KR20060022402A - Artificially cultivation method of paecilomyces tenuipes use protaetia orientalis larva - Google Patents

Abstract

The present invention relates to a method of artificially cultivating a snowfly Cordyceps sinensis by inoculating a snowhopper Cordyceps larva into a white spotted larva, causing an infection, and then forming a fruiting body from the flower larva infected with the bacterium. Pretreatment of the white spotted flower larva larvae were incubated in glutinous rice flour for 3 days and then washed with water; Inoculating the Cordyceps sinensis strain on the host formed by the step and culturing the mycelium; It is composed of the step of inducing fruiting bodies to the growth chamber in which white spots are filled with cordyceps fungi in the interior of the flower larva.

 White spotted flower larvae, Cordyceps sinensis, artificial cultivation

Description

Artificially cultivation method of Paecilomyces tenuipes use Protaetia orientalis larva}

The present invention relates to a method of artificial cultivation of Paecilomyces tenuipes based on the white-spotted larva ( Protaetia orientalis ) larva, more specifically, to inoculate the infection by inoculating the strains of the Snow creeper on the white-spotted larva It relates to a method for artificially cultivating snow Cordyceps sinensis by forming fruiting bodies from the flower larvae infected with the bacteria after induction.

In general, Cordyceps sinensis is derived from insects in winter and flower (grass) in summer. It is a type of sperm fungi in which fruiting bodies are formed. Insects live in the body of insects when temperature / humidity increases. It is a kind of medicinal mushroom that enters and develops, kills host insects, and forms fruiting bodies on the epidermis of insects.

Conventionally, Cordyceps sinensis refers to Cordyceps sinensis , which is a cordyceps from larvae of bat moths, and today it is collectively called insects from mushrooms, spiders and fungi as well as insects. Therefore, it can be classified into Cordyceps sinensis, Cordyceps militaris and Paecilomyces japonica . In addition, the pupa cordyceps and snowflower cordyceps may have a common point in that they can be based on the chrysalis (especially silkworm pupa) of the insect Lepidoptera.

Cordyceps sinensis is an expensive herbal medicine that has been used in China since ancient times as an antidote for tuberculosis, asthma, jaundice and opioid poisoning, as a remedial and tonic agent, and as an immune enhancer (Humber, RA 1990. Fungal pathogens of insects, spiders, and mites; isolation, preservation, and identification.USDA Agricultural Research Service; Jianzhe, Y., Xiaoloan, M., Qiming, M., Yichen, Z. and Huaan, W. 1989.Icons of medicinal fungi from China. China.p. 575; Kobayasi, Y. 1940.The genus and its allies.Sci.Rept.Tokyo Bunrika Daikaku, Sect.B., 5: 53-260; Shimiizu, D. 1994.Color iconography of vegetable sasps and plant worms.Seibundo Shinkosha.Japan.pp.38), with regard to fruiting bodies of Cordyceps sinensis, cordycepin, the main component of fruiting bodies, has been found to have anticancer activity (Cunningham, KG, W. Manson. FS Spring and SA Hutchinson. 1950. Cordycepin, a metabolic product from cultures of (Linn.) Link.nature, 166. 949), In China, fruiting bodies are used as royal dishes, while in Japan, cordyceps is used for medicinal purposes.

Although the cordyceps is collected in the natural state, the collection of fruiting bodies in the natural state is very difficult and the supply is limited, and as the weathering of the fruiting bodies becomes more difficult due to the phenomena due to the recent destruction of the ecosystem and pollution, Stable artificial production techniques are required (Basith, M., and Madelin, MF 1968. Studies on the production of perithecial stromata by in artificial culture. Canadian Journal of Botany. 46: 473-480). The fruiting body production technology of Cordyceps sinensis includes the method of collecting fruiting body in the natural state, spawn culture and cultivation method using silkworm pupa, and mass culture method using artificial medium.

However, as mentioned above, the collection of fruiting bodies in the natural state is becoming very difficult. Therefore, it is unable to adequately cope with the increase in domestic and foreign demand, and the spawn culture and cultivation method using silkworm pupa, and the mass cultivation method using artificial medium In recent years, research has been actively conducted.

However, it is currently difficult to form fruiting bodies, except for Paecilomyces japonica , in farmhouses, because first, the difference in fruiting ability is severe, and secondly, only the brown rice or silkworm chrysalis can be used independently or mixed to add the medium. There is a lack of research on third, and third, there was a problem that can not adequately provide cultivation conditions such as light (light) conditions, ventilation, humidity.

