CN109651524A - A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation - Google Patents
A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation Download PDFInfo
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- 238000000034 method Methods 0.000 title claims abstract description 38
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- 238000000855 fermentation Methods 0.000 title claims abstract description 27
- 230000004151 fermentation Effects 0.000 title claims abstract description 27
- 150000004676 glycans Chemical class 0.000 claims abstract description 54
- 229920001282 polysaccharide Polymers 0.000 claims abstract description 54
- 239000005017 polysaccharide Substances 0.000 claims abstract description 54
- 239000000284 extract Substances 0.000 claims abstract description 38
- 241001264174 Cordyceps militaris Species 0.000 claims abstract description 18
- 239000012141 concentrate Substances 0.000 claims abstract description 18
- 235000009508 confectionery Nutrition 0.000 claims abstract description 16
- 235000013399 edible fruits Nutrition 0.000 claims abstract description 15
- 238000012545 processing Methods 0.000 claims abstract description 14
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- 239000012153 distilled water Substances 0.000 claims description 45
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 44
- 239000000243 solution Substances 0.000 claims description 35
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- 241000209140 Triticum Species 0.000 claims description 11
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- 229940041514 candida albicans extract Drugs 0.000 claims description 10
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- 239000003153 chemical reaction reagent Substances 0.000 claims description 9
- BFSVOASYOCHEOV-UHFFFAOYSA-N 2-diethylaminoethanol Chemical compound CCN(CC)CCO BFSVOASYOCHEOV-UHFFFAOYSA-N 0.000 claims description 8
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- 238000001914 filtration Methods 0.000 claims description 7
- MYMOFIZGZYHOMD-UHFFFAOYSA-N Dioxygen Chemical compound O=O MYMOFIZGZYHOMD-UHFFFAOYSA-N 0.000 claims description 5
- 229910001882 dioxygen Inorganic materials 0.000 claims description 5
- 238000010992 reflux Methods 0.000 claims description 5
- 244000283482 Alloteropsis cimicina Species 0.000 claims 1
- 235000017898 Digitaria ciliaris Nutrition 0.000 claims 1
- 238000005342 ion exchange Methods 0.000 claims 1
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- 241001248610 Ophiocordyceps sinensis Species 0.000 description 8
- 150000002500 ions Chemical class 0.000 description 7
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- 238000011161 development Methods 0.000 description 5
- 102000004190 Enzymes Human genes 0.000 description 4
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- 241000190633 Cordyceps Species 0.000 description 3
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- OFEZSBMBBKLLBJ-UHFFFAOYSA-N cordycepine Natural products C1=NC=2C(N)=NC=NC=2N1C1OC(CO)CC1O OFEZSBMBBKLLBJ-UHFFFAOYSA-N 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 238000010998 test method Methods 0.000 description 2
- 239000002126 C01EB10 - Adenosine Substances 0.000 description 1
- 241001480006 Clavicipitaceae Species 0.000 description 1
- 206010011224 Cough Diseases 0.000 description 1
- SHZGCJCMOBCMKK-UHFFFAOYSA-N D-mannomethylose Natural products CC1OC(O)C(O)C(O)C1O SHZGCJCMOBCMKK-UHFFFAOYSA-N 0.000 description 1
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 description 1
- 208000000059 Dyspnea Diseases 0.000 description 1
- 206010013975 Dyspnoeas Diseases 0.000 description 1
- PNNNRSAQSRJVSB-SLPGGIOYSA-N Fucose Natural products C[C@H](O)[C@@H](O)[C@H](O)[C@H](O)C=O PNNNRSAQSRJVSB-SLPGGIOYSA-N 0.000 description 1
- 241000221775 Hypocreales Species 0.000 description 1
- SHZGCJCMOBCMKK-DHVFOXMCSA-N L-fucopyranose Chemical compound C[C@@H]1OC(O)[C@@H](O)[C@H](O)[C@@H]1O SHZGCJCMOBCMKK-DHVFOXMCSA-N 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 206010059013 Nocturnal emission Diseases 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- 240000006698 Spigelia anthelmia Species 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 229960005305 adenosine Drugs 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- PYMYPHUHKUWMLA-WDCZJNDASA-N arabinose Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)C=O PYMYPHUHKUWMLA-WDCZJNDASA-N 0.000 description 1
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 1
- SRBFZHDQGSBBOR-UHFFFAOYSA-N beta-D-Pyranose-Lyxose Natural products OC1COC(O)C(O)C1O SRBFZHDQGSBBOR-UHFFFAOYSA-N 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
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- 238000007796 conventional method Methods 0.000 description 1
- 239000006071 cream Substances 0.000 description 1
- SEGLCEQVOFDUPX-UHFFFAOYSA-N di-(2-ethylhexyl)phosphoric acid Chemical compound CCCCC(CC)COP(O)(=O)OCC(CC)CCCC SEGLCEQVOFDUPX-UHFFFAOYSA-N 0.000 description 1
- 238000004821 distillation Methods 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- -1 galactolipin Chemical compound 0.000 description 1
- 235000008216 herbs Nutrition 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 125000004435 hydrogen atom Chemical class [H]* 0.000 description 1
- 201000001881 impotence Diseases 0.000 description 1
- 238000009776 industrial production Methods 0.000 description 1
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- 239000011591 potassium Substances 0.000 description 1
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/0003—General processes for their isolation or fractionation, e.g. purification or extraction from biomass
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- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- General Health & Medical Sciences (AREA)
- Sustainable Development (AREA)
- Biochemistry (AREA)
- Materials Engineering (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Medicinal Chemistry (AREA)
- Polymers & Plastics (AREA)
- Organic Chemistry (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation, includes the following steps: A, Coragyceps militaris bacterium culture is obtained seed liquor；B, seed liquor solid fermentation obtains cordyceps militaris fruit body；C, cordyceps militaris fruit body crushes to obtain northern Chinese caterpillar Fungus mixed liquor；D, northern Chinese caterpillar Fungus mixed liquor extracts to obtain polysaccharide concentrate；E, polysaccharide concentrate centrifugal treating obtains northern Chinese caterpillar Fungus polyoses extract；F, northern Chinese caterpillar Fungus polyoses extract decolourizes to obtain northern Chinese caterpillar Fungus polyoses extract colourless solution；G, northern Chinese caterpillar Fungus polyoses extract colourless solution removes deproteinized, obtains northern Chinese caterpillar Fungus Thick many candies；H, northern Chinese caterpillar Fungus Thick many candies are further purified to obtain northern Chinese caterpillar Fungus polysaccharide.Extraction and purification process of the invention realizes culture, extraction, purifying continuous processing, and extraction efficiency is high, and polysaccharide yield is high, operates simpler.
