CN103636408A - Factory-like production method of silkworm cordyceps - Google Patents
Factory-like production method of silkworm cordyceps Download PDFInfo
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- CN103636408A CN103636408A CN201310698778.XA CN201310698778A CN103636408A CN 103636408 A CN103636408 A CN 103636408A CN 201310698778 A CN201310698778 A CN 201310698778A CN 103636408 A CN103636408 A CN 103636408A
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Abstract
The invention provides a factory-like production method of silkworm cordyceps. The method is characterized by comprising the steps as follows: separating fruiting bodies of cordyceps militaris, sterilizing and washing the fruiting bodies, putting the fruiting bodies on a slant culture medium, and placing the slant culture medium in an incubator for culture; placing a PDA (potato dextrose agar) culture medium in a shake flask, placing white mycelia in the shake flask for shake culture, inoculating the white mycelia on a rice culture medium, after the white mycelia grow over the rice culture medium, taking out the mycelia, adding a glucose liquid, mashing the mycelia, and obtaining a nutrition fungus suspension as an inoculation fluid; stabbing and injecting pleopod parts of silkworm larvae with a medical injector sucked with the inoculation liquid to obtain the inoculated silkworm larvae, obliquely placing the silkworm larvae on the sterilized rice culture medium, performing light-shielding variable-temperature culture on the silkworm larvae firstly, and performing light culture after white hyphae grow over the larvae; and collecting the silkworm cordyceps. The factory-like production method is simple in technology, the mass production can be realized, and the silkworm cordyceps with shapes similar to natural cordyceps sinensis and medium values basically identical with the cordyceps sinensis can be produced.
Description
Technical field
The present invention is relevant with the cultivation of Cordyceps sinensis indoors artificial, is that a kind of silkworm larvae body that utilizes goes out the silkworm Chinese caterpillar fungus factory production process of aweto in indoor production.The product that this explained hereafter goes out is the conjugant of polypide (silkworm larvae shape) and cursive script (fruit body), and, main component similar to the Cordyceps sinensis body of self-sow and medical value are basic identical.
Background technology
Publication number is that the Chinese patent of CN103262751A discloses a kind of method of utilizing Cordyceps militaris spawn infected silkworm larva to cultivate silkworm Chinese caterpillar fungus, and it comprises the steps: the selection of (1) silkworm: the silkworm of selection 3rd～5d in five ages is as host; (2) bacterial classification preparation: Cordyceps militaris spawn is inoculated in liquid nutrient medium, after concussion is cultivated, obtains liquid spawn after sterile gauze filtration sterilization ball; (3) preparation of Nutrition Soil: preparation water content 5%～20% soil, after autoclave sterilization as Nutrition Soil; (4) inoculation: silkworm is put into sterilizing 20～50min under ultraviolet, then inject the liquid spawn of 0.2～0.4ml in Silkworm, Bombyx mori; (5) cultivate management: postvaccinal silkworm is put into Nutrition Soil, be placed under 5 ℃～12 ℃ environment lucifuge and cultivate 5～12d, after to proceed to light intensity be 50～200lx, temperature is 21 ℃～23 ℃, humidity is to cultivate 35～60d under 86%～89% condition, obtains silkworm Chinese caterpillar fungus.The weak point of the method is, 1) silkworm is put into the silkworm pre-treatment time of sterilizing 20～50min under ultraviolet oversize; 2) do not have the liquid spawn of use 0.2～0.4ml with good conditionsi to be expelled in Silkworm, Bombyx mori, will make Silkworm, Bombyx mori flow out body fluid; 3) postvaccinal silkworm is put into Nutrition Soil, be placed under 5 ℃～12 ℃ environment lucifuge and cultivate, will make Silkworm, Bombyx mori flow out more body fluid and the skin of remaining silkworm body only.This patent is difficult to obtain silkworm Chinese caterpillar fungus.