The present invention, the special treatment of white spotted larva larva known to have excellent efficacy in the prevention and treatment of various geriatric diseases and improving the function of the liver, and based on the development of a method to artificially cultivate Paecilomyces tenuipes The purpose is to provide a method that can be easily produced in.

Characteristic technical configuration of the present invention for achieving the above object, the seed culture of snow Cordyceps sinensis using a liquid medium using a PDA (potato dextrose agar) or potato extract, the culture of inoculated mycelium is white spotted flower larva After pretreatment, the internal humidity is maintained at 70 ± 5%, inoculated with liquid seed and incubated for 6-8 days at 25 ± 2 ℃. In addition, the conditions for forming the fruiting body are at a temperature of 18 ± 2 ° C. and a humidity of 90 to 98%.

Referring to the artificial cultivation method of the snow Cordyceps sinensis based on the white spotted flower larva larva of the present invention having the characteristics as described above in detail as follows.

First, the white-spotted flower larva used in the present invention belongs to the caterpillar chafer family of taxonomy, and is a 1-environmental insect whose life cycle is completed once a year. After the age of June to July, the adult to allegory. White spotted flower larvae are known to be effective in improving liver disease and liver function, such as cirrhosis and liver cancer, in the private sector, and are also known to be effective in restorative effects and preventive treatment of various adult diseases.

Artificial cultivation method of the snowfly Cordyceps sinensis based on the white spotted flower larva larva according to the present invention having the above-mentioned efficacy, the first step of artificially cultivating the Cordyceps worms from the hostess white spotted flower larva Live stage larvae are incubated in glutinous rice flour for 3-5 days and then washed with water to maintain internal humidity at 70 ± 5%.

The reason for culturing the live larvae in glutinous rice flour is to supply nutrients necessary for internal growth and fruiting body formation, and to facilitate the excretion of residues left in the body of the larvae. In addition, the reason for incubating 3-5 days when incubating the larvae in the glutinous rice flour is to ensure a sufficient excretion period of the residues present in the larvae during this period to ensure complete excretion.

In addition, when the host's internal humidity is 75% or more, the bacteria are inhibited from sticking, and when the host's internal humidity is less than 65%, the fruiting body is poorly formed, so the host's internal humidity should be maintained at 70 ± 5%. In addition, after sterilization, the external humidity becomes excessive, which may act as a deterrent to the growth of the initial bacteria, and 5 to 10% of the chrysalis maintains 50 ± 5% of humidity.

In order to prevent excessive humidity after the sterilization, the reason why the pupa is maintained at 50 ± 5% of humidity is that even if the moisture is adjusted before sterilization, the internal / external humidity of the larva during sterilization may be excessive. Adding chrysalis containing 5% water can suppress this. In addition, the reason of adding 5 to 10% of the chrysalis, if less than 5% is difficult to control the humidity of the larva in the culture vessel, when more than 10% in the mycelial cultivation of the external growth rather than the internal growth of the fruiting body formation efficiency This is because there is a problem falling.

In the second step, the inoculation of Cordyceps sinensis on the host formed by the above step and incubating the mycelia, the liquid spawn is the edge of the mycelium cultured for 6-8 days at 25 ± 2 ℃ in PDA (Potato Dextrose Agar) medium Cut to 3 ± 1mm in diameter and inoculate the slice into 500 ± 5ml of potato extraction medium and incubate for 4-6 days at 160 ± 10rpm at 25 ± 2 ℃.

 In this case, the reason for using PDA (Potato Dextrose Agar) in preparing the liquid spawn is the most commonly used solid medium for the culture of Cordyceps sinensis mycelia, as reported in many papers. The reason for culturing at 25 ± 1 ° C. in the PDA medium is that the optimum temperature of Cordyceps mycelia culture is 25 ± 1 ° C., and the cut sections are inoculated in potato extraction medium and inoculated in potato extraction medium at 25 ± 2 ° C. The reason is that the potato extract medium is a natural product medium, which is cheaper than PDB (synthetic medium) medium, and is easy to cultivate liquid seed in large quantities by providing sufficient vitality during nutrition growth of mycelia.

The cultured liquid spawn was inoculated at a culture bottle in which 100 g of white spotted larvae were inoculated at ≤5% and then cultured for 7-8 days at a temperature of 25 ± 2 ° C., a humidity of less than 70%, and ventilation of 7-8 times / day. The reason why the larvae are inoculated in a cultured bottle with ≤5% is because inoculation of more than 5% may form a mass when the larvae are externally grown, which may inhibit the growth and deterioration of the number of feet when fruiting occurs. Doing so is a waste of liquid spawn. When the inoculation is performed, the temperature 25 ± 2 ° C., humidity less than 70%, and the ventilation 7 to 8 times / day is the reason why the optimal temperature and humidity for mycelial growth of Cordyceps fungi are less than 25 ° C. and 70% and high temperature and high humidity. Mycelial growth is slow and contamination rate by bacteria is high. In addition, poor ventilation results in a slow growth of the mycelium. Therefore, it is preferable to satisfy the above conditions.

