast glucan involves a combination of acid-base treatment and enzyme treatment. Chinese patent CN101570769A discloses a method for producing yeast glucan, mannan, and their production. Using bread or brewer's yeast cell walls as raw materials, the method involves treating the cells with alkaline protease and mannanase to separate the light and heavy phases. The heavy phase is then prepared into a 1%–10% solution, which undergoes alkali treatment, separation, acid treatment, and further separation. Finally, the resulting heavy phase is spray-dried at 100–180°C, yielding yeast glucan with a purity of over 70%.

[0006] Chinese patent CN101560269A discloses a method for preparing β-1,3-glucan, which uses beer lees, a major byproduct of the beer brewing industry, as raw material. The method involves treating the beer with α-amylase to hydrolyze α-glucan, followed by two consecutive alkaline extractions. The insoluble matter after separation is degreased and dehydrated using 1.5 to 20 times its volume of organic solvent, thereby obtaining yeast β-1,3-glucan with a purity of up to 80%. Summary of the Invention

[0007] The technical problem solved by this invention is that the preparation process of yeast β-glucan is complicated, requires high-quality equipment, faces great pressure from environmental protection, has high manufacturing costs, and produces glucan with low purity.

[0008] To address the aforementioned technical problems, this invention provides a yeast dextran and its preparation method.

[0009] Specifically, the present invention provides the following technical solution:

[0010] In a first aspect, the present invention provides a method for preparing yeast glucan, comprising the following steps:

[0011] Step 1: Add the first solvent to the yeast raw materials;

[0012] Step 2: Add an alkaline reagent to the solution obtained in Step 1 to carry out the extraction reaction;

[0013] Step 3: Separate the reaction solution obtained in Step 2 to obtain a solid-phase defatted cell wall;

[0014] Step 4: Add a second solvent to the solid defatted cell wall obtained in Step 3 to prepare a cell wall emulsion, and carry out the reaction;

[0015] Step 5: Add acid reagent to the reaction solution obtained in Step 4 to carry out the extraction reaction;

[0016] Step 6: Separate the reaction solution obtained in Step 5, collect the heavy phase, and obtain the yeast dextran product.

[0017] Preferably, in step 1, the yeast raw material includes dry yeast and / or yeast cell walls.

[0018] Preferably, in step 1, the first solvent is an organic solvent.

[0019] Preferably, in step 1, the first solvent contains C 1-4 Alcohol solvent.

[0020] More preferably, in step 1, the first solvent includes one or more of methanol, ethanol, n-propanol, isopropanol and n-butanol.

[0021] More preferably, in step 1, the mass of the first solvent is 5 to 15 times the mass of the yeast raw material.

[0022] Preferably, in step 2, the alkaline reagent includes one or two of sodium hydroxide, potassium hydroxide, lithium hydroxide, sodium ethoxide, and sodium methoxide.

[0023] Preferably, in step 2, the mass of the alkaline reagent is 5-12% of the mass of the yeast raw material.

[0024] Preferably, in step 2, the mass of the alkaline reagent is 5-11% of the mass of the yeast raw material.

[0025] Preferably, in step 2, the extraction temperature is 70–100°C.

[0026] Preferably, in step 2, the extraction temperature is 70–85°C.

[0027] Preferably, in step 2, the extraction time is 2 to 8 hours.

[0028] Preferably, in step 2, the extraction time is 3 to 5 hours.

[0029] Preferably, in step 4, the second solvent is water.

[0030] Preferably, in step 4, the temperature of the water is 95–100°C.

[0031] More preferably, in step 4, the dry matter mass percentage of the cell wall emulsion is 10% to 15%.

[0032] Preferably, in step 4, the temperature of the reaction is maintained at 95–100°C.

[0033] Preferably, in step 4, the reaction time is 0.15 to 5 hours.

[0034] Preferably, in step 4, the reaction time is 1 to 5 hours.

[0035] Preferably, in step 5, the solution after the reaction in step 4 is first cooled, then a third solvent is added for dilution, and finally an acid reagent is added for extraction.

[0036] Preferably, in step 5, the solution after the reaction in step 4 is cooled to 50–75°C.

