CN101338278A - Standard operation method for producing Cordycepsmilitaris of Chinese medicine - Google Patents
Standard operation method for producing Cordycepsmilitaris of Chinese medicine Download PDFInfo
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- CN101338278A CN101338278A CNA2008100217018A CN200810021701A CN101338278A CN 101338278 A CN101338278 A CN 101338278A CN A2008100217018 A CNA2008100217018 A CN A2008100217018A CN 200810021701 A CN200810021701 A CN 200810021701A CN 101338278 A CN101338278 A CN 101338278A
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Abstract
The invention discloses a method for culturing Chinese caterpillar fungus which is taken as Chinese medicinal plant in standardized scale culturing, comprising the using of the organic silkworm chrysalis. The silk cocoon is picked form the organic silkworm base. The silkworm chrysalis is obtained form the special silkworm harvest, storage and process sites, and is taken as the culture medium of Chinese medicinal plant Chinese caterpillar fungus. The strains should be adopted after being processed seed selection, freeze drying, preserving, activating and checking. The standardized culturing method should be adopted. The method comprises the pre culturing method and post culturing method. The standardized post treatment method is used, which comprises picking and drying method, packaging, storing and transporting methods. The Chinese medicinal plant Chinese caterpillar fungus quality guarantee method should be used, which comprises the Chinese medicinal plant Chinese caterpillar fungus quality standard and Chinese medicinal plant Chinese caterpillar fungus quality inspecting method, namely the Q/320691DJK01-2007 standard should be established. The method can produce the Chinese caterpillar fungus which is taken as Chinese medicinal plant in standardized way.
Description
Technical field:
The present invention relates to standard method and dedicated drying equipment that a kind of Chinese medicinal materials Cordyceps militaris (L.) Link. produces.
Background technology:
Cordyceps militaris (L.) Link. (Cordyceps militaris (L.) Link.) claims Cordyceps militaris (L.) Link., Cordyceps militaris, pupa grass again, is a kind of Chinese medicinal materials among developing.
Chinese caterpillar fungus is the bar-shaped fungi of a kind of length on polypide or pupal cell, and polypide (comprising larva and adult) or pupal cell are the substratum of this fungi.Chinese caterpillar fungus is the complex body of polypide (pupal cell) and fungi.Behind the insect infection Cordyceps fungus, Cordyceps fungus is progressively growth in its body, up to being full of larva (pupa) body, makes larva become sclerotium and profile is constant.Afterwards, grow sporophore, as careless shape, so the name Chinese caterpillar fungus from larva (pupa) body.
Chinese caterpillar fungus is seen in the domestic and foreign literature record kind more than 350.What have is grown on the larva body, and the length that has is on pupa, and what have is grown in on the polypide.Cordyceps militaris (L.) Link. claims pupa grass, Cordyceps militaris, Cordyceps militaris (L.) Link. again, and main product, is born on the pupal cell of the lepidopterous insects in broad-leaf forest or the hybridization woods topsoil among the mountain forest in Jilin in Yunnan more.Cordyceps militaris (L.) Link. and Cordyceps sinensis belong to Cordyceps (Cordyceps) together, and Cordyceps militaris (L.) Link. is the type species of this genus.The quality difference standard of Chinese caterpillar fungus mainly is the content of peculiar composition, as cordycepin, contain below 100 milligrams in per 100 grams of the Cordyceps sinensis that produce in Qinghai, and Cordyceps militaris (L.) Link. is generally more than the 200-300 milligram.
Over 30 years, the price increase of Cordyceps sinensis tens times.Current, the Cordyceps sinensis retail price of gift level has reached 500,000 yuan of per kilograms, and gradeless and uniformly-priced goods also reaches about 150,000 yuan.The reason of Cordyceps sinensis price increase comes from its therapeutic value.From the eighties from the beginning of, medical circle has rediscovered Cordyceps sinensis with the visual angle of modern biomedical.So far, there has been the vast amount of clinical report to show that Cordyceps sinensis has definite therapeutic action to multiple diseases such as renal failure, tuberculosis.The clinical meaning of Cordyceps sinensis not only is contraria, and is treatment, protection and coordinative role to each organ function of human body, relates to systems such as the heart, liver, lung, kidney, nerve, blood vessel, skin, bone, immunity.Therefore Cordyceps sinensis for heavily suffer from, old or difficult sufferer, particularly to the west of medical treatment be main, it is precious that the not good enough person of curative effect shows especially.
Why precious Cordyceps sinensis is, is because it possesses special effective ingredient after all.The specificity substance of Chinese caterpillar fungus is that cordycepin (cordycepin) is a kind of nucleosides material with anti-microbial activity, and the nuclear poly A polymerase is had very strong restraining effect.In DNA transcript mRNA process, make the mRNA dysmaturity, the growth of anticancer.Cordyceps polysaccharide is the branched polygalactomannan of a kind of height, can promote lymphocyte transformation, improves the anti-body contg and the body's immunological function of serum IgG, the anticancer ability that presses down cancer of enhancing body self.SOD (superoxide-dismutase) can eliminate superoxide radical in the body, has anti-ageing, the anticancer effect that presses down cancer.And why Cordyceps militaris (L.) Link. come into one's own and also be, the Cordyceps sinensis that its various effective ingredients are equivalent to, but wherein cordycepin content is higher than several times to tens times of Cordyceps sinensis.
In order to develop Cordyceps militaris (L.) Link., Chinese scholars has been carried out multi-faceted research work, relate to biology (Cordyceps militaris (L.) Link. form, ecological, distribute, the host, physiology etc.), the cultivation cultural method (produce by the Cordyceps militaris (L.) Link. liquid fermenting, in generation, expected solid culture, the ecological cultivation of biological host etc.), pharmaceutical chemistry (analyze by the Cordyceps militaris (L.) Link. effective ingredient, effective ingredient relatively waits), toxicology (Cordyceps militaris (L.) Link. acute toxicity, subacute, long term toxicity etc.), (Cordyceps militaris (L.) Link. is antitumor for pharmacology, uriniferous tubules is repaired, lowering blood glucose, immunomodulatory, the sexual function improving obstacle, antifatigue, antioxidant and anti-aging etc.), clinical treatment (Cordyceps militaris (L.) Link. treatment tumour, ephrosis, hepatic fibrosis, hypertension, diabetes, senile chronic bronchitis, regular flu, traumatic nerve recovery, exercise fatigue etc.), the result of these researchs shows that the pharmaceutical use of Cordyceps militaris (L.) Link. is definite, generally acknowledge, indubitable.So far, the development and use of Cordyceps militaris (L.) Link. 20 years, Cordyceps militaris (L.) Link. mycelia leavened prod (Cordyceps militaris (L.) Link. hypha capsule) has been obtained formal production authentication code (the accurate word Z20030034/35 of traditional Chinese medicines), and (the strong word G20060617 of state's food) is also granted for Chinese caterpillar fungus health food.As the property of Chinese medicine material, the Cordyceps militaris (L.) Link. prescription has also had 10 years at clinical prescription among the people or agreement side.From now on, the development Chinese patent medicine has become the inexorable trend of Cordyceps militaris (L.) Link. exploitation.
Over 20 years, the exploitation of Cordyceps militaris (L.) Link. has formed under the edible mushrooms notion instructs, under the microbiological pharmacy notion instructs and three different orientations under the Chinese medicinal materials notion instructs.
A: under the edible mushrooms notion instructs, be matrix to substitute culture material, Cordyceps militaris (L.) Link. is cultivated as a kind of mushroom.Showing as only is training objective with the sporophore, often is main raw material with the rice, Cordyccps-militaris-(L.)-link. Sporophore or directly edible, or and then be developed to food, medicated food or compound health-care food.
