CN103988712B - The cultural method of a kind of high yield Cordyceps sinensis polysaccharide Cordyceps militaris - Google Patents
The cultural method of a kind of high yield Cordyceps sinensis polysaccharide Cordyceps militaris Download PDFInfo
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- CN103988712B CN103988712B CN201410234859.9A CN201410234859A CN103988712B CN 103988712 B CN103988712 B CN 103988712B CN 201410234859 A CN201410234859 A CN 201410234859A CN 103988712 B CN103988712 B CN 103988712B
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- 241001264174 Cordyceps militaris Species 0.000 title claims abstract description 135
- 150000004676 glycans Chemical class 0.000 title claims abstract description 43
- 229920001282 polysaccharide Polymers 0.000 title claims abstract description 43
- 239000005017 polysaccharide Substances 0.000 title claims abstract description 43
- 238000000034 method Methods 0.000 title claims abstract description 30
- 241001248610 Ophiocordyceps sinensis Species 0.000 title claims description 21
- 239000001963 growth medium Substances 0.000 claims abstract description 47
- 238000011218 seed culture Methods 0.000 claims abstract description 23
- 239000000725 suspension Substances 0.000 claims abstract description 21
- 230000004913 activation Effects 0.000 claims abstract description 8
- 230000001954 sterilising effect Effects 0.000 claims description 31
- 239000000843 powder Substances 0.000 claims description 24
- 235000013312 flour Nutrition 0.000 claims description 22
- KFSLWBXXFJQRDL-UHFFFAOYSA-N Peracetic acid Chemical compound CC(=O)OO KFSLWBXXFJQRDL-UHFFFAOYSA-N 0.000 claims description 20
- 238000005286 illumination Methods 0.000 claims description 18
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 16
- 239000000203 mixture Substances 0.000 claims description 16
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 16
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- 239000008103 glucose Substances 0.000 claims description 15
- 238000004659 sterilization and disinfection Methods 0.000 claims description 15
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 14
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 14
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 14
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 14
- 239000001888 Peptone Substances 0.000 claims description 13
- 108010080698 Peptones Proteins 0.000 claims description 13
- 235000019319 peptone Nutrition 0.000 claims description 13
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- 235000019341 magnesium sulphate Nutrition 0.000 claims description 8
- 239000011780 sodium chloride Substances 0.000 claims description 7
- FYYHWMGAXLPEAU-UHFFFAOYSA-N Magnesium Chemical compound [Mg] FYYHWMGAXLPEAU-UHFFFAOYSA-N 0.000 claims description 6
- 238000010790 dilution Methods 0.000 claims description 6
- 239000012895 dilution Substances 0.000 claims description 6
- QEWYKACRFQMRMB-UHFFFAOYSA-N fluoroacetic acid Chemical compound OC(=O)CF QEWYKACRFQMRMB-UHFFFAOYSA-N 0.000 claims description 6
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- AIUDWMLXCFRVDR-UHFFFAOYSA-N dimethyl 2-(3-ethyl-3-methylpentyl)propanedioate Chemical class CCC(C)(CC)CCC(C(=O)OC)C(=O)OC AIUDWMLXCFRVDR-UHFFFAOYSA-N 0.000 description 3
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- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 2
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 2
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 description 2
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- 239000002126 C01EB10 - Adenosine Substances 0.000 description 1
- KQLDDLUWUFBQHP-UHFFFAOYSA-N Cordycepin Natural products C1=NC=2C(N)=NC=NC=2N1C1OCC(CO)C1O KQLDDLUWUFBQHP-UHFFFAOYSA-N 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
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- SRBFZHDQGSBBOR-LECHCGJUSA-N alpha-D-xylose Chemical compound O[C@@H]1CO[C@H](O)[C@H](O)[C@H]1O SRBFZHDQGSBBOR-LECHCGJUSA-N 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 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
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-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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- 150000001875 compounds Chemical class 0.000 description 1
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- OFEZSBMBBKLLBJ-BAJZRUMYSA-N cordycepin Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)C[C@H]1O OFEZSBMBBKLLBJ-BAJZRUMYSA-N 0.000 description 1
- OFEZSBMBBKLLBJ-UHFFFAOYSA-N cordycepine Natural products C1=NC=2C(N)=NC=NC=2N1C1OC(CO)CC1O OFEZSBMBBKLLBJ-UHFFFAOYSA-N 0.000 description 1
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- 238000011081 inoculation Methods 0.000 description 1
- 210000000265 leukocyte Anatomy 0.000 description 1
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- 239000000463 material Substances 0.000 description 1
- 239000013028 medium composition Substances 0.000 description 1
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- 235000019713 millet Nutrition 0.000 description 1
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Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G18/00—Cultivation of mushrooms
-
- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
- C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
Abstract
The present invention relates to field of edible fungus culture, the particularly cultural method of a kind of high polysaccharide Cordyceps militaris, comprises the following steps: by Cordyceps militaris spawn activation culture, make spore suspension; Ratio by spore suspension taking percentage by volume as 3-8% is inoculated in Cordyceps militaris seed culture medium and cultivates, and obtains seed liquor; Seed liquor is inoculated in Cordyceps militaris growth medium secretly cultivate successively, mycelia annesl cultivate and fructification cultivate, obtain fruiting bodies of cordyceps militaris. The cultural method of high polysaccharide Cordyceps militaris provided by the invention, spore suspension is provided in the Cordyceps militaris seed culture medium providing in the present invention and cultivates, and obtains seed liquor; Seed liquor is inoculated in Cordyceps militaris growth medium provided by the invention secretly cultivate successively, mycelia annesl cultivate and fructification cultivate, cultural method is simple, cultivation cycle is short, the fruiting bodies of cordyceps militaris Polysaccharides in Cultured Cordyceps militaris content obtaining is high, and the purity of the Polysaccharides in Cultured Cordyceps militaris extracting with this fruiting bodies of cordyceps militaris is high.