[0037] Preferably, in step 5, the solution after the reaction in step 4 is cooled to 60-75°C.

[0038] Preferably, in step 5, the third solvent is water.

[0039] Preferably, in step 5, the dry matter mass percentage in the diluted solution is 5% to 17%.

[0040] Preferably, in step 5, the mixture is diluted and mixed for 0.1 to 5 hours.

[0041] Preferably, in step 5, the acid reagent is selected from one or more of hydrochloric acid, sulfuric acid, and citric acid.

[0042] Preferably, in step 5, the pH of the suspension obtained in step 4 is adjusted to 4-6 using an acid reagent.

[0043] Preferably, in step 5, the extraction temperature is 50–100°C.

[0044] Preferably, in step 5, the extraction temperature is 70–100°C.

[0045] Preferably, in step 5, the extraction time is 0.5 to 8 hours.

[0046] Preferably, in step 6, the collected heavy phase is dried, and the drying is selected from spray drying, freeze drying or vacuum drying.

[0047] Preferably, in step 6, the collected heavy phase is dried by spray drying, with an inlet air temperature of 100-180°C and an outlet air temperature of 60-90°C.

[0048] Secondly, the present invention provides a yeast dextran, wherein the yeast dextran product is prepared by the yeast dextran preparation method described above.

[0049] Preferably, the purity of the yeast beta-glucan is ≥62%.

[0050] Preferably, the purity of the yeast beta-glucan is 62% to 91%.

[0051] More preferably, the purity of yeast beta-glucan is 79% to 91%.

[0052] More preferably, the purity of yeast beta-glucan is 82% to 91%.

[0053] Beneficial effects of the present invention

[0054] (1) The present invention uses yeast cell walls and / or dry yeast as raw materials, and adds an alkaline reagent to a low alcohol solvent to hydrolyze and remove fat-soluble impurities, dispersing and dissolving the fat-soluble impurities in the low alcohol solvent. This operation can replace the three steps of enzymatic hydrolysis, aqueous saponification and solvent treatment in the traditional process, shortening the process flow.

[0055] (2) In the preparation method of the present invention, only conventional separation equipment is needed to easily achieve solid-liquid separation of materials; the solvent can be recycled and reused, and the amount of alkaline waste liquid generated is greatly reduced, which greatly alleviates the pressure of environmental protection treatment and can effectively reduce production costs.

[0056] (3) In this invention, a low-grade alcohol organic solvent and an alkaline reagent are first used to act on the yeast raw material to obtain defatted yeast cell walls. Then, water is added to prepare a yeast cell wall emulsion for incubation reaction, followed by acid extraction. Finally, the purity of the yeast glucan obtained is higher, reaching up to 90.3%. Attached Figure Description

[0057] Figure 1 is a flowchart of the preparation process of yeast glucan of the present invention. Detailed Implementation

[0058] As described above, the present invention provides a yeast dextran and a method for preparing the same.

[0059] The lower alcohol solvent mentioned in this invention refers to C 1-4 alcohol solvent, "C 1-4 "" refers to a number of carbon atoms ranging from 1 to 4. (C) 1-4 The alcohol solvents include, but are not limited to, methanol, ethanol, n-propanol, isopropanol, n-butanol, and isobutanol.

[0060] The present invention provides the following technical solution:

[0061] Technical Solution 1: A method for preparing yeast glucan, characterized by comprising the following steps:

[0062] Step 1: Add the first solvent to the yeast raw materials;

[0063] Step 2: Add an alkaline reagent to the solution obtained in Step 1 to carry out the extraction reaction;

[0064] Step 3: Separate the reaction solution obtained in Step 2 to obtain a solid-phase defatted cell wall;

[0065] Step 4: Add a second solvent to the solid defatted cell wall obtained in Step 3 to prepare a cell wall emulsion, and carry out the reaction;

[0066] Step 5: Add acid reagent to the reaction solution obtained in Step 4 to carry out the extraction reaction;

[0067] Step 6: Separate the reaction solution obtained in Step 5, collect the heavy phase, and obtain the yeast dextran product.