Description
Technical field
The present invention relates to microcultures and technical field of natural product extraction, and in particular to a kind of based on solid fermentation
Northern Chinese caterpillar Fungus Polyose extraction purifying process.
Background technique
Northern Chinese caterpillar Fungus also known as Cordyceps militaris, Cordceps militaris belong to Ascomycotina, gang pyrenomycetes, Sphaerials, Clavicipitaceae, cordyceps sinensis
Belong to fungi.It is the type sepecies of Cordyceps, widely distributed, is recognized by countries in the world scholar and is received and educate under artificial condition
There is the effective components such as Cordyceps sinensis polysaccharide and the cordycepin of higher amount, unique pharmacology at complete stroma and northern Chinese caterpillar Fungus
Effect has increasingly caused the great attention of pharmacy circle.Since northern Chinese caterpillar Fungus has the advantages that above, become medicinal worm in Cordyceps
The outstanding person of careless bacterium.The success of the artificial culture technique of northern Chinese caterpillar Fungus opens rare medicinal herbs industrialization production again for China
A piece of sky.Currently, the artificial culture of domestic northern Chinese caterpillar Fungus has entered the industrialization stage.
Cordyceps sinensis polysaccharide or polysaccharide, one of the main active as northern Chinese caterpillar Fungus are contents in northern Chinese caterpillar Fungus
Highest pharmacological active substance, content can achieve 10%, be a kind of of great value potential novel dual-purpose of drug and food resources.Cordyceps sinensis
The polysaccharide that polysaccharide is made of mannose, cordycepin, adenosine, galactolipin, arabinose, glucose, fucose etc..It resists
Cancer activity and immunocompetence cause the great interest of people, in addition cordyceps sinensis is also used to control pulmonary tuberculosis, shortness of breath is breathed with cough, impotence,
Nocturnal emission, spontaneous sweating, soreness of waist and knee joint etc..
Cordyceps sinensis polysaccharide is mainly extracted from fruiting bodies of cordyceps militaris in the industrial production at present.But due to cordyceps militaris sporocarp
High production cost, it is high so as to cause the production cost of Cordyceps sinensis polysaccharide.
Mainly there are Hot water extraction, acidleach formulation, alkali method, ultrasonic wave extraction with the extracting method of Cordyceps sinensis polysaccharide at present
Method, microwave loss mechanisms etc..
In addition, being obtained using artificial fermentation's aweto and natural cordyceps chemical composition and pharmacology work
With essentially identical mycelium, become a kind of most viable method for solving worm grass resources shortage instantly.
But existing northern Chinese caterpillar Fungus polysaccharide extracting process, it can not achieve culture extraction purification continuous processing, extract effect
Rate is low, and northern Chinese caterpillar Fungus polysaccharide yield is low, and operating process is complicated, higher cost, and cannot further isolate and purify polysaccharide, nothing
Method obtains the higher polysaccharide component of purity, and the development and utilization of northern Chinese caterpillar Fungus polyose is made to receive limitation.
Based on the above situation, the invention proposes a kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation
Problem above can effectively be solved.
Summary of the invention
The purpose of the present invention is to provide a kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation.This hair
The bright northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation realizes culture, extraction, purifying continuous processing, mentions
Take high-efficient, northern Chinese caterpillar Fungus polysaccharide yield is high, and operating process is simpler, and cost is relatively low, and can further separate polysaccharide pure
Change, obtains the higher polysaccharide component of purity, be conducive to the development and utilization of northern Chinese caterpillar Fungus polyose.