Description
Technical field
The present invention relates to field of edible fungus culture, many in particular to a kind of high yield Chinese caterpillar fungusThe cultural method of sugar Cordyceps militaris.
Background technology
Cordyceps militaris claims again northern Chinese caterpillar fungus, northern Chinese caterpillar Fungus, is that fungi autoeciousness is in insects such as LepidopterasThe upper entomogenous fungi complex forming of polypide, be a kind of important and nutriment with tonic effect.The multiple efficacy of a drug effects such as that Cordyceps militaris has is antitumor, anti-inflammatory, it in East Asia Region by wideGeneral as invigorant and medicinal fungus use, the Ministry of Public Health of China is formal on March 16th, 2009Classified as new resource food.
The wherein polysaccharide in Cordyceps militaris is mainly by mannose, cordycepin, adenosine, galaThe polysaccharide of sugar, arabinose, wood sugar essence, glucose, fucose composition. Experiment showed,Cordyceps sinensis polysaccharide can improve immune function of human body, and increasing leukocyte improves respiratory system, can makeAdrenal gland weight, blood plasma cortisol, aldosterone and adrenal gland inner cholesterol content increase, and havePromote adrenal effect, suppress tumor growth, and there is antitumor, radioresistance, fall bloodSugar and lipoprotein, cough-relieving, reduce phlegm, moistening lung and the pharmacological action such as delay senility, clinically useIn treatment malignant tumour, therefore, Cordyceps sinensis polysaccharide has important value.
At present, Polysaccharides in Cultured Cordyceps militaris content mainly concentrates on by liquid deep layer fermenting and improves,But what submerged fermentation obtained is cordyceps mycelium, it comprises outside Cordyceps militaris self, goes backThere are some culture medium residues, affect the purity of Polysaccharides in Cultured Cordyceps militaris.
Summary of the invention
The object of the present invention is to provide Cordyceps militaris seed culture medium, Cordyceps militaris growth mediumAnd the cultural method of a kind of high yield Cordyceps sinensis polysaccharide Cordyceps militaris, to solve the above problems.
Cordyceps militaris seed culture medium is provided in an embodiment of the present invention, by weight,Comprise following composition: glucose 8-12 part, peptone 2-4 part, dusty yeast 4-6 part, phosphorusAcid dihydride potassium 0.02-0.06 part, magnesium sulfate 0.02-0.06 part, sodium chloride 4-6 part, vitaminB10.05-0.15 part, yam flour 8-12 part, water 800-1200 part.
Cordyceps militaris growth medium is also provided in an embodiment of the present invention, by weight,Comprise following composition: corn flour 12-18 part, wheat powder 5-10 part, yam flour 3-8 part,Dry Ribes burejense powder 2-5 part, peptone 0.02-0.05 part, potassium dihydrogen phosphate 0.02-0.04 part, sulphurAcid magnesium 0.02-0.03 part, glucose 0.02-0.08 part, vitaminB10 .0005-0.001Part, water 30-40 part.
A cultural method for high yield Cordyceps sinensis polysaccharide Cordyceps militaris, comprises the following steps:
(a), by Cordyceps militaris spawn activation culture, making concentration is 1-3 × 107CFU/ml'sSpore suspension;
(b) by described spore suspension, the ratio taking percentage by volume as 3-8% is inoculated in rightRequire to cultivate in the Cordyceps militaris seed culture medium described in 1, obtain seed liquor;
(c) described seed liquor is inoculated in to Cordyceps militaris grown cultures claimed in claim 2In base, secretly cultivate successively, mycelia annesl cultivate and fructification cultivate, obtain Cordyceps militarisEntity.
Preferably, in described step (b), condition of culture is: 25-30 DEG C, 240-260Rpm/min shaken cultivation.
Preferably, in described step (b), cultivate 3-5d, obtain described seed liquor.
Preferably, by after 20-30 times of described seed liquor dilution, taking percentage by volume as 5-8%Inoculate.
Preferably, in described step (c), described dark cultivation is entered between the cultivation of having sterilizedRow is cultivated, and condition of culture is: temperature 20-25 DEG C, humidity 60-70%, cultivates 6-8d.