[0068] Technical Solution 2. The method for preparing yeast glucan according to Technical Solution 1, wherein, in step 1, the yeast raw material includes dry yeast and / or yeast cell wall;

[0069] Preferably, the yeast used in the dry yeast and / or yeast cell wall is selected from one or more of Saccharomyces cerevisiae, Candida albicans, Gastrodia elata, Rhodotorula rubra, and Cryptococcus pluvialis.

[0070] And / or, preferably, the present invention does not limit the source of the dry yeast in any way, and commercially available dry yeast is suitable for use in the present invention; more preferably, the number of live yeast cells in the dry yeast is >2.0 x 10⁻⁶. 10 CFU / g, and / or, moisture <6.0%; more preferably, the number of viable yeast cells in the dry yeast is 2.0 x 10⁻⁶. 10 -5.0x10 10 CFU / g, and / or, moisture 1.0%-6.0%;

[0071] And / or, the present invention does not limit the source of yeast cell walls in any way, and commercially available yeast cell walls are suitable for use in the present invention; preferably, the moisture content is ≤8.0%, mannan >20.0%, and / or, β-glucan 20.0-40.0%; and / or, more preferably, the moisture content is 1.0%-8.0%, mannan 20.0%-25.0%, and / or, β-glucan 20.0-40.0%.

[0072] Technical Solution 3. The method for preparing yeast glucan according to Technical Solution 1 or 2, wherein, in step 1, the first solvent is an organic solvent;

[0073] Preferably, the first solvent contains C 1-4 The alcohol solvent; more preferably, the volume percentage of alcohol in the first solvent is 60% to 100%; even more preferably, the volume percentage of alcohol in the organic solvent is 70% to 100%;

[0074] And / or, more preferably, the first solvent includes one or more of methanol, ethanol, n-propanol, isopropanol and n-butanol;

[0075] More preferably, the mass of the first solvent is 5 to 15 times or 6 to 10 times the mass of the yeast raw material; most preferably, in some specific embodiments, the mass of the first solvent can be 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or 15 times the mass of the yeast raw material, or the mass of the first solvent within the numerical range formed by any two of the above specific values ​​as endpoints is a multiple of the mass of the yeast raw material.

[0076] Technical Solution 4. The method for preparing yeast glucan according to any one of Technical Solutions 1-3, wherein, in step 2, the alkaline reagent includes one or two of sodium hydroxide, potassium hydroxide, lithium hydroxide, sodium ethoxide, and sodium methoxide;

[0077] Preferably, the mass of the alkaline reagent is 5% to 12% of the mass of the yeast raw material;

[0078] More preferably, the mass of the alkaline reagent is 5% to 11% or 8% to 11% of the mass of the yeast raw material;

[0079] More preferably, the mass of the alkali reagent can be 5%, 6%, 7%, 8%, 9%, 10%, 11%, or 12% of the mass of the yeast raw material, or the mass of the alkali reagent within the numerical range formed by any two of the above specific values ​​as endpoints is a percentage of the mass of the yeast raw material.

[0080] Technical Solution 5. The method for preparing yeast glucan according to any one of Technical Solutions 1-4, wherein, in step 2, the extraction temperature is 70-100℃; and / or the extraction time is 2-8h;

[0081] Preferably, the extraction temperature is 70–85°C; more preferably, the extraction temperature is 75–85°C; and / or the extraction time is 3–5 hours.

[0082] More preferably, the extraction temperature can be 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100°C, or an extraction temperature within the numerical range formed by any two of the above specific values ​​as endpoints;

[0083] And / or, the extraction time can be 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5 or 8 hours, or the extraction time can be within the numerical range formed by any two of the above specific values ​​as endpoints.

[0084] Technical Solution 6. The method for preparing yeast glucan according to any one of Technical Solutions 1-5, wherein, in step 4, the second solvent is water;

[0085] Preferably, the temperature of the water is 95 to 100°C; more preferably, in some specific embodiments, the temperature of the water can be 95, 96, 97, 98, 99 or 100°C, or the temperature of the water within the range of any two of the above specific values ​​as endpoints.