B: under the microbiological pharmacy notion instructs, be means with the liquid fermentation tank, be training objective with the Cordyceps militaris (L.) Link. fungus filament, and then be developed to medicine, protective foods.
C: under the Chinese medicinal materials notion instructs, being matrix with polypide or worm pupa, is training objective with complete Cordyceps militaris (L.) Link. herb, or directly edible, medicinal, or and then is developed to food, medicated food or protective foods.
With worm or pupa is that matrix is carried out Cordyceps militaris (L.) Link. and cultivated, and does three developing direction in current also the branch: be matrix with the silkworm, be matrix with the tussah or be matrix with other insects (as pine moth, tenebrio molitor).Because the difference of matrix insect, its pharmaceutical use is variant.
With the silkworm is that matrix is carried out the Cordyceps militaris (L.) Link. cultivation, currently also develops into four different branches: be matrix, be matrix, be matrix or add other compositions with the extract of bright or dried silkworm chrysalis and form synthetic medium with the oven dry mulberry silkworm chrysalis with fresh and alive mulberry silkworm chrysalis (cuing open the cocoon pupa) with aged silkworm larvae.These differences cause corresponding Cordyceps militaris (L.) Link. production technique difference, and the production cost difference also exists different between its pharmaceutical use.
Over 20 years, Cordyccps-militaris-(L.)-link. Sporophore is cultivated as a kind of mushroom, accumulated quite abundant cultivation experience, formed a whole set of methodology, and developed into the scale production standardized management.The Cordyceps militaris (L.) Link. mycelia is cultivated as a kind of pharmacy intermediate, also formed tens of tons throughput, its product is as market on the new drug.But, be used as under the guidance of Chinese medicinal materials notion, be that the culturing purposes thing carries out mass-producing, the standardization producer with complete Cordyceps militaris (L.) Link. herb, the present invention still belongs to the first.
What is Cordyceps militaris (L.) Link. (claiming Cordyceps militaris again)? different people provides different answers; Is which type of Cordyceps militaris (L.) Link. the Chinese medicinal materials Cordyceps militaris (L.) Link.? answer is also different.In the industry even such conception of species occurs: same Cordyceps militaris spawn, longly on rice, be Cordyceps militaris, longly on pupa, cry Cordyceps militaris (L.) Link., long on the cocoon chrysalis and long not distinguishing on mulberry silkworm chrysalis, or the like.In the face of a kind of like this trouble waters, the pharmacist of traditional Chinese medicine how follows, and the traditional Chinese physician how dares to use! As for how judgement is wherein good and bad, which is genuine, then there are no laws to abide by especially, do not know where to begin.
GAP standard operating procedure (SOP) is the basis and the prerequisite of the genuine Chinese medicinal materials of development.For the Cordyceps militaris (L.) Link. among developing, the meaning of SOP still promotes the prerequisite of clinical use of Cordyceps militaris (L.) Link. Chinese medicinal materials and further exploitation compound Cordyceps militaris (L.) Link. Chinese patent medicine except the needs of Cordyceps militaris (L.) Link. tunnelization, and this also is the certainty of Cordyceps militaris (L.) Link. development process.Set up the Chinese medicinal materials production standard working specification (SOP) of Cordyceps militaris (L.) Link., will help to make the standardization of Cordyceps militaris (L.) Link. cultivation and production undoubtedly, and then promote Cordyceps militaris (L.) Link. to become the process of Chinese medicinal materials new drug.
Summary of the invention:
The object of the present invention is to provide a kind of the standardization to produce standard method and the dedicated drying equipment of Chinese medicinal materials Cordyceps militaris (L.) Link..
Technical solution of the present invention is:
A kind of scale cultivation that standardizes is used as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials, it is characterized in that: comprise the following steps:
A: adopt organic mulberry silkworm chrysalis, comprising: obtain silk cocoon from organic silkworm and mulberry base, receive, store up and processing place acquisition silkworm chrysalis from silkworm chrysalis then, as the used substratum of Chinese medicinal materials Cordyceps militaris (L.) Link.;
B: adopt being fit to the bacterial classification of Chinese medicinal materials Cordyceps militaris (L.) Link., is through seed selection, freeze-drying preservation, activation, bacterial classification by the bacterial classification check;
C: the code requirement cultural method comprises: preceding cultural method, back cultural method;
D: the code requirement post-treating method comprises: the drying means of gathering, packing conveying method;
E: adopt Chinese medicinal materials Cordyceps militaris (L.) Link. method for guaranteeing quality, comprising: Chinese medicinal materials Cordyceps militaris (L.) Link. quality standard, Chinese medicinal materials Cordyceps militaris (L.) Link. quality determining method, promptly set up the Q/320691DJK01-2007 standard.
In the steps A, silk cocoon is cutd open cocoon, detects the silkworm chrysalis heavy metal, weigh the silk cocoon quality: plumbous (in Pb, dry weight, down with) be not higher than 1mg/kg, arsenic (in As) is not higher than 0.5mg/kg, mercury (in Hg) is not higher than 0.2mg/kg;
The used substratum of Chinese medicinal materials Cordyceps militaris (L.) Link. adopts the complete silkworm chrysalis of wet (or through freezing) behind the mulberry cocoon filature; Wet silkworm chrysalis process by following given process behind the described filature: a level sortation-two level sortation-three level sortation-cleaning-blanching-cool off-be used for production or quick-frozen-packing is kept in; Behind work program such as filature, this silkworm chrysalis heavy metal contamination index still meets lead and is not higher than 1mg/kg, arsenic and is not higher than 0.5mg/kg, and mercury is not higher than the standard of 0.2mg/kg.
Among the step B through seed selection, freeze-drying preservation, activate supervisor gained bacterial classification and whether can be used for the Chinese medicinal materials Cordyceps militaris (L.) Link. and produce, depend on whether accord with Q/320691DJK01-2007 standard of final gained Cordyceps militaris (L.) Link..
Cultivate before among the step C and be: silkworm chrysalis or through the complete silkworm chrysalis of the refrigerated cuvette of packing into will wet, sterilize, inoculate, culturing room to preceding cultivation finishes before placing then, charge amount is to account for below 3/5 (L/L) in the transparent small vessels, inoculate with the liquid spawn that is no more than 48 hours cell ages during inoculation, on the silkworm chrysalis of the bacterium of under 100 grades of purification conditions the liquid spawn sprinkle being gone out.The inoculation than 10: 1 to 10: 3 (silkworm chrysalis: liquid spawn, V/V); Preceding culture temperature is controlled between 10 ℃ to 25 ℃ 20 days to 60 days; The back culture vessel adopts the light-passing plastic basket among the step C, and the transparent plastics basket serves as a contrast with five of food fresh keeping membranes or six bread, back culture environment temperature is controlled between 15 ℃ to 30 ℃, 10 ℃ of temperature difference per day, the humidity that also will control environment is 85%, natural scattering light or source of artificial light, illumination 200 to 2000 Luxs; Cultivate gained Chinese medicinal materials Cordyceps militaris (L.) Link., accord with Q/320691DJK01-2007 standard.
The drying of step D adopts low temperature flow-through type warm air drying, and hot blast temperature is controlled between 55 ℃ to 65 ℃, dry 6 to 8 hours.
Described low temperature flow-through type warm air drying adopts simple and easy low temperature flow-through type hot-air drier, comprise blower fan, interchanger is set on the blower fan, the multilayer charging tray is set on the interchanger, charging tray top is provided with extraction hood, is connected to a thermal head on the exchanger heat water pump and stretches into temperature controller in the charging tray.
Use the double-deck two envelope weather-proof dressings of plastics bag during the accumulating of step D packing, lucifuge, refrigeration below 5 ℃.
After handling through step D or again after preserving more than 2 years, Cordyceps militaris (L.) Link. is accord with Q/320691DJK01-2007 standard still.
