CN114774493A - Method for extracting and purifying phellinus igniarius active substance polysaccharide - Google Patents
Method for extracting and purifying phellinus igniarius active substance polysaccharide Download PDFInfo
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Abstract
The invention discloses a method for extracting and purifying phellinus igniarius active substance polysaccharide, which comprises the following steps: freeze-drying and crushing at low temperature, performing enzymolysis after ultraviolet irradiation, performing hot water-ultrasonic extraction, extracting crude phellinus igniarius polysaccharides, degreasing, decoloring, deproteinizing, and dialyzing to finally obtain high-purity phellinus igniarius active substance polysaccharides. The invention extracts phellinus igniarius polysaccharides to the maximum extent by ultraviolet irradiation treatment and combined extraction of enzymolysis and ultrasound, and finally obtains the high-purity phellinus igniarius polysaccharides by degreasing, depigmentation and deproteinization treatment. The method provided by the invention has the advantages of high extraction rate, short extraction time, convenient operation, high purity of the obtained phellinus igniarius polysaccharides and great medicinal value.
Description
Technical Field
The invention belongs to the technical field of active substance extraction, and particularly relates to a method for extracting and purifying phellinus igniarius active substance polysaccharide.
Background
Phellinus igniarius is a large precious medicinal fungus belonging to Basidiomycetes class, and is sweet, pungent and slightly bitter in taste. Modern researches show that phellinus igniarius has the efficacies of resisting tumors, resisting oxidation and the like, the efficacy of phellinus igniarius is mainly that active substances of phellinus igniarius play a role, and polysaccharide is the most abundant in the active substances. At present, phellinus igniarius polysaccharide products are used for treating cancers, but the price is high in the market, and the application of phellinus igniarius polysaccharide as an immune type medicine to clinic is a new direction for treating tumors in recent years, and the phellinus igniarius polysaccharide participates in metabolic regulation activities of organisms with multiple functions, so that the phellinus igniarius polysaccharide is more and more emphasized in recent years, and the research on the phellinus igniarius polysaccharide is gradually increased.
At present, research on phellinus igniarius polysaccharides mainly focuses on phellinus igniarius sporophores, the phellinus igniarius growth period is long, the obtained sporophores are more time-consuming and raw materials, the phellinus igniarius polysaccharides are expensive in price and not widely used in production and medical treatment. If the content of the phellinus linteus mycelium polysaccharide is slightly different from the components and the fruit body, the polysaccharide in the mycelium can be extracted and processed, so that a large amount of economic cost can be reduced, and the phellinus linteus mycelium polysaccharide can be applied to the market. The traditional sample pretreatment mainly adopts a water boiling method, the extraction time is more than 6 hours, the energy consumption is high, and the yield of crude polysaccharide is low and is only 2 percent of the crude drug. Then the crude phellinus linteus polysaccharide is purified by a gel filtration technology, the process is complicated, the time consumption is long, the product purity is low, generally about 25 percent, and the phellinus linteus polysaccharide product with higher purity is needed as a pharmaceutical preparation.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides a method for extracting and purifying phellinus igniarius active substance polysaccharide so as to overcome the technical defects of low yield and low purity of phellinus igniarius polysaccharide in the prior art.