[0086] More preferably, the dry matter mass percentage in the cell wall emulsion is 10% to 15% (preferably, the dry matter mass percentage in the cell wall emulsion is 11% to 14%); even more preferably, in some specific embodiments, the dry matter mass percentage in the cell wall emulsion is 10%, 11%, 12%, 13%, 14% or 15%, or the dry matter mass percentage in the cell wall emulsion is within the numerical range formed by any two of the above specific values ​​as endpoints;

[0087] More preferably, the reaction temperature is maintained at 95–100°C; and / or the reaction time is 0.15–5 h; preferably, the reaction time is 1–5 h; more preferably, the reaction time is 3–5 h;

[0088] More preferably, in some specific embodiments, the reaction temperature is maintained at 95, 96, 97, 98, 99 or 100°C, or a reaction temperature within the numerical range formed by any two of the above specific values ​​as endpoints; and / or, the reaction time can be 0.15, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5 or 5 hours, or a reaction time within the numerical range formed by any two of the above specific values ​​as endpoints;

[0089] And / or, more preferably, water is added to the solid defatted cell wall to prepare a cell wall emulsion, the water temperature is 95-100°C, the mixture is stirred evenly to obtain a suspension, and the reaction is continued at 95-100°C for 0.15-5 hours.

[0090] Technical Solution 7. The method for preparing yeast glucan according to any one of Technical Solutions 1-6, wherein in step 5, the solution after the reaction in step 4 is first cooled, then a third solvent is added for dilution, and finally an acid reagent is added for extraction reaction;

[0091] Preferably, the solution after the reaction in step 4 is cooled to 50–75°C; more preferably, the solution after the reaction in step 4 is cooled to 60–75°C; even more preferably, in some specific embodiments, the solution after the reaction in step 4 is cooled to 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74 or 75°C, or a cooling temperature within the numerical range formed by any two of the above specific values ​​as endpoints;

[0092] And / or, preferably, the third solvent is water;

[0093] And / or, preferably, the dry matter percentage in the diluted solution is 5% to 17% (preferably, the dry matter percentage in the diluted solution is 5% to 7%); more preferably, in some specific embodiments, the dry matter percentage in the diluted solution is 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16% or 17%, or the dry matter percentage in the diluted solution within the numerical range formed by any two of the above specific values ​​as endpoints;

[0094] And / or, preferably, the dilution and mixing time is 0.1 to 5 hours; more preferably, in some specific embodiments, the dilution and mixing time is 0.1, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5 or 5 hours, or the dilution and mixing time is within the numerical range formed by any two of the above specific values ​​as endpoints.

[0095] Technical Solution 8. A method for preparing yeast glucan according to any one of technical solutions 1-7, wherein, in step 5, the acid reagent is selected from one or more of hydrochloric acid, sulfuric acid and citric acid;

[0096] Preferably, the suspension obtained in step 4 is adjusted to a pH of 4 to 6 using an acid reagent; more preferably, the suspension obtained in step 4 is adjusted to a pH of 4, 4.5, 5 or 6 using an acid reagent, or to a pH within the range of any two of the above specific values ​​as endpoints.

[0097] And / or, the extraction temperature is 50–100°C; preferably, the extraction temperature is 70–100°C; and / or, the extraction time is 0.5–8 h; more preferably, the extraction temperature is 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, The extraction temperature is 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100℃, or within the numerical range formed by any two of the above specific values ​​as endpoints; and / or the extraction time is 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5 or 8h, or within the numerical range formed by any two of the above specific values ​​as endpoints.

[0098] Technical Solution 9. A method for preparing yeast glucan according to any one of Technical Solutions 1-8, wherein in step 6, the collected heavy phase is dried, and the drying is selected from spray drying, freeze drying or vacuum drying;

[0099] Preferably, the drying is spray drying, with an inlet air temperature of 100-180°C and an outlet air temperature of 60-90°C.

[0100] Technical Solution 10. A yeast glucan, characterized in that the yeast glucan is prepared by any one of the yeast glucan preparation methods described in Technical Solutions 1-9.

[0101] Technical Solution 11. The yeast glucan according to Technical Solution 10, wherein the purity of the yeast glucan is ≥62%; preferably, the purity of the yeast glucan is 62% to 91%; more preferably, the purity of the yeast glucan is 79% to 91%.

[0102] Steps 1 and 2 of this invention are saponification reactions. The lower alcohol solution promotes the miscibility of oils and alkali reagents, making them a homogeneous system, thereby accelerating the saponification reaction.

