CN110882285A - Efficient preparation method of active substances in phellinus igniarius - Google Patents
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Abstract
The invention discloses an efficient preparation method of active substances in phellinus igniarius, which comprises the following steps of S1, raw material pretreatment: drying and crushing phellinus igniarius sporocarp to form phellinus igniarius powder; s2, MgO and tetrahydrofuran treatment: adding MgO accounting for 40-60% of the weight of the phellinus igniarius powder and tetrahydrofuran accounting for 20-40 times of the weight of the phellinus igniarius powder into the phellinus igniarius powder, uniformly stirring, placing the phellinus igniarius powder into a reaction kettle, and extracting for 30-60 minutes at the temperature of 200 ℃ and 260 ℃; s3, ultrasonic treatment: cooling to room temperature after reaction, adding water 1-2 times of the volume of the mixture into the mixture, performing ultrasonic treatment at 30-50 deg.C for 10-30 min, and filtering the mixture to obtain filtrate containing Phellinus linteus polyphenol and Phellinus linteus polysaccharide. The filtrate can be further separated to obtain Phellinus linteus polyphenol and Phellinus linteus polysaccharide, respectively. The method can simultaneously prepare the phellinus igniarius polyphenol and the phellinus igniarius polysaccharide, improves the extraction rate of the phellinus igniarius polysaccharide and the phellinus igniarius polyphenol, has simple process, and is suitable for large-scale production.
Description
Technical Field
The invention relates to the field of edible and medicinal fungi processing, and particularly relates to an efficient preparation method of active substances in phellinus igniarius.
Background
Phellinus linteus is a precious edible and medicinal fungus and is called forest gold in the name of gold. Phellinus linteus is a traditional Chinese medicinal material in China, is recorded in Shennong Bai Cao Jing and Li Shizhen Ben Cao gang mu for the earliest time, is mainly used for relieving diarrhea, metrorrhagia, leukorrhagia, spleen deficiency and diarrhea, and can also benefit five internal organs, expel toxin and activate blood. According to research and statistics, more than 20 pharmacological functions of the phellinus igniarius include bacteriostasis, inflammation diminishing, oxidation resistance, tumor resistance, body immunity enhancement, liver protection, blood sugar reduction, blood fat reduction and pneumonia resistance.
The edible and medicinal fungi can not achieve the treatment effect when being directly eaten, and the efficacy can be improved and the pharmacological action can be exerted only by extracting, separating and purifying the effective components. In the preparation process of the active ingredients, extraction is the first step and the most critical step, and how to release the active ingredients to the maximum extent is of great significance.
Chinese patent document CN108610391A discloses a method for extracting polysaccharide and adenosine from phellinus igniarius sporocarp, comprising the following steps: (1) crushing phellinus igniarius sporocarp into phellinus igniarius sporocarp particles, and putting the phellinus igniarius sporocarp particles into an extraction tank; (2) adding water with the volume 6-10 times of that of phellinus igniarius sporocarp particles into an extraction tank, continuously refluxing for 1.5-3 hours, performing suction filtration, continuously refluxing filter residues for 2-3 hours by using water with the volume 4-5 times of that of the filter residues, and performing suction filtration; (3) mixing the filtrates obtained in the step (2) by suction filtration twice, and concentrating to obtain 0.3-0.5g/ml phellinus igniarius solution; (4) adjusting the pH value of the phellinus igniarius solution in the step (3) to 9-10 by using a sodium hydroxide solution, loading the phellinus igniarius solution with the adjusted pH value into a stainless steel chromatographic column filled with macroporous resin, and controlling the flow rate of an effluent liquid, wherein the obtained effluent liquid is a crude phellinus igniarius polysaccharide solution; (5) regulating the pH of the crude phellinus igniarius polysaccharide solution obtained in the step (4) to be neutral by using a sulfuric acid solution, filtering out macromolecules such as protein and the like by using a first ultrafiltration membrane, collecting filtrate, passing through a second ultrafiltration membrane for removing micromolecular impurities, and concentrating the obtained trapped fluid in a reduced pressure vacuum manner to obtain phellinus igniarius polysaccharide; (6) eluting the adsorbate adsorbed in the macroporous resin in the step (4) by using an ethanol solution, and concentrating the obtained eluent under reduced pressure and vacuum to obtain an extract; (7) and (3) mixing the extract obtained in the step (6) with silica gel by a dry method according to the mass ratio of 1:1.2, loading the mixture into a chromatographic column, eluting by using ethyl acetate with the volume of 1-2 columns, eluting by using a mixed eluent of ethyl acetate and acetone, collecting one fraction per 1500ml, detecting a liquid phase, combining 24 th-29 th fractions in the fractions, carrying out vacuum concentration under reduced pressure, combining the precipitates, filtering to obtain precipitates, standing the filtrate, filtering out after crystallization, mixing the filtered precipitates with the original precipitates, recrystallizing the mixed precipitates in an acetone-ethanol solution with the volume ratio of 6:4 to obtain colorless needle-like crystal adenosine, carrying out suction filtration, and carrying out vacuum drying in a vacuum drier at 50 ℃ to obtain an adenosine finished product.