In order to achieve the purpose, the invention provides the following technical scheme:
a method for extracting and purifying phellinus igniarius active substance polysaccharide comprises the following steps:
(1) freeze-drying and crushing at low temperature: crushing the cleaned phellinus igniarius mycelium at low temperature, and sieving to obtain phellinus igniarius powder;
(2) enzymolysis after ultraviolet irradiation: placing the phellinus linteus powder obtained in the step (1) into an ultraviolet irradiation field for irradiation, then adding deionized water and a complex enzyme, and filtering after enzymolysis to obtain an enzymolysis filtrate and filter residues;
(3) leaching: adding deionized water into the filter residue obtained in the step (2), performing ultrasonic extraction, centrifuging after extraction is finished, and taking supernatant to obtain an extract and residue;
(4) extracting crude phellinus igniarius polysaccharides: mixing the enzymolysis filtrate obtained in the step (2) and the leaching liquor obtained in the step (3), heating, concentrating, centrifuging, removing precipitate, adding 95% ethanol into the supernatant, standing for 24h, centrifuging, collecting precipitate, and drying to obtain crude phellinus igniarius polysaccharide;
(5) degreasing, depigmenting and deproteinizing: adding petroleum ether into the crude phellinus igniarius polysaccharide obtained in the step (4), centrifuging, sucking and removing the petroleum ether, repeating for 2 times, and removing lipid substances; adding hydrogen peroxide into the degreased polysaccharide to the concentration of 20%, performing water bath reaction, removing pigments, and adjusting the pH value of the polysaccharide solution to 7.0 by using ammonia water; adding trichloroacetic acid into the decolored crude polysaccharide sample until the concentration is 5%, standing, centrifuging, discarding the precipitate, and repeating for 3 times to obtain a polysaccharide solution which is degreased, decolored and deproteinized;
(6) and (3) dialysis: and (4) selecting a 7000D dialysis bag, dialyzing the polysaccharide solution subjected to degreasing, depigmentation and deproteinization obtained in the step (5) by using the dialysis bag to obtain a purified phellinus igniarius polysaccharide solution, and then concentrating under reduced pressure to obtain the purified phellinus igniarius polysaccharide.
Preferably, the crushing temperature in the step (1) is-65 to-45 ℃, and the sieving mesh number is 40 to 80 meshes.
Preferably, the ultraviolet irradiation dose in the step (2) is 1000-1500 mJ/cm2(ii) a The compound enzyme is pectinThe enzyme, the cellulase and the papain are mixed to prepare the enzyme-enzyme feed, and the mass ratio of the three enzymes is 1: 1: 1.
preferably, the feed-liquid ratio of the phellinus linteus powder to the deionized water in the step (2) is 1 g: 45-50 mL; the mass ratio of the phellinus igniarius powder to the complex enzyme is 10: 1, the enzymolysis temperature is 25-30 ℃, and the enzymolysis time is 5-10 min.
Preferably, the material-liquid ratio of the filter residue to the deionized water in the step (3) is 1 g: 45mL, wherein the ultrasonic extraction temperature is 50-70 ℃, the ultrasonic extraction time is 15-30 min, the centrifugation speed is 4000-5000 r/min, and the centrifugation time is 15-30 min.
Preferably, the centrifugation speed in the step (5) is 1800-2000 r/min, and the centrifugation time is 15-20 min.
Preferably, in the step (5), the concentration of the hydrogen peroxide is 5%, the water bath temperature is 45-50 ℃, and the water bath time is 1-2 hours.
Preferably, the concentration of the trichloroacetic acid in the step (5) is 1.5mol/L, the standing temperature is 60 ℃, and the standing time is 30 min.
Preferably, the dialysis time in the step (6) is 24 hours, and water is changed every 2-3 hours.
Compared with the prior art, the invention has the following beneficial effects:
(1) the method comprises the following steps of firstly, freezing and crushing phellinus igniarius mycelia at a low temperature, wherein cell sap in cell walls can be in a frozen state at the low temperature, and the cell sap cannot remain on the inner wall of a machine during crushing; meanwhile, the phellinus igniarius hyphae in a low-temperature state are high in brittleness and easy to crush, and the crushing efficiency is effectively improved; the crushed phellinus igniarius powder is processed at a lower temperature, is immediately put into a eutectic solvent water solution at the normal temperature, and under the action of thermal expansion, cooling and permeation, cell walls of phellinus igniarius mycelia are damaged, and phellinus igniarius active substance polysaccharide is extracted more quickly.
(2) The method has the advantages that the active substance polysaccharide in the phellinus igniarius mycelia is extracted by applying the ultraviolet irradiation technology, so that the extraction rate of the polysaccharide is improved, and meanwhile, under the combined extraction action of enzymolysis and ultrasonic waves, the extraction time is greatly shortened, and the extraction efficiency is improved; finally, the polysaccharide is subjected to degreasing, depigmentation and deproteinization treatment, so that the purity of the polysaccharide is improved, and the prepared phellinus igniarius active substance polysaccharide has high purity and great medicinal value.
