CN103725730B - A kind of process for refining of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides - Google Patents
A kind of process for refining of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides Download PDFInfo
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Abstract
The present invention relates to a kind of process for refining of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides.It and common hericium erinaceum polysaccharide have larger different on structure forms, at present for the rare report of correlative study of Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium polysaccharides.The Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium utilizing the present invention to produce, in mycelial process of growth, add selenium element, after extracting mycelium polysaccharides, the biological action of Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium polysaccharides class material can either be played, meanwhile, the micro-regulatory function of selenium element can also be played.So Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides prepared by the present invention can have broad application prospects in food antimicrobial additive, hypoglycemic drug additive, Organism immunoregulation healthcare products etc.
Description
Technical field
The present invention relates to a kind of process for refining of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides, belong to microbial fermentation engineering technical field.
Background technology
As one of main component that living matter is formed, for the research of polysaccharide in organism carried out very deep, the function of polysaccharide molecule is mainly concentrated both ways, on the one hand, polysaccharide molecule can carry out the transmission of internal body bioinformation as informational molecule, therefore, this kind of polysaccharide molecule have that kind is many, content is few, complex structure, the feature that contains much information.On the other hand, polysaccharide molecule can as the main composition composition of cell and extracellular matrix, and this saccharoidal kind is less, but the content of often kind of polysaccharide molecule is more, and its tactical rule is complicated, and granular size has macroscopical unhomogeneity.Meanwhile, polysaccharide is also a class natural high moleculer eompound, and it is the polymer linked together by glycosidic link by aldose or ketose, is the requisite compositions of all lived organisms, closely related with the function maintaining organism.Prove through nearly two large quantity research during the last ten years, polysaccharide has biologic activity widely, as antitumor, immunomodulatory, anti-infective, antiviral, anticoagulation, hypoglycemic, radioprotective, reducing blood-fat, promotion nucleic acid and protein biosynthesizing, control cell fission and differentiation, adjustment cell growth with old and feeble etc., and polysaccharide is minimum as the toxicity of medicinal application process, thus, the activity research of polysaccharide has caused the great interest of people.
At present, the research object of active polysaccharide mainly comprises plant, animal class, Mycophyta, bacterium class, lichens, algae, pollen class polysaccharide etc., research method relates to various chemical process and instrument analytical method, research range relates to the separation and purification of polysaccharide, physico-chemical property, structural analysis, immunology, toxicology, pharmacodynamics and clinical medicine treatment use etc., can be described as polysaccharide material Quality Research reached of a great variety, research deeply, widely used degree.Carry out the research of the aspect such as analysis, polysaccharide pharmacological action of the exploitation of polysaccharide resource, polysaccharide molecule structure, for enrich further polysaccharide material species resource, explore polysaccharide molecule biologic activity all tool be of great significance, thus, the research of polysaccharide has become one of hot research content in the field such as molecular biology, pharmaceutics.Hericium is in Basidiomycota, hedgehog hydnum Cordycepps, and be famous medicine-food two-purpose fungi, it is nutritious, has the laudatory title of " mountain delicacy hedgehog hydnum, seafood delights bird's nest ".It is generally acknowledged, the main active substances of Hericium erinaceus (Bull. Ex Fr.) Pers. is polysaccharide, polypeptide and aliphatics acyl class material, and wherein Hericium erinaceus polysaccharide is the focus of research always.Hericium erinaceus polysaccharide belongs to the one of food and medicament dual-purpose fungus polysaccharide; pharmacological research proves; Hericium erinaceus Polysaccharides can strengthen peritoneal macrophage phagocytic function, improve body's hypoxia tolerance, improve body immunity, improve painstaking effort work outpuies, inhibition tumor cell growth, anti-ageing, anti-mutation, anti-oxidant; there is provide protection to radiation injury, also have better protecting effect to stomach mucous membrane.Thus, carry out careful research for Hericium erinaceus polysaccharide and there is very great meaning.
