CN103621308A - Culture medium for producing isaria tenuipes entities and industrialized cultural method for isaria tenuipes entities - Google Patents
Culture medium for producing isaria tenuipes entities and industrialized cultural method for isaria tenuipes entities Download PDFInfo
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Abstract
The invention relates to the technical field of microorganism, in particular to a culture medium for producing isaria tenuipes entities and an industrialized cultural method for the isaria tenuipes entities. The industrialized cultural method for the isaria tenuipes entities includes the steps of culturing seeds in a slope, seeds in a shaking flask and seeds in a fermentation tank, using wheat or barley as main materials of the culture medium, using silkworm chrysalis powder as auxiliary materials of the culture medium, carrying out sterilization in a material steaming tank, cooling, obtaining the solid fermentation culture medium, adding the seed solution into the material steaming tank, evenly mixing the seed solution with the solid culture medium, subpackaging the mixture into solid fermentation culture boxes through an automatic subpackaging machine, covering the solid fermentation culture boxes with films for sealing, moving the solid fermentation culture boxes into a culture room, controlling conditions such as the temperature, the illumination, the humidity and the ventilation in the culture process, and culturing the isaria tenuipes entities. Large-scale industrialized production of the isaria tenuipes entities can be achieved. The industrialized cultural method for the isaria tenuipes entities is high in yield, good in product quality and low in product price, and has the good industrial production and medical treatment application prospects.
Description
Technical field
The present invention relates to microbial technology field, be specifically related to medium and industrial method for culturing that a kind of thin pin Isaria fruit body is produced.
Background technology
Thin pin Isaria (Isaria tenuipes), is once called as thin pin Paecilomyces varioti (Paecilomyces tenuipes), is the phorozoon of mountain, Kaohsiung Chinese caterpillar fungus (Coryceps takaomantana).Thin pin Isaria culture have immunoregulatory activity, antitumor, hypoglycemic, reduce cholesterol level in blood fat, the invasion and attack that suppress scytitis, prevention and treatment exogenous chemical substances and virus infection, anti-oxidant, improve the effects such as normal pressure hypoxia-bearing capability and calmness, analgesia.
The host of thin pin Isaria is lepidopterous larvae or pupa under field conditions (factors), although this bacterial classification is worldwide distribution, distributed quantity is also few, and a large amount of collection has difficulties.Artificial cultivation and the application study thereof of thin pin Isaria in China, Japan, Korea S, have been carried out.Chen Zhuan has reported that thin pin Isaria cultivates proterties (fungi journal, 1989,8 (3): 214-220).In Japan and Korea S, about the research of thin pin Isaria, more lay particular emphasis on and take silkworm or other insect and manually cultivate (Sang-Duk Ji.Mycobiology, 2011,39 (3): 158-163) as live body host.But the silkworm living or other insects source are subject to geography and season limit, and single silkworm is expensive, are not suitable for large-scale industry and cultivate, and therefore are also difficult to be widely used in the exploitation of downstream product.
Summary of the invention
The object of the invention is to, overcome the defect of prior art, disclose and can realize thin pin Isaria medium and the thin pin Isaria fruit body industrial method for culturing that thin pin Isaria fruit body is cultivated.The present invention passes through slant medium, shake-flask seed medium, fermentation tank seed culture medium is prepared respectively slant pore, first order seed, and secondary seed, take wheat or barley is major ingredient simultaneously, take dried silkworm chrysalis meal as auxiliary material, adopt material steaming tank boiling sterilization, secondary seed is entered to material steaming tank, by material steaming tank, rotate and mix, then adopt automatic packer to divide and install in cultivation box, by vibrated bed, medium is vibrated smooth again, overlay film sealing, move in culturing room and carry out solid fermentation cultivation, control temperature in cultivation process, illumination, humidity, the conditions such as ventilation, cultivate and obtain thin pin Isaria fruit body, realize boiling, sterilizing, cultivate integrated and automation, be applicable to the suitability for industrialized production of thin pin Isaria fruit body.