Detailed Description
The technical solutions of the present invention will be described below clearly and completely in conjunction with the embodiments, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1
A method for extracting and purifying phellinus igniarius active substance polysaccharide comprises the following steps:
(1) low-temperature freeze-drying and crushing: pulverizing cleaned Phellinus Linteus mycelium at-65 deg.C, and sieving with 40 mesh sieve to obtain Phellinus Linteus powder;
(2) carrying out enzymolysis after ultraviolet irradiation: putting the phellinus igniarius powder (10g) obtained in the step (1) into an ultraviolet radiation field for radiation, wherein the radiation dose is 1000mJ/cm2Adding 450mL of deionized water and 1g of complex enzyme (the mass ratio of pectinase to cellulase to papain is 1: 1: 1), carrying out enzymolysis for 5min at 25 ℃, and filtering to obtain enzymolysis filtrate and filter residue;
(3) leaching: adding 450mL of deionized water into the filter residue (10g) obtained in the step (2), performing ultrasonic leaching at 50 ℃ for 15min, centrifuging at the speed of 4000r/min for 15min after the completion of the extraction, and taking supernatant to obtain leaching liquor and residues;
(4) extracting crude phellinus igniarius polysaccharide: mixing the enzymolysis filtrate obtained in the step (2) and the leaching liquor obtained in the step (3), heating and concentrating, centrifuging, removing precipitate, adding equal volume of 95% ethanol into supernate, standing for 24h, centrifuging, collecting precipitate, and drying to obtain crude phellinus igniarius polysaccharide;
(5) degreasing, depigmenting and deproteinizing: adding 50mL of petroleum ether into the crude phellinus igniarius polysaccharide obtained in the step (4), centrifuging at 1800r/min for 15min, sucking and removing the petroleum ether, repeating for 2 times, and removing lipid substances; adding 5% hydrogen peroxide to polysaccharide after degreasing to polysaccharide with polysaccharide concentration of 20%, performing water bath reaction at 45 deg.C for 1h, removing pigment, and adjusting pH of polysaccharide solution to 7.0 with ammonia water; adding 1.5mol/L trichloroacetic acid to the decolored crude polysaccharide sample until the concentration of the most sugar is 5%, standing for 30min at 60 ℃, centrifuging, discarding the precipitate, and repeating for 3 times to obtain a polysaccharide solution which is degreased, decolored and deproteinized;
(6) and (3) dialysis: and (3) selecting a 7000D dialysis bag, dialyzing the polysaccharide solution subjected to degreasing, depigmentation and deproteinization obtained in the step (5) by using the dialysis bag, wherein the dialysis time is 24 hours, changing water every 2 hours to obtain a purified phellinus igniarius polysaccharide solution, and then carrying out reduced pressure concentration to obtain the purified phellinus igniarius polysaccharide.
Example 2
A method for extracting and purifying phellinus igniarius active substance polysaccharide comprises the following steps:
(1) freeze-drying and crushing at low temperature: pulverizing cleaned Phellinus Linteus mycelium at-55 deg.C, and sieving with 60 mesh sieve to obtain Phellinus Linteus powder;
(2) carrying out enzymolysis after ultraviolet irradiation: putting the phellinus igniarius powder (10g) obtained in the step (1) into an ultraviolet irradiation field for irradiation, wherein the irradiation dose is 1200mJ/cm2Then 480mL of deionized water and 1g of complex enzyme (the mass ratio of pectinase to cellulase to papain is 1: 1: 1) are added, enzymolysis is carried out for 8min at 27 ℃, and an enzymolysis filtrate and filter residue are obtained after filtration;
(3) leaching: adding 450mL of deionized water into the filter residue (10g) obtained in the step (2), performing ultrasonic leaching at 60 ℃ for 20min, centrifuging at the speed of 4500r/min for 20min after the extraction is finished, and taking supernatant to obtain leaching liquor and residue;
(4) extracting crude phellinus igniarius polysaccharides: mixing the enzymolysis filtrate obtained in the step (2) and the leaching liquor obtained in the step (3), heating, concentrating, centrifuging, removing precipitate, adding equal volume of 95% ethanol into the supernatant, standing for 24h, centrifuging, collecting precipitate, and drying to obtain crude polysaccharide of Phellinus igniarius;