In addition, about the exploitation of rich selenium product, be also a focus direction of scientific rersearch at present in food, medicine, healthcare products.Selenium element is the active centre of the multiple enzyme of human body, and it is also one of various trace elements of needed by human.Multinomial result of study shows, the amino-acid compound of selenium can play in the vital movement of human body resists disease, to effect that is anti-ageing, that strengthen immune function of human body.Such as, selenium element is the active centre of Selenoperoxidase family and thioredoxin reductase family, after being combined with selenium element by Selenoperoxidase family and thioredoxin reductase family, having stronger scavenging activated oxygen, prevent the function of lipid peroxdation, is the important component part of body antioxidant system.In addition; selenium element has certain supporting role for the cellular immunization of body and the maintenance of humoral immunization ability; the effect that selenium element has protection thymus gland, maintains the lymphocytic activity of body and stimulate antibody to produce, thus achieve the function of enhancing body immunological competence.
At present, studying more is the biological activity of hericium erinaceum polysaccharide, and the correlative study for Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium polysaccharides is less.Therefore, in the mycelial process of growth of Hericium erinaceus (Bull. Ex Fr.) Pers., add selenium element, after extracting mycelium polysaccharides, the biological activity of polysaccharose substance can either be played, also can play the regulatory function of the trace element of selenium simultaneously.So Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides prepared by the present invention can have broad application prospects in food, medicine, healthcare products.
Summary of the invention
The object of the present invention is to provide a kind of process for refining of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides.
The process for refining of above-mentioned Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides, comprises the steps:
(1) activation of Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification: be numbered 5.0058 Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classifications in China General Microbiological culture presevation administrative center by buying, PDA solid test-tube culture medium is utilized to activate, culture condition is quiescent culture 96 hours-144 hours in 25 DEG C of-28 DEG C of incubators, obtains the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after activation;
(2) the liquid primary fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.: by the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after the activation in step (1), transfer under aseptic condition in liquid primary fermentation substratum, every 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation substratum contains glucose 20g, peptone 0.3g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o1.5g, Sodium Selenite 0.125g, adding distil water, to 1L, uses after packing sterilizing, carries out liquid primary fermentation and cultivate 15 days-20 days after switching in 25 DEG C of-28 DEG C of incubators, obtains Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product;
(3) the liquid large scale fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.; The Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product that step (2) is obtained, aseptically picking one ferfas silk sheet, transfer and be equipped with in the 500mL triangular flask of 50mL Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum in one, 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum contains glucose 25g, peptone 0.4g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o1.5g, Sodium Selenite 0.25g, adding distil water, to 1L, uses after packing sterilizing, after switching in 25 DEG C of-28 DEG C of incubators quiescent culture 15 days-20 days, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product;
(4) the mycelial collection of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product that step (3) is obtained, 8 layers of gauze are aseptically utilized to carry out collecting by filtration mycelium, recycling distilled water cleaning 3-5 time, and unwatering as much as possible, then, Hericium erinaceus (Bull. Ex Fr.) Pers. mycelia sheet is placed in 35 DEG C-40 DEG C desiccation culture casees and carries out drying treatment 24 hours-48 hours, obtain the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after drying treatment;
(5) the mycelial pulverizing of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after the drying treatment obtain step (4), utilizes clean mortar to carry out milled processed, be ground to become thinner Powdered till, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder;
(6) extraction of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides: take the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder 5g that step (5) obtains, the distilled water of 150mL-250mL is added in the glass beaker of 500mL, 50 DEG C of-70 DEG C of extracting at constant temperature 3h-5h are carried out in thermostat water bath, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out Crude polysaccharides, then, 10000rpm collected by centrifugation Crude polysaccharides, finally be deposited in the drying treatment of carrying out Crude polysaccharides in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides;
(7) Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides is refining: Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides 2g step (6) obtained, phenol chloroform-primary isoamyl alcohol mixed solution the 20mL of the proportions that utilization is 25:24:1 according to volume ratio, carry out soaking at room temperature process 3h-5h, afterwards mixed solution is carried out 10000rpm centrifugal, collect supernatant liquid, the dialysis tubing utilizing molecular retention ability to be 3500KDa is to distill water dialysis 24h, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out polysaccharide, then, 10000rpm collected by centrifugation polysaccharide, finally be deposited in the drying treatment of carrying out polysaccharide in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide.
Advantage of the present invention:
The present invention relates to a kind of process for refining of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides.The Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide prepared by the method both can give full play to the immunomodulatory of Hericium erinaceus (Bull. Ex Fr.) Pers. mycelia polysaccharide and anti-oxidant, hypoglycemic biological action, the micro-regulatory function of selenium element can be played again, therefore, it can become the main component of healthcare products trace mineral supplement.Meanwhile, the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide of acquisition, also has good using value in the exploitation of hypoglycemic medicine.