Publication number is that the Chinese patent of CN1304995 discloses a kind of method (application number 01103275.8 of producing Cordyceps sinensis of cultivating with silkworm, Lian Gen etc. during inventor), this disclosure of the invention is to cultivate the method for producing Cordyceps sinensis with silkworm (B ombyx mori L.).It is aweto to be done to, after physics and chemistry biological processes, be inoculated in Silkworm, Bombyx mori, then simulates natural surroundings, makes aweto infect parasitic Silkworm, Bombyx mori and finally grow stroma, cultivates aweto.This patent weak point is, aweto is done after physics and chemistry biological processes, and the bacterial classification that is inoculated in Silkworm, Bombyx mori is difficult at silkworm tumor growth, and success rate is extremely low, can not form batch production.
The weak point of above-mentioned patent is do not have real investigation not take corresponding measure to the growth key point of aweto, so can not mass-produce aweto.
Summary of the invention
The object of the invention is to utilize and in silkworm larvae Ti factory, manually produce, medical value essentially identical silkworm Chinese caterpillar fungus factory production process similar to natural Cordyceps sinensis body.
In order to achieve the above object, the invention provides a kind of silkworm Chinese caterpillar fungus industrial production method, it is characterized in that, comprise the steps:
The first step: Workplace is carried out disinfection; By live body silkworm sterilization afterflush, dry, standby; Preparation PDA medium and rice medium;
Second step: the fruit body of separated Cordyceps militaris, after fruit body sterilization, rinsing, chopping, sets aside on slant medium, is put into and in incubator, cultivates 15-25 days;
The 3rd step: PDA medium is placed in to shaking flask, puts white hypha body in shaking flask concussion and cultivate, after 4-8 days, white hypha body is inoculated on rice medium, after covering with white hypha body, takes out mycelium and add Glucose Liquid, mycelia is smashed to pieces, obtained dietetic bacterial suspension as inoculation liquid;
The 4th step: with inhaling the abdominal foot portion that has the injector for medical purpose of inoculation liquid thorn note silkworm larvae body, the silkworm larvae body that obtains having inoculated, silkworm larvae body is tiltedly placed on the rice medium that sterilizing is good, first carried out lucifuge Fluctuation temperature culture, treated that pure white mycelia covers with polypide, seen that light cultivates, treat white hypha flavescence, proceed to see that light cultivates, treat that fruit body grows to 5～8 centimeters, fruit body surface is maturation before having yellow powder to occur;
The 5th step: gather, be emitted in dish airing or dry, will dry in 50～55 ℃ of temperature, make polypide water content not higher than 10%, to prevent moldy metamorphism; With bag film sealing, packing is preserved, or is positioned in the container of dry sealing tight building.
Preferably, in described the 4th step, treat white hypha flavescence, sand is covered on silkworm larvae body, a mouth that only exposes worm, proceeds to see light cultivation, in the 5th step, while gathering, along fruit body, with tweezers, push sand aside, take out polypide and fruit body, with clear water, wash away the sand on Chinese caterpillar fungus.
Preferably, in the described first step, by live body silkworm sterilization afterflush, dry, standby concrete steps are: the live body silkworm of 4 ± 2 days after choosing the 5th and casting off a skin, under gnotobasis, with 75% ethanolic solution erasing polypide, carry out disinfection, the polypide of sterilizing with aseptic water washing, under gnotobasis, dry polypide, the silkworm larvae body drying saves backup at-2 ℃ ± 1 ℃ low temperature.
Preferably, PDA medium in the described first step is mixed to get by sterile distilled water, peeling potato, glucose and agar, and the ratio of described sterile distilled water, peeling potato, glucose and agar is 1000mL: 150-250g: 10-30g: 10-30g.
Preferably, the rice medium in the described first step is obtained by rice and water mixing sterilizing, and described rice and the ratio of water are 30-40 gram: 50ml.