The present invention is achieved through the following technical solutions:
A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation, includes the following steps:
A, Coragyceps militaris bacterium is inoculated on northern Chinese caterpillar Fungus seed culture medium, puts shaking table culture case into, cultivated
2.5-4d, the composition of the northern Chinese caterpillar Fungus seed culture medium include the raw material of following parts by weight: 20-30 parts of sucrose, yeast extract
1.5-3 parts, 0.5-0.7 parts of magnesium sulfate of 0.8-1.1 parts of potassium dihydrogen phosphate, 1000 parts of distilled water；
B, the seed liquor for obtaining step A is inoculated on the northern Chinese caterpillar Fungus solid medium of sterilized processing, inoculum concentration
It for 8-11%, puts into constant temperature and humidity incubator, under the conditions of 28.5 DEG C of temperature, ferments 5-8 days, it is real to obtain northern Chinese caterpillar Fungus
Body；The northern Chinese caterpillar Fungus solid medium includes the raw material of following parts by weight: 1000 parts of wheat flour, 190-215 parts of mealy potato,
25-35 parts of glucose sugar, 2.1-3.5 parts of potassium dihydrogen phosphate, 2.5-3.6 parts of peptone, 0.9-1.3 parts of magnesium sulfate, distilled water 450-
600 parts；
C, the obtained cordyceps militaris fruit body of step B is put into wall-breaking machine, the distilled water of 10-12 times of quality is added,
5-15min is crushed, northern Chinese caterpillar Fungus mixed liquor is obtained；
D, the northern Chinese caterpillar Fungus mixed liquor for obtaining step C, ultrasonication 40-50min flow back at 70-80 DEG C and mention
3-4.5h is taken, is concentrated after extracting solution filtering, obtains polysaccharide concentrate；
E, the dehydrated alcohol of 2.8-3.4 times of volume is added in the polysaccharide concentrate that step D is obtained, is cooled to 4 after stirring
DEG C, 18-36 hours are stood, carries out centrifugal treating, and precipitating drying is obtained into northern Chinese caterpillar Fungus polyoses extract；
F, the northern Chinese caterpillar Fungus polyoses extract that step E is obtained adds distilled water to dissolve, and adjusts pH value to 8-9, dioxygen is added dropwise
For aqueous solution to colourless, 50-60 DEG C of heat preservation 2-3h obtains northern Chinese caterpillar Fungus polyoses extract colourless solution；
G, northern Chinese caterpillar Fungus polyoses extract colourless solution that step F is obtained is mixed with papain and amylase molten
Liquid mixing, the volume ratio of the two are 1:(0.45-0.6), 60-65 DEG C of enzymatic hydrolysis 1-2h；20% volume is added in enzymolysis liquid
Sevage reagent, shaken cultivation 0.5-1h, then repeatedly centrifugation takes supernatant until no albumen precipitation is precipitated, and dries, obtains
To northern Chinese caterpillar Fungus Thick many candies；
H, upper DEAE Ago-Gel FF ion is handed over after dissolving the northern Chinese caterpillar Fungus Thick many candies that step G is obtained with distilled water
Chromatographic column is changed, gradient elution is carried out with sodium chloride solution, obtains northern Chinese caterpillar Fungus polysaccharide.
Northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation of the invention, realizes culture, extraction, purifying
Continuous processing, extraction efficiency are high, and northern Chinese caterpillar Fungus polysaccharide yield is high, and operating process is simpler, and cost is relatively low, and can by polysaccharide into
One step isolates and purifies, and obtains the higher polysaccharide component of purity, is conducive to the development and utilization of northern Chinese caterpillar Fungus polyose.
Preferably, the composition of northern Chinese caterpillar Fungus seed culture medium described in step B includes the raw material of following parts by weight: sucrose
25 parts, 2.5 parts of yeast extract, 0.9 part of potassium dihydrogen phosphate, 0.6 part of magnesium sulfate, 1000 parts of distilled water.
Preferably, northern Chinese caterpillar Fungus solid medium described in step B includes the raw material of following parts by weight: wheat flour 1000
Part, 205 parts of mealy potato, glucose sugared 30 parts, 2.8 parts of potassium dihydrogen phosphate, 3.2 parts of peptone, 1.1 parts of magnesium sulfate, distilled water 520
Part.
Preferably, the condition of culture control of shaking table culture case described in step B is the revolving speed of 110-125r/min, 28 DEG C
Environment temperature.
Preferably, concentration described in step D is concentrated at 60 DEG C.
Preferably, hydrogen peroxide solution described in step F, volumetric concentration 25%.
Preferably, in papain described in step G and amylase mixed solution papain and amylase matter
Amount is than being 0.8:(1.2-1.4).
Preferably, centrifugation described in step G is 6000r/min centrifugation 10min.
Compared with prior art, the present invention have the following advantages that and the utility model has the advantages that
1, the northern Chinese caterpillar Fungus Polyose extraction purifying process of the invention based on solid fermentation realizes culture, extraction, pure
Change continuous processing, extraction efficiency is high, and northern Chinese caterpillar Fungus polysaccharide yield is high, and operating process is simpler, and cost is relatively low, and can be by polysaccharide
It further isolates and purifies, obtains the higher polysaccharide component of purity, be conducive to the development and utilization of northern Chinese caterpillar Fungus polyose.
Specific embodiment
The present invention is described in further detail below with reference to embodiment, embodiments of the present invention are not limited thereto.
Test method described in following embodiments or test method are unless otherwise specified conventional method；The reagent
And material is obtained from routine business approach, or prepare in conventional manner unless otherwise specified.