Embodiment
Below in conjunction with specific embodiment, further detailed description is done to the present invention.
The low-temperature purifying method technique of embodiment 1 one kinds of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides.
(1) activation of Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification: be numbered 5.0058 Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classifications in China General Microbiological culture presevation administrative center by buying, PDA solid test-tube culture medium is utilized to activate, culture condition is quiescent culture 96 hours-144 hours in 25 DEG C of-28 DEG C of incubators, obtains the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after activation;
(2) the liquid primary fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.: by the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after the activation in step (1), transfer under aseptic condition in liquid primary fermentation substratum, every 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation substratum contains glucose 20g, peptone 0.3g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o1.5g, Sodium Selenite 0.125g, adding distil water, to 1L, uses after packing sterilizing, carries out liquid primary fermentation and cultivate 15 days-20 days after switching in 25 DEG C of-28 DEG C of incubators, obtains Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product;
(3) the liquid large scale fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.; The Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product that step (2) is obtained, aseptically picking one ferfas silk sheet, transfer and be equipped with in the 500mL triangular flask of 50mL Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum in one, 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum contains glucose 25g, peptone 0.4g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o1.5g, Sodium Selenite 0.25g, adding distil water, to 1L, uses after packing sterilizing, after switching in 25 DEG C of-28 DEG C of incubators quiescent culture 15 days-20 days, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product;
(4) the mycelial collection of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product that step (3) is obtained, 8 layers of gauze are aseptically utilized to carry out collecting by filtration mycelium, recycling distilled water cleaning 3-5 time, and unwatering as much as possible, then, Hericium erinaceus (Bull. Ex Fr.) Pers. mycelia sheet is placed in 35 DEG C-40 DEG C desiccation culture casees and carries out drying treatment 24 hours-48 hours, obtain the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after drying treatment;
(5) the mycelial pulverizing of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after the drying treatment obtain step (4), utilizes clean mortar to carry out milled processed, be ground to become thinner Powdered till, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder;
(6) extraction of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides: take the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder 5g that step (5) obtains, the distilled water of 150mL is added in the glass beaker of 500mL, 50 DEG C of extracting at constant temperature 4h are carried out in thermostat water bath, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out Crude polysaccharides, then, 10000rpm collected by centrifugation Crude polysaccharides, finally be deposited in the drying treatment of carrying out Crude polysaccharides in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides;
(7) Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides is refining: Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides 2g step (6) obtained, be the phenol chloroform-primary isoamyl alcohol mixed solution 20mL of the proportions of 25:24:1 according to volume ratio, carry out soaking at room temperature process 3h-5h, afterwards mixed solution is carried out 10000rpm centrifugal, collect supernatant liquid, the dialysis tubing utilizing molecular retention ability to be 3500KDa is to distill water dialysis 24h, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out polysaccharide, then, 10000rpm collected by centrifugation polysaccharide, finally be deposited in the drying treatment of carrying out polysaccharide in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide.
Under the treatment condition of this extraction process, the yield of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide is 8.32%.
The middle temperature process for refining of embodiment 2 one kinds of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides.