Chinese patent document CN108542922A discloses a high-efficiency preparation method of active ingredients in phellinus linteus, which comprises the following steps: pretreatment of raw materials: (1) drying the phellinus igniarius sporocarp at 50-60 ℃, crushing, and sieving by a 40-60-mesh sieve to obtain phellinus igniarius powder; (2) mixing the phellinus igniarius powder obtained in the step one with deionized water according to a mass-volume ratio of 1: 20-1: 50, adding cellulase with the mass being 1-2% of the mass of the phellinus linteus powder after uniformly mixing, uniformly mixing again, and treating for 1-2 hours at the temperature of 45-55 ℃ to obtain a reaction mixture; (3) adding the reaction mixture obtained in the step two into an extraction kettle for extraction, wherein the pressure in the extraction kettle is 5-20 mPa, the extraction temperature is 100-200 ℃, the extraction time is 30-90 min, and an extraction liquid is obtained after the extraction is finished; (4) centrifuging the extract obtained in the third step for 10-20 min at the rotating speed of 4500-7000 r/min, and separating supernatant to obtain first supernatant; (5) adding chlorohexadecyl pyridine into the first supernatant, shaking uniformly, standing for 24-36 h at normal temperature, and centrifuging for 10-15 min at the rotating speed of 3500-7000 r/min to obtain a first precipitate and a second supernatant; (6) adding the first precipitate obtained in the fifth step into an ethanol solution of NaCl to obtain a precipitate mixed solution, placing the precipitate mixed solution in a shaking table, shaking for 1-1.5 h, centrifuging at a rotating speed of 3500-7000 r/min for 10-15 min to obtain a precipitate after centrifugation, washing the precipitate with 80% ethanol, and freeze-drying to obtain high-activity phellinus igniarius polysaccharide SP 1; (7) and (4) concentrating the second supernatant obtained in the fifth step under reduced pressure, and freeze-drying to obtain the extract rich in phellinus linteus triterpenes and flavones.
Chinese patent document CN106491662A discloses a method for simultaneously extracting polysaccharides and flavones from phellinus igniarius cultivation waste, comprising the following steps: (1) drying and crushing the phellinus igniarius cultivation waste to obtain phellinus igniarius cultivation waste dry powder; (2) mixing dry powder of phellinus igniarius cultivation waste with an alkaline ethanol solution according to the mass ratio of 1:10-1: 30; (3) carrying out ultrasonic treatment on the mixed solution in the step (2) for 20-40 minutes, and filtering to obtain filtrate and residues; (4) adding deionized water 8-12 times the weight of the residue into the residue in the step (3), uniformly mixing, adjusting the pH value to 4-6 to obtain a mixed solution, adding a complex enzyme accounting for 0.2-0.5% of the weight of the mixed solution, mixing the complex enzyme with cellulase and glucose oxidase, carrying out enzymolysis at 45-55 ℃ for 40-80 minutes, and carrying out centrifugal treatment on the enzymolysis solution to obtain a supernatant and a precipitate; the complex enzyme is cellulase: glucose oxidase 2-3: 1; (5) performing rotary evaporation on the filtrate obtained in the step (3), removing ethanol, adding ethyl acetate for extraction, separating liquid, and performing reduced pressure concentration and drying on the upper layer liquid to obtain phellinus igniarius flavone; (6) and (4) carrying out alcohol precipitation on the supernatant obtained in the step (4) after carrying out reduced pressure concentration by using a rotary evaporator, and centrifuging to obtain the supernatant and a precipitate, wherein the precipitate is phellinus igniarius polysaccharide.
