CN109797179B - Green preparation method of yeast glucan - Google Patents
Green preparation method of yeast glucan Download PDFInfo
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- CN109797179B CN109797179B CN201910216107.2A CN201910216107A CN109797179B CN 109797179 B CN109797179 B CN 109797179B CN 201910216107 A CN201910216107 A CN 201910216107A CN 109797179 B CN109797179 B CN 109797179B
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Abstract
The invention belongs to the technical field of glucan, and particularly relates to a green preparation method of yeast glucan, which comprises the following steps: step 1, adding common yeast cell walls into distilled water, stirring uniformly to form a suspension, heating at a constant temperature for 2-4h, then adding tourmaline, adding hydrogen peroxide, sealing, cooking for 2-4h, preserving heat for 2-6h, taking out the tourmaline, and centrifuging to obtain a precipitate; step 2, adding the precipitate into distilled water, stirring uniformly, adding a dispersing agent, heating and stirring to form a suspension, and then adding alkali liquor to adjust the pH to 9-11 to obtain an alkaline suspension; step 3, adding protease and tourmaline into the alkaline suspension, carrying out enzymolysis for 4-6h at constant temperature, taking out the tourmaline, centrifuging, and washing with water to obtain insoluble substances; and 4, adding the insoluble substances into acetone, carrying out microwave reaction for 2-5h, filtering, and carrying out freeze drying to obtain yeast glucan powder. The invention takes yeast cell walls as production raw materials, takes tourmaline as an auxiliary catalyst and an auxiliary oxidant, and greatly improves the yield of yeast glucan.
Description
Technical Field
The invention belongs to the technical field of glucan, and particularly relates to a green preparation method of yeast glucan.
Background
At present, yeast beta-D-glucan (yeast beta-glucan or yeast glucan for short) is an immunomodulator (non-specific immunopotentiator) with good physiological functions, and has the physiological effects of enhancing immunity, resisting tumors, resisting bacteria, diminishing inflammation, resisting oxidation, promoting wound healing, resisting radiation, reducing blood fat and the like, but the yeast beta-D-glucan is insoluble in water and most other solvents and only slightly soluble in dimethyl sulfoxide, so that the application of the yeast glucan is severely limited.
In order to improve the solubility of the yeast glucan, domestic and foreign scholars perform modification and modification researches on the yeast glucan, including chemical methods and physical methods. The main chemical modifications are phosphorylation, carboxymethylation, sulfation and sulfomodification, which increase the solubility of dextran by attaching these polar groups. However, such reactions are usually carried out in liquid homogeneous or heterogeneous systems, which require a large amount of solvents, even unsafe organic solvents, acids, bases, etc., and thus, not only are the costs high and the pollution serious, but also potential safety hazards may be caused. The main physical methods are sonication and oxidative degradation, which increase the solubility of dextran by reducing its molecular weight. However, the molecular weight of glucan can only be reduced to a certain limit value by ultrasonic depolymerization, and impurities such as an oxidant are introduced into the glucan by oxidative degradation.
Disclosure of Invention
Aiming at the problems in the prior art, the invention provides a green preparation method of yeast glucan, which takes yeast cell walls as a production raw material and tourmaline as an auxiliary catalyst and an auxiliary oxidant, thereby greatly improving the yield of the yeast glucan.
In order to achieve the technical purpose, the technical scheme of the invention is as follows:
a green preparation method of yeast glucan comprises the following steps:
step 1, adding common yeast cell walls into distilled water, stirring uniformly to form a suspension, heating at a constant temperature for 2-4h, then adding tourmaline, adding hydrogen peroxide, sealing, cooking for 2-4h, preserving heat for 2-6h, taking out the tourmaline, and centrifuging to obtain a precipitate;
step 2, adding the precipitate into distilled water, stirring uniformly, adding a dispersing agent, heating and stirring to form a suspension, and then adding alkali liquor to adjust the pH to 9-11 to obtain an alkaline suspension;
step 3, adding protease and tourmaline into the alkaline suspension, carrying out enzymolysis for 4-6h at constant temperature, taking out the tourmaline, centrifuging, and washing with water to obtain insoluble substances;
and 4, adding the insoluble substances into acetone, carrying out microwave reaction for 2-5h, filtering, and carrying out freeze drying to obtain yeast glucan powder.
The concentration of the yeast cell walls in the distilled water in the step 1 is 15-45g/L, the stirring speed for uniformly stirring is 500-1000r/min, and the temperature for constant-temperature heating is 90-100 ℃.