The industrial method for culturing that the invention discloses a kind of thin pin Isaria fruit body, concrete steps are as follows:
1) actication of culture: thin pin Isaria is inoculated on slant medium and is cultivated, obtain slant pore, collect slant pore, preparation spore liquid;
2) preparation of seed liquor:
A. the preparation of primary seed solution: the spore liquid access shake-flask seed medium of previous step is cultivated, obtained primary seed solution;
B. the preparation of secondary seed solution: by the primary seed solution access fermentation tank seed culture medium of previous step, fermented and cultured obtains secondary seed solution;
3) preparation of solid fermentation medium: solid culture based raw material is added in material steaming tank, sterilizing, cooling, obtain for cultivating the solid culture medium of thin pin Isaria fruit body;
4) by step 2) secondary seed solution prepared adds in the solid culture medium in step 3) material steaming tank, the mode of rotating by material steaming tank, solid culture medium is mixed with seed liquor, by automatic packer, divide and be filled to solid fermentation cultivation box, overlay film sealing, by vibrated bed, the medium vibration of cultivating in box is smooth;
5) solid fermentation: step 4) medium is vibrated to smooth cultivation box and move into culturing room, cultivate and obtain thin pin Isaria fruit body.
Preferably, described in step 1), slant medium formula is as follows:
In described culture medium prescription, the processing method of potato is potato boiling water boiling 30min, and filtered through gauze is got filtrate and mixed with other components; The processing method of cicada pupa is cicada pupa boiling water boiling 30min, and filtered through gauze is got filtrate and mixed with other components.
Preferably, the condition of culture of step 1) slant pore is: 23 ~ 27 ℃, and dark culturing 10 ~ 15 days.
Described spore liquid is to obtain with the slant pore on aqua sterilisa wash-out slant medium.
Preferably, described in step 1), the concentration of spore liquid is 106 ~ 107 spore/ml.
Preferably, step 2) described shake-flask seed culture medium prescription is as follows:
Potato 10～20wt%
Sucrose 1 ~ 2wt%
Water surplus.
Preferably, step 2) inoculum concentration of miospore liquid access shake-flask seed medium is 5 ~ 10v/v%.
More excellent, step 2) cultivation temperature of first order seed is 23 ~ 27 ℃, and rotating speed is 120 ~ 180rpm, cultivates 36 ~ 48h.
Preferably, step 2) formula of described fermentation tank seed culture medium is as follows:
Preferably, step 2), the inoculum concentration of first order seed access fermentation tank seed culture medium is 5 ~ 10v/v%.
Preferably, the cultivation temperature of secondary seed is 23 ~ 27 ℃, throughput 1.5 ~ 2.5vvm, and stir speed (S.S.) 200 ~ 300rpm, cultivates 48 ~ 72 hours.
Preferably, described in step 3), solid culture based formulas is: wheat and/or barley: dried silkworm chrysalis meal: the mass ratio of water is 1～1.5:0.05～0.3:1～2.5.
Preferably, solid culture medium sterilizing 40 ~ 60 minutes at 121 ℃, keeps material steaming tank rotation during sterilizing described in step 3), and cooling, the medium preparing is as the solid culture medium of cultivating thin pin Isaria fruit body.
Preferably, in step 4), the inoculum concentration of secondary seed solution access solid culture medium is 5 ~ 10v/w%.
More excellent, step 4) at room temperature, by the mode of rotation, mixes solid culture medium with mycelium.
More excellent, step 4) is divided and is filled to solid fermentation cultivation box, overlay film sealing by automatic packer.
More excellent, step 4) is smooth by the medium vibration of cultivating in box in vibrating bed vibration.
Preferably, to cultivate the temperature of fruit body be 23～27 ℃ to solid fermentation described in step 5), relative moisture 40～80%, and intensity of illumination 50～300Lux, ventilates once, cultivates 15～25 days for 8～12 hours.