Preferably, the compound method of the slant medium in described second step is: by peeling potatoes extractive, add agar in potato juice, add water to 1000mL, be respectively charged in test tube, tampon, after sterilizing, is cooled to 60 ℃ beyond the Great Wall, be placed on inclined-plane, wherein, the weight content of potato juice is 20%, and the weight content of agar is 1.7%.
Preferably, the temperature of the incubator in described second step is 20-30 ℃.
Preferably, the temperature that in described the 3rd step, concussion is cultivated is 15-25 ℃.
Preferably, the concrete steps of the lucifuge Fluctuation temperature culture in described the 4th step are: in dark surrounds, be first placed under the condition of 0 ± 4 ℃ of temperature, humidity 85% and cultivate more than 10 days, then cultivate 10-20 days in 10 ℃ ± 2 ℃ environment.
Preferably, the concrete steps that light is cultivated of seeing in described the 4th step are: lucifuge Fluctuation temperature culture is covered with to the polypide of mycelia after 5 days, at illuminance 1000lx, at 8～10 hours every day, in 20 ℃ ± 2 ℃ of temperature, humidity 65% environment, cultivate.
Preferably, in described the 4th step, proceed to see that when light is cultivated, illuminance is 150-1500lx, every day 6-8 hour, incubation time is 15-25 days.
Compared with prior art, the invention has the beneficial effects as follows:
Technique of the present invention is simple, can large-scale production, and can produce, medical value essentially identical silkworm Chinese caterpillar fungus similar to natural Cordyceps sinensis body.
Embodiment
Below in conjunction with embodiment, illustrate the present invention.
Embodiment 1
Workplace is keeped in repair to arrangement, water supply and drainage system is cleared up to sterilization, maintenance and inspection Circuits System, cleaning ventilating system, keeps in repair arrangement to culturing rack, use equipment etc., pasteurised completely, inoculation Fang He culturing room disinfection standard is the highest.
Prepare silkworm larvae body: select the healthy and strong live body silkworm that the 5th is casted off a skin latter 5 days; Under gnotobasis, first use 75% ethanolic solution erasing polypide, carry out disinfection; The polypide of sterilizing with aseptic water washing again; Then under gnotobasis, dry polypide; The polypide of drying is preserved in-2 ℃ of low temperature, standby.
Prepare PDA medium: clean the potato of 200g peeling, add sterile distilled water 1000mL, add glucose 20g, add agar 20g, dipping, after sterilization, cooling standby.
Prepare rice medium: clean 36 grams of rice, join in 380ml jam jar, add 50ml water simultaneously, lid lid, after conventional autoclaving, cooling standby.
Prepare inoculation liquid: commercially available Cordyceps militaris (purchased from Xuzhou Hong Yu agricultural science and technology Co., Ltd, one-level (mulberry) silkworm chrysalis Cordyceps sinensis) is isolated to fruit body; With 75% ethanolic solution, flood cordyceps militaris sporocarp 5 minutes, then with sterile water, repeatedly rinse 3 times; Dry, with scalpel, fruit body is cut into grain of rice size; (its compound method is by peeling potatoes extractive to set aside to 45 slant mediums of spending, in potato juice, add agar, add water to 1000mL, be respectively charged in test tube, beyond the Great Wall tampon, after sterilizing, be cooled to 60 ℃, be placed on inclined-plane, wherein, the weight content of potato juice is 20%, and the weight content of agar is 1.7%) on; Put it in 26 ℃ of incubators and cultivate 20 days; PDA medium is placed in to shaking flask, and extracting waste mycelium 2cmX2cm puts in the shaking flask of PDA liquid nutrient medium 65ml, at 20 ℃, shakes 6 days continuously; Take off shaking flask, with moving liquid instrument, draw 0.1ml concussion liquid, be inoculated on rice medium, treat that white hypha body covers with; Taking-up mycelia 10ml adds 20% Glucose Liquid 100ml, with tissue mashing machine, mycelia is fully smashed to pieces; Make dietetic bacterial suspension and be placed in refrigerator and preserve as inoculation liquid, stand-by.