The mass of the tourmaline in the step 1 is 2-10% of the cell wall of the common yeast, the addition amount of the hydrogen peroxide is 110-150% of the mass of the tourmaline, the temperature of the sealed cooking is 120-150 ℃, and the temperature of the heat preservation is 80-90 ℃.
The concentration of the precipitate in the step 2 in distilled water is 190g/L, the dispersant adopts polyvinylpyrrolidone, the adding amount is 2-5% of the mass of the precipitate, the heating and stirring temperature is 50-70 ℃, the stirring speed is 300r/min, and the alkali liquor adopts sodium hydroxide solution.
The protease adding amount in the step 3 is 0.1-0.3% of the mass of the precipitate, and the tourmaline in the step 1 is adopted as the tourmaline.
The temperature of the constant temperature enzymolysis in the step 3 is 30-40 ℃.
The concentration of the insoluble substances in the acetone in the step 4 is 10-30g/L, the temperature of the microwave reaction is 20-30 ℃, and the power is 300-500W.
The freeze drying in the step 4 comprises pre-freezing at-70 ℃ for 22 hours and freeze drying at-60 ℃ for 6-10 hours.
Step 1, adding common yeast cell walls into distilled water to form suspension, performing high-temperature extraction in a constant-temperature heating mode, and opening pores of the cell walls; meanwhile, due to the cooperation of tourmaline and hydrogen peroxide, the pyroelectricity of tourmaline is fully used to generate negative ions, and the negative ions are matched with peroxide groups of hydrogen peroxide, so that cell walls can be fully opened, and the extraction yield is greatly improved.
Step 2, adding the precipitate into distilled water and a dispersing agent to form a fully stirred solution, and forming an alkaline suspension under an alkaline condition, wherein the dispersing agent is used to further improve the dispersion of the precipitate.
And 3, performing constant-temperature enzymolysis reaction on the protease and the tourmaline, wherein the tourmaline fully utilizes negative ions generated by the pyroelectricity of the tourmaline to stimulate active groups of the protease, so that the enzymolysis reaction is effectively promoted, and the negative ions play a role in promoting the active enzymolysis groups in the precipitate.
And 4, adding the insoluble substances into acetone, and effectively improving the impurity removal rate by using the kinetic energy of the impurities in the insoluble substances through microwave reaction.
From the above description, it can be seen that the present invention has the following advantages:
1. the invention takes yeast cell walls as production raw materials, takes tourmaline as an auxiliary catalyst and an auxiliary oxidant, and greatly improves the yield of yeast glucan.
2. According to the invention, tourmaline is matched with hydrogen peroxide, yeast cell walls are opened in a cooking mode, the release of yeast glucan can be effectively improved, and the problem of low release rate of the traditional extraction process is solved.
3. The invention takes the tourmaline as an auxiliary catalyst, and promotes the enzymolysis efficiency of protease by the thermoelectricity of the tourmaline, thereby not only improving the enzymolysis speed, but also improving the enzymolysis depth.
4. The invention adopts a microwave reaction mode to promote the rapid dissolution of impurities in insoluble substances and reduce the residue of the impurities.
Detailed Description
The present invention is described in detail with reference to examples, but the present invention is not limited to the claims.
Example 1
A green preparation method of yeast glucan comprises the following steps:
step 1, adding common yeast cell walls into distilled water, stirring uniformly to form a suspension, heating at a constant temperature for 2 hours, then adding tourmaline, adding hydrogen peroxide, sealing, cooking for 2 hours, preserving heat for 2 hours, taking out the tourmaline, and centrifuging to obtain a precipitate;
step 2, adding the precipitate into distilled water, uniformly stirring, adding a dispersing agent, heating and stirring to form a suspension, and then adding alkali liquor to adjust the pH to 9 to obtain an alkaline suspension;
step 3, adding protease and tourmaline into the alkaline suspension, carrying out enzymolysis for 4h at constant temperature, taking out the tourmaline, centrifuging, and washing with water to obtain insoluble substances;
and 4, adding the insoluble substances into acetone, carrying out microwave reaction for 2 hours, filtering, and carrying out freeze drying to obtain yeast glucan powder.
The concentration of the yeast cell walls in the distilled water in the step 1 is 15g/L, the stirring speed for uniformly stirring is 500r/min, and the constant-temperature heating temperature is 90 ℃.
The mass of the tourmaline in the step 1 is 2% of the cell wall of the common yeast, the addition amount of the hydrogen peroxide is 110% of the mass of the tourmaline, the temperature of the sealed cooking is 120 ℃, and the temperature of the heat preservation is 80 ℃.