In the prior art, no literature report for simultaneously extracting the phellinus igniarius polyphenol and the phellinus igniarius polysaccharide is found.
Disclosure of Invention
The inventor has analyzed that it is considered that the reason for this is that the fruiting body of Phellinus linteus has high lignification degree and complex structure, and the active substance is generally bound to lignin by physical or chemical bonds, and it is difficult to extract the effective components of Phellinus linteus polyphenol and Phellinus linteus polysaccharide simultaneously by the general method. Thus, breaking the bond between the active substance and lignin by removing lignin is also an effective means of extraction.
In view of the above, the present invention aims to provide a method for efficiently preparing an active substance from phellinus linteus, which can simultaneously and efficiently extract phellinus linteus polyphenol and phellinus linteus polysaccharide, and release the active ingredients phellinus linteus flavone and phellinus linteus polysaccharide to the maximum extent.
The adopted technical scheme is as follows:
a high-efficiency preparation method of active substances in phellinus igniarius comprises the following steps:
s1, raw material pretreatment: drying and crushing phellinus igniarius sporocarp to form phellinus igniarius powder;
s2, MgO and tetrahydrofuran treatment: adding MgO accounting for 40-60% of the weight of the phellinus igniarius powder and tetrahydrofuran accounting for 20-40 times of the weight of the phellinus igniarius powder into the phellinus igniarius powder, uniformly stirring, placing the phellinus igniarius powder into a reaction kettle, and extracting for 30-60 minutes at the temperature of 200 ℃ and 260 ℃;
s3, ultrasonic treatment: cooling to room temperature after reaction, adding water 1-2 times of the volume of the mixture into the mixture, performing ultrasonic treatment at 30-50 deg.C for 10-30 min, and filtering the mixture to obtain filtrate containing Phellinus linteus polyphenol and Phellinus linteus polysaccharide.
In the technical scheme, on one hand, tetrahydrofuran is low in toxicity, MgO is used as a catalyst, and lignin in phellinus igniarius sporocarp can be dissolved under the action of the catalyst MgO, so that more phellinus igniarius flavone and phellinus igniarius polysaccharide are released, and the extraction rate is improved.
On the other hand, tetrahydrofuran is used for dissolving lignin of phellinus igniarius sporocarp under the catalysis of MgO, and then ultrasonic treatment is carried out, so that effective components of phellinus igniarius flavone and phellinus igniarius polysaccharide are released to the maximum degree at the same time.
And further, S4, adding dichloromethane with the same volume to the filtrate for extraction, and drying an organic layer to obtain phellinus igniarius polyphenol. The yield of phellinus igniarius polyphenol is 2-4.7%.
Further comprises S5. concentrating the water layer, adding anhydrous alcohol to make the alcohol concentration reach 70-80% by volume fraction, collecting the precipitate to obtain Phellinus linteus polysaccharide. The yield of the phellinus igniarius polysaccharide is 8.7 to 11.2 percent.
Further, in S2, MgO in an amount of 50% by weight of the phellinus linteus powder and tetrahydrofuran in an amount of 20 to 40 times by weight of the phellinus linteus powder are added to the phellinus linteus powder.