Beneficial effect of the present invention is: the invention provides a kind of by a large amount of industrial culture methods of producing thin pin Isaria fruit body of artificial method of cultivating, by inclined-plane seed, shake-flask seed, fermentation tank seed expands cultivation, take wheat or barley as medium major ingredient, take dried silkworm chrysalis meal as medium auxiliary material, adopt material steaming tank boiling sterilization, secondary seed is entered to material steaming tank to be rotated and is mixed by material steaming tank, then adopt automatic packer to divide and install in cultivation box, by vibrated bed, medium is vibrated smooth again, overlay film sealing, move in culturing room, control temperature in cultivation process, illumination, humidity, the conditions such as ventilation, cultivate and obtain thin pin Isaria fruit body, can realize the suitability for industrialized production of thin pin Isaria, the yield of the thin pin Isaria fruit body that the inventive method is cultivated is that 15 ~ 20%(1Kg that often feeds intake can produce 150 ~ 200 grams of fruit bodys), 9% the output higher than prior art, adenosine content reaches 1.2mg/g, far above the 0.33mg/g(Hong In-Pyo of bibliographical information, Mycobiology, 2007,35 (4): 215-218), and, compare with the thin pin Isaria fruit body of cultivating with silkworm, the cultural method output of the thin pin Isaria of the present invention fruit body is high, superior product quality, and product price is relatively cheap, realize the heavy industrialization of thin pin Isaria fruit body and cultivated, there is good industry and medical use prospect.
Accompanying drawing explanation
Fig. 1: thin pin Isaria solid culture fruit body
Embodiment
Below by specific embodiment, further describe technical scheme of the present invention.Should be understood that these embodiment are only not used in and limit the scope of the invention for the present invention is described.
The industrialization of embodiment 1 thin pin Isaria fruit body is cultivated
Method one:
1. the preparation of medium:
The preparation of 1.1 slant mediums: take 100 grams of potatos, boiling water boiling 30 minutes, filtered through gauze obtains filtrate, with 20 grams of sucrose, is settled to 500ml; Take 100 grams of silkworm chrysalises, boiling water boiling 30 minutes, filtered through gauze obtains filtrate, is settled to 500ml; Two kinds of filtrates of preparation are mixed, add 20 grams, agar, dissolve, packing, sterilizing, cooling acquisition slant medium is standby.
1.2 shake-flask seed medium: take 200 grams of potatoes, boiling water boiling 30 minutes, filtered through gauze obtains filtrate, with 20 grams of sucrose, is settled to 1000ml, every bottled amount 200ml, sterilizing obtains shake-flask seed medium.
1.3 fermentation tank seed culture mediums: take peptone 450g, yeast extract 300g, maltose 450g, glucose 300g, be dissolved in 30L water, pH is adjusted to 7.0, sterilizing in 50 liters of fermentation tanks, obtains fermentation tank seed culture medium.
1.4 get wheat 50Kg, dried silkworm chrysalis meal 2.5Kg, water 125Kg, add cooker, and 121 ℃ of sterilizings 40 minutes are cooling, as the solid culture medium of cultivating fruit body.
2. the cultural method of fruit body
1) actication of culture: by thin pin Isaria bacterium access slant medium, 23 ℃ of dark culturing 15 days, with aqua sterilisa by spore from inclined-plane wash-out, preparation concentration is 10
6~ 10
7the spore liquid of individual spore/ml;
2) preparation of primary seed solution: spore liquid is accessed to shake-flask seed medium according to the inoculum concentration of 10v/v%, shaking flask condition of culture, 180rpm, cultivates 48 hours for 23 ℃, obtains primary seed solution.
3) preparation of secondary seed solution: according to 10v/v% inoculum concentration, primary seed solution is accessed in fermentation tank to fermentation condition: 23 ℃, throughput 1.5vvm, stir speed (S.S.) 200rpm, cultivates 72 hours, obtains secondary seed solution.
4) the mixing of seed liquor and solid culture medium, packing and vibration: secondary seed is entered in material steaming tank by 10v/w% inoculum concentration, the mode of rotating by material steaming tank, solid culture medium is mixed with mycelium, by automatic packer, divide and be filled to solid fermentation cultivation box, overlay film sealing; Then by vibrated bed, the medium vibration of cultivating in box is smooth.