The concentration of the precipitate in the step 2 in distilled water is 100g/L, the dispersant adopts polyvinylpyrrolidone, the adding amount is 2% of the mass of the precipitate, the heating and stirring temperature is 50 ℃, the stirring speed is 100r/min, and the alkali liquor adopts sodium hydroxide solution.
The adding amount of the protease in the step 3 is 0.1 percent of the mass of the precipitate, and the tourmaline in the step 1 is adopted as the tourmaline.
The temperature of the constant temperature enzymolysis in the step 3 is 30 ℃.
The concentration of the insoluble substances in the acetone in the step 4 is 10g/L, the temperature of the microwave reaction is 20 ℃, and the power is 300W.
The freeze drying in the step 4 comprises pre-freezing at-70 ℃ for 22 hours and freeze drying at-60 ℃ for 6 hours.
Example 2
A green preparation method of yeast glucan comprises the following steps:
step 1, adding common yeast cell walls into distilled water, stirring uniformly to form a suspension, heating at a constant temperature for 4 hours, then adding tourmaline, adding hydrogen peroxide, sealing, cooking for 4 hours, preserving heat for 6 hours, taking out the tourmaline, and centrifuging to obtain a precipitate;
step 2, adding the precipitate into distilled water, uniformly stirring, adding a dispersing agent, heating and stirring to form a suspension, and then adding alkali liquor to adjust the pH to 11 to obtain an alkaline suspension;
step 3, adding protease and tourmaline into the alkaline suspension, carrying out enzymolysis for 6h at constant temperature, taking out the tourmaline, centrifuging, and washing with water to obtain insoluble substances;
and 4, adding the insoluble substances into acetone, carrying out microwave reaction for 5 hours, filtering, and carrying out freeze drying to obtain yeast glucan powder.
The concentration of the yeast cell walls in the distilled water in the step 1 is 45g/L, the stirring speed for uniformly stirring is 1000r/min, and the constant-temperature heating temperature is 100 ℃.
The mass of the tourmaline in the step 1 is 10% of the cell wall of the common yeast, the addition amount of the hydrogen peroxide is 150% of the mass of the tourmaline, the temperature of the sealed cooking is 150 ℃, and the temperature of the heat preservation is 90 ℃.
The concentration of the precipitate in the step 2 in distilled water is 190g/L, the dispersant adopts polyvinylpyrrolidone, the adding amount is 5% of the mass of the precipitate, the heating and stirring temperature is 70 ℃, the stirring speed is 300r/min, and the alkali liquor adopts sodium hydroxide solution.
The adding amount of the protease in the step 3 is 0.3 percent of the mass of the precipitate, and the tourmaline in the step 1 is adopted as the tourmaline.
The temperature of the constant temperature enzymolysis in the step 3 is 40 ℃.
The concentration of the insoluble substances in the acetone in the step 4 is 30g/L, the temperature of the microwave reaction is 30 ℃, and the power is 500W.
The freeze drying in the step 4 comprises pre-freezing at-70 ℃ for 22h and freeze drying at-60 ℃ for 10 h.
Example 3
A green preparation method of yeast glucan comprises the following steps:
step 1, adding common yeast cell walls into distilled water, stirring uniformly to form a suspension, heating at a constant temperature for 3 hours, then adding tourmaline, adding hydrogen peroxide, sealing, steaming for 3 hours, preserving heat for 4 hours, taking out the tourmaline, and centrifuging to obtain a precipitate;
step 2, adding the precipitate into distilled water, uniformly stirring, adding a dispersing agent, heating and stirring to form a suspension, and then adding alkali liquor to adjust the pH to 10 to obtain an alkaline suspension;
step 3, adding protease and tourmaline into the alkaline suspension, carrying out enzymolysis for 5h at constant temperature, taking out the tourmaline, centrifuging, and washing with water to obtain insoluble substances;
and 4, adding the insoluble substances into acetone, carrying out microwave reaction for 4 hours, filtering, and carrying out freeze drying to obtain yeast glucan powder.
The concentration of the yeast cell walls in the distilled water in the step 1 is 30g/L, the stirring speed for uniformly stirring is 800r/min, and the constant-temperature heating temperature is 100 ℃.
The mass of the tourmaline in the step 1 is 8% of the cell wall mass of the common yeast, the addition amount of the hydrogen peroxide is 130% of the mass of the tourmaline, the temperature of the sealed cooking is 140 ℃, and the temperature of the heat preservation is 85 ℃.
The concentration of the precipitate in the step 2 in distilled water is 160g/L, the dispersant adopts polyvinylpyrrolidone, the adding amount is 4% of the mass of the precipitate, the heating and stirring temperature is 60 ℃, the stirring speed is 200r/min, and the alkali liquor adopts sodium hydroxide solution.