(5) degreasing, depigmenting and deproteinizing: adding 50mL of petroleum ether into the crude polysaccharide of the phellinus igniarius obtained in the step (4), centrifuging at 1900r/min for 18min, sucking and removing the petroleum ether, repeating for 2 times, and removing lipid substances; adding 5% hydrogen peroxide to polysaccharide after degreasing till the polysaccharide concentration is 20%, performing water bath reaction at 48 ℃ for 1.5h, removing pigment, and adjusting the pH of the polysaccharide solution to 7.0 by using ammonia water; adding 1.5mol/L trichloroacetic acid to the decolorized crude polysaccharide sample until the polysaccharide concentration is 5%, standing at 60 deg.C for 30min, centrifuging, removing precipitate, and repeating for 3 times to obtain defatted, decolorized, and deproteinized polysaccharide solution;
(6) and (3) dialysis: and (4) selecting a 7000D dialysis bag, dialyzing the polysaccharide solution subjected to degreasing, depigmentation and deproteinization obtained in the step (5) by using the dialysis bag, wherein the dialysis time is 24 hours, water is changed every 3 hours to obtain a purified phellinus igniarius polysaccharide solution, and then carrying out reduced pressure concentration to obtain the purified phellinus igniarius polysaccharide.
Example 3
A method for extracting and purifying phellinus igniarius active substance polysaccharide comprises the following steps:
(1) freeze-drying and crushing at low temperature: pulverizing cleaned Phellinus Linteus mycelium at-45 deg.C, and sieving with 80 mesh sieve to obtain Phellinus Linteus powder;
(2) enzymolysis after ultraviolet irradiation: placing 10g of phellinus linteus powder obtained in the step (1) into an ultraviolet irradiation field for irradiation, wherein the irradiation dose is 1500mJ/cm2Then adding 500mL of deionized water and 1g of complex enzyme (the mass ratio of pectinase to cellulase to papain is 1: 1: 1), carrying out enzymolysis for 10min at 30 ℃, and filtering to obtain enzymolysis filtrate and filter residue;
(3) leaching: adding 450mL of deionized water into the filter residue (10g) obtained in the step (2), performing ultrasonic leaching at 70 ℃ for 30min, centrifuging at the speed of 5000r/min for 30min after extraction is finished, and taking supernatant to obtain leaching liquor and residue;
(4) extracting crude phellinus igniarius polysaccharide: mixing the enzymolysis filtrate obtained in the step (2) and the leaching liquor obtained in the step (3), heating, concentrating, centrifuging, removing precipitate, adding equal volume of 95% ethanol into the supernatant, standing for 24h, centrifuging, collecting precipitate, and drying to obtain crude polysaccharide of Phellinus igniarius;
(5) degreasing, depigmenting and deproteinizing: adding 50mL of petroleum ether into the crude phellinus igniarius polysaccharide obtained in the step (4), centrifuging at 2000r/min for 20min, sucking and removing the petroleum ether, repeating for 2 times, and removing lipid substances; adding 5% hydrogen peroxide to polysaccharide after degreasing to polysaccharide with polysaccharide concentration of 20%, performing water bath reaction at 50 deg.C for 2 hr, removing pigment, and adjusting pH of polysaccharide solution to 7.0 with ammonia water; adding 1.5mol/L trichloroacetic acid to the decolorized crude polysaccharide sample until the polysaccharide concentration is 5%, standing at 60 deg.C for 30min, centrifuging, removing precipitate, and repeating for 3 times to obtain defatted, decolorized, and deproteinized polysaccharide solution;
(6) and (3) dialysis: and (4) selecting a 7000D dialysis bag, dialyzing the polysaccharide solution subjected to degreasing, depigmentation and deproteinization obtained in the step (5) by using the dialysis bag, wherein the dialysis time is 24 hours, water is changed every 3 hours to obtain a purified phellinus igniarius polysaccharide solution, and then carrying out reduced pressure concentration to obtain the purified phellinus igniarius polysaccharide.