[0103] And / or, the solid defatted cell wall obtained in step 3 includes components such as dextran, mannan, and proteins.

[0104] Unless otherwise stated, all reagents / instruments used in the embodiments and comparative examples of this invention are conventional commercially available products. Information on the experimental materials and instruments used in this invention is provided in the table below:

[0105] Table 1 Experimental Materials / Instruments and Manufacturers

[0106] The detection of the number of live cells in dry yeast was carried out in accordance with the People's Republic of China National Standard GB 7300.501-2021 Feed Additives Part 5: Microbial Saccharomyces cerevisiae Appendix B Method B.2 Second Staining Method for Determination of Live Cell Count in Yeast.

[0107] The moisture content was determined in accordance with the People's Republic of China National Standard GB / T 6435-2014, Determination of Moisture in Feed.

[0108] The detection of β-glucan content was performed according to the detection of yeast β-glucan in Example 1. The detection procedure for mannan content was the same as that for yeast β-glucan in Example 1. Since β-glucan and mannan have different retention times in the chromatogram, the content of β-glucan and mannan can be calculated based on the peak area of ​​the corresponding chromatographic peaks.

[0109] To better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0110] Example 1

[0111] (1) Preparation of yeast β-glucan

[0112] Step 1: Add 500g of anhydrous ethanol to 100g of dry yeast;

[0113] Step 2: Add 5g of sodium ethoxide, stir, heat to 80℃, and keep warm while stirring for 3 hours;

[0114] Step 3: After filtration and separation, a solid-phase defatted cell wall is obtained, and the filtrate is concentrated under reduced pressure to recover ethanol;

[0115] Step 4: Add 450 mL of water (water temperature is 95℃) to the solid defatted cell wall to prepare a cell wall emulsion, wherein the dry matter mass percentage is 10%, mix well, and keep warm and stirring at 95℃ for 3 hours.

[0116] Step 5: After the temperature drops to 70℃, dilute with water to make the dry matter percentage 5%, stir for 0.5 hours, and mix thoroughly. Add concentrated sulfuric acid to adjust the pH to approximately 5.5, and heat to 75℃ and stir for 1 hour.

[0117] Step 6: Centrifuge at 8000 r / min for 10 min. Wash the separated heavy phase with water until neutral, collect the heavy phase, and place it in a spray drying tower. The inlet air temperature is 180℃ and the outlet air temperature is 80℃. The solid obtained after drying is β-glucan. Its purity was determined to be 83.91% by liquid chromatography, and the yield was 39.01%.

[0118] (2) Detection of yeast β-glucan

[0119] Sample solution preparation: Accurately weigh 0.4 g (to the nearest 0.0002 g) of sample into a 25 mL stoppered glass test tube, accurately add 6.0 mL of hydrochloric acid (37%), tighten the cap, and mix with a vortex mixer to obtain a homogeneous suspension. Place the test tube in a 30 °C water bath for 45 minutes, mixing with a vortex mixer every 15 minutes. Transfer the entire suspension to a 250 mL hydrolysis flask, wash the test tube several times with 100–120 mL of water, and add the washings to the hydrolysis flask. Treat at 121 °C for 60 minutes. After hydrolysis, cool to room temperature in a water bath or ice bath. Add 9 mL of sodium hydroxide solution to adjust the pH of the solution to 6–7, transfer to a 200 mL volumetric flask, and dilute to the mark with water. Filter using a 0.45 μm pore size cellulose acetate membrane for later use. After acid treatment of the sample, test according to the specified chromatographic conditions, and calculate the results quantitatively using the external standard method.

[0120] The chromatographic conditions are as follows:

[0121] Chromatographic column: Aminex HPX-87H ion Exclusion (7.8mm×300mm BIO-RAD 1);

[0122] Mobile phase: 0.005 mol / L sulfuric acid solution. Use a 1 mL graduated pipette to pipette 0.55 mL of concentrated sulfuric acid into a 5 L beaker, add 2 L of water, and stir until homogeneous.

[0123] Flow rate: 0.6 mL / min;

[0124] Column temperature: 65℃;

[0125] Injection volume: 20 μL.