Preferably, in described step (c), the condition of culture that described mycelia annesl is cultivated is:Temperature 15-20 DEG C, humidity 60-70%, the 1.5-2.5h that ventilates every day, illumination 8-12h, lightBe 250-300Lux according to intensity, cultivate 5-8d.
Preferably, in described step (c), the condition of culture that described fructification is cultivated is:Ventilative cultivation, temperature is 20 DEG C ± 2 DEG C, humidity is 80-90%, the 1.5-2.5h that ventilates every day,Illumination 6-8h, intensity of illumination is 250-300Lux, being cultured to fructification length is 6-10cm,Gather and obtain described fruiting bodies of cordyceps militaris.
Preferably, described sterilization is: the container that peracetic acid soln is housed is placed in to 75-85 DEG CWater-bath in stifling to sterilizing between described cultivation, fumigation time is 90-100min, described mistakeEvery cubic metre of space consumption of fluoroacetic acid is 3-3.2g; During sterilization, cultivate room temperature and be not less than20 DEG C, humidity is 60%-80%.
The cultural method of the high yield Cordyceps sinensis polysaccharide Cordyceps militaris that the embodiment of the present invention provides, willSpore suspension is inoculated in the Cordyceps militaris seed culture medium providing in the present invention and cultivates,To seed liquor; Seed liquor is inoculated in Cordyceps militaris growth medium provided by the invention and is complied withInferiorly secretly cultivate, mycelia annesl cultivates and fructification is cultivated, cultural method is simple, cultivatesCycle is short, and the fruiting bodies of cordyceps militaris Polysaccharides in Cultured Cordyceps militaris content obtaining is high, and with this Cordyceps militarisThe purity of the Polysaccharides in Cultured Cordyceps militaris of entity extraction is high.
Detailed description of the invention
Below by specific embodiment, the present invention is described in further detail.
Cordyceps militaris seed culture medium is provided in an embodiment of the present invention, by weight,Comprise following composition: glucose 8-12 part, peptone 2-4 part, dusty yeast 4-6 part, phosphorusAcid dihydride potassium 0.02-0.06 part, magnesium sulfate 0.02-0.06 part, sodium chloride 4-6 part, vitaminB10.05-0.15 part, yam flour 8-12 part, water 800-1200 part.
Further, by weight, comprise following composition: glucose 9-10 part, albumenPeptone 3-4 part, dusty yeast 5-6 part, potassium dihydrogen phosphate 0.03-0.05 part, magnesium sulfate 0.03-0.05Part, sodium chloride 4-5 part, vitaminB10 .05-0.15 part, yam flour 9-10 part, water 1000Part.
The Cordyceps militaris seed culture medium comprehensive nutrition obtaining, medium component concentration and proportioning are closedSuitable; The spore suspension of Cordyceps militaris is seeded to basal culture medium, and part spore grows into mycelium,Spore vitality is vigorous, and spore count amplification is fast, and obtains part mycelium, the kind obtainingSub-liquid contains spore and mycelium, is beneficial to subsequent growth.
Cordyceps militaris growth medium is also provided in an embodiment of the present invention, by weight,Comprise following composition: corn flour 12-18 part, wheat powder 5-10 part, yam flour 3-8 part,Dry Ribes burejense powder 2-5 part, peptone 0.02-0.05 part, potassium dihydrogen phosphate 0.02-0.04 part, sulphurAcid magnesium 0.02-0.03 part, glucose 0.02-0.08 part, vitaminB10 .0005-0.001Part, water 30-40 part.
Further, by weight, comprise following composition: corn flour 14-16 part, wheatSub-powder 6-8 part, yam flour 4-6 part, dry Ribes burejense powder 3-4 part, peptone 0.03-0.04 part,Potassium dihydrogen phosphate 0.02-0.04 part, magnesium sulfate 0.02-0.03 part, glucose 0.03-0.06 part,VitaminB10 .0005-0.001 part, water 30-40 part.
The Cordyceps militaris growth medium comprehensive nutrition obtaining, medium component concentration and proportioning are closedSuitable; The seed liquor of Cordyceps militaris is seeded to basal culture medium, is beneficial to spore and is grown to mycelium,Obtain more mycelium, be beneficial to and obtain more fruiting bodies of cordyceps militaris, and the pupa worm obtainingGrass seed entity Polysaccharides in Cultured Cordyceps militaris content is high.
Wherein, the corn flour relating in Cordyceps militaris seed culture medium and Cordyceps militaris growth mediumBe 1-3mm with the granularity of wheat powder, yam flour and dry Ribes burejense powder are made in the following waysStandby: fresh Chinese yam to be dried, then pulverize, obtain the yam flour that granularity is 0.8-1.5mm;Fresh Rosa roxburghii is dried, go to pulverize after seed, obtaining granularity is the dry Ribes burejense powder of 0.8-1.5mm.The corn flour, wheat powder, yam flour and the dry Ribes burejense powder that use this granularity, it is in sterilizingIn process, can be evenly distributed, the component distributing homogeneous in the culture medium obtaining after sterilizing.