Prepare sterilization sand: choose thin river sand, requiring humidity is between 50-60%, carries out conventional sterilization, cooling standby.
Prepare other material: Dispoable medical syringe, black paper and so on article etc., and carry out disinfection, standby.
Each place, various material are prepared to disinfect, and carry out inoculated and cultured.With the syringe of preparing, draw the abdominal foot portion of the silkworm larvae body of 0.01ml inoculation liquid thorn note preparation; The silkworm larvae of having inoculated angle at 45 ° is neatly tiltedly placed on ready rice medium, adds a cover and cultivate, first covered ready black paper, be placed in the environment of temperature-2 ℃, humidity 85%, cultivate 10 days, then cultivate 15 days in 10 ℃ of environment, treat that pure white mycelia covers with polypide; Then in 20 ℃ of temperature, humidity 65% environment, adopt illuminance 1000lx, see every day under the light condition of 9 hours and cultivate, mycelia to be seen is light yellow by white annesl, just the sand of preparation is covered to polypide upper, only exposes Tou Hezui.Proceed to see that light cultivates, a few days ago luminous intensity is strong, the time is long, later gradually a little less than, the time is shorter, cultivates 20 days, the head of worm may have grown into the tangerine look point of fruit body after growing as seen.Continue to cultivate.When fruit body is grown to 8 centimetres of surfaces and will be had yellow powder to occur, gather in time.Gather and will with tweezers, push gently sand aside along fruit body, dig out together with worm band grass, take care to break.Taking-up silkworm Chinese caterpillar fungus is washed away after sand with clear water, be emitted in the dish of preparation and carry out airing.For ensuring the quality of products, the Chinese caterpillar fungus of drying is dried in 50 ℃ of temperature, make polypide water content not higher than 10%, to prevent moldy metamorphism.For founding brand, product will carry out classification, carry out respectively cordycepin, Cordyceps sinensis polysaccharide, cordycepic acid, protein, the vitamin of silkworm Chinese caterpillar fungus, the mensuration of organized enzyme equal size.After mensuration, according to different product data, carry out classification, pack respectively label.Be positioned in the dry container sealing and preserve.
According to technique of the present invention, the silkworm Chinese caterpillar fungus of can batch production producing, similar to natural Cordyceps sinensis body, medical value is basic identical.The cordycepin content of product of the present invention is 0.92%, and the content of Cordyceps sinensis polysaccharide is 7.6%, and the content of the cordycepin of natural Cordyceps sinensis is 0.93%, and the content of Cordyceps sinensis polysaccharide is 5.3%.
Embodiment 2
Workplace is keeped in repair to arrangement, identical with embodiment 1.
Prepare silkworm larvae body, prepare PDA nutrient solution cultivation base, prepare rice medium, preparation inoculation liquid and prepare other material all identical with embodiment 1.Just do not prepare the sand of sterilizing.Concrete steps are as follows:
Each place, various material are prepared to disinfect, and carry out inoculated and cultured.With the syringe of preparing, draw the abdominal foot portion of the silkworm larvae body of 0.01ml inoculation liquid thorn note preparation; The silkworm larvae of having inoculated angle at 45 ° is neatly tiltedly placed on ready rice medium, adds a cover and cultivate.First covered ready black paper, be placed in the environment of temperature-2 ℃, humidity 85%, cultivate 10 days, then cultivate 15 days in 10 ℃ of environment, treated that pure white mycelia covers with polypide; Then in 20 ℃ of temperature, humidity 65% environment, adopt illuminance 1000lx, see every day under the light condition of 9 hours and cultivate, mycelia to be seen is light yellow by white annesl, proceeds to see light cultivation.A few days ago luminous intensity is strong, the time long, later gradually a little less than, the time is shorter, cultivates 20 days, each minister of worm may have grown into a plurality of tangerine look points of fruit body after going out as seen.Continue to cultivate.When fruit body is grown to 8 centimetres of surfaces and will be had yellow powder to occur, gather in time.Gathering will connect worm band grass and together trip out, and takes care to break.By taking out silkworm Chinese caterpillar fungus and be emitted in the dish of preparation, carry out airing or dry.For ensuring the quality of products, the Chinese caterpillar fungus of airing is dried in 50 ℃ of temperature, make polypide water content not higher than 10%.Then be positioned in the dry container sealing and preserve, to prevent moldy metamorphism.