[0126] The formula for calculating the β-glucan content w (also known as the purity of β-glucan): expressed as a mass fraction (%), is as follows:

[0127] Where: w - the content of β-glucan in the sample, %; A - the concentration of glucose calculated from the standard curve corresponding to the peak area of ​​20 μL sample solution, in milligrams per liter (mg / L); m - the mass of the sample, in grams (g); 0.2 - the volume of the sample after treatment, in liters; 0.9 - the coefficient for converting glucose to β-glucan; F - the empirical compensation coefficient for the lower result caused by the destruction of glucose during acid hydrolysis of the sample, which is 1.14.

[0128] Example 2

[0129] Step 1: Add 800g of 90% ethanol (by volume) to 100g of yeast cell walls;

[0130] Step 2: Add 10g of potassium hydroxide, stir and heat to 80℃, keep warm and stir for 3 hours;

[0131] Step 3: After filtration and separation, a solid-phase defatted cell wall is obtained. In addition, the filtrate is concentrated under reduced pressure to recover ethanol.

[0132] Step 4: Add 400 mL of water (at a temperature of 95°C) to the solid defatted cell wall, wherein the dry matter mass percentage is 12.5%, mix well and keep warm at 95°C for 5 hours;

[0133] Step 5: After the temperature drops to 65℃, dilute with water to a dry matter mass percentage of 6.25%, and stir for 0.5 hours. Add hydrochloric acid to adjust the pH to approximately 4, and heat to 70℃ while stirring for 1 hour.

[0134] Step 6: Centrifuge at 8000 rpm for 10 min. Wash the separated heavy phase with water until neutral, collect the heavy phase, and place it in a spray drying tower with an inlet air temperature of 180℃ and an outlet air temperature of 90℃. The solid obtained after drying is β-glucan. The detection method is the same as in Example 1. After testing, the purity of β-glucan is 90.30%, and the yield is 36.06%.

[0135] Example 3

[0136] Step 1: Add 1500g of 90% methanol (volume percentage) to 100g of yeast cell walls;

[0137] Step 2: Add another 10g of potassium hydroxide, heat to 70℃ while stirring, and keep warm and stirring for 5 hours;

[0138] Step 3: After filtration and separation, a solid-phase cell wall is obtained. In addition, the filtrate is concentrated under reduced pressure to recover methanol.

[0139] Step 4: Add 550 mL of water (water temperature is 95℃) to the solid phase to prepare a cell wall emulsion, in which the dry matter mass percentage is 15%, mix well and keep warm and stirred at 95℃ for 1 hour;

[0140] Step 5: After the temperature drops to 60℃, dilute with water to make the dry matter content 7.5% by mass, and stir for 0.5 hours. Add citric acid to adjust the pH to approximately 4.5, and heat to 70℃ while stirring for 1 hour.

[0141] Step 6: Centrifuge at 5000 rpm for 30 min. Wash the separated heavy phase with water until neutral, collect the heavy phase, and place it in a spray drying tower with an inlet air temperature of 120°C and an outlet air temperature of 75°C. The solid obtained after drying is β-glucan. The detection method is the same as in Example 1. After testing, the purity of β-glucan is 82.90%, and the yield is 34.20%.

[0142] Example 4

[0143] Step 1: Add 800g of 90% isopropanol (volume percentage) to 100g of yeast cell walls;

[0144] Step 2: Add 12g of potassium hydroxide, heat to 90℃ while stirring, and keep warm and stirring for 2 hours;

[0145] Step 3: After filtration and separation, a solid cell wall is obtained, and the filtrate is concentrated under reduced pressure to recover isopropanol;

[0146] Step 4: Add 350 mL of water (water temperature is 95℃) to the solid phase, wherein the dry matter mass percentage is 12.5%, mix well and keep warm and stirring at 95℃ for 5 hours;

[0147] Step 5: After the temperature drops to 70℃, dilute with water to a dry matter mass percentage of 6.25%, and stir for 0.5 hours. Add hydrochloric acid to adjust the pH to approximately 5.2, and maintain the temperature at 70℃ while stirring for 1 hour.

