KR100405990B1 - Processs for Preparing Organic Germanium from Cordyceps militaris - Google Patents

Abstract

The present invention is a cordyceps militaris mycelium containing a high concentration of organic germanium by liquid culture and pupa culture of Cordyceps sinensis and

The present invention relates to a method for producing a fruiting body. Cicada cordyceps mycelia were prepared by liquid culture, and inorganic germanium (GeO2) was added to the liquid culture medium (glucose 40g / L, yeast extract 10g / L, peptone 10g / L) to cultivate cicada cordyceps. In the chrysalis medium (chrysalis 200g), 100 ppm of inorganic germanium (GeO2) was added and cultured cicada cordyceps to prepare fruiting bodies and mycelium containing high concentration of organic germanium.

Description

Process for Preparing Organic Germanium Cordyceps sinensis {Processs for Preparing Organic Germanium from Cordyceps militaris}

Technology Field: Microorganism (Central Cordyceps) Cultivation Technology and Organic Germanium Production Technology

Prior art: To date, Cordyceps sinensis in medium without added inorganic germanium (GeO2)

Incubation failed to produce cordyceps containing organic germanium.

Cordyceps sinensis is a word that means insects in winter and mushrooms in summer. Actually, the spores of the fungus, Cordyceps fungus, present in soil, etc., live in the respiratory, digestive, joints, etc. When the host (host) insect dies while entering the body and eating nutrients and developing growth, after a while, a fruit-shaped fruit-shaped body is formed on the epidermis of insects to spread the offspring. Insects that Cordyceps sinensis uses for its own growth are characterized by maintaining a shape like a mummy without decay until the fruiting body (mushroom) of Cordyceps is released. Cordyceps sinensis is a kind of insect parasitic medicinal (edible) mushroom, and belongs to the genus Cordyceps sinus fungus. Recent studies have shown excellent efficacy in anti-cancer, immune enhancement, chronic fatigue, and anti-aging. In Japan, it is sold as a drug for lung disease and pleurisy. Cordyceps attracts attention because it is highly likely to be developed as a pollution-free pesticide, an anticancer drug or a health supplement food.In Korea, the pharmacological effects were jointly researched by the Rural Development Administration and the Seoul National University Natural Products Science Research Institute. It turns out to be excellent. Cordyceps sinensis contains a very important substance called cordycepin (3'-deoxy-adenosine), which has been identified as an isomer of quinic acid. It is activated to prevent normal cells from becoming cancer cells. Cordyceps sinensis is recognized not only as a medicine but also as a dietary supplement, and various powder and liquid products are currently on the market. In addition, research on pollution-free microbial pesticides by using Cordyceps sinensis to control the density of insects in the natural ecosystem. The development of such natural biological pesticides has attracted great attention because microorganisms, which are natural enemies of pests, can solve not only pest control but also environmental pollution problems caused by chemical pesticides.

Organo germanium can be used for intracellular oxygen supply, blood purification and blood circulation improvement, activation of immune NK cells and macrophages, induction of immune function by inducing interferon secretion, enhancement of natural healing ability, inhibition of cancer cell development and metastasis, reduction of osteoporosis, It is known to be effective in preventing hypertension, lowering cholesterol, preventing hepatitis, delaying aging, dementia, analgesic and anti-inflammatory effects, brain cell activity, and fatigue recovery.In developed countries such as the United States and Japan, organic germanium is used for cancer and heart disease. Research to treat incurable diseases such as back is actively being conducted.

Clinical studies have also been reported to be effective in brain tumors, lung cancer, arthritis, leukemia, Kaposi's sarcoma, alcoholism, rheumatoid arthritis in children, and about 20-30% increase in cell activity in skin cell activation effect experiments. It is effective and does not appear to be toxic in the cytotoxicity test at all, and is expected to be used for skin aging prevention and wrinkle removal.

To date, no report has been made on the production of organic germanium using Cordyceps sinensis.However, other methods of producing organic germanium using microorganisms include adding selenium to culture medium of edible yeast to produce organic selenium. A method is disclosed, and Japanese Patent Laid-Open No. 53-130483 discloses only a yeast containing germanium and a manufacturing method thereof. Patent No. 10-1997-138078 discloses a new technology and method for preparing a medium for cordyceps cultivation.

Therefore, in order to develop edible mushrooms containing new pharmacologically active substances by containing (combining) organic germanium in Cordyceps sinensis, which is currently spotlighted as a dietary supplement, Cordyceps sinensis is stronger than other microorganisms, and in terms of yield of cell growth. Focusing on its superiority, Cordyceps militaris , a cicada cordyceps isolated from Korea, was adapted to medium concentrations of inorganic germanium, and then grown on high concentrations of germanium-containing medium to make germanium into mycelia and fruiting bodies. A large amount of ions were transferred to produce mycelia and fruiting bodies containing high concentrations of organo germanium.