The composition respectively Cordyceps militaris seed culture medium and Cordyceps militaris growth medium being contained separatelyTake rear mixing, all adopt autoclave sterilization, be specially 121 DEG C, sterilizing 20-30min,After sterilizing, be cooled to room temperature, obtain being directly used in the culture medium of cultivating bacterial strain.
The cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris, comprises the following steps:
(a), by Cordyceps militaris spawn activation culture, making concentration is 1-3 × 107CFU/ml'sSpore suspension;
(b) by described spore suspension, the ratio taking percentage by volume as 3-8% is inoculated in rightRequire to cultivate in the Cordyceps militaris seed culture medium described in 1, obtain seed liquor;
(c) described seed liquor is inoculated in to Cordyceps militaris grown cultures claimed in claim 2In base, secretly cultivate successively, mycelia annesl cultivate and fructification cultivate, obtain Cordyceps militarisEntity.
Adopt Cordyceps militaris seed culture medium provided by the invention and Cordyceps militaris growth medium to carry outThe cultivation of Cordyceps militaris, cultural method is simple, and cultivation cycle is short, and cost is low, the pupa worm obtainingGrass seed entity Polysaccharides in Cultured Cordyceps militaris content is high; In addition, available technology adopting liquid fermentation and cultureCordyceps militaris, the major part of Cordyceps militaris is in zymotic fluid, and the materials such as the secretion that self produces are mixedMix in zymotic fluid, thereby the Cordyceps militaris the obtaining composition that contains zymotic fluid, and the present invention carriesThe Cordyceps militaris growth medium of confession is solid medium, adopts solid culture Cordyceps militaris, plucksThe fruiting bodies of cordyceps militaris obtaining, the impurity while having avoided adopting liquid fermentation and culture in culture mediumSneak into the defect in fruiting bodies of cordyceps militaris, the Cordyceps militaris impurity obtaining still less, therefore, follow-upThe purity of the polysaccharide that Cordyceps militaris is extracted is higher.
Cordyceps militaris spawn activation culture, Cordyceps militaris spawn is purchased from Chinese Typical Representative culture collectionThe heart, preserving number is CCTCCM2013056, and Cordyceps militaris spawn is inoculated in to potato plugCulture medium, cultivates 6-8d for 27 DEG C ± 2 DEG C, obtains the spore of Cordyceps militaris, with containing 0.05%The aseptic washing of Tween 80 under spore, making concentration is 1-3 × 107The spore of CFU/mlSub-suspension. Wherein, potato slant medium composition comprises potato 100g, glucose10g, potassium dihydrogen phosphate 1g, agar 16g, water 1000ml; Compound method is: getThe potato 100g of peeling, is cut into small pieces, and the 1000m1 that adds water, boils 20min, uses 4-6Layer filtered through gauze, then supplies dehydration to 1000m1, adds agar 16g to dissolve, thenAdd again glucose 10g, potassium dihydrogen phosphate 1g, packing test tube, 121 DEG C of autoclavings25-30min, prepares potato slant medium. Employing potato slant medium is livedThe cordyceps species of pupating, method is simple, and the spore obtaining is many. Telling of employing 0.05%It is 1-3 × 10 that spore is made concentration by the sterilized water of temperature 807The spore suspension of CFU/ml,The spore suspension spore making is evenly distributed; Then the ratio taking percentage by volume as 3-8% connectsPlant to Cordyceps militaris seed culture medium, inoculum concentration is moderate, is beneficial at Cordyceps militaris seed culture medium fastFast-growing is long.
Preferably, in described step (b), condition of culture is: 25-30 DEG C, 240-260Rpm/min shaken cultivation. Empirical tests, cultivation temperature is 25-30 DEG C, Cordyceps militaris is Cordyceps militarisIn seed culture medium, growth fast; Rotating speed 240-260rpm/min, is not damaging Cordyceps militaris sporeIn the situation of son, increase dissolved oxygen amount, be beneficial to spore and mycelial growth, simultaneously can be byCordyceps militaris seed culture medium mixes, to prevent that local Cordyceps militaris seed culture medium nutrition from becomingDivide not enough and affect spore and mycelial growth, therefore, Cordyceps militaris under this condition of cultureSpore fast growth, the mycelium of formation is more, save incubation time.
Preferably, in described step (b), cultivate 3-5d, obtain described seed liquor.Cultivate 3-5d, the seed liquor thalline content obtaining is high, and it is vigorous to grow.
Preferably, by after 20-30 times of described seed liquor dilution, taking percentage by volume as 5-8%Inoculate. In Cordyceps militaris growth medium, inoculate appropriate seed liquor, be beneficial to and obtain pupaCordyceps militaris sporocarp; In order to prevent the rear distributing inhomogeneity of inoculum concentration and inoculation, first by seed liquorDilute, then inoculate taking percentage by volume as 5-8%, can obtain seed liquor and divideThe uniform Cordyceps militaris growth medium of cloth, is beneficial to the growth of thalline.