(1) activation of Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification: be numbered 5.0058 Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classifications in China General Microbiological culture presevation administrative center by buying, PDA solid test-tube culture medium is utilized to activate, culture condition is quiescent culture 96 hours-144 hours in 25 DEG C of-28 DEG C of incubators, obtains the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after activation;
(2) the liquid primary fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.: by the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after the activation in step (1), transfer under aseptic condition in liquid primary fermentation substratum, every 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation substratum contains glucose 20g, peptone 0.3g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o1.5g, Sodium Selenite 0.125g, adding distil water, to 1L, uses after packing sterilizing, carries out liquid primary fermentation and cultivate 15 days-20 days after switching in 25 DEG C of-28 DEG C of incubators, obtains Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product;
(3) the liquid large scale fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.; The Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product that step (2) is obtained, aseptically picking one ferfas silk sheet, transfer and be equipped with in the 500mL triangular flask of 50mL Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum in one, 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum contains glucose 25g, peptone 0.4g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o1.5g, Sodium Selenite 0.25g, adding distil water, to 1L, uses after packing sterilizing, after switching in 25 DEG C of-28 DEG C of incubators quiescent culture 15 days-20 days, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product;
(4) the mycelial collection of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product that step (3) is obtained, 8 layers of gauze are aseptically utilized to carry out collecting by filtration mycelium, recycling distilled water cleaning 3-5 time, and unwatering as much as possible, then, Hericium erinaceus (Bull. Ex Fr.) Pers. mycelia sheet is placed in 35 DEG C-40 DEG C desiccation culture casees and carries out drying treatment 24 hours-48 hours, obtain the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after drying treatment;
(5) the mycelial pulverizing of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after the drying treatment obtain step (4), utilizes clean mortar to carry out milled processed, be ground to become thinner Powdered till, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder;
(6) extraction of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides: take the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder 5g that step (5) obtains, the distilled water of 150mL-250mL is added in the glass beaker of 500mL, 60 DEG C of extracting at constant temperature 4h are carried out in thermostat water bath, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out Crude polysaccharides, then, 10000rpm collected by centrifugation Crude polysaccharides, finally be deposited in the drying treatment of carrying out Crude polysaccharides in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides;
(7) Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides is refining: Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides 2g step (6) obtained, phenol chloroform-primary isoamyl alcohol mixed solution the 20mL of the proportions that utilization is 25:24:1 according to volume ratio, carry out soaking at room temperature process 3h-5h, afterwards mixed solution is carried out 10000rpm centrifugal, collect supernatant liquid, the dialysis tubing utilizing molecular retention ability to be 3500KDa is to distill water dialysis 24h, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out polysaccharide, then, 10000rpm collected by centrifugation polysaccharide, finally be deposited in the drying treatment of carrying out polysaccharide in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide.
Under the treatment condition of this extraction process, the yield of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide is 9.36%.
The high-temperature refined craft of embodiment 3 one kinds of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides.。
(1) activation of Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification: be numbered 5.0058 Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classifications in China General Microbiological culture presevation administrative center by buying, PDA solid test-tube culture medium is utilized to activate, culture condition is quiescent culture 96 hours-144 hours in 25 DEG C of-28 DEG C of incubators, obtains the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after activation;
(2) the liquid primary fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.: by the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after the activation in step (1), transfer under aseptic condition in liquid primary fermentation substratum, every 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation substratum contains glucose 20g, peptone 0.3g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o1.5g, Sodium Selenite 0.125g, adding distil water, to 1L, uses after packing sterilizing, carries out liquid primary fermentation and cultivate 15 days-20 days after switching in 25 DEG C of-28 DEG C of incubators, obtains Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product;
(3) the liquid large scale fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.; The Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product that step (2) is obtained, aseptically picking one ferfas silk sheet, transfer and be equipped with in the 500mL triangular flask of 50mL Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum in one, 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum contains glucose 25g, peptone 0.4g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o1.5g, Sodium Selenite 0.25g, adding distil water, to 1L, uses after packing sterilizing, after switching in 25 DEG C of-28 DEG C of incubators quiescent culture 15 days-20 days, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product;
(4) the mycelial collection of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product that step (3) is obtained, 8 layers of gauze are aseptically utilized to carry out collecting by filtration mycelium, recycling distilled water cleaning 3-5 time, and unwatering as much as possible, then, Hericium erinaceus (Bull. Ex Fr.) Pers. mycelia sheet is placed in 35 DEG C-40 DEG C desiccation culture casees and carries out drying treatment 24 hours-48 hours, obtain the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after drying treatment;
(5) the mycelial pulverizing of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after the drying treatment obtain step (4), utilizes clean mortar to carry out milled processed, be ground to become thinner Powdered till, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder;
(6) extraction of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides: take the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder 5g that step (5) obtains, the distilled water of 150mL-250mL is added in the glass beaker of 500mL, 70 DEG C of extracting at constant temperature 3h-5h are carried out in thermostat water bath, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out Crude polysaccharides, then, 10000rpm collected by centrifugation Crude polysaccharides, finally be deposited in the drying treatment of carrying out Crude polysaccharides in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides;
(7) Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides is refining: Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides 2g step (6) obtained, phenol chloroform-primary isoamyl alcohol mixed solution the 20mL of the proportions that utilization is 25:24:1 according to volume ratio, carry out soaking at room temperature process 3h-5h, afterwards mixed solution is carried out 10000rpm centrifugal, collect supernatant liquid, the dialysis tubing utilizing molecular retention ability to be 3500KDa is to distill water dialysis 24h, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out polysaccharide, then, 10000rpm collected by centrifugation polysaccharide, finally be deposited in the drying treatment of carrying out polysaccharide in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide.