Comparative example 1
A method for extracting and purifying phellinus igniarius active substance polysaccharide comprises the following steps:
(1) low-temperature freeze-drying and crushing: pulverizing cleaned Phellinus Linteus mycelium at-65 deg.C, and sieving with 40 mesh sieve to obtain Phellinus Linteus powder;
(2) enzymolysis: adding 450mL of deionized water and 1g of complex enzyme (the mass ratio of pectinase to cellulase to papain is 1: 1: 1) into 10g of the phellinus linteus powder obtained in the step (1), carrying out enzymolysis for 5min at 25 ℃, and filtering to obtain enzymolysis filtrate and filter residue;
(3) leaching: adding 450mL of deionized water into the filter residue (10g) obtained in the step (2), performing ultrasonic leaching at 50 ℃ for 15min, centrifuging at the speed of 4000r/min for 15min after the completion of the extraction, and taking supernatant to obtain leaching liquor and residues;
(4) extracting crude phellinus igniarius polysaccharide: mixing the enzymolysis filtrate obtained in the step (2) and the leaching liquor obtained in the step (3), heating, concentrating, centrifuging, removing precipitate, adding equal volume of 95% ethanol into the supernatant, standing for 24h, centrifuging, collecting precipitate, and drying to obtain crude polysaccharide of Phellinus igniarius;
(5) degreasing, depigmenting and deproteinizing: adding 50mL of petroleum ether into the crude phellinus igniarius polysaccharide obtained in the step (4), centrifuging at 1800r/min for 15min, sucking and removing the petroleum ether, repeating for 2 times, and removing lipid substances; adding 5% hydrogen peroxide to polysaccharide after degreasing to polysaccharide with the polysaccharide concentration of 20%, performing water bath reaction at 45 deg.C for 1h, removing pigment, and adjusting pH of polysaccharide solution to 7.0 with ammonia water; adding 1.5mol/L trichloroacetic acid to the decolorized crude polysaccharide sample until the polysaccharide concentration is 5%, standing at 60 deg.C for 30min, centrifuging, removing precipitate, and repeating for 3 times to obtain defatted, decolorized, and deproteinized polysaccharide solution;
(6) and (3) dialysis: and (3) selecting a 7000D dialysis bag, dialyzing the polysaccharide solution subjected to degreasing, depigmentation and deproteinization obtained in the step (5) by using the dialysis bag, wherein the dialysis time is 24 hours, changing water every 2 hours to obtain a purified phellinus igniarius polysaccharide solution, and then carrying out reduced pressure concentration to obtain the purified phellinus igniarius polysaccharide.
Comparative example 2
A method for extracting and purifying phellinus igniarius active substance polysaccharide comprises the following steps:
(1) low-temperature freeze-drying and crushing: pulverizing cleaned Phellinus Linteus mycelium at-65 deg.C, and sieving with 40 mesh sieve to obtain Phellinus Linteus powder;
(2) ultraviolet irradiation: putting the phellinus igniarius powder (10g) obtained in the step (1) into an ultraviolet radiation field for radiation, wherein the radiation dose is 1000mJ/cm2Then adding 450mL of deionized water, stirring at 25 ℃ for 5min, and filtering to obtain filtrate and filter residue;
(3) leaching: adding 450mL of deionized water into the filter residue (10g) obtained in the step (2), performing ultrasonic leaching at 50 ℃ for 15min, centrifuging at the speed of 4000r/min for 15min after the completion of the extraction, and taking supernatant to obtain leaching liquor and residues;
(4) extracting crude phellinus igniarius polysaccharide: mixing the filtrate obtained in the step (2) and the leaching liquor obtained in the step (3), heating and concentrating, centrifuging, removing precipitates, adding 95% ethanol with the same volume into supernate, standing for 24h, centrifuging, collecting precipitates, and drying to obtain crude phellinus igniarius polysaccharides;
(5) degreasing, depigmenting and deproteinizing: adding 50mL of petroleum ether into the crude phellinus igniarius polysaccharide obtained in the step (4), centrifuging at 1800r/min for 15min, sucking and removing the petroleum ether, repeating for 2 times, and removing lipid substances; adding 5% hydrogen peroxide to polysaccharide after degreasing to polysaccharide with polysaccharide concentration of 20%, performing water bath reaction at 45 deg.C for 1h, removing pigment, and adjusting pH of polysaccharide solution to 7.0 with ammonia water; adding 1.5mol/L trichloroacetic acid to the decolored crude polysaccharide sample until the concentration of the most sugar is 5%, standing for 30min at 60 ℃, centrifuging, discarding the precipitate, and repeating for 3 times to obtain a polysaccharide solution which is degreased, decolored and deproteinized;
(6) and (3) dialysis: and (3) selecting a 7000D dialysis bag, dialyzing the polysaccharide solution subjected to degreasing, depigmentation and deproteinization obtained in the step (5) by using the dialysis bag, wherein the dialysis time is 24 hours, changing water every 2 hours to obtain a purified phellinus igniarius polysaccharide solution, and then carrying out reduced pressure concentration to obtain the purified phellinus igniarius polysaccharide.