Further, in S5, after concentrating the water layer, adding absolute ethyl alcohol to make the concentration of the ethyl alcohol reach 75% by volume fraction, and collecting the precipitate to obtain the phellinus igniarius polysaccharides.
Further, in S1, Phellinus linteus fruiting body is dried at 50-60 deg.C and then pulverized into superfine powder.
Similarly, from the perspective of products, the invention can also provide a mixed solution containing phellinus igniarius polyphenol and phellinus igniarius polysaccharide at the same time, and the mixed solution is prepared by the efficient preparation method in the scheme.
Further, the mixed solution is added with dichloromethane with the same volume for extraction, and the organic layer is dried to obtain the phellinus igniarius polyphenol, wherein the yield of the phellinus igniarius polyphenol is 2-4.7%.
Further, after the water layer is concentrated, absolute ethyl alcohol is added to ensure that the volume fraction of the ethyl alcohol reaches 70-80%, and the precipitate is collected to obtain the phellinus igniarius polysaccharides, wherein the yield of the phellinus igniarius polysaccharides is 8.7-11.2%.
The invention has the beneficial effects that:
the efficient preparation method of the active substances in the phellinus igniarius can be used for simultaneously preparing the phellinus igniarius polyphenol and the phellinus igniarius polysaccharide, improves the extraction rate of the phellinus igniarius polysaccharide and the phellinus igniarius polyphenol, is simple in process, and is suitable for large-scale production.
Detailed Description
The present invention is described in detail below with reference to specific examples, but the use and purpose of these exemplary embodiments are merely to exemplify the present invention, and do not set forth any limitation on the actual scope of the present invention in any form, and the scope of the present invention is not limited thereto.
Example 1
A high-efficiency preparation method of active substances in Phellinus linteus comprises the following steps,
s1, raw material pretreatment: drying and crushing phellinus igniarius sporocarp;
s2, MgO and tetrahydrofuran treatment: adding MgO accounting for 50 percent of the weight of the phellinus linteus powder and tetrahydrofuran accounting for 20 times of the weight of the phellinus linteus powder into the phellinus linteus powder, uniformly stirring the mixture, putting the mixture into a reaction kettle, and extracting the mixture for 30 minutes at 200 ℃;
s3, after the reaction is finished and the temperature is reduced to the room temperature, adding water with the volume being 1 time of that of the mixture into the mixture, carrying out ultrasonic treatment at the temperature of 30 ℃ for 10 minutes, and filtering the mixture;
s4, adding dichloromethane with the same volume into the filtrate for extraction, and drying the organic layer to obtain phellinus igniarius polyphenol, wherein the yield is 2.1%.
S5, adding absolute ethyl alcohol after the water layer is concentrated to enable the concentration of the ethyl alcohol to reach 75% by volume fraction, and collecting precipitates to obtain phellinus igniarius polysaccharides. The yield of phellinus igniarius polysaccharides is 8.9%.
Example 2
A high-efficiency preparation method of active substances in Phellinus linteus comprises the following steps,
s1, raw material pretreatment: drying and crushing phellinus igniarius sporocarp;
s2, MgO and tetrahydrofuran treatment: adding MgO accounting for 50 percent of the weight of the phellinus linteus powder and tetrahydrofuran accounting for 30 times of the weight of the phellinus linteus powder into the phellinus linteus powder, uniformly stirring the mixture, putting the mixture into a reaction kettle, and extracting the mixture for 40 minutes at 220 ℃;
D. after the reaction is finished and the temperature is reduced to room temperature, adding water with the volume 1.5 times of that of the mixture, carrying out ultrasonic treatment at 40 ℃ for 20 minutes, and filtering the mixture;
E. the filtrate is added with dichloromethane with the same volume for extraction, and the organic layer is dried to obtain phellinus igniarius polyphenol with the yield of 2.65 percent.
F. Concentrating the water layer, adding anhydrous ethanol to make ethanol concentration reach 75 vol%, and collecting precipitate to obtain Phellinus linteus polysaccharide. The yield of phellinus linteus polysaccharide is 9.47%.