Under the treatment condition of this extraction process, the yield of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide is 11.84%.
Claims (1)
1. a process for refining for Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides, is characterized in that, comprises the following steps:
(1) activation of Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification: be numbered 5.0058 Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classifications in China General Microbiological culture presevation administrative center by buying, PDA solid test-tube culture medium is utilized to activate, culture condition is quiescent culture 96 hours-144 hours in 25 DEG C of-28 DEG C of incubators, obtains the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after activation;
(2) the liquid primary fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.: by the Hericium erinaceus (Bull. Ex Fr.) Pers. bacterial classification after the activation in step (1), transfer under aseptic condition in liquid primary fermentation substratum, 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation substratum contains glucose 20g, peptone 0.3g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o 1.5g, Sodium Selenite 0.125g, adding distil water, to 1L, uses after packing sterilizing, carries out liquid primary fermentation and cultivate 15 days-20 days after switching in 25 DEG C of-28 DEG C of incubators, obtains Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product;
(3) the liquid large scale fermentation of Hericium erinaceus (Bull. Ex Fr.) Pers.; The Hericium erinaceus (Bull. Ex Fr.) Pers. liquid primary fermentation product that step (2) is obtained, aseptically picking one ferfas silk sheet, transfer and be equipped with in the 500mL triangular flask of 50mL Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum in one, 1L Hericium erinaceus (Bull. Ex Fr.) Pers. liquid large scale fermentation substratum contains glucose 25g, peptone 0.4g, yeast extract paste 0.6g, KH
2pO
42g, MgSO
47H
2o 1.5g, Sodium Selenite 0.25g, adding distil water, to 1L, uses after packing sterilizing, after switching in 25 DEG C of-28 DEG C of incubators quiescent culture 15 days-20 days, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product;
(4) the mycelial collection of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium liquid large scale fermentation product that step (3) is obtained, gauze is aseptically utilized to carry out collecting by filtration mycelium, recycling distilled water cleaning 3-5 time, and unwatering as much as possible, then, Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium is placed in 35 DEG C-40 DEG C desiccation culture casees and carries out drying treatment 24 hours-48 hours, obtain the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after drying treatment;
(5) the mycelial pulverizing of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium: the Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium after the drying treatment obtain step (4), utilizes clean mortar to carry out milled processed, be ground to become thinner Powdered till, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder;
(6) extraction of Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides: take the Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium powder 5g that step (5) obtains, the distilled water of 150mL-250mL is added in the glass beaker of 500mL, 50 DEG C of-70 DEG C of extracting at constant temperature 3h-5h are carried out in thermostat water bath, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out Crude polysaccharides, then, 10000rpm collected by centrifugation Crude polysaccharides, finally be deposited in the drying treatment of carrying out Crude polysaccharides in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides;
(7) Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium polysaccharides is refining: Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium Crude polysaccharides 2g step (6) obtained, be the phenol chloroform-primary isoamyl alcohol mixed solution 20mL of the proportions of 25:24:1 according to volume ratio, carry out soaking at room temperature process 3h-5h, afterwards mixed solution is carried out 10000rpm centrifugal, collect supernatant liquid, the dialysis tubing utilizing molecular retention ability to be 3500KDa is to distill water dialysis 24h, afterwards, the ratio being 1:1 according to volume ratio adds the precipitation that dehydrated alcohol carries out polysaccharide, then, 10000rpm collected by centrifugation polysaccharide, finally be deposited in the drying treatment of carrying out polysaccharide in the constant incubator of 35 DEG C-40 DEG C, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. Rich in selenium mycelium refined polysaccharide.
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| CN108753625B (en) * | 2018-05-24 | 2021-11-30 | 富乐顿生物工程科技（北京）有限公司 | Polysaccharide-producing space Hericium coralloides ST21-3 and application thereof in improving biological immune activity |
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| CN102643359A (en) * | 2012-04-10 | 2012-08-22 | 上海市农业科学院 | Hericium erinaceus polysaccharide and preparation method thereof |
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| CN102643359A (en) * | 2012-04-10 | 2012-08-22 | 上海市农业科学院 | Hericium erinaceus polysaccharide and preparation method thereof |
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