The extraction rate and the purity of the purified polysaccharide in examples 1 to 3 and comparative examples 1 to 2 were calculated respectively according to the following formula, and the results are summarized in Table 1.
The extraction yield (g/g) is equal to the amount of crude phellinus igniarius polysaccharide/phellinus igniarius powder multiplied by 100%.
Taking 10mL of purified polysaccharide solution, and obtaining the polysaccharide by using alcohol precipitation, centrifugation and drying methods. The concentration of the crude phellinus igniarius polysaccharide is calculated by adopting a phenol-sulfuric acid color development method, and the purity of the crude phellinus igniarius polysaccharide after purification is further calculated.
Purity (%) of purified polysaccharide C1/C2%
In the formula: c1 is the concentration of pure polysaccharide in the purified polysaccharide; c2 is the concentration of polysaccharide before purification.
TABLE 1 Phellinus linteus polysaccharide extraction rate and purity test results
| Extraction ratio of crude Phellinus Linteus polysaccharide (%) | Phellinus linteus polysaccharide purity (%) | |
| Example 1 | 6.53 | 73.6 |
| Example 2 | 6.21 | 68.5 |
| Example 3 | 6.48 | 72.1 |
| Comparative example 1 | 3.19 | 36.1 |
| Comparative example 2 | 2.94 | 34.9 |
As shown in the above results, the extraction rate of crude Phellinus linteus polysaccharides in comparative examples 1-2 is lower than that in examples 1-3, which shows that the extraction rate of Phellinus linteus polysaccharides can be improved by the extraction method of the present invention, and the purity of Phellinus linteus polysaccharides in examples 1-3 is higher than that in comparative examples 1-2, which shows that the purity of Phellinus linteus polysaccharides can be improved by the purification method of the present invention.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (9)
1. A method for extracting and purifying phellinus igniarius active substance polysaccharide is characterized by comprising the following steps:
(1) low-temperature freeze-drying and crushing: pulverizing cleaned Phellinus Linteus mycelium at low temperature, and sieving to obtain Phellinus Linteus powder;
(2) carrying out enzymolysis after ultraviolet irradiation: placing the phellinus linteus powder obtained in the step (1) into an ultraviolet irradiation field for irradiation, then adding deionized water and a complex enzyme, and filtering after enzymolysis to obtain an enzymolysis filtrate and filter residues;
(3) hot water-ultrasonic extraction: adding deionized water into the filter residue obtained in the step (2), performing ultrasonic extraction, centrifuging after extraction is finished, and taking supernatant to obtain an extract and residue;
(4) extracting crude phellinus igniarius polysaccharide: mixing the enzymolysis filtrate obtained in the step (2) and the leaching liquor obtained in the step (3), heating and concentrating, centrifuging, removing precipitate, adding 95% ethanol into supernate, standing for 24h, centrifuging, collecting precipitate, and drying to obtain crude phellinus igniarius polysaccharide;
(5) degreasing, depigmenting and deproteinizing: adding petroleum ether into the crude phellinus igniarius polysaccharide obtained in the step (4), centrifuging, sucking and removing the petroleum ether, repeating for 2 times, and removing lipid substances; adding hydrogen peroxide into the degreased polysaccharide to the concentration of 20%, performing water bath reaction, removing pigments, and adjusting the pH value of the polysaccharide solution to 7.0 by using ammonia water; adding trichloroacetic acid into the decolored crude polysaccharide sample until the concentration is 5%, standing, centrifuging, discarding the precipitate, and repeating for 3 times to obtain a polysaccharide solution which is degreased, decolored and deproteinized;
(6) and (3) dialysis: and (4) selecting a 7000D dialysis bag, dialyzing the polysaccharide solution subjected to degreasing, depigmentation and deproteinization obtained in the step (5) by using the dialysis bag to obtain a purified phellinus igniarius polysaccharide solution, and then concentrating under reduced pressure to obtain the purified phellinus igniarius polysaccharide.