As one of index that detects the Chinese medicinal materials Cordyceps militaris (L.) Link., cordycepin content is not less than 1mg/g with cordycepin content.
Advantage of the present invention:
One, the culture medium in the GAP of solution Cordyceps militaris (L.) Link. Chinese medicinal materials is selected problem
The GAP culture medium of Cordyceps militaris (L.) Link. Chinese medicinal materials will meet the following conditions: can satisfy under scale production, from the horse's mouth, the quality controllable prerequisite, make the gained Cordyceps militaris (L.) Link. possess complete, Chinese medicinal materials and differentiate external appearance characteristic and more rich effective component content.
1. present method is different with the Cordyceps militaris (L.) Link. cultivating method in following patent of invention and the similar known technology: as " cultural method of northeast Chinese caterpillar fungus and goods thereof (C093117581.X) ", " cultivating method of disease-resisting high-yield cordyceps (C096115115) ", " cultural method of cordyceps militaris fruit body (C099124200.9) ".Aforesaid method only is the cultivation target with the sporophore, is main raw material with the rice, can not get Chinese medicinal materials Cordyceps militaris (L.) Link. integral outer appearance, and effective component content is lower.
2. present method is different with the cordyceps culturing method in patent of invention " the artificial culture Cordyceps militaris (L.) Link. improves the method (C002109304.0) of its content of physiologically active substance " and the similar known technology.Aforesaid method is under liquid fermentation condition, is the culturing purposes thing to produce the Cordyceps militaris (L.) Link. fungus powder, can not get the Cordyceps militaris (L.) Link. of diagnostic characteristics.
3. present method is different with the Cordyceps militaris (L.) Link. cultivating method in patent of invention " a kind of method of being used as medicine with the Antheraea pernyi Geurin Meneville culturing Cordyceps militaris (L.) Link. (C094106100) ", " silkworm chrysalis Cordyceps sinensis industrialized preparing process (C00130381.3) " and the similar known technology.Above-mentioned technological method is to make substratum with fresh and alive tussah (or silkworm) pupa, will cut silk cocoon open and obtain fresh and alive tussah (or silkworm) pupa, causes unnecessary cost allowance.
4. present method is different with the Cordyceps militaris (L.) Link. cultivating method in patent of invention " method of using baked silkworm chrysalis for artificially breeding Chinese caterpillar fungus (C093102809) " and the similar known technology.Above-mentioned technological method is to make substratum with the mulberry silkworm chrysalis of oven dry.Commercially available dried silkworm chrysalis or dry naturally or coal fire oven dry and getting is used for feed more, and the former many impurity, polyacid lose, and many lead contents of latter height should not use.
5. present method is different with the Cordyceps militaris (L.) Link. cultivating method in patent of invention " the artificial culture methods (C097113190) of Chinese caterpillar funguses such as Cordyceps militaris (L.) Link. " and the similar known technology.This method adopts the extract of pupa to add other compositions composition artificial mediums, finally can not get the Cordyceps militaris (L.) Link. of outward appearance diagnostic characteristics.
One of purpose of the present invention, be the shortcoming of evading above raw material, select suitable cultivation raw material, make culture that complete Cordyceps militaris (L.) Link. outward appearance diagnostic characteristics be arranged, effective ingredient (as cordycepin) content is higher, and quality controllable number people, can satisfy needs a large amount of, scale production.Take this to solve the selection problem of the culture medium material in the GAP of Cordyceps militaris (L.) Link. Chinese medicinal materials.
Two, standard is as the Chinese caterpillar fungus culture medium material source of Chinese medicinal materials
As mentioned above, mulberry silkworm chrysalis was as cultivating raw material after the inventive method adopted complete filature.This raw material has its plurality of advantages, but in order to guarantee to produce qualified Chinese medicinal materials Cordyceps militaris (L.) Link., also it must be limited in nuisanceless green or the organic scope.
Current, except special for the cultivation Cordyceps militaris (L.) Link. painstakingly cultivate, various silkworms (pupa) comprise mulberry silkworm chrysalis behind aged silkworm, bright pupa, dry silkworm chrysalis, the filature, all have the source uncertain.With mulberry silkworm chrysalis behind the filature is example, the silk cocoon shelves mouth of silk mill (batch) often change because of various reasons, as for the ambient conditions in silk cocoon country of origin, whether be subjected to certain pollution, then the no one can tell Chu.This present situation keeps the stable very unfavorable of Chinese medicinal materials Cordyceps militaris (L.) Link. quality to the scale production of Chinese medicinal materials Cordyceps militaris (L.) Link..
For this reason, be necessary to design a whole set of feasible management method and corresponding technical measures, originate as Chinese medicinal materials Cordyceps militaris (L.) Link. cultivated material in order to standard.
Three, improve bacterial classification and the bacterial classification using method that is used for above culture material
Chinese caterpillar fungus class fungi is parasitic and saprophytic facultative fungi.In the process of natural infection insect, carry out parasitic life earlier, then (after insect death) carries out saprogenesis.Silkworm chrysalis live body surface seeding or the inoculation of silkworm chrysalis living body puncture are the parasiticss of utilizing it, are also utilizing the natural resistance against diseases of live body self (opposing living contaminants).The present invention utilizes mulberry silkworm chrysalis behind the complete filature, has the water content height, problem such as become sour easily, and do not have natural resistance against diseases to utilize, Cordyceps militaris (L.) Link. raise craft condition that therefore need be different with other culture materials.
The present invention is different with the Cordyceps militaris (L.) Link. cultivating method in " the artificial culture methods (C097113190) of Chinese caterpillar funguses such as Cordyceps militaris (L.) Link. " and the similar known technology.Cultivate Cordyceps militaris (L.) Link. with live body silkworm (or pupa) in this type of technology, need be with solid spawn or liquid spawn, be seeded on the live body silkworm (or pupa) in injection under the surface sterilization condition, perhaps the live body silkworm (or pupa) of surface sterilization rendered in solid spawn or the liquid spawn, make its natural infection.This with live body silkworm (or pupa) be the inoculation technique of core because can not thoroughly sterilize, so can not get rid of latent infection, the intersection that also is difficult to get rid of between incubation period infects again.The live body inoculation technique is unsuitable for mulberry silkworm chrysalis behind the filature.
Four, solve the cultivating method problem that is applicable in the Chinese medicinal materials Cordyceps militaris (L.) Link.
The GAP standard operating procedure (SOP) of Cordyceps militaris (L.) Link. Chinese medicinal materials, the Cordyceps militaris (L.) Link. that requires to turn out keeps Chinese medicinal materials discriminating external appearance characteristic, and the effective ingredient ratio content is higher.For this reason, propose the cultivation of Chinese medicinal materials Cordyceps militaris (L.) Link. is divided into " preceding cultivation " and " cultivate the back " two stages.Prior art does not relate to this special method.
Five, proposition is applicable to the method for quality control of the GAP of Chinese medicinal materials Cordyceps militaris (L.) Link.
Prior art does not relate to Chinese medicinal materials Cordyceps militaris (L.) Link. quality standard and detection method problem thereof yet.
We have set up Chinese medicinal materials Cordyceps militaris (L.) Link. quality standard, and that has established Cordyceps militaris (L.) Link. enters the Chinese medicinal materials sequence, promote the quality base of Cordyceps militaris (L.) Link. Chinese patent medicine development.
Six, proposition is applicable to the post-treating method of the GAP of Chinese medicinal materials Cordyceps militaris (L.) Link.