5) solid fermentation: will cultivate box and move into culturing room's cultivation, 23～25 ℃ of cultivation temperature, relative moisture 40～50%, intensity of illumination 50 ~ 200Lux, ventilates once, cultivates the fruit body that can obtain artificial cultivation for 25 days for every 12 hours.
6) fruit body is gathered: culture is poured into harvester, and fruit body is separated with residual media, and 40～50 ℃ of oven dry obtain dry fruit body.The output of cultured products is thin pin Isaria fruit body dry weight 160g ± 5g/1kg solid culture medium.
Method two
1. the preparation of medium:
The preparation of 1.1 slant mediums: take 150 grams of potatos, boiling water boiling 30 minutes, filtered through gauze obtains filtrate; Take 75 grams of silkworm chrysalises, boiling water boiling 30 minutes, filtered through gauze obtains filtrate; Potato filtrate and silkworm chrysalis filtrate are mixed, with 20 grams of sucrose, be settled to 1000ml; Add 15 grams, agar, dissolve, packing, sterilizing, cooling acquisition slant medium is standby.
1.2 shake-flask seed medium: take 150 grams of potatoes, boiling water boiling 30 minutes, filtered through gauze obtains filtrate, with 10 grams of sucrose, is settled to 1000ml, every bottled amount 200ml, sterilizing obtains shake-flask seed medium.
1.3 fermentation tank seed culture mediums: take peptone 300g, yeast extract 600g, maltose 900g, glucose 450g, be dissolved in 30L water, pH is adjusted to 6.0, sterilizing in 50 liters of fermentation tanks, obtains fermentation tank seed culture medium.
1.4 get wheat 150Kg, dried silkworm chrysalis meal 30Kg, water 100Kg, add cooker, and 121 ℃ of sterilizings 50 minutes are cooling, as the solid culture medium of cultivating fruit body.
2. the cultural method of fruit body
1) actication of culture: by thin pin Isaria bacterium access slant medium, 25 ℃ of dark culturing 10 days, with aqua sterilisa by spore from inclined-plane wash-out, preparation concentration is 10
6~ 10
7the spore liquid of individual spore/ml;
2) preparation of primary seed solution: spore liquid is accessed to shake-flask seed medium according to the inoculum concentration of 5v/v%, shaking flask condition of culture, 150rpm, cultivates 40 hours for 25 ℃, obtains primary seed solution.
3) preparation of secondary seed solution: according to 5v/v% inoculum concentration, primary seed solution is accessed in fermentation tank to fermentation condition: 25 ℃, throughput 2.5vvm, stir speed (S.S.) 250rpm, cultivates 48 hours, obtains secondary seed solution.
4) the mixing of seed liquor and solid culture medium, packing and vibration: secondary seed is entered in material steaming tank by 7.5v/w% inoculum concentration, the mode of rotating by material steaming tank, solid culture medium is mixed with mycelium, by automatic packer, divide and be filled to solid fermentation cultivation box, overlay film sealing; Then by vibrated bed, the medium vibration of cultivating in box is smooth.
5) solid fermentation: will cultivate box, and move into culturing room and cultivate, 24～26 ℃ of cultivation temperature, relative moisture 70～80%, intensity of illumination 200～300Lux, ventilates once, cultivates the fruit body that can obtain artificial cultivation for 15 days for every 10 hours.
6) fruit body of gathering: culture is poured into harvester, and fruit body is separated with residual media, and 40～50 ℃ of oven dry obtain dry fruit body.The output of cultured products is: thin pin Isaria fruit body dry weight 200g ± 4g/1kg solid culture medium.
Method three
1. the preparation of medium:
The preparation of 1.1 slant mediums: take 200 grams of potatos, boiling water boiling 30 minutes, filtered through gauze obtains filtrate; Take 50 grams of silkworm chrysalises, boiling water boiling 30 minutes, filtered through gauze obtains filtrate; Potato filtrate and silkworm chrysalis filtrate are mixed, with 10 grams of sucrose, be settled to 1000ml; Add 15 grams, agar, dissolve, packing, sterilizing, cooling acquisition slant medium is standby.