Embodiment 1:
A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation, includes the following steps:
A, Coragyceps militaris bacterium is inoculated on northern Chinese caterpillar Fungus seed culture medium, puts shaking table culture case into, cultivated
2.5-4d, the composition of the northern Chinese caterpillar Fungus seed culture medium include the raw material of following parts by weight: 20-30 parts of sucrose, yeast extract
1.5-3 parts, 0.5-0.7 parts of magnesium sulfate of 0.8-1.1 parts of potassium dihydrogen phosphate, 1000 parts of distilled water；
B, the seed liquor for obtaining step A is inoculated on the northern Chinese caterpillar Fungus solid medium of sterilized processing, inoculum concentration
It for 8-11%, puts into constant temperature and humidity incubator, under the conditions of 28.5 DEG C of temperature, ferments 5-8 days, it is real to obtain northern Chinese caterpillar Fungus
Body；The northern Chinese caterpillar Fungus solid medium includes the raw material of following parts by weight: 1000 parts of wheat flour, 190-215 parts of mealy potato,
25-35 parts of 20g glucose sugar, 2.1-3.5 parts of potassium dihydrogen phosphate, 2.5-3.6 parts of peptone, 0.9-1.3 parts of magnesium sulfate, distilled water
450-600 parts；
C, the obtained cordyceps militaris fruit body of step B is put into wall-breaking machine, the distilled water of 10-12 times of quality is added,
5-15min is crushed, northern Chinese caterpillar Fungus mixed liquor is obtained；
D, the northern Chinese caterpillar Fungus mixed liquor for obtaining step C, ultrasonication 40-50min flow back at 70-80 DEG C and mention
3-4.5h is taken, is concentrated after extracting solution filtering, obtains polysaccharide concentrate；
E, the dehydrated alcohol of 2.8-3.4 times of volume is added in the polysaccharide concentrate that step D is obtained, is cooled to 4 after stirring
DEG C, 18-36 hours are stood, carries out centrifugal treating, and precipitating drying is obtained into northern Chinese caterpillar Fungus polyoses extract；
F, the northern Chinese caterpillar Fungus polyoses extract that step E is obtained adds distilled water to dissolve, and adjusts pH value to 8-9, dioxygen is added dropwise
For aqueous solution to colourless, 50-60 DEG C of heat preservation 2-3h obtains northern Chinese caterpillar Fungus polyoses extract colourless solution；
G, northern Chinese caterpillar Fungus polyoses extract colourless solution that step F is obtained is mixed with papain and amylase molten
Liquid mixing, the volume ratio of the two are 1:(0.45-0.6), 60-65 DEG C of enzymatic hydrolysis 1-2h；20% volume is added in enzymolysis liquid
Sevage reagent, shaken cultivation 0.5-1h, then repeatedly centrifugation takes supernatant until no albumen precipitation is precipitated, and dries, obtains
To northern Chinese caterpillar Fungus Thick many candies；
H, upper DEAE Ago-Gel FF ion is handed over after dissolving the northern Chinese caterpillar Fungus Thick many candies that step G is obtained with distilled water
Chromatographic column is changed, gradient elution is carried out with sodium chloride solution, obtains northern Chinese caterpillar Fungus polysaccharide.
Embodiment 2:
A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation, includes the following steps:
A, Coragyceps militaris bacterium is inoculated on northern Chinese caterpillar Fungus seed culture medium, puts shaking table culture case into, cultivated
2.5-4d, the composition of the northern Chinese caterpillar Fungus seed culture medium include the raw material of following parts by weight: 20-30 parts of sucrose, yeast extract
1.5-3 parts, 0.5-0.7 parts of magnesium sulfate of 0.8-1.1 parts of potassium dihydrogen phosphate, 1000 parts of distilled water；
B, the seed liquor for obtaining step A is inoculated on the northern Chinese caterpillar Fungus solid medium of sterilized processing, inoculum concentration
It for 8-11%, puts into constant temperature and humidity incubator, under the conditions of 28.5 DEG C of temperature, ferments 5-8 days, it is real to obtain northern Chinese caterpillar Fungus
Body；The northern Chinese caterpillar Fungus solid medium includes the raw material of following parts by weight: 1000 parts of wheat flour, 190-215 parts of mealy potato,
25-35 parts of 20g glucose sugar, 2.1-3.5 parts of potassium dihydrogen phosphate, 2.5-3.6 parts of peptone, 0.9-1.3 parts of magnesium sulfate, distilled water
450-600 parts；
C, the obtained cordyceps militaris fruit body of step B is put into wall-breaking machine, the distilled water of 10-12 times of quality is added,
5-15min is crushed, northern Chinese caterpillar Fungus mixed liquor is obtained；
D, the northern Chinese caterpillar Fungus mixed liquor for obtaining step C, ultrasonication 40-50min flow back at 70-80 DEG C and mention
3-4.5h is taken, is concentrated after extracting solution filtering, obtains polysaccharide concentrate；
E, the dehydrated alcohol of 2.8-3.4 times of volume is added in the polysaccharide concentrate that step D is obtained, is cooled to 4 after stirring
DEG C, 18-36 hours are stood, carries out centrifugal treating, and precipitating drying is obtained into northern Chinese caterpillar Fungus polyoses extract；
F, the northern Chinese caterpillar Fungus polyoses extract that step E is obtained adds distilled water to dissolve, and adjusts pH value to 8-9, dioxygen is added dropwise
For aqueous solution to colourless, 50-60 DEG C of heat preservation 2-3h obtains northern Chinese caterpillar Fungus polyoses extract colourless solution；
G, northern Chinese caterpillar Fungus polyoses extract colourless solution that step F is obtained is mixed with papain and amylase molten
Liquid mixing, the volume ratio of the two are 1:(0.45-0.6), 60-65 DEG C of enzymatic hydrolysis 1-2h；20% volume is added in enzymolysis liquid
Sevage reagent, shaken cultivation 0.5-1h, then repeatedly centrifugation takes supernatant until no albumen precipitation is precipitated, and dries, obtains
To northern Chinese caterpillar Fungus Thick many candies；
H, upper DEAE Ago-Gel FF ion is handed over after dissolving the northern Chinese caterpillar Fungus Thick many candies that step G is obtained with distilled water
Chromatographic column is changed, gradient elution is carried out with sodium chloride solution, obtains northern Chinese caterpillar Fungus polysaccharide.