The Chinese medicinal materials Cordyceps militaris (L.) Link. needs good post-treating method.The Chinese medicinal materials Cordyceps militaris (L.) Link. is longer than Cordyccps-militaris-(L.)-link. Sporophore internal moisture diffusion length, usually has drying not up to standard, take care of variable color for a long time in addition mildew, change powder (?evil).Although the Chinese medicinal materials Cordyceps militaris (L.) Link. is lower than Cordyceps sinensis price, be a kind of valuable ingredient of traditional Chinese medicine after all, therefore should not come keeping, store and transportation with the Chinese medicinal materials ordinary method.For this reason the present invention intend from the Chinese medicinal materials Cordyceps militaris (L.) Link. gather, drying plant and Cordyceps militaris (L.) Link. preserve three angles and address this problem.
Seven, reduce the production cost of Chinese medicinal materials Cordyceps militaris (L.) Link.
In order to make Cordyceps militaris (L.) Link. Chinese medicinal materials and Chinese patent medicine popular, popular, improve the market competitiveness of Chinese medicinal materials Cordyceps militaris (L.) Link., the present invention improves technology by changing raw material, has further reduced cost.
In a word, the present invention will provide core technology for implementing " the GAP standard operating instructions (SOP) of Cordyceps militaris (L.) Link. Chinese medicinal materials ".
In conjunction with the present invention, set up Chinese medicinal materials Cordyceps militaris (L.) Link. production standard working method (SOP) and comprising:
A: the method for setting up organic mulberry silkworm chrysalis supply system.
B: Chinese medicinal materials Cordyceps militaris spawn security system and method.
C: Chinese medicinal materials Cordyceps militaris (L.) Link. quality assurance system and method.
D: the cultivating method in the Chinese medicinal materials Cordyceps militaris (L.) Link. batch production scale production.
E: Chinese medicinal materials Cordyceps militaris (L.) Link. treatment system and method.
Specifically can be:
One: the method for setting up organic mulberry silkworm chrysalis supply system
1: organic silkworm and mulberry base standard
2: managing of mulberry field and sericulture rules
3: the special-purpose silk cocoon processing rules of gathering
4: special-purpose silkworm chrysalis processing rules
5: special-purpose silkworm chrysalis storage and transport regulations
6: organic silkworm and mulberry base internal examination rules
Two: Chinese medicinal materials Cordyceps militaris spawn security system and method:
1: screen Cordyceps militaris (L.) Link. from wild cordyceps militaris or individual plant with the conventional organization separation method
2: use conventional strain breeding method, it is stable that many generations are repeated to proterties
3: with conventional low temperature culture presevation, running check is also done the fruiting experiment, prevents to degenerate
4: routine is manufactured liquid spawn
5: the bacterial classification testing procedures that is fit to the Chinese medicinal materials Cordyceps militaris (L.) Link.;
Three: Chinese medicinal materials Cordyceps militaris (L.) Link. quality assurance system and method:
1: Chinese medicinal materials Cordyceps militaris (L.) Link. quality standard
2: Chinese medicinal materials Cordyceps militaris (L.) Link. physical and chemical index cordycepin is differentiated, content assaying method (TLC HPLC method) (annex 2)
Four: the cultivating method in the Chinese medicinal materials Cordyceps militaris (L.) Link. batch production scale production:
1: the preceding cultural method of Chinese medicinal materials Cordyceps militaris (L.) Link.,
Cultivating before among the present invention, is to cultivate raw material to finish the whole process of sending out bacterium (walking bacterium) in cuvette.
1.1 it is qualified that substratum adopts, wet silkworm chrysalis or again through refrigerated watered mulberry leaf silkworm chrysalis behind the filature.
1.2 during the watered mulberry leaf silkworm chrysalis was packed transparent small vessels into behind the filature, charge amount was to account for below 3/5 (L/L).Cuvette can be a vial, also can be Plastic Bottle or bag.
1.3 routine is sealed, sterilization, cooling.
1.4 with liquid spawn or solid spawn, under 100 grades of purification conditions, the method with drenching, spreading is seeded on the watered mulberry leaf silkworm chrysalis.
1.5 the strain cultivation cycle of using among the present invention is no more than 48 hours.The solid spawn inoculation was than 1: 20 to 1: 30 (w/w), and liquid-spawn inoculation was than 1: 10 to 1: 30 (ml/ml).
1.6 preceding culture temperature is controlled between 10 ℃ to 25 ℃, 20 days to 60 days.
2: cultural method behind the Chinese medicinal materials Cordyceps militaris (L.) Link..
Back cultivation among the present invention is meant preceding cultured products is cultivated into the Chinese medicinal materials Cordyceps militaris (L.) Link. again, possesses qualified appearance character and Cordyceps militaris (L.) Link. and differentiates other features of evaluation.
2.1 after the preceding cultivation, mulberry silkworm chrysalis is full of the Cordyceps militaris (L.) Link. mycelia, and forms agglomerate because of the expansion of mycelia.The present invention at first is distributed to the mulberry silkworm chrysalis after the preceding cultivation in the incubator of back.
2.2. disperse and in relatively cleaner environment, to carry out, finish by craft.The staff will do the healthy prevention and prevent that foreign material from sneaking into.
2.3 the back incubator among the present invention adopts special light-passing plastic case, plastic box serves as a contrast with five of semi-transparent gas plastic films (as food fresh keeping membrane) or six bread.
2.4 the back is cultivated the culturing room of usefulness and is required scattered light is arranged, natural scattering light or source of artificial light, illumination 200 to 2000 Luxs (Lux).
2.5 back culturing room temperature is controlled between 15 ℃ to 30 ℃, 10 ℃ of the temperature difference.
2.6 back culturing room ambient moisture is controlled at more than 85%.
3: the process control of the forward and backward cultural method of Chinese medicinal materials Cordyceps militaris (L.) Link.
3.1 take place as emergency situation, must report without delay, must not take treatment measures without authorization.
3.2 when if the setting up of change in any system or new equipment arranged, producing the responsible official should report the developments the very first time.
Five: Chinese medicinal materials Cordyceps militaris (L.) Link. post-treating method:
1: the Chinese medicinal materials Cordyceps militaris (L.) Link. is gathered, dry, Packaging Method.
1.1 Chinese medicinal materials Cordyceps militaris (L.) Link. what is called is gathered, and just sophisticated Cordyceps militaris (L.) Link. is shifted out incubator, or removes the lateral plastic film of incubator, moves on in the ventilating chamber or adds forced ventilation, to reduce surperficial moisture content.
1.2 Chinese medicinal materials Cordyceps militaris (L.) Link. drying means can adopt cryodrying as required, low-temperature vacuum drying or freeze-drying.
1.3 adopt low temperature flow-through type warm air drying, hot blast temperature is controlled between 55 ℃ to 65 ℃, 6 to 8 hours.
1.4 designed a kind of low temperature flow-through type hot-air drier, schema is seen (accompanying drawing 2).
1.5 the dry standard of Chinese medicinal materials Cordyceps militaris (L.) Link., water content is controlled at below 7%.
1.6 dried Cordyceps militaris (L.) Link. is packed into the double-layer plastic bag, seal, require plastics bag to want safety non-toxic evil, thickness is not less than 2.5, otherwise pierces through easily, loses efficacy.
2: the storage of Chinese medicinal materials Cordyceps militaris (L.) Link., transportation method.
Want 5 ℃ of lucifuge refrigerations 2.1 be encapsulated into the Cordyceps militaris (L.) Link. of plastics bag.
2.2 when storage, transportation, outer with plastic box or carton splendid attire, prevent the plastics bag incised wound, galling.
3: the Chinese medicinal materials Cordyceps militaris (L.) Link. is gathered, the process control of packing, accumulating
4.1 take place as emergency situation, must report without delay, must not take treatment measures without authorization.
4.2 when if the setting up of change in any system or new equipment arranged, producing the responsible official should report the developments the very first time.