Preferably, in described step (c), described dark cultivation is entered between the cultivation of having sterilizedRow is cultivated, and condition of culture is: temperature 20-25 DEG C, humidity 60-70%, cultivates 6-8d. ShouldUnder condition of culture, thalli growth is quick, covers with mycelia in incubator, and cultivation cycle is short.
Preferably, in described step (c), the condition of culture that described mycelia annesl is cultivated is:Temperature 15-20 DEG C, humidity 60-70%, the 1.5-2.5h that ventilates every day, illumination 8-12h, lightBe 250-300Lux according to intensity, cultivate 5-8d. Under this condition of culture, thalli growth is quick,Mycelia starts to form the former base of millet shape in media surface, and mycelium annesl is effective, and trainingThe cycle of supporting is short.
Preferably, in described step (c), the condition of culture that described fructification is cultivated is:Ventilative cultivation, temperature is 20 DEG C ± 2 DEG C, humidity is 80-90%, the 1.5-2.5h that ventilates every day,Illumination 6-8h, intensity of illumination is 250-300Lux, being cultured to fructification length is 6-10cm,Gather and obtain described fruiting bodies of cordyceps militaris. Ventilative cultivation is: on the film of culture vessel mouthPrick several holes, be beneficial to circulation of air, be beneficial to culture growth. And at this condition of cultureUnder, thalli growth is quick, and cultivation cycle is short, the fruiting bodies of cordyceps militaris Polysaccharides in Cultured Cordyceps militaris obtainingContent is high.
Preferably, described sterilization is: the container that peracetic acid soln is housed is placed in to 75-85 DEG CWater-bath in stifling to sterilizing between described cultivation, fumigation time is 90-100min, described mistakeEvery cubic metre of space consumption of fluoroacetic acid is 3-3.2g; During sterilization, cultivate room temperature and be not less than20 DEG C, humidity is 60%-80%. Space to Cordyceps militaris growth carries out disinfection, to prevent itBe subject to disease, and prevent the pollution of other microorganisms; Adopt Peracetic acid to disappear between cultivating to stiflingPoison, sterilization is thorough, and to the injury of Cordyceps militaris culture nothing itself, in the training period, pupa wormGrass culture growth conditions is good, and the fruiting bodies of cordyceps militaris that obtains is pollution-free and the Cordyceps militaris that obtainsFructification Polysaccharides in Cultured Cordyceps militaris content is high.
The cultural method of the high yield Cordyceps sinensis polysaccharide Cordyceps militaris that the embodiment of the present invention provides, willSpore suspension is inoculated in the Cordyceps militaris seed culture medium providing in the present invention and cultivates,To seed liquor; Seed liquor is inoculated in Cordyceps militaris growth medium provided by the invention and is complied withInferiorly secretly cultivate, mycelia annesl cultivates and fructification is cultivated, cultural method is simple, cultivatesCycle is short, and the fruiting bodies of cordyceps militaris Polysaccharides in Cultured Cordyceps militaris content obtaining is high, and with this Cordyceps militarisThe purity of the Polysaccharides in Cultured Cordyceps militaris of entity extraction is high.
Embodiment 1
By weight, take following composition: 8 parts of glucose, 2 parts of peptones, yeast4 parts, powder, 0.02 part of potassium dihydrogen phosphate, 0.02 part, magnesium sulfate, 4 parts, sodium chloride, vitaminB10.05 part, 8 parts of yam flours, 800 parts, water, then 121 DEG C, sterilizing 25min, goes outAfter bacterium, be cooled to room temperature, obtain Cordyceps militaris seed culture medium;
By weight, take following composition: 12 parts of corn flour, 5 parts, wheat powder, Chinese yam3 parts, powder, 2 parts of dry Ribes burejense powders, 0.02 part of peptone, 0.02 part of potassium dihydrogen phosphate, sulfuric acid30 parts, 0.02 part, magnesium, 0.02 part of glucose, vitaminB10 .0005 part, water, then121 DEG C, sterilizing 25min, is cooled to room temperature after sterilizing, obtain Cordyceps militaris growth medium;
The cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris, comprises the following steps:
(a), by Cordyceps militaris spawn activation culture, making concentration is 1 × 107The spore of CFU/mlSub-suspension;
(b) spore suspension is inoculated in to Cordyceps militaris kind taking percentage by volume as 8% ratioIn sub-culture medium, cultivate, condition of culture is: 25 DEG C, 240rpm/min shaken cultivation, cultivates3d, obtains seed liquor;
(c) by after 20 times of seed liquor dilutions, taking percentage by volume as 5%, seed liquor is inoculatedIn Cordyceps militaris growth medium, secretly cultivate successively, mycelia annesl cultivates and fructification trainingSupport;
Dark cultivation cultivated between the cultivation of having sterilized, and condition of culture is: 20 DEG C of temperature,Humidity 60%, cultivates 6d;
The condition of culture that mycelia annesl is cultivated is: 15 DEG C of temperature, and humidity 60%, ventilate to every day1.5h, illumination 8h, intensity of illumination is 250Lux, cultivates 5d;
After mycelia annesl has been cultivated, culture is carried out to fructification cultivation, condition of culture is:Ventilative cultivation, temperature is 20 DEG C ± 2 DEG C, humidity is 80%, the 1.5h that ventilates every day, illumination6h, intensity of illumination is 250Lux, being cultured to fructification length is 6-10cm, gathers and obtainsFruiting bodies of cordyceps militaris;
Wherein, between cultivating, carry out disinfection in the following ways: peracetic acid soln will be housedContainer to be placed in the water-bath of 75 DEG C stifling to sterilizing between described cultivation, fumigation time is 100Min, every cubic metre of space consumption of Peracetic acid is 3-3.2g; During sterilization, chamber between cultivationTemperature is not less than 20 DEG C, and humidity is 60%-80%.