The adding amount of the protease in the step 3 is 0.2 percent of the mass of the precipitate, and the tourmaline in the step 1 is adopted as the tourmaline.
The temperature of the constant temperature enzymolysis in the step 3 is 35 ℃.
The concentration of the insoluble substances in the acetone in the step 4 is 20g/L, the temperature of the microwave reaction is 25 ℃, and the power is 400W.
The freeze drying in the step 4 comprises pre-freezing at-70 ℃ for 22h and freeze drying at-60 ℃ for 8 h.
Performance detection
| Example 1 | Example 2 | Example 3 | |
| Sample phase | Light yellow powder | Light yellow powder | Light yellow powder |
| Dextran content | 80% | 85% | 83% |
In summary, the invention has the following advantages:
1. the invention takes yeast cell walls as production raw materials, takes tourmaline as an auxiliary catalyst and an auxiliary oxidant, and greatly improves the yield of yeast glucan.
2. According to the invention, tourmaline is matched with hydrogen peroxide, yeast cell walls are opened in a cooking mode, the release of yeast glucan can be effectively improved, and the problem of low release rate of the traditional extraction process is solved.
3. The invention takes the tourmaline as an auxiliary catalyst, and promotes the enzymolysis efficiency of protease by the thermoelectricity of the tourmaline, thereby not only improving the enzymolysis speed, but also improving the enzymolysis depth.
4. The invention adopts a microwave reaction mode to promote the rapid dissolution of impurities in insoluble substances and reduce the residue of the impurities.
It should be understood that the detailed description of the invention is merely illustrative of the invention and is not intended to limit the invention to the specific embodiments described. It will be appreciated by those skilled in the art that the present invention may be modified or substituted equally as well to achieve the same technical result; as long as the use requirements are met, the method is within the protection scope of the invention.
Claims (6)
1. A green preparation method of yeast glucan is characterized by comprising the following steps: the method comprises the following steps:
step 1, adding common yeast cell walls into distilled water, stirring uniformly to form a suspension, heating at a constant temperature for 2-4h, then adding tourmaline, adding hydrogen peroxide, sealing, cooking for 2-4h, preserving heat for 2-6h, taking out the tourmaline, and centrifuging to obtain a precipitate; the concentration of the yeast cell walls in the distilled water is 15-45g/L, the stirring speed for uniform stirring is 500-1000r/min, the constant-temperature heating temperature is 90-100 ℃, the mass of the tourmaline is 2-10% of the mass of the common yeast cell walls, the addition amount of the hydrogen peroxide is 110-150% of the mass of the tourmaline, the temperature for sealed cooking is 120-150 ℃, and the heat preservation temperature is 80-90 ℃;
step 2, adding the precipitate into distilled water, stirring uniformly, adding a dispersing agent, heating and stirring to form a suspension, and then adding alkali liquor to adjust the pH to 9-11 to obtain an alkaline suspension;
step 3, adding protease and tourmaline into the alkaline suspension, carrying out enzymolysis for 4-6h at constant temperature, taking out the tourmaline, centrifuging, and washing with water to obtain insoluble substances;
and 4, adding the insoluble substances into acetone, carrying out microwave reaction for 2-5h, filtering, and carrying out freeze drying to obtain yeast glucan powder.
2. The green process for preparing yeast glucan according to claim 1, wherein: the concentration of the precipitate in the step 2 in distilled water is 190g/L, the dispersant adopts polyvinylpyrrolidone, the adding amount is 2-5% of the mass of the precipitate, the heating and stirring temperature is 50-70 ℃, the stirring speed is 300r/min, and the alkali liquor adopts sodium hydroxide solution.
3. The green process for preparing yeast glucan according to claim 1, wherein: the protease adding amount in the step 3 is 0.1-0.3% of the mass of the precipitate, and the tourmaline in the step 1 is adopted as the tourmaline.
4. The green process for preparing yeast glucan according to claim 1, wherein: the temperature of the constant temperature enzymolysis in the step 3 is 30-40 ℃.
5. The green process for preparing yeast glucan according to claim 1, wherein: the concentration of the insoluble substances in the acetone in the step 4 is 10-30g/L, the temperature of the microwave reaction is 20-30 ℃, and the power is 300-500W.
6. The green process for preparing yeast glucan according to claim 1, wherein: the freeze drying in the step 4 comprises pre-freezing at-70 ℃ for 22 hours and freeze drying at-60 ℃ for 6-10 hours.
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