The present invention be advantageous in that:
One: the GAP standard operating procedure (SOP) of Cordyceps militaris (L.) Link. Chinese medicinal materials is the prerequisite of the genuine Cordyceps militaris (L.) Link. Chinese medicinal materials of development, also is the certainty of Cordyceps militaris (L.) Link. development process.Prior art does not relate to GAP standard operating procedure (SOP) problem of setting up the Cordyceps militaris (L.) Link. Chinese medicinal materials.The present invention provides core technology for the GAP standard operating procedure (SOP) of Cordyceps militaris (L.) Link. Chinese medicinal materials.
Two: traditional Chinese medicine circle thinks that " worm (pupa) " of Cordyceps sinensis is cloudy, and " grass " is sun, and yin yang complementarity share to good.Utilization is a matrix to substitute culture material, or is means with the liquid fermentation tank, can not produce the Cordyceps militaris (L.) Link. that meets Chinese medicine Cordyceps sinensis traditional appearance notion, promptly has " worm (pupa) " that the Cordyceps militaris (L.) Link. of " grass " is arranged.And the Cordyceps militaris (L.) Link. that the inventive method is turned out can obtain the approval of Chinese medicine circle, and this is a principal advantage of the present invention.
Three: set up nuisanceless, organic mulberry silkworm chrysalis supply system, standard is as the Cordyceps militaris (L.) Link. cultivated material source of Chinese medicinal materials
" first workshop " that the Chinese medicinal materials Cordyceps militaris (L.) Link. produces is used as in nuisanceless, organic silkworm and mulberry base, " second workshop " that the Chinese medicinal materials Cordyceps militaris (L.) Link. produces is used as in special reel silk from cocoons, selected, special storage, this is the basis of standardization scale cultivation Chinese medicinal materials Cordyceps militaris (L.) Link..So the ambient conditions in silk cocoon country of origin is understood, the clear wash rice of work flow, agricultural residual problem and heavy metals exceeding standard problem can take preventive measures, and this is to the scale production of Chinese medicinal materials Cordyceps militaris (L.) Link., keep the stable of Chinese medicinal materials Cordyceps militaris (L.) Link. quality, it is very favourable to establish the genuine Chinese medicinal materials brand of Cordyceps militaris (L.) Link..
Four: the technical superiority in the material choice
1: the utilization cocoon chrysalis is a matrix, or the Cordyceps militaris (L.) Link. of turning out with other insect larvaes or pupa, and its property of medicine and mulberry silkworm chrysalis are that the Cordyceps militaris (L.) Link. property of medicine of matrix is different.In principle, " cocoon chrysalis Cordyceps militaris (L.) Link. " or " XX worm Cordyceps militaris (L.) Link. " should be considered as another kind of Chinese medicinal materials, and proved that the pharmaceutical use of mulberry silkworm chrysalis Cordyceps militaris (L.) Link. is better than the cocoon chrysalis Cordyceps militaris (L.) Link..
2: compare with other insects as raw material, silkworm obtains easily in batches, thereby easy control source, quality and quantity.
3: with compare as raw material with the silkworm mature larva, or with compare as raw material with live body (conventional or aseptic) mulberry silkworm chrysalis, can already not contend over raw materials with mulberry silkworm chrysalis with silk, can not destroy silk in other words, thereby reduce cost, and reduced of the restriction of sericulture cycle the Cordyceps militaris (L.) Link. cultivation.
4: commercially available oven dry mulberry silkworm chrysalis Rong Yi Li goes into the foreign material foul, and heavy metal content is usually higher, has increased the possibility that quality accident takes place.Compare as raw material with the oven dry mulberry silkworm chrysalis, reduced the operation of punchinging, also do not need supplement the nutrients liquid or water, reduced running cost, also reduced the possibility that quality accident takes place simultaneously with mulberry silkworm chrysalis cultivation Cordyceps militaris (L.) Link. behind the filature.
5: the extract with silkworm chrysalis adds other compositions composition artificial mediums, can not get meeting the complete Cordyceps militaris (L.) Link. of Chinese medicinal materials idea.
In a word, the advantage compared with above-mentioned technological method of the present invention is: (or through refrigerated) complete mulberry silkworm chrysalis has satisfied the raw materials requirement of cultivating qualified Chinese medicinal materials Cordyceps militaris (L.) Link. after utilizing filature.
Five: the technical superiority in Chinese medicinal materials Cordyceps militaris spawn security system and the method
Chinese caterpillar fungus class fungi is parasitic and saprophytic facultative fungi.In the process of natural infection insect, carry out parasitic life earlier, then (after insect death) carries out saprogenesis.Silkworm chrysalis live body surface seeding or the inoculation of silkworm chrysalis living body puncture are the parasiticss of utilizing it, and the present invention utilizes the saprophytic principle design of Cordyceps.
1: have 2 with the existing relevant inoculation technique of utilizing mulberry silkworm chrysalis to cultivate Cordyceps militaris (L.) Link.:
A:, be seeded on the live body silkworm (or pupa) in injection under the surface sterilization condition with solid spawn or liquid spawn.This method is because the sterilization of can not revealing what is at the bottom of sth, and infect before especially can not getting rid of silkworm-raising jobs alive, so pollution rate is higher, and success ratio is lower.
B: under the surface sterilization condition,, make its natural infection to render in solid spawn or the liquid spawn on the live body silkworm (or pupa).Because can not infuse end sterilization, especially can not get rid of and take place to infect before the silkworm-raising jobs alive or by parasitism, this method pollution rate is higher, success ratio is lower.The intersection that this method also is difficult to get rid of between incubation period infects again.
The present invention is with liquid spawn or solid spawn, is seeded under aseptic condition behind the complete filature on bright (freezing) watered mulberry leaf silkworm chrysalis.The difference of comparing with art methods is: be not " is on the liquid nutrient medium of main component in the moiety with silkworm chrysalis with inoculation ", neither be inoculated in " on the silkworm chrysalis that obtains with the artificial diet aseptic breeding of silkworm ".On this basis, present method is different with following patent of invention: " the artificial culture methods (C097113190) of Chinese caterpillar funguses such as Cordyceps militaris (L.) Link. ".
Six: the technical superiority in the cultivating method
1. cultivating before among the present invention, is to cultivate raw material to finish the whole process of sending out bacterium in cuvette.The employing cuvette has a lot of advantages.
Cultivation is finished the sterilization back to sending out bacterium (walking bacterium) end before among the present invention in cuvette.Emphasize that charge amount is to account for below 2/5 volume in the transparent small vessels.Cuvette can be a vial, also can be Plastic Bottle or bag.Requiring transparent purpose is incubation growth situation before observing for convenience, and requiring vessel volume will be for convenience of in the inoculation operation for a short time, is convenient to handle abnormal conditions simultaneously, reduces the crossed contamination loss.
2. wet silkworm chrysalis behind the filature is packed in the transparent small vessels, charge amount is to account for below 3/5 (L/L), and advantage is to keep the mixing space, helps the contact of bacterium liquid balanced, sends out bacterium synchronously.
3. be no more than the liquid spawn cell age of 48 hours cell ages and suitable inoculation ratio, help to form growth vigor, also help to send out synchronously bacterium.
4. preceding culture temperature is controlled between 10 ℃ to 25 ℃, 20 days to 60 days.It is few that low temperature is sent out bacterium (walking bacterium) pollution, quality better.
5. the back culture vessel adopts the special light-passing plastic case with semi-transparent gas plastic film cylinder-packing.Making full use of back culturing room spatial simultaneously, ensureing the needed abundant and appropriate illumination of Cordyceps militaris (L.) Link. around here, competent gaseous interchange also is provided.
6. back culturing room temperature is controlled between 15 ℃ to 30 ℃ 10 ℃ of temperature difference per day.The bigger temperature difference has been simulated the woodland natural condition, helps the formation of Cordyccps-militaris-(L.)-link. Sporophore and synthesizing of cordycepin.