According to this technique, the silkworm Chinese caterpillar fungus of can batch production producing, although body and natural Cordyceps sinensis have bigger difference, fruit body (grass) president is at each position of a silkworm Chinese caterpillar fungus, medical value difference is little.The cordycepin content of product of the present invention is 0.91%, and the content of Cordyceps sinensis polysaccharide is 7.7%, and the content of the cordycepin of natural Cordyceps sinensis is 0.93%, and the content of Cordyceps sinensis polysaccharide is 5.3%.
In sum, advantage of the present invention is: according to technique of the present invention, can batch production produce, medical value essentially identical silkworm Chinese caterpillar fungus similar to natural Cordyceps sinensis body; Also can batch production produce with natural Cordyceps sinensis medical value is basic identical but the silkworm Chinese caterpillar fungus that body differs greatly.
Claims (9)
1. a silkworm Chinese caterpillar fungus industrial production method, is characterized in that, comprises the steps:
The first step: Workplace is carried out disinfection; By live body silkworm sterilization afterflush, dry, standby; Preparation PDA medium and rice medium;
Second step: the fruit body of separated Cordyceps militaris, after fruit body sterilization, rinsing, chopping, sets aside on slant medium, is put into and in incubator, cultivates 15-25 days;
The 3rd step: PDA medium is placed in to shaking flask, puts white hypha body in shaking flask concussion and cultivate, after 4-8 days, white hypha body is inoculated on rice medium, after covering with white hypha body, takes out mycelium and add Glucose Liquid, mycelia is smashed to pieces, obtained dietetic bacterial suspension as inoculation liquid;
The 4th step: with inhaling the abdominal foot portion that has the injector for medical purpose of inoculation liquid thorn note silkworm larvae body, the silkworm larvae body that obtains having inoculated, silkworm larvae body is tiltedly placed on the rice medium that sterilizing is good, first carried out lucifuge Fluctuation temperature culture, treated that pure white mycelia covers with polypide, seen that light cultivates, treat white hypha flavescence, proceed to see that light cultivates, treat that fruit body grows to 5～8 centimeters, fruit body surface is maturation before having yellow powder to occur;
The 5th step: gather, be emitted in dish airing or dry, will dry in 50～55 ℃ of temperature, make polypide water content not higher than 10%, to prevent moldy metamorphism; With bag film sealing, packing is preserved, or is positioned in the container of dry sealing tight building.
2. silkworm Chinese caterpillar fungus industrial production method as claimed in claim 1, is characterized in that, in described the 4th step, treat white hypha flavescence, sand is covered on silkworm larvae body, only expose a mouth for worm, proceed to see light cultivation, in the 5th step, while gathering, along fruit body, with tweezers, push sand aside, take out polypide and fruit body, with clear water, wash away the sand on Chinese caterpillar fungus.
3. silkworm Chinese caterpillar fungus industrial production method as claimed in claim 1, it is characterized in that, in the described first step, by live body silkworm sterilization afterflush, dry, standby concrete steps are: the live body silkworm of 4 ± 2 days after choosing the 5th and casting off a skin, under gnotobasis, with 75% ethanolic solution erasing polypide, carry out disinfection, the polypide of sterilizing with aseptic water washing is dried polypide under gnotobasis, and the silkworm larvae body drying saves backup at-2 ℃ ± 1 ℃ low temperature.
4. silkworm Chinese caterpillar fungus industrial production method as claimed in claim 1, it is characterized in that, PDA medium in the described first step is mixed to get by sterile distilled water, peeling potato, glucose and agar, and the ratio of described sterile distilled water, peeling potato, glucose and agar is 1000mL: 150-250g: 10-30g: 10-30g.