It is golden yellow that the fruiting bodies of cordyceps militaris obtaining is, anthrone colorimetric method for determining fruiting bodies of cordyceps militarisTotal sugar content, the Polysaccharides in Cultured Cordyceps militaris content obtaining is 18%.
Embodiment 2
By weight, take following composition: 10 parts of glucose, 3 parts of peptones, yeast5 parts, powder, 0.04 part of potassium dihydrogen phosphate, 0.04 part, magnesium sulfate, 5 parts, sodium chloride, vitaminB10.10 part, 10 parts of yam flours, 1000 parts, water, then 121 DEG C, sterilizing 30min,After sterilizing, be cooled to room temperature, obtain Cordyceps militaris seed culture medium;
By weight, take following composition: 15 parts of corn flour, 8 parts, wheat powder, Chinese yam5 parts, powder, 4 parts of dry Ribes burejense powders, 0.04 part of peptone, 0.03 part of potassium dihydrogen phosphate, sulfuric acid35 parts, 0.02 part, magnesium, 0.05 part of glucose, vitaminB10 .0008 part, water, then121 DEG C, sterilizing 30min, is cooled to room temperature after sterilizing, obtain Cordyceps militaris growth medium;
The cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris, comprises the following steps:
(a), by Cordyceps militaris spawn activation culture, making concentration is 2 × 107The spore of CFU/mlSub-suspension;
(b) spore suspension is inoculated in to Cordyceps militaris kind taking percentage by volume as 5% ratioIn sub-culture medium, cultivate, condition of culture is: 27 DEG C, 250rpm/min shaken cultivation, cultivates4d, obtains seed liquor;
(c) by after 25 times of seed liquor dilutions, taking percentage by volume as 6%, seed liquor is inoculatedIn Cordyceps militaris growth medium, secretly cultivate successively, mycelia annesl cultivates and fructification trainingSupport;
Dark cultivation cultivated between the cultivation of having sterilized, and condition of culture is: 22 DEG C of temperature,Humidity 65%, cultivates 7d;
The condition of culture that mycelia annesl is cultivated is: 18 DEG C of temperature, and humidity 65%, ventilate to every day2.0h, illumination 10h, intensity of illumination is 280Lux, cultivates 7d;
After mycelia annesl has been cultivated, culture is carried out to fructification cultivation, condition of culture is:Ventilative cultivation, temperature is 20 DEG C ± 2 DEG C, humidity is 85%, the 2.0h that ventilates every day, illumination7h, intensity of illumination is 280Lux, being cultured to fructification length is 6-10cm, gathers and obtainsFruiting bodies of cordyceps militaris;
Wherein, between cultivating, carry out disinfection in the following ways: peracetic acid soln will be housedContainer to be placed in the water-bath of 80 DEG C stifling to sterilizing between described cultivation, fumigation time is 95Min, every cubic metre of space consumption of described Peracetic acid is 3-3.2g; During sterilization, cultivateRoom temperature is not less than 20 DEG C, and humidity is 60%-80%.
It is golden yellow that the fruiting bodies of cordyceps militaris obtaining is, anthrone colorimetric method for determining fruiting bodies of cordyceps militarisTotal sugar content, the Polysaccharides in Cultured Cordyceps militaris content obtaining is 20%.