7. the Chinese medicinal materials cordyceps culturing method that utilizes among the present invention to be provided, its effect finally shows, can turn out appearance character and meet the Chinese materia medica notion, there is worm that the Cordyceps militaris (L.) Link. of grass is arranged, its effective ingredient (cordycepin) be higher than additive method several times to tens times, meet Cordyceps militaris (L.) Link. and differentiate to identify the various features that possessed.
Seven: the technical superiority in the quality control
1: the GAP standard operating procedure (SOP) of Cordyceps militaris (L.) Link. Chinese medicinal materials is the precondition of Cordyceps militaris (L.) Link. Chinese patent medicine exploitation, also is the certainty of Cordyceps militaris (L.) Link. development process.The present invention is a technological core of setting up the GAP standard operating procedure of Cordyceps militaris (L.) Link. Chinese medicinal materials (SOP).Prior art does not relate to this level.
2: in order to set up the GAP standard operating procedure (SOP) of Cordyceps militaris (L.) Link. Chinese medicinal materials, propose, the GAP of Cordyceps militaris (L.) Link. Chinese medicinal materials needs to set up " stable raw material is specialized in system, comprising: organic silkworm and mulberry base, special-purpose silkworm chrysalis receipts, method for storing ".Prior art does not relate to stable raw material and specializes in system problem.
3: in order to set up the GAP standard operating procedure (SOP) of Cordyceps militaris (L.) Link. Chinese medicinal materials, propose, set up " Chinese medicinal materials Cordyceps militaris (L.) Link. quality assurance system comprises: Chinese medicinal materials Cordyceps militaris (L.) Link. quality standard Q/320691DJK01-2007, Chinese medicinal materials Cordyceps militaris (L.) Link. quality determining method ".Established the Chinese medicinal materials Cordyceps militaris (L.) Link. form standard that combines with sporophore with sclerotium first; Having established first is the Chinese medicinal materials Cordyceps militaris (L.) Link. physical and chemical index of leading indicator with the cordycepin content.Prior art does not relate to Chinese medicinal materials Cordyceps militaris (L.) Link. quality standard and detection method thereof.
Eight: the technical superiority of Chinese medicinal materials Cordyceps militaris (L.) Link. aftertreatment
1: the low temperature flow-through type hot-air drier that the present invention shows, hot blast temperature is controlled between 55 ℃ to 65 ℃, 6 to 8 hours.Its advantage be structure can letter can be numerous, cost is lower, high efficiency.
2: in the Chinese medicinal materials Cordyceps militaris (L.) Link. packing conveying method, Cordyceps militaris (L.) Link. is sealed protection against the tide, lucifuge, 5 ℃ of refrigerations with double-deck pair of plastics bag.Facts have proved that this method is respond well.Preserve the Chinese medicinal materials Cordyceps militaris (L.) Link. more than 2 years, its physical and chemical index and external appearance characteristic do not have obvious variation.
Nine: the technical superiority in the accounting cost control
1: implement the present invention and cultivate in the Chinese medicinal materials Cordyceps militaris (L.) Link., cordycepin content exceeds tens times or hundred times up to 13.5mg/g.Carry out price comparison with cordycepin content: Cordyceps sinensis gradeless and uniformly-priced goods (cordycepin content 1mg/g) is 100,000 yuan/kg of market price at present, and the market value of the Cordyceps militaris (L.) Link. that the present invention produces should be 1,350,000 yuan/kg; Otherwise as if the 2000 yuan/kg of market value of the Cordyceps militaris (L.) Link. that produces in the present invention, the present market price of Cordyceps sinensis gradeless and uniformly-priced goods should be 150 yuan/kg; And the liquid fermentation tank Cordyceps fungus powder bulk drug that to be means produce (cordycepin content 0.1mg/g is following) at present market price should be 15 yuan/kg, and should not be 150 yuan/kg-200 unit/kg.Cordyceps fungus powder commodity (medicine or protective foods) market price at present should be 100 yuan/kg-300 unit/kg, and should not be 2000 yuan/kg-5000 unit/kg.Enforcement of the present invention, will make vast salary layer enjoy Chinese caterpillar fungus becomes possibility, and its social benefit is seen some from this.
2: the invention solves with fresh and alive mulberry silkworm chrysalis cultivation Cordyceps militaris (L.) Link. and silkworm and already fight for the silk problem of resource.Utilization live body mulberry silkworm chrysalis is a matrix, is that matrix is produced Cordyceps militaris (L.) Link. with silkworm larvae perhaps, certainly will will destroy silk cocoon, influences the production (the silk cocoon filature of cutting off is cheap) of silk, and the present invention has saved the silk resource, reduces the Cordyceps militaris (L.) Link. raw materials cost.
3: Cordyceps militaris (L.) Link. cultivation thing is to have " worm (pupa) " that " grass " arranged, the Cordyceps militaris (L.) Link. of whole strain, so the unit's of pressing processing volume conversion, the Chinese medicinal materials Cordyceps militaris (L.) Link. rate ratio of implementing the present invention's production is that matrix improves more than 20 times to substitute culture material, its direct production cost so the several times that descend.
The present invention is the management of Cordyceps militaris (L.) Link. Chinese patent medicine cost of developing, and popular, and popular market operation provides favourable condition, has improved the cost advantage and the market competitiveness of Chinese medicinal materials Cordyceps militaris (L.) Link..
Below in conjunction with drawings and Examples this is described further:
Fig. 1 is a schema of the present invention.
Fig. 2 is the configuration diagram of dedicated drying equipment of the present invention.
Embodiment:
A kind of scale cultivation that standardizes is used as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials, comprises the following steps:
A: adopt organic mulberry silkworm chrysalis, comprising: obtain silk cocoon from organic silkworm and mulberry base, receive, store up and processing place acquisition silkworm chrysalis from silkworm chrysalis then, as the used substratum of Chinese medicinal materials Cordyceps militaris (L.) Link.;
B: adopt being fit to the bacterial classification of Chinese medicinal materials Cordyceps militaris (L.) Link., is through seed selection, freeze-drying preservation, activation, bacterial classification by the bacterial classification check;
C: the code requirement cultural method comprises: preceding cultural method, back cultural method;
D: the code requirement post-treating method comprises: the drying means of gathering, packing conveying method;
E: adopt Chinese medicinal materials Cordyceps militaris (L.) Link. method for guaranteeing quality, comprising: Chinese medicinal materials Cordyceps militaris (L.) Link. quality standard, Chinese medicinal materials Cordyceps militaris (L.) Link. quality determining method, promptly set up the Q/320691DJK01-2007 standard.
In the steps A, silk cocoon is cutd open cocoon, detects the silkworm chrysalis heavy metal, weigh the silk cocoon quality: plumbous (in Pb, dry weight, down with) be not higher than 1mg/kg, arsenic (in As) is not higher than 0.5mg/kg, mercury (in Hg) is not higher than 0.2mg/kg;
The used substratum of Chinese medicinal materials Cordyceps militaris (L.) Link. adopts the complete silkworm chrysalis of wet (or through freezing) behind the mulberry cocoon filature; Wet silkworm chrysalis process by following given process behind the described filature: a level sortation-two level sortation-three level sortation-cleaning-blanching-cool off-be used for production or quick-frozen-packing is kept in; Behind work program such as filature, this silkworm chrysalis heavy metal contamination index still meets lead and is not higher than 1mg/kg, arsenic and is not higher than 0.5mg/kg, and mercury is not higher than the standard of 0.2mg/kg.
Among the step B through seed selection, freeze-drying preservation, activate supervisor gained bacterial classification and whether can be used for the Chinese medicinal materials Cordyceps militaris (L.) Link. and produce, depend on whether accord with Q/320691DJK01-2007 standard of final gained Cordyceps militaris (L.) Link..