The composition of northern Chinese caterpillar Fungus seed culture medium described in step B includes the raw material of following parts by weight: 25 parts of sucrose, ferment
2.5 parts of female cream, 0.9 part of potassium dihydrogen phosphate, 0.6 part of magnesium sulfate, 1000 parts of distilled water.
Northern Chinese caterpillar Fungus solid medium described in step B includes the raw material of following parts by weight: 1000 parts of wheat flour, potato
205 parts of powder, 20g glucose sugared 30 parts, 2.8 parts of potassium dihydrogen phosphate, 3.2 parts of peptone, 1.1 parts of magnesium sulfate, 520 parts of distilled water.
The condition of culture control of shaking table culture case described in step B is the revolving speed of 110-125r/min, 28 DEG C of environment temperature
Degree.
Concentration described in step D is concentrated at 60 DEG C.
Hydrogen peroxide solution described in step F, volumetric concentration 25%.
The mass ratio of papain and amylase is in papain described in step G and amylase mixed solution
0.8:(1.2-1.4).
Centrifugation described in step G is 6000r/min centrifugation 10min.
Embodiment 3:
A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation, includes the following steps:
A, Coragyceps militaris bacterium is inoculated on northern Chinese caterpillar Fungus seed culture medium, puts shaking table culture case into, cultivated
2.5d, the northern Chinese caterpillar Fungus seed culture medium composition include following parts by weight raw material: 20 parts of sucrose, 1.5 parts of yeast extract,
0.8 part of potassium dihydrogen phosphate, 0.5 part of magnesium sulfate, 1000 parts of distilled water；
B, the seed liquor for obtaining step A is inoculated on the northern Chinese caterpillar Fungus solid medium of sterilized processing, inoculum concentration
It is 11%, puts into constant temperature and humidity incubator, under the conditions of 28.5 DEG C of temperature, ferments 5 days, obtain cordyceps militaris fruit body；
The northern Chinese caterpillar Fungus solid medium includes the raw material of following parts by weight: 1000 parts of wheat flour, 190 parts of mealy potato, glucose sugar
25 parts, 2.1 parts of potassium dihydrogen phosphate, 2.5 parts of peptone, 0.9 part of magnesium sulfate, 450 parts of distilled water；
C, the obtained cordyceps militaris fruit body of step B is put into wall-breaking machine, the distilled water of 10 times of quality is added, crushed
5min obtains northern Chinese caterpillar Fungus mixed liquor；
D, the northern Chinese caterpillar Fungus mixed liquor for obtaining step C, ultrasonication 40min, the refluxing extraction 4.5h at 70 DEG C,
It is concentrated after extracting solution filtering, obtains polysaccharide concentrate；
E, the dehydrated alcohol of 2.8 times of volumes is added in the polysaccharide concentrate that step D is obtained, is cooled to 4 DEG C after stirring, it is quiet
It sets 18 hours, carries out centrifugal treating, and precipitating drying is obtained into northern Chinese caterpillar Fungus polyoses extract；
F, the northern Chinese caterpillar Fungus polyoses extract that step E is obtained adds distilled water to dissolve, and adjusts pH value to 8, hydrogen peroxide is added dropwise
For solution to colourless, 50 DEG C of heat preservation 3h obtain northern Chinese caterpillar Fungus polyoses extract colourless solution；
G, northern Chinese caterpillar Fungus polyoses extract colourless solution that step F is obtained is mixed with papain and amylase molten
Liquid mixing, the volume ratio of the two are 1:0.45,60 DEG C of enzymatic hydrolysis 2h；The Sevage reagent of 20% volume is added in enzymolysis liquid, shakes
Culture 0.5h is swung, then repeatedly centrifugation takes supernatant until no albumen precipitation is precipitated, and dries, it is thick to obtain northern Chinese caterpillar Fungus
Polysaccharide；
H, upper DEAE Ago-Gel FF ion is handed over after dissolving the northern Chinese caterpillar Fungus Thick many candies that step G is obtained with distilled water
Chromatographic column is changed, gradient elution is carried out with sodium chloride solution, obtains northern Chinese caterpillar Fungus polysaccharide.
In the present embodiment, the condition of culture control of shaking table culture case described in step B is the revolving speed of 110r/min, 28 DEG C
Environment temperature.
In the present embodiment, concentration described in step D is concentrated at 60 DEG C.
In the present embodiment, hydrogen peroxide solution described in step F, volumetric concentration 25%.
In the present embodiment, papain and starch in papain described in step G and amylase mixed solution
The mass ratio of enzyme is 0.8:1.2.
In the present embodiment, centrifugation described in step G is 6000r/min centrifugation 10min.