2. The extraction method according to claim 1, wherein the crushing temperature in the step (1) is-65 to-45 ℃, and the sieving mesh number is 40 to 80 meshes.
3. The extraction method according to claim 1, wherein the ultraviolet irradiation dose in the step (2) is 1000 to 1500mJ/cm2(ii) a The compound enzyme is prepared by mixing pectinase, cellulase and papain, wherein the mass ratio of the three enzymes is 1: 1: 1.
4. the extraction method according to claim 1, wherein the feed-liquid ratio of the phellinus linteus powder to deionized water in step (2) is 1 g: 45-50 mL; the mass ratio of the phellinus linteus powder to the complex enzyme is 10: 1, the enzymolysis temperature is 25-30 ℃, and the enzymolysis time is 5-10 min.
5. The extraction method according to claim 1, wherein the feed-liquid ratio of the filter residue to the deionized water in the step (3) is 1 g: 45mL, wherein the ultrasonic extraction temperature is 50-70 ℃, the ultrasonic extraction time is 15-30 min, the centrifugation speed is 4000-5000 r/min, and the centrifugation time is 15-30 min.
6. The method of claim 1, wherein the centrifugation speed in step (5) is 1800-2000 r/min and the centrifugation time is 15-20 min.
7. The extraction method according to claim 1, wherein the concentration of the hydrogen peroxide in the step (5) is 5%, the temperature of the water bath is 45-50 ℃, and the time of the water bath is 1-2 h.
8. The extraction method according to claim 1, wherein the concentration of trichloroacetic acid in the step (5) is 1.5mol/L, the standing temperature is 60 ℃, and the standing time is 30 min.
9. The extraction method according to claim 1, wherein the dialysis time in step (6) is 24 hours, and the water is changed every 2-3 hours.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116217748A (en) * | 2023-03-01 | 2023-06-06 | 宁波御菌生物技术有限公司 | Method for extracting polysaccharide from Phellinus linteus fruiting body |
| CN116535533A (en) * | 2023-04-28 | 2023-08-04 | 宁波御菌生物技术有限公司 | Extraction process of selenium-containing Phellinus linteus polysaccharide |
| CN116535533B (en) * | 2023-04-28 | 2024-06-07 | 宁波御菌生物技术有限公司 | Extraction process of selenium-containing Phellinus linteus polysaccharide |
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2022
- 2022-04-25 CN CN202210439931.6A patent/CN114774493A/en active Pending
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116217748A (en) * | 2023-03-01 | 2023-06-06 | 宁波御菌生物技术有限公司 | Method for extracting polysaccharide from Phellinus linteus fruiting body |
| CN116217748B (en) * | 2023-03-01 | 2024-05-14 | 宁波御菌生物技术有限公司 | Method for extracting polysaccharide from Phellinus linteus fruiting body |
| CN116535533A (en) * | 2023-04-28 | 2023-08-04 | 宁波御菌生物技术有限公司 | Extraction process of selenium-containing Phellinus linteus polysaccharide |
| CN116535533B (en) * | 2023-04-28 | 2024-06-07 | 宁波御菌生物技术有限公司 | Extraction process of selenium-containing Phellinus linteus polysaccharide |
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