Cultivate before among the step C and be: silkworm chrysalis or through the complete silkworm chrysalis of the refrigerated cuvette of packing into will wet, sterilize, inoculate, culturing room to preceding cultivation finishes before placing then, charge amount is to account for below 3/5 (L/L) in the transparent small vessels, inoculate with the liquid spawn that is no more than 48 hours cell ages during inoculation, on the silkworm chrysalis of the bacterium of under 100 grades of purification conditions the liquid spawn sprinkle being gone out.The inoculation than 10: 1 to 10: 3 (silkworm chrysalis: liquid spawn, V/V); Preceding culture temperature is controlled between 10 ℃ to 25 ℃ 20 days to 60 days; The back culture vessel adopts the light-passing plastic basket among the step C, and the transparent plastics basket serves as a contrast with five of food fresh keeping membranes or six bread, back culture environment temperature is controlled between 15 ℃ to 30 ℃, 10 ℃ of temperature difference per day, the humidity that also will control environment is 85%, natural scattering light or source of artificial light, illumination 200 to 2000 Luxs; Cultivate gained Chinese medicinal materials Cordyceps militaris (L.) Link., accord with Q/320691DJK01-2007 standard.
The drying of step D adopts low temperature flow-through type warm air drying, and hot blast temperature is controlled between 55 ℃ to 65 ℃, dry 6 to 8 hours.
Use the double-deck two envelope weather-proof dressings of plastics bag during the accumulating of step D packing, lucifuge, refrigeration below 5 ℃.
After handling through step D or again after preserving more than 2 years, Cordyceps militaris (L.) Link. is accord with Q/320691DJK01-2007 standard still.
The Cordyceps militaris (L.) Link. appearance character that the present invention obtains is the complex body of elongated stroma and sclerotium, is covered with mycelia around the sclerotium, and most mycelia presents orange, depth inequality; In the Cordyceps militaris (L.) Link. physical and chemical index, carry out cordycepin with HBLC and differentiate that the positive, content are between the 1.0mg/g to 15.0mg/g.
Dedicated drying equipment comprises blower fan 10, interchanger 9 is set on the blower fan, multilayer charging tray 1,2,3,4,5,6,7,8 is set on the interchanger, and charging tray top is provided with extraction hood 0, is connected to a thermal head 13 on interchanger hot water (vapour) pipe 16 and stretches into temperature controller 11 in the charging tray.Also have supply lead 12, material among the figure every sieve 14, handle 15, backwater (vapour) pipe 17.
When dry: 1: equipment is installed, and oven dry is prepared:
1.1 connect the output line of temperature controller (11) to hot water pump (10).
1.2 connect the temperature controller supply lead.
1.3 connect fan power line (12).
1.4 connect temperature controller heat-sensitive line (13).
1.5 check whether temperature controller (11) work is normal.
1.6 the hot well tank rising pipe is connected the hot water inlet (16) of scatterer (9).
1.7 the hot well tank return water pipe is connected the water return outlet (17) of scatterer (9).
1.8 connection water heater power supply, or vapour pipe.
2: oven dry:
2.1 remove or the air exhaustion pipe cowl of slinging (0).
2.2 move down charging tray (1)-(8).
2.3 be with emgloves, the Cordyceps militaris (L.) Link. lay that will wet is to charging tray (8) ... (1), individual layer lay, slightly space.
2.4 charging tray is built, is built air exhaustion pipe cowl (0).
2.5 open with the by-pass valve control between hot well tank rising pipe and radiator heat water inlet.
2.6 open the temperature controller play button.
2.7 the temperature controller thermal head is placed the bed of material of charging tray (7).
2.8 record, timing.
2.96 about hour, open air exhaustion pipe cowl (0), check the interior Cordyceps militaris (L.) Link. drying regime of charging tray (8).
2.10 the water content inspection, be lower than 8% after, the Cordyceps militaris (L.) Link. dry product in rewinding dish (8)-(1).
Use this method, in June, 2000 is in Jiangsu, Nantong, and 2002 to 2007 in Jiangsu, the Wuxi, the amount of finish 8kg of class, Cordyceps militaris (L.) Link. is hard, crisp, and perverse feel is arranged, and the profile outward appearance is good, and observed watercut is below 7%.
In conjunction with the present invention, can set up following standard:
Chinese medicinal materials Cordyceps militaris (L.) Link. standard
1. Chinese medicinal materials Cordyceps militaris (L.) Link. proterties standard
This product is the complex body of elongated stroma and sclerotium.
The elongated garden of stroma cylindricality bears from sclerotium, and it is uncertain living position, and single giving birth to or all living creatures is upright or crooked, long 0.5-15CM. diameter 0.1-0.5CM, the surface is orange-yellow or orange red, and few branch has thin vertical wrinkle, head expands into ellipse garden shape slightly, blunt garden, top, long 0.5-5CM, diameter 0.2-0.7CM.Matter is pliable and tough, and section is coarse, and the edge is orange-yellow or orange red; Yellow in it to Vandyke brown, the little raw meat of gas, it is little salty to distinguish the flavor of.
The sclerotium profile keeps the pupa shape, and mycelia is seen by mycelia or internode in the surface, and mycelia is yellowish-white, and section is coarse, is full of thread mycelia to orange, and white is to yellow-white; The little raw meat of gas, it is little salty to distinguish the flavor of.
2. Chinese medicinal materials Cordyceps militaris (L.) Link. physical and chemical index
Table 1 physical and chemical index
| Cordycepin is differentiated | Positive |
| Cordycepin mg/100g 〉= | 100 |
| Moisture %≤ | 8.0 |
| Ash content %≤ | 6.0 |
| Plumbous (in Pb) mg/kg≤ | 1.0 |
| Arsenic (in As) mg/kg≤ | 0.5 |
| Mercury (in Hg) mg/kg≤ | 0.2 |
Chinese medicinal materials Cordyceps militaris (L.) Link. physical and chemical index cordycepin is differentiated, content assaying method
(TLC HPLC method)
1 principle and method
Cordycepin contrast crystalline substance and Cordyceps militaris (L.) Link. sample liquid are distinguished point sample in the GF254 thin layer plate, after acetone one chloroform one propyl carbinol developping agent launches, under the UV-light reflectometer, observe spot, with the speckle displacement of reference substance, carry out cordycepin qualitative identification in the sample per sample.
2 key instruments and reagent (summary)
The preparation of 3 reference substances
Precision takes by weighing cordycepin contrast 4.5mg in the 10ml measuring bottle, and methanol constant volume shakes up to scale.
The preparation of 4 samples
Get an amount of Cordyceps militaris (L.) Link., put the dry 1h of 60～C in the loft drier, precision takes by weighing 0.40g, put in the apparatus,Soxhlet's, 50ml adds diethyl ether, 60 ' C Soxhlet is extracted 15h, and 2h volatilizes ether under the taking-up content normal temperature, puts in the 100ml round-bottomed flask, adding the backflow of 95% ethanol boiling water (adds amount of alcohol and is respectively 30ml for three times, 20ml, 20ml) the 1h suction filtration that at every turn refluxes, merging filtrate reclaims ethanol to about 5ml, is transferred in the 25ml measuring bottle, repeatedly wash with distilled water and to make content be transferred in the measuring bottle fully, distilled water is settled to scale, shakes up rearmounted refrigerator cold-storage 12h, the refrigeration centrifugal 20min of liquid (4500r/min).Get supernatant liquor 4ml and boil off solvent, with about 2ml dissolve with methanol.
5 sample thin-layer qualitative are differentiated
5.1 developping agent: acetone one chloroform one propyl carbinol--25% ammoniacal liquor (30: 30: 40; 10).