Embodiment 3
By weight, take following composition: 12 parts of glucose, 4 parts of peptones, yeast6 parts, powder, 0.06 part of potassium dihydrogen phosphate, 0.06 part, magnesium sulfate, 6 parts, sodium chloride, vitaminB10.15 part, 12 parts of yam flours, 1200 parts, water, then 121 DEG C, sterilizing 35min,After sterilizing, be cooled to room temperature, obtain Cordyceps militaris seed culture medium;
By weight, take following composition: 18 parts of corn flour, 10 parts, wheat powder, mountain8 parts, medicinal powder, 5 parts of dry Ribes burejense powders, 0.05 part of peptone, 0.04 part of potassium dihydrogen phosphate, sulphur40 parts, 0.03 part, magnesium of acid, 0.08 part of glucose, vitaminB10 .001 part, water, then121 DEG C, sterilizing 35min, is cooled to room temperature after sterilizing, obtain Cordyceps militaris growth medium;
The cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris, comprises the following steps:
(a), by Cordyceps militaris spawn activation culture, making concentration is 3 × 107The spore of CFU/mlSub-suspension;
(b) spore suspension is inoculated in to Cordyceps militaris kind taking percentage by volume as 3% ratioIn sub-culture medium, cultivate, condition of culture is: 30 DEG C, 260rpm/min shaken cultivation, cultivates5d, obtains seed liquor;
(c) by after 30 times of seed liquor dilutions, taking percentage by volume as 8%, seed liquor is inoculatedIn Cordyceps militaris growth medium, secretly cultivate successively, mycelia annesl cultivates and fructification trainingSupport;
Dark cultivation cultivated between the cultivation of having sterilized, and condition of culture is: 25 DEG C of temperature,Humidity 70%, cultivates 8d;
The condition of culture that mycelia annesl is cultivated is: 20 DEG C of temperature, and humidity 70%, ventilate to every day2.5h, illumination 12h, intensity of illumination is 300Lux, cultivates 8d;
After mycelia annesl has been cultivated, culture is carried out to fructification cultivation, condition of culture is:Ventilative cultivation, temperature is 20 DEG C ± 2 DEG C, humidity is 90%, the 2.5h that ventilates every day, illumination8h, intensity of illumination is 300Lux, being cultured to fructification length is 6-10cm, gathers and obtainsFruiting bodies of cordyceps militaris;
Wherein, between cultivating, carry out disinfection in the following ways: peracetic acid soln will be housedContainer to be placed in the water-bath of 85 DEG C stifling to sterilizing between described cultivation, fumigation time is 90Min, every cubic metre of space consumption of described Peracetic acid is 3-3.2g; During sterilization, cultivateRoom temperature is not less than 20 DEG C, and humidity is 60%-80%.
It is golden yellow that the fruiting bodies of cordyceps militaris obtaining is, anthrone colorimetric method for determining fruiting bodies of cordyceps militarisTotal sugar content, the Polysaccharides in Cultured Cordyceps militaris content obtaining is 25%.
The cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris provided by the invention, cultural method is simple,Adopt different culture mediums to cultivate at different cultivation stages, with guarantee bacterial classification vigor andAdapt to the demand of cultivation target; Condition of culture, the environmental condition of each cultivation stage are strictly controlledSystem, is beneficial to standardization and standardization that Cordyceps militaris is cultivated and manages; Cordyceps militaris obtaining is realBody Polysaccharides in Cultured Cordyceps militaris content is high, is 18-25%, and the pupa worm of extracting with this fruiting bodies of cordyceps militarisThe purity of grass polysaccharide is high.
The foregoing is only the preferred embodiments of the present invention, be not limited to the present invention,For a person skilled in the art, the present invention can have various modifications and variations. AllWithin the spirit and principles in the present invention, any amendment of doing, be equal to replacement, improvement etc.,Within all should being included in protection scope of the present invention.
Claims (9)
1. Cordyceps militaris growth medium, is characterized in that, by weight, comprises followingComposition: corn flour 12-18 part, wheat powder 5-10 part, yam flour 3-8 part, dry Ribes burejense powder2-5 part, peptone 0.02-0.05 part, potassium dihydrogen phosphate 0.02-0.04 part, magnesium sulfate0.02-0.03 part, glucose 0.02-0.08 part, vitaminB10 .0005-0.001 part, water30-40 part.
2. a cultural method for high yield Cordyceps sinensis polysaccharide Cordyceps militaris, is characterized in that, comprisesFollowing steps:
(a), by Cordyceps militaris spawn activation culture, making concentration is 1-3 × 107CFU/ml'sSpore suspension;
(b) by described spore suspension, the ratio taking percentage by volume as 3-8% is inoculated in pupa wormIn grass seed culture medium, cultivate, obtain seed liquor;
(c) described seed liquor is inoculated in to Cordyceps militaris grown cultures claimed in claim 1In base, secretly cultivate successively, mycelia annesl cultivate and fructification cultivate, obtain Cordyceps militarisEntity;
Described Cordyceps militaris seed culture medium, by weight, comprises following composition: glucose8-12 part, peptone 2-4 part, dusty yeast 4-6 part, potassium dihydrogen phosphate 0.02-0.06 part,Magnesium sulfate 0.02-0.06 part, sodium chloride 4-6 part, vitaminB10 .05-0.15 part, Chinese yamPowder 8-12 part, water 800-1200 part.
3. the cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris according to claim 2,It is characterized in that, in described step (b), condition of culture is: 25-30 DEG C, 240-260Rpm/min shaken cultivation.
4. the cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris according to claim 3,It is characterized in that, in described step (b), cultivate 3-5d, obtain described seed liquor.
5. the cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris according to claim 4,It is characterized in that, in described step (c), by after 20-30 times of described seed liquor dilution,Inoculate taking percentage by volume as 5-8%.
6. the cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris according to claim 5,It is characterized in that, in described step (c), described dark cultivation is entered between the cultivation of having sterilizedRow is cultivated, and condition of culture is: temperature 20-25 DEG C, humidity 60-70%, cultivates 6-8d.