1.2 shake-flask seed medium: take 100 grams of potatoes, boiling water boiling 30 minutes, filtered through gauze obtains filtrate, with 15 grams of sucrose, is settled to 1000ml, every bottled amount 200ml, sterilizing obtains shake-flask seed medium.
1.3 fermentation tank seed culture mediums: take peptone 900g, yeast extract 450g, maltose 300g, glucose 900g, be dissolved in 30L water, pH is adjusted to 6.5, sterilizing in 50 liters of fermentation tanks, obtains fermentation tank seed culture medium.
1.4 get wheat 125Kg, dried silkworm chrysalis meal 37.5Kg, water 100Kg, add cooker, and 121 ℃ of sterilizings 60 minutes are cooling, as the solid culture medium of cultivating fruit body.
2. the cultural method of fruit body
1) actication of culture: by thin pin Isaria bacterium access slant medium, 27 ℃ of dark culturing 13 days, with aqua sterilisa by spore from inclined-plane wash-out, preparation concentration is 10
6~ 10
7the spore liquid of individual spore/ml;
2) preparation of primary seed solution: spore liquid is accessed to shake-flask seed medium according to the inoculum concentration of 7.5v/v%, shaking flask condition of culture, 120rpm, cultivates 36 hours for 27 ℃, obtains primary seed solution.
3) preparation of secondary seed solution: according to 7.5v/v% inoculum concentration, primary seed solution is accessed in fermentation tank to fermentation condition: 27 ℃, throughput 2.0vvm, stir speed (S.S.) 300rpm, cultivates 60 hours, obtains secondary seed solution.
4) the mixing of seed liquor and solid culture medium, packing and vibration: secondary seed is entered in material steaming tank by 5v/w% inoculum concentration, the mode of rotating by material steaming tank, solid culture medium is mixed with mycelium, by automatic packer, divide and be filled to solid fermentation cultivation box, overlay film sealing; Then by vibrated bed, the medium vibration of cultivating in box is smooth.
5) solid fermentation: will cultivate box, and move into culturing room and cultivate, 25～27 ℃ of cultivation temperature, relative moisture 40～80%, intensity of illumination 100～250Lux, ventilates once, cultivates the fruit body that can obtain artificial cultivation for 20 days for every 8 hours.
6) fruit body of gathering: culture is poured into harvester, fruit body is separated with residual media, dry and obtain dry fruit body.The present embodiment is cultivated the fruit body of gained and is seen Fig. 1, and the output of cultured products is: thin pin Isaria fruit body dry weight 180g ± 5.5g/1kg solid culture medium.
Embodiment 2
Active ingredient to the artificial cultured products of thin pin Paecilomyces varioti: adenosine and cordycepic acid content detect, sample is taken from embodiment 1 method three.
(1) Determination of Adenosine
Adenosine is measured with reference to the method for version Chinese pharmacopoeia in 2010 and is carried out.
Instrument and reagent: Waters high performance liquid chromatograph (1525BINARY HPLC PUMP, 2998 Photodiode Array Detector, U.S. Waters company); Acetonitrile (HPLC level, Fisher); Potassium dihydrogen phosphate (ARJi， Chemical Reagent Co., Ltd., Sinopharm Group); Benzinum (AR level, 60-90 ℃， Chemical Reagent Co., Ltd., Sinopharm Group); Adenosine (A9251-1G, Sigma company).Chromatographic condition, chromatographic column: XBridge C18 chromatographic column (Waters, 4.6mm * 250mm, 5 μ m); Mobile phase: acetonitrile-0.04mol/L potassium dihydrogen phosphate (5:95); Flow velocity: 1.0mL/min; Detect wavelength: 260nm; Column temperature: 35 ℃; Sample size: 20 μ L.
(2) cordycepic acid content is measured
Instrument and reagent:
Ultraviolet-uisible spectrophotometer; Electronic balance; Centrifuge H-1650; Electric heat constant temp. water tank CU600 type; Adjustable closed electric furnace.Mannitol standard items, ammonium acetate, acetylacetone,2,4-pentanedione, glacial acetic acid, potassium metaperiodate, L-rhamnose, be ARJi, Chemical Reagent Co., Ltd., Sinopharm Group and produce.