Example 3
A high-efficiency preparation method of active substances in Phellinus linteus comprises the following steps,
s1, raw material pretreatment: drying and crushing phellinus igniarius sporocarp;
s2, MgO and tetrahydrofuran treatment: adding MgO accounting for 50 percent of the weight of the phellinus linteus powder and tetrahydrofuran accounting for 40 times of the weight of the phellinus linteus powder into the phellinus linteus powder, uniformly stirring the mixture, putting the mixture into a reaction kettle, and extracting the mixture for 50 minutes at 240 ℃;
s3, after the reaction is finished and the temperature is reduced to the room temperature, adding water with the volume being 1.8 times of that of the mixture into the mixture, carrying out ultrasonic treatment at 45 ℃ for 25 minutes, and filtering the mixture;
s4, adding dichloromethane with the same volume into the filtrate for extraction, and drying the organic layer to obtain phellinus igniarius polyphenol, wherein the yield is 3.46%.
S5, adding absolute ethyl alcohol after the water layer is concentrated to enable the concentration of the ethyl alcohol to reach 75% by volume fraction, and collecting precipitates to obtain phellinus igniarius polysaccharides. The yield of phellinus igniarius polysaccharides is 10.05%.
Example 4
A high-efficiency preparation method of active substances in Phellinus linteus comprises the following steps,
s1, raw material pretreatment: drying Phellinus linteus fruiting body at 55 deg.C, and micronizing;
s2, MgO and tetrahydrofuran treatment: adding MgO accounting for 50 percent of the weight of the phellinus linteus powder and tetrahydrofuran accounting for 35 times of the weight of the phellinus linteus powder into the phellinus linteus powder, uniformly stirring the mixture, putting the mixture into a reaction kettle, and extracting the mixture for 60 minutes at 250 ℃;
s3, after the reaction is finished and the temperature is reduced to the room temperature, adding water with the volume 2 times of that of the mixture into the mixture, carrying out ultrasonic treatment at the temperature of 50 ℃ for 30 minutes, and filtering the mixture;
s4, adding dichloromethane with the same volume into the filtrate for extraction, and drying the organic layer to obtain phellinus igniarius polyphenol, wherein the yield is 3.89%.
S5, adding absolute ethyl alcohol after the water layer is concentrated to enable the concentration of the ethyl alcohol to reach 75% by volume fraction, and collecting precipitates to obtain phellinus igniarius polysaccharides. The yield of phellinus igniarius polysaccharides is 10.54%.
Example 5
A high-efficiency preparation method of active substances in Phellinus linteus comprises the following steps,
s1, raw material pretreatment: drying and crushing phellinus igniarius sporocarp;
s2, MgO and tetrahydrofuran treatment: adding MgO accounting for 50 percent of the weight of the phellinus linteus powder and tetrahydrofuran accounting for 40 times of the weight of the phellinus linteus powder into the phellinus linteus powder, uniformly stirring the mixture, putting the mixture into a reaction kettle, and extracting the mixture for 50 minutes at 260 ℃;
s3, after the reaction is finished and the temperature is reduced to the room temperature, adding water with the volume 1.6 times of that of the mixture into the mixture, carrying out ultrasonic treatment at 45 ℃ for 25 minutes, and filtering the mixture;
s4, adding dichloromethane with the same volume into the filtrate for extraction, and drying the organic layer to obtain phellinus igniarius polyphenol, wherein the yield is 3.55%.
S5, adding absolute ethyl alcohol after the water layer is concentrated to enable the concentration of the ethyl alcohol to reach 75% by volume fraction, and collecting precipitates to obtain phellinus igniarius polysaccharides. The yield of phellinus linteus polysaccharide is 9.98%.
The above-listed detailed description is only a specific description of a possible embodiment of the present invention, and they are not intended to limit the scope of the present invention, and equivalent embodiments or modifications made without departing from the technical spirit of the present invention should be included in the scope of the present invention.