Embodiment 4:
A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation, includes the following steps:
A, Coragyceps militaris bacterium is inoculated on northern Chinese caterpillar Fungus seed culture medium, puts shaking table culture case into, cultivate 4d,
The composition of the northern Chinese caterpillar Fungus seed culture medium includes the raw material of following parts by weight: 30 parts of sucrose, 3 parts of yeast extract, di(2-ethylhexyl)phosphate
1.1 parts of hydrogen potassium, 0.7 part of magnesium sulfate, 1000 parts of distilled water；
B, the seed liquor for obtaining step A is inoculated on the northern Chinese caterpillar Fungus solid medium of sterilized processing, inoculum concentration
It is 8%, puts into constant temperature and humidity incubator, under the conditions of 28.5 DEG C of temperature, ferments 8 days, obtain cordyceps militaris fruit body；Institute
State the raw material that northern Chinese caterpillar Fungus solid medium includes following parts by weight: 1000 parts of wheat flour, 215 parts of mealy potato, glucose sugar 35
Part, 3.5 parts of potassium dihydrogen phosphate, 3.6 parts of peptone, 1.3 parts of magnesium sulfate, 600 parts of distilled water；
C, the obtained cordyceps militaris fruit body of step B is put into wall-breaking machine, the distilled water of 12 times of quality is added, crushed
15min obtains northern Chinese caterpillar Fungus mixed liquor；
D, the northern Chinese caterpillar Fungus mixed liquor for obtaining step C, ultrasonication 50min, refluxing extraction 3h, mentions at 80 DEG C
It is concentrated after taking liquid to filter, obtains polysaccharide concentrate；
E, the dehydrated alcohol of 3.4 times of volumes is added in the polysaccharide concentrate that step D is obtained, is cooled to 4 DEG C after stirring, it is quiet
It sets 36 hours, carries out centrifugal treating, and precipitating drying is obtained into northern Chinese caterpillar Fungus polyoses extract；
F, the northern Chinese caterpillar Fungus polyoses extract that step E is obtained adds distilled water to dissolve, and adjusts pH value to 9, hydrogen peroxide is added dropwise
For solution to colourless, 60 DEG C of heat preservation 2h obtain northern Chinese caterpillar Fungus polyoses extract colourless solution；
G, northern Chinese caterpillar Fungus polyoses extract colourless solution that step F is obtained is mixed with papain and amylase molten
Liquid mixing, the volume ratio of the two are 1:0.6,65 DEG C of enzymatic hydrolysis 1h；The Sevage reagent of 20% volume is added in enzymolysis liquid, vibrates
1h is cultivated, then repeatedly centrifugation takes supernatant until no albumen precipitation is precipitated, and dries, obtains northern Chinese caterpillar Fungus Thick many candies；
H, upper DEAE Ago-Gel FF ion is handed over after dissolving the northern Chinese caterpillar Fungus Thick many candies that step G is obtained with distilled water
Chromatographic column is changed, gradient elution is carried out with sodium chloride solution, obtains northern Chinese caterpillar Fungus polysaccharide.
In the present embodiment, the condition of culture control of shaking table culture case described in step B is the revolving speed of 125r/min, 28 DEG C
Environment temperature.
In the present embodiment, concentration described in step D is concentrated at 60 DEG C.
In the present embodiment, hydrogen peroxide solution described in step F, volumetric concentration 25%.
In the present embodiment, papain and starch in papain described in step G and amylase mixed solution
The mass ratio of enzyme is 0.8:1.4.
In the present embodiment, centrifugation described in step G is 6000r/min centrifugation 10min.
Embodiment 5:
A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation, includes the following steps:
A, Coragyceps militaris bacterium is inoculated on northern Chinese caterpillar Fungus seed culture medium, puts shaking table culture case into, cultivated
3.5d, the northern Chinese caterpillar Fungus seed culture medium composition include following parts by weight raw material: 28 parts of sucrose, 2.2 parts of yeast extract,
1 part of potassium dihydrogen phosphate, 0.65 part of magnesium sulfate, 1000 parts of distilled water；
B, the seed liquor for obtaining step A is inoculated on the northern Chinese caterpillar Fungus solid medium of sterilized processing, inoculum concentration
It is 9%, puts into constant temperature and humidity incubator, under the conditions of 28.5 DEG C of temperature, ferments 7 days, obtain cordyceps militaris fruit body；Institute
State the raw material that northern Chinese caterpillar Fungus solid medium includes following parts by weight: 1000 parts of wheat flour, 205 parts of mealy potato, glucose sugar 32
Part, 3.3 parts of potassium dihydrogen phosphate, 2.9 parts of peptone, 1.2 parts of magnesium sulfate, 525 parts of distilled water；
C, the obtained cordyceps militaris fruit body of step B is put into wall-breaking machine, the distilled water of 11.5 times of quality, powder is added
Broken 13min obtains northern Chinese caterpillar Fungus mixed liquor；
D, the northern Chinese caterpillar Fungus mixed liquor for obtaining step C, ultrasonication 48min, the refluxing extraction 3.8h at 73 DEG C,
It is concentrated after extracting solution filtering, obtains polysaccharide concentrate；
E, the dehydrated alcohol of 3.3 times of volumes is added in the polysaccharide concentrate that step D is obtained, is cooled to 4 DEG C after stirring, it is quiet
It sets 28 hours, carries out centrifugal treating, and precipitating drying is obtained into northern Chinese caterpillar Fungus polyoses extract；
F, the northern Chinese caterpillar Fungus polyoses extract that step E is obtained adds distilled water to dissolve, and adjusts pH value to 8, hydrogen peroxide is added dropwise
For solution to colourless, 52 DEG C of heat preservation 2.8h obtain northern Chinese caterpillar Fungus polyoses extract colourless solution；
G, northern Chinese caterpillar Fungus polyoses extract colourless solution that step F is obtained is mixed with papain and amylase molten
Liquid mixing, the volume ratio of the two are 1:0.58,63 DEG C of enzymatic hydrolysis 2h；The Sevage reagent of 20% volume is added in enzymolysis liquid, shakes
Culture 0.6h is swung, then repeatedly centrifugation takes supernatant until no albumen precipitation is precipitated, and dries, it is thick to obtain northern Chinese caterpillar Fungus
Polysaccharide；
H, upper DEAE Ago-Gel FF ion is handed over after dissolving the northern Chinese caterpillar Fungus Thick many candies that step G is obtained with distilled water
Chromatographic column is changed, gradient elution is carried out with sodium chloride solution, obtains northern Chinese caterpillar Fungus polysaccharide.