[0148] Step 6: Centrifuge at 7000 rpm for 20 minutes. Wash the separated heavy phase with water until neutral, collect the heavy phase, and place it in a spray drying tower with an inlet air temperature of 180°C and an outlet air temperature of 80°C. The solid obtained after drying is β-glucan. The detection method is the same as in Example 1. After testing, the purity of β-glucan is 79.33%, and the yield is 34.39%.

[0149] Example 5

[0150] Step 1: Add 800g of 90% isopropanol (volume percentage) to 100g of yeast cell walls;

[0151] Step 2: Add 12g of potassium hydroxide, heat to 90℃ while stirring, and keep warm and stirring for 2 hours;

[0152] Step 3: After filtration and separation, a solid-phase cell wall is obtained. In addition, the filtrate is concentrated under reduced pressure to recover isopropanol.

[0153] Step 4: Add 350 mL of water (water temperature is 95℃) to the solid phase, wherein the dry matter mass percentage is 12.5%, mix well and keep warm and stirring at 95℃ for 5 hours;

[0154] Step 5: After the temperature drops to 50℃, dilute with water to a dry matter mass percentage of 6.25%, and stir for 0.5 hours. Add hydrochloric acid to adjust the pH to approximately 4, and maintain the temperature at 50℃ while stirring for 1 hour.

[0155] Step 6: Centrifuge at 8000 rpm for 10 minutes. Wash the separated heavy phase with water until neutral, collect the heavy phase, and place it in a spray drying tower with an inlet air temperature of 180°C and an outlet air temperature of 90°C. The solid obtained after drying is β-glucan. The detection method is the same as in Example 1. After testing, the purity of β-glucan is 69.33%, and the yield is 37.12%.

[0156] Example 6

[0157] Step 1: Add 800g of acetonitrile to 100g of yeast cell wall;

[0158] Step 2: Add 12g of potassium hydroxide, heat to 90℃ while stirring, and keep warm and stirring for 2 hours;

[0159] Step 3: After filtration and separation, a solid-phase cell wall is obtained. In addition, the filtrate is concentrated under reduced pressure to recover acetonitrile.

[0160] Step 4: Add 350 mL of water (at 100 °C) to the solid phase, wherein the dry matter mass percentage is 12.5%, mix well and keep warm and stirring at 100 °C for 3 hours;

[0161] Step 5: After the temperature drops to 60℃, dilute with water to a dry matter mass percentage of 6.25%, and stir for 0.5 hours. Add hydrochloric acid to adjust the pH to approximately 5, and heat to 70℃ while stirring for 1 hour.

[0162] Step 6: Centrifuge at 8000 rpm for 10 minutes. Wash the separated heavy phase with water until neutral, collect the heavy phase, and place it in a spray drying tower with an inlet air temperature of 150°C and an outlet air temperature of 75°C. The solid obtained after drying is β-glucan. The detection method is the same as in Example 1. After testing, the purity of β-glucan is 62.8%, and the yield is 41.39%.

[0163] Comparative Example 1

[0164] Step 1: Add 450mL of water to 50g of yeast cell wall to control the dry matter content to about 10%, and mix well;

[0165] Step 2: Add 11.28g of sodium hydroxide, heat to 90℃ while stirring, and keep warm and stirring for 3 hours;

[0166] Step 3: After filtration and separation, a solid phase is obtained;

[0167] Step 4: Dilute with water to achieve a solid-phase dry matter percentage of 5%, maintaining a temperature of 70°C and stirring for 0.5 hours. Add sulfuric acid solution to adjust the pH to approximately 5.2, and heat to 95°C while stirring for 1 hour.

[0168] Step 5: Centrifuge at 8000 rpm for 10 min. Wash the separated heavy phase with water until neutral, collect the heavy phase, and place it in a spray drying tower with an inlet air temperature of 180°C and an outlet air temperature of 80°C. The solid obtained after drying is β-glucan. The detection method is the same as in Example 1. After testing, the purity of β-glucan is 52.8%, and the yield is 45.6%.

[0169] While the specific embodiments of the present invention have been described above, they are not intended to limit the scope of protection of the present invention. Based on the technical solutions of the present invention, various modifications or variations that can be made by those skilled in the art without creative effort are still within the scope of protection of the present invention.