5.2 the brilliant point sample 5ul of standard, sample spot 5ul, each dot spacing 2cm, put and be deployed into 8cm place (about 35min) in the separation chamber, taking-up is put room temperature and is dried, put under the UV-light reflectometer and to observe (go into=254nm), the result is presented at each sample of the place of identical Rf value with reference substance all same blob.
6 cordycepin contents
6.1 principle and method
Adopt high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia version in 2002) to measure cordycepin content.With octadecylsilane chemically bonded silica is weighting agent: methyl alcohol one water (12: 88) is moving phase: the detection wavelength is 260nib.Number of theoretical plate calculates by the cordycepin peak should be not less than 3000.Measure content of cordycepin in the Cordyceps militaris (L.) Link. with the external standard two-point method.
6.2 key instrument and reagent (summary)
6.3 the preparation of standard solution
The preparation precision of reference substance solution takes by weighing cordycepin reference substance 4.5mg, puts in the 10ml measuring bottle, adds the moving phase constant volume, shake up, accurate absorption 0.1,1.5ml are in the 10ml measuring bottle, and moving phase is settled to scale, shake up, promptly get (every ml contains cordycepin 4.50ug and 67.5ug).
6.4 the preparation of need testing solution
Get an amount of Cordyceps militaris (L.) Link., put the dry 1h of 60 ' C in the loft drier, precision takes by weighing 0.40g, put in the apparatus,Soxhlet's, 50ml adds diethyl ether, 60 ' C Soxhlet is extracted 1.5h, and 1h flings to ether under the taking-up content normal temperature, puts in the 100ml round-bottomed flask, adding the backflow of 90% ethanol boiling water (adds amount of alcohol and is respectively 30ml for three times, 20ml, 20ml) the 1h suction filtration that at every turn refluxes, merging filtrate reclaims ethanol to about 5mL, is transferred in the 25ml measuring bottle, repeatedly wash with distilled water and content to be transferred to fully be in the bottle, distilled water is settled to scale, shakes up rearmounted refrigerator cold-storage 12h, the refrigeration centrifugal 10min of liquid (4500r/min).Get supernatant liquor and cross the 0.45um millipore filtration, promptly.
6.5 measure
Accurate respectively reference substance solution and each 10ml of trial-product of drawing injects hplc determination promptly.
6.6 the result calculates
Press cordycepin content formula in the typical curve calculation sample:
X(％)＝(C*25/W*105)*100％
In the formula:
The percentage composition of cordycepin in X--sample, unit are milligram every hectogram (mg/100g);
The cordycepin amount that C-two points external standard method calculates, unit is every milliliter of microgram (ug/ml):
The sampling amount of w-sample, unit is gram (g).
Calculation result keeps 4 position effective digitals.
Claims (10)
1, a kind of scale cultivation that standardizes is used as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials, it is characterized in that: comprise the following steps:
A: adopt organic mulberry silkworm chrysalis, comprising: obtain silk cocoon from organic silkworm and mulberry base, obtain silkworm chrysalis from special-purpose silkworm chrysalis receipts, storage and processing place then, as the used substratum of Chinese medicinal materials Cordyceps militaris (L.) Link.;
B: adopt being fit to the bacterial classification of Chinese medicinal materials Cordyceps militaris (L.) Link., is through seed selection, freeze-drying preservation, activation, bacterial classification by the bacterial classification check;
C: the code requirement cultural method comprises: preceding cultural method, back cultural method;
D: the code requirement post-treating method comprises: the drying means of gathering, packing conveying method;
E: adopt Chinese medicinal materials Cordyceps militaris (L.) Link. method for guaranteeing quality, comprising: Chinese medicinal materials Cordyceps militaris (L.) Link. quality standard, Chinese medicinal materials Cordyceps militaris (L.) Link. quality determining method, promptly set up the Q/320691DJK01-2007 standard.
2, standardized cultivation according to claim 1 is as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials, it is characterized in that: in the steps A, silk cocoon is cutd open cocoon, detects the silkworm chrysalis heavy metal, weigh the silk cocoon quality: plumbous (in Pb, dry weight, down with) be not higher than 1mg/kg, arsenic (in As) is not higher than 0.5mg/kg, mercury (in Hg) is not higher than 0.2mg/kg.
3, standardized cultivation according to claim 1 is characterized in that as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials: the used substratum of Chinese medicinal materials Cordyceps militaris (L.) Link. adopts the complete silkworm chrysalis of wet (or through freezing) behind the mulberry cocoon filature; Wet silkworm chrysalis process by following given process behind the described filature: a level sortation-two level sortation-three level sortation-cleaning-blanching-cool off-be used for production or quick-frozen-packing is kept in; Behind work program such as filature, this silkworm chrysalis heavy metal contamination index still meets lead and is not higher than 1mg/kg, arsenic and is not higher than 0.5mg/kg, and mercury is not higher than the standard of 0.2mg/kg.
4, standardized cultivation according to claim 1 is as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials, it is characterized in that: among the step B through seed selection, freeze-drying preservation, activate supervisor gained bacterial classification and whether can be used for the Chinese medicinal materials Cordyceps militaris (L.) Link. and produce, depend on whether accord with Q/320691DJK01-2007 standard of final gained Cordyceps militaris (L.) Link..
5, standardized cultivation according to claim 1 is as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials, it is characterized in that: cultivate before among the step C and be: silkworm chrysalis or through the complete silkworm chrysalis of the refrigerated cuvette of packing into will wet, sterilize, inoculate, culturing room to preceding cultivation finishes before placing then, charge amount is to account for below 3/5 (V/V) in the transparent small vessels, inoculate with the liquid spawn that is no more than 48 hours cell ages during inoculation, on the silkworm chrysalis of the bacterium of under 100 grades of purification conditions the liquid spawn sprinkle being gone out.The inoculation than 10: 1 to 10: 3 (silkworm chrysalis: liquid spawn, V/V); Preceding culture temperature is controlled between 10 ℃ to 25 ℃ 20 days to 60 days; The back culture vessel adopts the light-passing plastic basket among the step C, and the transparent plastics basket serves as a contrast with five of food fresh keeping membranes or six bread, back culture environment temperature is controlled between 15 ℃ to 30 ℃, 10 ℃ of temperature difference per day, the humidity that also will control environment is 85%, natural scattering light or source of artificial light, illumination 200 to 2000 Luxs; Cultivate gained Chinese medicinal materials Cordyceps militaris (L.) Link., accord with Q/320691DJK01-2007 standard.
6, standardized cultivation according to claim 1 is characterized in that as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials: the drying of step D adopts low temperature flow-through type warm air drying, and hot blast temperature is controlled between 55 ℃ to 65 ℃, dry 6 to 8 hours.
7, standardized cultivation according to claim 6 is as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials, it is characterized in that: described low temperature flow-through type warm air drying adopts simple and easy low temperature flow-through type hot-air drier, comprise blower fan, interchanger is set on the blower fan, the multilayer charging tray is set on the interchanger, charging tray top is provided with extraction hood, is connected to a thermal head on the exchanger heat water pipe pump and stretches into temperature controller in the charging tray.
8, standardized cultivation according to claim 1 is characterized in that as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials: use the double-deck two envelope weather-proof dressings of plastics bag during the accumulating of step D packing, lucifuge, refrigeration below 5 ℃.
9, standardized cultivation according to claim 1 is characterized in that as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials: after handling through step D or again after preserving more than 2 years, Cordyceps militaris (L.) Link. is accord with Q/320691DJK01-2007 standard still.
10, standardized cultivation according to claim 1 is characterized in that as the method for the Cordyceps militaris (L.) Link. of Chinese medicinal materials: as one of index that detects the Chinese medicinal materials Cordyceps militaris (L.) Link., cordycepin content is not less than 1mg/g with cordycepin content.
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