Reagent preparation:
Potassium metaperiodate solution: 15mmol (being 3.45g) potassium metaperiodate is dissolved in 1L0.12mol/L hydrochloric acid solution; Nash reagent: 150g ammonium acetate+2mL glacial acetic acid+2mL acetylacetone,2,4-pentanedione, with distilled water diluting to 1L (matching while using); L-rhamnose solution: L-rhamnose 100mg, is settled to 100mL with distilled water.
Accurately take 1.0034g sample, be placed in dry 500mL conical flask, record the gross mass m of conical flask and sample
1, in conical flask, add boiling distillated water 100mL, be positioned on electric furnace and make after its constantly boiling 15min, be put in rapidly in cold water and be cooled to room temperature, take out, dry the globule of bottle outer wall, adding distilled water to final mass is m
1+ 100g, shakes up, and filters, and gets filtrate, is testing sample ,-20 ℃ of preservations.
Precision takes PEARLITOL 25C standard items 0.1g in beaker, add a small amount of distilled water dissolve complete, be transferred to constant volume in 100mL volumetric flask, be mixed with the mannitol solution of 1mg/mL, after diluting, obtain that mass concentration is respectively 10,50,90,130, the mannitol standard liquid of 170mg/L.Get above-mentioned each 1mL of concentration standard product mannitol solution, split in different test tubes, then add respectively 1mL sodium periodate solution, mix, room temperature is placed 10min, add 2mL0.1%L-rhamnose solution to remove too much periodate, vibration mixes, and adds the freshly prepared Nash reagent of 4mL, and 53 ℃ of heating water bath 15min make its colour developing, be quickly cooled to room temperature, at 412nm wavelength, place measures its absorbance.With distilled water, replace mannitol standard liquid, use the same method operation in contrast, measure its absorbance.Take concentration of standard solution as abscissa, and absorbance is ordinate, and drawing standard curve, obtains regression equation.
By 8 times of dilutions of test sample extract adding distil water, the sample of getting after 1ml dilution is placed in tool plug test tube, measure light absorption value, every pipe sample is surveyed 3 Abs, obtain mean value, by regression equation, obtain liquid cordycepic acid concentration c to be measured, by the long-pending v of known tracer liquid, sample quality m, obtains test sample cordycepic acid content (mg/g) according to following formula.
Result shows, adenosine calibration curve, adenosine calibration curve y=7.08e+004x-1.05e+004, R
2=0.999978, good at 1～100 μ g/mL and peak area linear relation.
Cordycepic acid calibration curve, the standard items mannitol concentration (mg/mL) of take is abscissa, Abs is ordinate, obtains regression equation: y=0.0826x+0.0019, R
2=0.9998, description standard is savored mannitol amount within the scope of 0～21.25mg/L, is good linear relation with Abs.
In the thin artificial cultured products of pin Paecilomyces varioti, adenosine content reaches 1.2mg/g as calculated, and cordycepic acid content reaches 35.0mg/g.
Compare with the thin pin Isaria fruit body of cultivating with silkworm, thin pin Isaria fruit body of the present invention can be carried out large-scale industrialization cultivation under manual control condition, the high and quality better of output, and product price is relatively cheap, has good prospects for commercial application.
Claims (10)
1. an industrial method for culturing for thin pin Isaria fruit body, concrete steps are as follows:
1) actication of culture: thin pin Isaria is inoculated on slant medium and is cultivated, obtain slant pore, collect slant pore, preparation spore liquid;
2) preparation of seed liquor:
A. the preparation of primary seed solution: the spore liquid access shake-flask seed medium of previous step is cultivated, obtained primary seed solution;
B. the preparation of secondary seed solution: by the primary seed solution access fermentation tank seed culture medium of previous step, fermented and cultured obtains secondary seed solution;
3) preparation of solid fermentation medium: solid culture based raw material is added in material steaming tank, sterilizing, cooling, obtain for cultivating the solid culture medium of thin pin Isaria fruit body;
4) by step 2) secondary seed solution prepared adds in the solid culture medium in step 3) material steaming tank, the mode of rotating by material steaming tank, solid culture medium is mixed with seed liquor, by automatic packer, divide and be filled to solid fermentation cultivation box, overlay film sealing, by vibrated bed, the medium vibration of cultivating in box is smooth;
5) solid fermentation: step 4) medium is vibrated to smooth cultivation box and move into culturing room, cultivate and obtain thin pin Isaria fruit body.