In the present embodiment, the condition of culture control of shaking table culture case described in step B is the revolving speed of 115r/min, 28 DEG C
Environment temperature.
In the present embodiment, concentration described in step D is concentrated at 60 DEG C.
In the present embodiment, hydrogen peroxide solution described in step F, volumetric concentration 25%.
In the present embodiment, papain and starch in papain described in step G and amylase mixed solution
The mass ratio of enzyme is 0.8:1.25.
In the present embodiment, centrifugation described in step G is 6000r/min centrifugation 10min.
Embodiment 6:
A kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation, includes the following steps:
A, Coragyceps militaris bacterium is inoculated on northern Chinese caterpillar Fungus seed culture medium, puts shaking table culture case into, cultivate 3d；
B, the seed liquor for obtaining step A is inoculated on the northern Chinese caterpillar Fungus solid medium of sterilized processing, inoculum concentration
It is 10%, puts into constant temperature and humidity incubator, under the conditions of 28.5 DEG C of temperature, ferments 7 days, obtain cordyceps militaris fruit body；
C, the obtained cordyceps militaris fruit body of step B is put into wall-breaking machine, the distilled water of 11 times of quality is added, crushed
12min obtains northern Chinese caterpillar Fungus mixed liquor；
D, the northern Chinese caterpillar Fungus mixed liquor for obtaining step C, ultrasonication 45min, the refluxing extraction 4.2h at 75 DEG C,
It is concentrated after extracting solution filtering, obtains polysaccharide concentrate；
E, the dehydrated alcohol of 2.9 times of volumes is added in the polysaccharide concentrate that step D is obtained, is cooled to 4 DEG C after stirring, it is quiet
It sets 20 hours, carries out centrifugal treating, and precipitating drying is obtained into northern Chinese caterpillar Fungus polyoses extract；
F, the northern Chinese caterpillar Fungus polyoses extract that step E is obtained adds distilled water to dissolve, and adjusts pH value to 8.5, dioxygen is added dropwise
For aqueous solution to colourless, 55 DEG C of heat preservation 2.4h obtain northern Chinese caterpillar Fungus polyoses extract colourless solution；
G, northern Chinese caterpillar Fungus polyoses extract colourless solution that step F is obtained is mixed with papain and amylase molten
Liquid mixing, the volume ratio of the two are 1:0.55,62 DEG C of enzymatic hydrolysis 1.5h；The Sevage reagent of 20% volume is added in enzymolysis liquid,
Shaken cultivation 0.8h, then repeatedly centrifugation takes supernatant until no albumen precipitation is precipitated, and dries, obtains northern Chinese caterpillar Fungus
Thick many candies；
H, upper DEAE Ago-Gel FF ion is handed over after dissolving the northern Chinese caterpillar Fungus Thick many candies that step G is obtained with distilled water
Chromatographic column is changed, gradient elution is carried out with sodium chloride solution, obtains northern Chinese caterpillar Fungus polysaccharide.
In the present embodiment, the composition of northern Chinese caterpillar Fungus seed culture medium described in step B includes the original of following parts by weight
Material: 25 parts of sucrose, 2.5 parts of yeast extract, 0.9 part of potassium dihydrogen phosphate, 0.6 part of magnesium sulfate, 1000 parts of distilled water.
In the present embodiment, northern Chinese caterpillar Fungus solid medium described in step B includes the raw material of following parts by weight: wheat
1000 parts of powder, 205 parts of mealy potato, 30 parts of glucose sugar, 2.8 parts of potassium dihydrogen phosphate, 3.2 parts of peptone, 1.1 parts of magnesium sulfate, distillation
520 parts of water.
In the present embodiment, the condition of culture control of shaking table culture case described in step B is the revolving speed of 120r/min, 28 DEG C
Environment temperature.
In the present embodiment, concentration described in step D is concentrated at 60 DEG C.
In the present embodiment, hydrogen peroxide solution described in step F, volumetric concentration 25%.
In the present embodiment, papain and starch in papain described in step G and amylase mixed solution
The mass ratio of enzyme is 0.8:1.3.
In the present embodiment, centrifugation described in step G is 6000r/min centrifugation 10min.
Compared with prior art, the present invention have the following advantages that and the utility model has the advantages that
Northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation of the invention, realizes culture, extraction, purifying
Continuous processing, extraction efficiency are high, and northern Chinese caterpillar Fungus polysaccharide yield is high, and operating process is simpler, and cost is relatively low, and can by polysaccharide into
One step isolates and purifies, and obtains the higher polysaccharide component of purity, is conducive to the development and utilization of northern Chinese caterpillar Fungus polyose.