Claims

1. A method for preparing a yeast glucan, characterized by, The method comprises the following steps: Step 1: adding a first solvent to a yeast raw material; Step 2: adding a base reagent to the solution obtained in Step 1 to perform an extraction reaction; Step 3: separating the reaction solution obtained in Step 2 to obtain a solid-phase defatted cell wall; Step 4: adding a second solvent to the solid-phase defatted cell wall obtained in Step 3 to prepare a cell wall milk, and performing a reaction; Step 5: adding an acid reagent to the reaction solution obtained in Step 4 to perform an extraction reaction; Step 6: separating the reaction solution obtained in Step 5 to collect a heavy phase, and obtaining a yeast glucan product.

2. The method of preparing yeast glucans according to claim 1, wherein, In Step 1, the yeast raw material comprises dry yeast and / or yeast cell walls.

3. The method for preparing yeast glucans according to claim 1 or 2, wherein, In Step 1, the first solvent is an organic solvent; Preferably, the first solvent is an alcohol solvent containing C 1-4 of formula (I). More preferably, the first solvent comprises one or more than two of methanol, ethanol, n-propanol, isopropanol, and n-butanol; Further more preferably, the mass of the first solvent is 5-15 times the mass of the yeast raw material.

4. The method of preparing yeast glucans according to any one of claims 1-3, wherein, In Step 2, the base reagent comprises one or more than two of sodium hydroxide, potassium hydroxide, lithium hydroxide, sodium ethoxide, and sodium methoxide; Preferably, the mass of the base reagent is 5%-12% of the mass of the yeast raw material; More preferably, the mass of the base reagent is 5%-11% of the mass of the yeast raw material.

5. The method of preparing yeast glucans according to any one of claims 1-4, wherein, In Step 2, the extraction temperature is 70-100°C; and / or the extraction time is 2-8h; Preferably, the extraction temperature is 70-85°C; and / or the extraction time is 3-5h.

6. The method of preparing yeast glucans according to any one of claims 1-5, wherein, In Step 4, the second solvent is water; Preferably, the temperature of the water is 95-100°C; More preferably, the mass percentage of dry matter in the cell wall milk is 10%-15%; Further more preferably, the reaction temperature is maintained at 95-100°C; and / or the reaction time is 0.15-5h; preferably, the reaction time is 1-5h.

7. The method of preparing yeast glucans according to any one of claims 1-6, wherein, In Step 5, the solution after Step 4 is cooled first, then a third solvent is added for dilution, and finally an acid reagent is added for an extraction reaction; Preferably, the solution after Step 4 is cooled to 50-75°C; more preferably, the solution after Step 4 is cooled to 60-75°C; And / or, preferably, the third solvent is water; And / or, preferably, the mass percentage of dry matter in the diluted solution is 5%-17%; And / or, preferably, the mixing time after dilution is 0.1-5h.

8. The method of preparing yeast glucans according to any one of claims 1-7, wherein, In Step 5, the acid reagent is selected from one or more than two of hydrochloric acid, sulfuric acid, and citric acid; Preferably, the suspension obtained in Step 4 is adjusted to a pH of 4-6 with the acid reagent; And / or, the extraction temperature is 50-100°C; preferably, the extraction temperature is 70-100°C; and / or, the extraction time is 0.5-8h.

9. The method of preparing yeast glucans according to any one of claims 1-8, wherein, In Step 6, the collected heavy phase is dried, and the drying is selected from spray drying, freeze drying, or vacuum drying; Preferably, the drying is spray drying, the inlet air temperature is 100-180°C, and the outlet air temperature is 60-90°C.

10. A yeast glucan, characterized in that, The yeast glucan is prepared by the method for preparing yeast glucan according to any one of claims 1-9.

11. The yeast glucan of claim 10, wherein, The purity of the yeast glucan is > 62%; preferably, the purity of the yeast glucan is between 62% and 91%; more preferably, the purity of the yeast glucan is between 79% and 91%. The purity of the yeast glucan is > 62%; preferably, the purity of the yeast glucan is between 62% and 91%; more preferably, the purity of the yeast glucan is between 79% and 91%. The purity of the yeast glucan is > 62%;

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