7. the cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris according to claim 6,It is characterized in that, in described step (c), the condition of culture that described mycelia annesl is cultivated is:Temperature 15-20 DEG C, humidity 60-70%, the 1.5-2.5h that ventilates every day, illumination 8-12h, lightBe 250-300Lux according to intensity, cultivate 5-8d.
8. the cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris according to claim 7,It is characterized in that, in described step (c), the condition of culture that described fructification is cultivated is:Ventilative cultivation, temperature is 20 DEG C ± 2 DEG C, humidity is 80-90%, the 1.5-2.5h that ventilates every day,Illumination 6-8h, intensity of illumination is 250-300Lux, being cultured to fructification length is 6-10cm,Gather and obtain described fruiting bodies of cordyceps militaris.
9. the cultural method of high yield Cordyceps sinensis polysaccharide Cordyceps militaris according to claim 6,It is characterized in that, described sterilization is: the container that peracetic acid soln is housed is placed in to 75-85 DEG CWater-bath in stifling to sterilizing between described cultivation, fumigation time is 90-100min, described mistakeEvery cubic metre of space consumption of fluoroacetic acid is 3-3.2g; During sterilization, cultivate room temperature and be not less than20 DEG C, humidity is 60%-80%.
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| CN103988712B (en) * | 2014-05-29 | 2016-05-11 | 熊艳 | The cultural method of a kind of high yield Cordyceps sinensis polysaccharide Cordyceps militaris |
| CN104756759B (en) * | 2015-03-17 | 2017-01-04 | 沁阳市西向食用菌研究所 | A kind of Rhizoma Dioscoreae Cordyceps and cultural method thereof |
| CN105418192A (en) * | 2015-12-04 | 2016-03-23 | 正源堂（天津）生物科技有限公司 | Cordyceps militaris strain culture medium and preparation method thereof |
| CN105503375A (en) * | 2015-12-05 | 2016-04-20 | 正源堂（天津）生物科技有限公司 | Efficient cordyceps militaris strain medium |
| CN106867791A (en) * | 2017-04-13 | 2017-06-20 | 刘勇 | The preparation method of Cordceps militaris health liquor |
| CN108633616A (en) * | 2018-05-11 | 2018-10-12 | 江苏圣福来生态农业有限公司 | A kind of culture implantation methods improving activity substance content in Cordyceps militaris |
| KR101981132B1 (en) * | 2018-11-30 | 2019-05-22 | 박성웅 | Attachable Trap for Culture of Cordyceps spp. and Cultivating Method Thereof |
| CN110129207B (en) * | 2019-04-22 | 2024-04-05 | 江苏农林职业技术学院 | Liquid fermentation medium for high-yield antioxidant cordyceps sobolifera mycelium and production method of antioxidant cordyceps sobolifera mycelium electuary |
| CN110122193B (en) * | 2019-06-17 | 2021-07-09 | 山西万海澳生物科技有限责任公司 | Cordyceps militaris cultivation method capable of stabilizing high-content polysaccharide |
| CN110522684A (en) * | 2019-07-22 | 2019-12-03 | 北京京瑜科技有限公司 | A kind of two-way liquid fermentation process of Cordyceps militaris-mung bean |
| CN110663455B (en) * | 2019-10-22 | 2022-07-29 | 安发（福建）生物科技有限公司 | Cordyceps militaris cultivation inoculation method |
| CN110643518A (en) * | 2019-10-22 | 2020-01-03 | 安发（福建）生物科技有限公司 | Method for culturing hirsutella sinensis |
| CN112314325A (en) * | 2020-09-14 | 2021-02-05 | 上海国宝企业发展中心 | Method for culturing artificial cordyceps militaris sporocarp |
| CN112352945A (en) * | 2020-09-14 | 2021-02-12 | 上海国宝企业发展中心 | Artificial cordyceps militaris food with effects of resisting fatigue and improving immunity |
| CN112273142B (en) * | 2020-11-12 | 2022-04-29 | 江苏康能生物工程股份有限公司 | Culture method for increasing content of crude polysaccharide in cordyceps militaris sporocarp |
| CN112568056A (en) * | 2020-12-08 | 2021-03-30 | 锦州聚红农业科技有限公司 | High-adenosine cordyceps militaris cultivation method |
| CN112602534A (en) * | 2020-12-15 | 2021-04-06 | 青海珠峰冬虫夏草工程技术研究有限公司 | Method for high-yield tending of cordyceps sinensis in original land |
| CN112514797B (en) * | 2020-12-22 | 2021-06-11 | 甘肃省农业科学院林果花卉研究所 | Peach stock culture medium and use method thereof |
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| CN115093972B (en) * | 2022-04-26 | 2023-06-30 | 上海市农业科学院 | Cordyceps militaris strain and application thereof |
| CN115053750A (en) * | 2022-06-10 | 2022-09-16 | 扬州大学 | Method for artificially cultivating Clanis bilineata tsingtauica cordyceps militaris by inoculating live bean worms |
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