The above is only presently preferred embodiments of the present invention, not does limitation in any form to the present invention, it is all according to
According to technical spirit any simple modification to the above embodiments of the invention, equivalent variations, protection of the invention is each fallen within
Within the scope of.
Claims (8)
1. a kind of northern Chinese caterpillar Fungus Polyose extraction purifying process based on solid fermentation, characterized by the following steps:
A, Coragyceps militaris bacterium is inoculated on northern Chinese caterpillar Fungus seed culture medium, puts shaking table culture case into, cultivate 2.5-4d,
The northern Chinese caterpillar Fungus seed culture medium composition include following parts by weight raw material: 20-30 parts of sucrose, 1.5-3 parts of yeast extract,
0.5-0.7 parts of magnesium sulfate of 0.8-1.1 parts of potassium dihydrogen phosphate, 1000 parts of distilled water；
B, the seed liquor for obtaining step A is inoculated on the northern Chinese caterpillar Fungus solid medium of sterilized processing, inoculum concentration 8-
11%, it puts into constant temperature and humidity incubator, under the conditions of 28.5 DEG C of temperature, ferments 5-8 days, obtain cordyceps militaris fruit body；
The northern Chinese caterpillar Fungus solid medium includes the raw material of following parts by weight: 1000 parts of wheat flour, 190-215 parts of mealy potato, Portugal
Sugared 25-35 parts of sugar, 2.1-3.5 parts of potassium dihydrogen phosphate, 2.5-3.6 parts of peptone, 0.9-1.3 parts of magnesium sulfate, distilled water 450-600
Part；
C, the obtained cordyceps militaris fruit body of step B is put into wall-breaking machine, the distilled water of 10-12 times of quality is added, crushed
5-15min obtains northern Chinese caterpillar Fungus mixed liquor；
D, the northern Chinese caterpillar Fungus mixed liquor for obtaining step C, ultrasonication 40-50min, the refluxing extraction 3- at 70-80 DEG C
4.5h is concentrated after extracting solution filtering, obtains polysaccharide concentrate；
E, the dehydrated alcohol of 2.8-3.4 times of volume is added in the polysaccharide concentrate that step D is obtained, is cooled to 4 DEG C after stirring, it is quiet
It sets 18-36 hours, carries out centrifugal treating, and precipitating drying is obtained into northern Chinese caterpillar Fungus polyoses extract；
F, the northern Chinese caterpillar Fungus polyoses extract that step E is obtained adds distilled water to dissolve, and adjusts pH value to 8-9, it is water-soluble that dioxygen is added dropwise
For liquid to colourless, 50-60 DEG C of heat preservation 2-3h obtains northern Chinese caterpillar Fungus polyoses extract colourless solution；
G, the northern Chinese caterpillar Fungus polyoses extract colourless solution that step F is obtained is mixed with papain and amylase mixed solution
Close, the volume ratio of the two is 1:(0.45-0.6), 60-65 DEG C of enzymatic hydrolysis 1-2h；The Sevage of 20% volume is added in enzymolysis liquid
Reagent, shaken cultivation 0.5-1h, then repeatedly centrifugation takes supernatant until no albumen precipitation is precipitated, and dries, obtains the northern winter
Worm summer grass Thick many candies；
H, upper DEAE Ago-Gel FF ion exchange layer after dissolving the northern Chinese caterpillar Fungus Thick many candies that step G is obtained with distilled water
Column is analysed, gradient elution is carried out with sodium chloride solution, obtains northern Chinese caterpillar Fungus polysaccharide.
2. the northern Chinese caterpillar Fungus Polyose extraction purifying process according to claim 1 based on solid fermentation, it is characterised in that:
The composition of northern Chinese caterpillar Fungus seed culture medium described in step A includes the raw material of following parts by weight: 25 parts of sucrose, yeast extract 2.5
Part, 0.9 part of potassium dihydrogen phosphate, 0.6 part of magnesium sulfate, 1000 parts of distilled water.
3. the northern Chinese caterpillar Fungus Polyose extraction purifying process according to claim 1 based on solid fermentation, it is characterised in that:
Northern Chinese caterpillar Fungus solid medium described in step B includes the raw material of following parts by weight: 1000 parts of wheat flour, 205 parts of mealy potato,
Glucose sugar 30 parts, 2.8 parts of potassium dihydrogen phosphate, 3.2 parts of peptone, 1.1 parts of magnesium sulfate, 520 parts of distilled water.
4. the northern Chinese caterpillar Fungus Polyose extraction purifying process according to claim 1 based on solid fermentation, it is characterised in that:
The condition of culture control of shaking table culture case described in step A is the revolving speed of 110-125r/min, 28 DEG C of environment temperature.
5. the northern Chinese caterpillar Fungus Polyose extraction purifying process according to claim 1 based on solid fermentation, it is characterised in that:
Concentration described in step D is concentrated at 60 DEG C.
6. the northern Chinese caterpillar Fungus Polyose extraction purifying process according to claim 1 based on solid fermentation, it is characterised in that:
Hydrogen peroxide solution described in step F, volumetric concentration 25%.
7. the northern Chinese caterpillar Fungus Polyose extraction purifying process according to claim 1 based on solid fermentation, it is characterised in that:
The mass ratio of papain and amylase is 0.8:(1.2- in papain described in step G and amylase mixed solution
1.4)。
8. the northern Chinese caterpillar Fungus Polyose extraction purifying process according to claim 1 based on solid fermentation, it is characterised in that:
Centrifugation described in step G is 6000r/min centrifugation 10min.
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