5. silkworm Chinese caterpillar fungus industrial production method as claimed in claim 1, is characterized in that, the rice medium in the described first step is obtained by rice and water mixing sterilizing, and described rice and the ratio of water are 30-40 gram: 50ml.
6. silkworm Chinese caterpillar fungus industrial production method as claimed in claim 1, is characterized in that, the compound method of the slant medium in described second step is: by peeling potatoes extractive, in potato juice, add agar, add water to 1000mL, be respectively charged in test tube, beyond the Great Wall tampon, after sterilizing, be cooled to 60 ℃, be placed on inclined-plane, wherein, the weight content of potato juice is 20%, and the weight content of agar is 1.7%.
7. silkworm Chinese caterpillar fungus industrial production method as claimed in claim 1, it is characterized in that, the concrete steps of the lucifuge Fluctuation temperature culture in described the 4th step are: in dark surrounds, first be placed under the condition of 0 ± 4 ℃ of temperature, humidity 85% and cultivate more than 10 days, then cultivate 10-20 days in 10 ℃ ± 2 ℃ environment.
8. silkworm Chinese caterpillar fungus industrial production method as claimed in claim 1, it is characterized in that, the concrete steps that light is cultivated of seeing in described the 4th step are: lucifuge Fluctuation temperature culture is covered with to the polypide of mycelia after 5 days, at illuminance 1000lx, at 8～10 hours every day, in 20 ℃ ± 2 ℃ of temperature, humidity 65% environment, cultivate.
9. silkworm Chinese caterpillar fungus industrial production method as claimed in claim 1, is characterized in that, proceeds to see that when light is cultivated, illuminance is 150-1500lx in described the 4th step, every day 6-8 hour, incubation time is 15-25 days.
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Cited By (6)
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| CN103960055A (en) * | 2014-05-15 | 2014-08-06 | 高青和润丝绸有限公司 | Method for cultivating silkworm cordyceps |
| CN103999690A (en) * | 2014-05-13 | 2014-08-27 | 陈洪佳 | Artificial cultivation method for cordyceps militaris |
| CN105660189A (en) * | 2016-02-23 | 2016-06-15 | 张家港顺泰元生物科技有限公司 | Cultivation method for cordyceps militaris |
| CN105706738A (en) * | 2016-02-23 | 2016-06-29 | 张家港顺泰元生物科技有限公司 | Cordycepsmilitaris cultivation method |
| CN105779300A (en) * | 2016-02-22 | 2016-07-20 | 嘉兴市潜福食品有限公司 | Solid culture medium and cultivation method of cordyceps militaris fruiting bodies |
| CN107980472A (en) * | 2017-11-28 | 2018-05-04 | 藤县友强淮山种植专业合作社 | A kind of implantation methods of cordyceps sinensis |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103999690A (en) * | 2014-05-13 | 2014-08-27 | 陈洪佳 | Artificial cultivation method for cordyceps militaris |
| CN103960055A (en) * | 2014-05-15 | 2014-08-06 | 高青和润丝绸有限公司 | Method for cultivating silkworm cordyceps |
| CN105779300A (en) * | 2016-02-22 | 2016-07-20 | 嘉兴市潜福食品有限公司 | Solid culture medium and cultivation method of cordyceps militaris fruiting bodies |
| CN105660189A (en) * | 2016-02-23 | 2016-06-15 | 张家港顺泰元生物科技有限公司 | Cultivation method for cordyceps militaris |
| CN105706738A (en) * | 2016-02-23 | 2016-06-29 | 张家港顺泰元生物科技有限公司 | Cordycepsmilitaris cultivation method |
| CN107980472A (en) * | 2017-11-28 | 2018-05-04 | 藤县友强淮山种植专业合作社 | A kind of implantation methods of cordyceps sinensis |
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Application publication date: 20140319 |