In the third step, the fruiting body is formed, and the white spotted insecticidal fungus is filled with white fungus inside the larva to induce the fruiting body formation. The basic conditions of culture in the growth room are maintained at temperature 18 ± 2 ℃, humidity 90 ± 8%, brightness 250 ± 5Lux and ventilation is performed 3-5 times per day.

After 2-3 days of cultivation in the growth room, open the lid of the cultivation bottle and fully humidify it with humidity ≥95% for 3 days and confirm the formation of the fruiting body. Incubate until mature enough. The maturation period of the fruiting body takes about 3 weeks, and after 40 days of the first inoculation, it produces fully grown Cordyceps sinensis.

The reason for maintaining the temperature of the culture conditions of 18 ± 2 ℃, humidity 90 ± 8%, luminous intensity 250 ± 10Lux, the temperature for inducing the formation of the fruiting body is 18 ℃, if the lower can be feet but enough Not only does it grow, it grows slowly. At higher temperatures, fruit body formation is not induced. In case of humidity, more than 95% of humidity is required at the initial stage (about 3 days), and it is enough to keep it between 90 ~ 98% after the progression. If it is less than 90%, even if the fruiting body is formed does not grow enough or agglomeration occurs, and the spores are formed early.

In addition, in the case of matt or excessive light conditions, the growth of the fruiting body is slow or stops in the middle of growth, white aerial mycelium is formed, spores are formed early.

Mycelial culture medium

PDA badge Liquid spawn medium -Potatoes, Infusion from 200g / L-Dextrose: 2%-Agar: 2%                                                                                                                                      -Potato extract: 300ml / L-Sugar: 3% ※ Potato extract is chopped 200g of potatoes in 1 liter of water and boiled for 30 minutes, then use the supernatant from which potatoes are removed.

As described above, the present invention is known that the white spotted larva larvae host is known to have a good effect on the improvement of liver function and disease, Cordyceps grown as a substrate is grown from insects containing the nutrients Various bioactive substances that come out can be used in various ways for the treatment of human health and disease, and also the more stable cultivation / production of Cordyceps sinensis based on white spotted larvae.

Claims (6)

Pretreatment step of the white spotted flower larva which is incubated for 3 days in the live larvae in glutinous rice flour and then washed with water; Inoculating the Cordyceps sinensis strain on the host formed by the step and culturing the mycelium; An artificial cultivation method of snow cordyceps, based on white spotted flower larvae, comprising the step of inducing the fruiting body formation of the white spotted insecticidal fungus inside the state of the white larvae.  According to claim 1, White spotted pupa to maintain the humidity of 50 ± 5% in order to maintain the internal humidity of the host at 70 ± 5% in the pretreatment of the flower larva, and to prevent excessive external humidity after sterilization An artificial cultivation method of Snow Cordyceps sinensis based on white spotted flower larvae, characterized by adding 5 to 10% of powder.  The method of claim 1, inoculating the Cordyceps sinensis strain on the host and culturing the mycelium, the liquid spawn cuts the edge of the mycelium cultured at 25 ± 1 ° C. in a PDA (Potato Dextrose Agar) medium and then removes the slices from the potato. Method of artificial cultivation of Snow Cordyceps sinensis based on white spotted flower larvae, inoculated into the medium and shake cultured at 25 ± 2 ° C.  The method of claim 1, wherein inoculating the Cordyceps sinensis strain on the host and culturing the mycelium, the cultured liquid spawn is inoculated at ≤5% in a culture bottle infected with white spotted flower larvae, followed by a temperature of 25 ± 2 ° C. A method of artificial cultivation of snowworm Cordyceps sinensis based on white spotted flower larvae, characterized by cultivation with less than 70% humidity and ventilation.  The white spotted flower according to claim 1, wherein the culture conditions in the growth chamber in the step of forming the fruiting body are performed by maintaining ventilation at a temperature of 18 ± 2 ° C., a humidity of 90 ± 8%, and a brightness of 250 ± 10 Lux. Artificial cultivation method of Snow Cordyceps sinensis based on plain larvae. Cordyceps sinensis cultivated mainly from white spotted flower larvae.