2. cultural method as claimed in claim 1, is characterized in that, slant medium formula is as follows described in step 1):
3. cultural method as claimed in claim 1, is characterized in that, the condition of culture of step 1) slant pore is: 23～27 ℃, and dark culturing 10～15 days.
4. cultural method as claimed in claim 1, is characterized in that step 2) described shake-flask seed culture medium prescription is as follows:
Potato 10～20wt%
Sucrose 1～2wt%
Water surplus.
5. cultural method as claimed in claim 1, is characterized in that step 2) cultivation temperature of described first order seed is 23～27 ℃, rotating speed is 120～180rpm, cultivates 36～48h.
6. cultural method as claimed in claim 1, is characterized in that step 2) described fermentation tank seed culture based formulas is as follows:
7. cultural method as claimed in claim 1, is characterized in that step 2) cultivation temperature of described secondary seed is 23～27 ℃, throughput 1.5～2.5vvm, stir speed (S.S.) 200～300rpm, cultivates 48～72 hours.
8. cultural method as claimed in claim 1, is characterized in that, solid culture based formulas is described in step 3): wheat and/or barley: dried silkworm chrysalis meal: the mass ratio of water is 1～1.5:0.05～0.3:1～2.5.
9. cultural method as claimed in claim 1, is characterized in that, the condition of sterilizing is sterilizing 40 ~ 60 minutes at 121 ℃ described in step 3).
10. cultural method as claimed in claim 1, is characterized in that, to cultivate the temperature of fruit body be 23～27 ℃ to solid fermentation described in step 5), relative moisture 40～80%, and intensity of illumination 50～300Lux, ventilates once, cultivates 15～25 days for 8～12 hours.
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| CN107475130A (en) * | 2017-09-19 | 2017-12-15 | 广东省微生物研究所 | Thin handle Isaria novel bacterial and its cultural method and purposes |
| CN109266557A (en) * | 2018-10-17 | 2019-01-25 | 上海市农业科学院 | The preparation method of a kind of thin foot Isaria cordyceps sinensis aggregate species strain with health role |
| CN112314330A (en) * | 2020-11-13 | 2021-02-05 | 浙江泛亚生物医药股份有限公司 | Method for culturing Roberts' cordyceps sinensis |
| CN115039633A (en) * | 2021-03-09 | 2022-09-13 | 鲁东大学 | Artificial culture method for sporocarp of Isaria japonica |
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Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1221031A (en) * | 1998-08-26 | 1999-06-30 | 刘放 | Continuous transfer new technology for microbe industrial fermentation |
| CN1563348A (en) * | 2004-04-16 | 2005-01-12 | 浙江省农业科学院 | New strain APC-20 of Paecilomyces cicadae and fementation process for artificial culture |
| KR20050004992A (en) * | 2003-07-01 | 2005-01-13 | 한영환 | Methods of the mycelial culture products of mushrooms using desalted deep seawater for increasing the extracellular glycoprotein |
| KR20060022402A (en) * | 2004-09-07 | 2006-03-10 | 까치마을영농조합법인 | Artificially cultivation method of paecilomyces tenuipes use protaetia orientalis larva |
| JP2006141392A (en) * | 2004-10-21 | 2006-06-08 | Sumitomo Chemical Co Ltd | Method for producing conidium of microorganism belonging to paecilomyces |
| CN101195805A (en) * | 2007-11-01 | 2008-06-11 | 华富生物科技(上海)有限公司 | Cordyceps sinensis epiphyte and artificial cultivation method thereof |