Claims (9)
1. A high-efficiency preparation method of active substances in phellinus igniarius is characterized by comprising the following steps:
s1, raw material pretreatment: drying and crushing phellinus igniarius sporocarp to form phellinus igniarius powder;
s2, MgO and tetrahydrofuran treatment: adding MgO accounting for 40-60% of the weight of the phellinus igniarius powder and tetrahydrofuran accounting for 20-40 times of the weight of the phellinus igniarius powder into the phellinus igniarius powder, uniformly stirring, placing the phellinus igniarius powder into a reaction kettle, and extracting for 30-60 minutes at the temperature of 200 ℃ and 260 ℃;
s3, ultrasonic treatment: cooling to room temperature after reaction, adding water 1-2 times of the volume of the mixture into the mixture, performing ultrasonic treatment at 30-50 deg.C for 10-30 min, and filtering the mixture to obtain filtrate containing Phellinus linteus polyphenol and Phellinus linteus polysaccharide.
2. The method for efficiently preparing active substances in phellinus igniarius according to claim 1, further comprising S4. adding dichloromethane with the same volume as the filtrate for extraction, and drying the organic layer to obtain phellinus igniarius polyphenol.
3. The method for efficiently preparing an active substance in Phellinus linteus according to claim 2, further comprising S5. concentrating the water layer, adding absolute ethanol to make the ethanol concentration reach 70-80% by volume fraction, and collecting the precipitate to obtain Phellinus linteus polysaccharide.
4. The method for efficiently producing an active substance in Phellinus linteus according to claim 3, wherein MgO is added to Phellinus linteus powder in an amount of 50% by weight of Phellinus linteus powder and tetrahydrofuran is added in an amount of 20-40 times by weight of Phellinus linteus powder in S2.
5. The method for efficiently producing an active substance from Phellinus linteus according to claim 3, wherein in S5, the water layer is concentrated and then absolute ethanol is added to make the ethanol concentration reach 75% by volume, and the precipitate is collected to obtain Phellinus linteus polysaccharide.
6. The method for efficiently producing an active substance in Phellinus linteus according to claim 3, wherein in S1, Phellinus linteus fruiting body is dried at 50-60 deg.C and then pulverized.
7. A mixed solution containing both Phellinus linteus polyphenol and Phellinus linteus polysaccharide, which is prepared by the efficient preparation method according to claim 1.
8. The mixed solution containing both Phellinus linteus polyphenol and Phellinus linteus polysaccharide as claimed in claim 7, wherein the same volume of dichloromethane is added for extraction, and organic layer is dried to obtain Phellinus linteus polyphenol with yield of 2% -4.7%.
9. The mixed solution containing both Phellinus linteus polyphenol and Phellinus linteus polysaccharide as claimed in claim 7, wherein the water layer is concentrated, and then anhydrous ethanol is added to make ethanol concentration reach 70-80% by volume, and precipitate is collected to obtain Phellinus linteus polysaccharide, and yield of Phellinus linteus polysaccharide is 8.7% -11.2%.
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| CN111925458A (en) * | 2020-08-24 | 2020-11-13 | 浙江省农业科学院 | Efficient preparation method of phellinus igniarius polysaccharides |
| CN112110886A (en) * | 2020-10-10 | 2020-12-22 | 长春师范大学 | Method for separating polyphenol compounds in phellinus igniarius by utilizing high-efficiency countercurrent chromatography |
| CN114832022A (en) * | 2022-06-16 | 2022-08-02 | 湖北省农业科学院农产品加工与核农技术研究所 | Preparation of phellinus igniarius sporocarp phenolic active substance and application of phellinus igniarius sporocarp phenolic active substance in regulation of intestinal flora and uric acid metabolism |
| CN114832022B (en) * | 2022-06-16 | 2023-09-15 | 湖北省农业科学院农产品加工与核农技术研究所 | Preparation of Phellinus linteus fruiting body phenol active substances and application thereof in regulating intestinal flora and uric acid metabolism |
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