| CN102242154A (en) * | 2010-05-14 | 2011-11-16 | 浙江泛亚生物医药股份有限公司 | Liquid fermentation method for producing paecilomyces cicadae mycelia and application of culture product |
| CN102533567A (en) * | 2011-12-20 | 2012-07-04 | 吉林大学 | Paecilomyces tenuipes mutagenesis bacterial strain and culturing method thereof |
-
2012
- 2012-08-22 CN CN201210300985.0A patent/CN103621308B/en active Active
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1221031A (en) * | 1998-08-26 | 1999-06-30 | 刘放 | Continuous transfer new technology for microbe industrial fermentation |
| KR20050004992A (en) * | 2003-07-01 | 2005-01-13 | 한영환 | Methods of the mycelial culture products of mushrooms using desalted deep seawater for increasing the extracellular glycoprotein |
| CN1563348A (en) * | 2004-04-16 | 2005-01-12 | 浙江省农业科学院 | New strain APC-20 of Paecilomyces cicadae and fementation process for artificial culture |
| KR20060022402A (en) * | 2004-09-07 | 2006-03-10 | 까치마을영농조합법인 | Artificially cultivation method of paecilomyces tenuipes use protaetia orientalis larva |
| JP2006141392A (en) * | 2004-10-21 | 2006-06-08 | Sumitomo Chemical Co Ltd | Method for producing conidium of microorganism belonging to paecilomyces |
| CN101195805A (en) * | 2007-11-01 | 2008-06-11 | 华富生物科技(上海)有限公司 | Cordyceps sinensis epiphyte and artificial cultivation method thereof |
| CN102242154A (en) * | 2010-05-14 | 2011-11-16 | 浙江泛亚生物医药股份有限公司 | Liquid fermentation method for producing paecilomyces cicadae mycelia and application of culture product |
| CN102533567A (en) * | 2011-12-20 | 2012-07-04 | 吉林大学 | Paecilomyces tenuipes mutagenesis bacterial strain and culturing method thereof |
Non-Patent Citations (3)
| Title |
|---|
| 蒲顺昌等: "高雄山虫草无性型-细脚拟青霉原生质体制备及其再生的初步研究", 《安徽农业大学学报》 * |
| 陈祝安等: "细脚拟青霉培养性状和药理作用的初步研究", 《菌物学报》 * |
| 韩燕峰等: "高雄山虫草人工栽培菌株筛选及搔菌对其子实体生物量的影响", 《食用菌学报》 * |
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| CN105985150A (en) * | 2015-01-28 | 2016-10-05 | 浙江泛亚生物医药股份有限公司 | Isaria cicadae coremium liquid culture medium |
| CN105985150B (en) * | 2015-01-28 | 2020-10-23 | 浙江泛亚生物医药股份有限公司 | Cordyceps sobolifera sporostalk bundle liquid culture medium |
| CN107475130A (en) * | 2017-09-19 | 2017-12-15 | 广东省微生物研究所 | Thin handle Isaria novel bacterial and its cultural method and purposes |
| CN107475130B (en) * | 2017-09-19 | 2019-09-17 | 广东省微生物研究所 | Thin handle Isaria novel bacterial and its cultural method and purposes |
| CN109266557A (en) * | 2018-10-17 | 2019-01-25 | 上海市农业科学院 | The preparation method of a kind of thin foot Isaria cordyceps sinensis aggregate species strain with health role |
| CN112314330A (en) * | 2020-11-13 | 2021-02-05 | 浙江泛亚生物医药股份有限公司 | Method for culturing Roberts' cordyceps sinensis |
| WO2022100701A1 (en) * | 2020-11-13 | 2022-05-19 | 浙江泛亚生物医药股份有限公司 | Method for cultivating ophiocordyceps robertsii |
| CN115039633A (en) * | 2021-03-09 | 2022-09-13 | 鲁东大学 | Artificial culture method for sporocarp of Isaria japonica |
| CN115044477A (en) * | 2021-03-09 | 2022-09-13 | 张清洋 | Isaria japonica strain and application thereof |
| CN115044477B (en) * | 2021-03-09 | 2023-10-13 | 张清洋 | Isaria japonica strain and application thereof |
| CN115039633B (en) * | 2021-03-09 | 2024-05-31 | 鲁东大学 | Artificial culture method for sporophore of Isaria japonica |
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