Therefore, in one aspect, the present invention is aseptically inoculated mycelium of Cordyceps militaris into a liquid culture medium containing 10 g / L yeast extract, 10 g / L peptone, 40 g / L glucose and 100 ppm of inorganic germanium. Shaking cultured at 25 ° C. for 170 hours at 170 rpm for 120 hours to allow the mycelium to adapt to germanium and to increase the amount of inorganic germanium to 500ppm, 1000ppm and 5000ppm in the same medium. In the same manner, the present invention provides a method for producing a mycelium containing a high concentration of organic germanium, comprising repeating culturing to induce high concentration of organic germanium in the mycelia and fruiting bodies. in, the mycelium of the cicada Cordyceps (Cordyceps militaris) yeast extract 10g / L, peptone 10g / L Aseptically inoculating a liquid culture medium containing 40 g / L of glucose, shaking culture the mycelium at 170 rpm for about 120 hours at 25 ° C, and autoclaving the liquid culture mycelium at 121 ° C for 1 hour. And inoculating a solid pupa medium containing inorganic germanium (100 ppm, 0.144 g / L), incubating the medium for 5 days at 25 ° C. until the white mycelium is spread throughout the pupa, and the medium is cultured. It provides a method for producing mycelium and fruiting body containing a high concentration of organic germanium, characterized in that it comprises the step of culturing about 60 days until the orange fruiting body is lowered to below 20 ℃. It is distributed in the form of follicular spores or conidia of the insects, and when the temperature and humidity are high from June to September, Occurs on the soft skin of the extinguisher, node, etc. They are mostly found in valleys and in wetlands with relatively small weeds. As the Cordyceps sinensis is a fungus that occurs mainly in the natural state, it was difficult to cultivate the strain by artificial culture. However, artificial cultures have been carried out in laboratory conditions for some time, and mass cultures have recently become possible. The components and conditions of the general artificial culture medium of Cordyceps sinensis are well known, and the closer to nature, the easier it is to cultivate Cordyceps sinensis. In this patent, by using a method of adding inorganic germanium, which has not been reported so far, to produce a Cordyceps herb culture medium, the production yield of mycelia is high, and it is intended to produce Cordyceps sinensis containing high concentration of organic germanium in the mycelium and fruiting bodies.

Referring to the method of cultivating the cultivated Militaris, which is the strain of the present invention, and the results thereof are as follows.

1. Production of Mycelia by Liquid Culture: Cordyceps

militaris ) yeast extract 10g / L, peptone 10g / L, glucose 40g / L, inorganic germanium

Aseptically inoculated into a liquid medium containing 0.144g / L (100ppm) to germanium

Adaptation was made. The strain grown by adapting to 100ppm concentration

Inoculate and culture in 500ppm, 1000ppm, 5000ppm concentration medium

Induced to contain a high concentration of germanium. After culture, only mycelium is collected

After thoroughly washing and drying, the content of organic germanium was measured.

Organic of various natural products known to contain organic germanium

Germanium content and organic germanium in Cordyceps mycelium prepared by this method

Content is as Table 1 and Table 2.

Table 1 Organic Germanium Content of Various Natural Products

Botanical name ppm / g Botanical name ppm / g Ashidaba 4.6 ppm Wolfberry 1.24 ppm Ginseng 2.5-3.2 ppm Company 1.52 ppm Polyphora Mushroom 8 ppm Twill 2.39ppm garlic 7.54 ppm Class 1.08ppm Ganoderma lucidum mushroom 3.5 ppm Rule 0.5 ppm Triceps 2.57ppm Aloe 0.72 ppm Go 2.62ppm reimbursement 0.72 ppm

[Table 2] In Mycelial Dried Mycelia of Liquid Cultured at Various Inorganic Germanium Concentrations

Organic Germanium Content

Inorganic Germanium Concentration (ppm) Content (ppm / g) 0 0 ppm 100 442.4 ppm 500 853.6 ppm 1000 1404.2 ppm 5000 4509.7 ppm

As shown in Tables 1 and 2, the amount of organic germanium contained in various natural products and the amount of germanium contained in the liquid cultured Cordyceps mycelium by artificially adding inorganic germanium were found to contain the most organic germanium. Cordyceps sinensis-dried mycelium, which had been liquid-cultured than polypora mushrooms, was found to contain as little as 55 to 560 times organic germanium.

2. Production of Mycelium and Fruiting Body by Chrysalis of Chrysalis: A small amount of liquid culture medium containing appropriate amount of mycelium was inoculated into chrysalis medium containing inorganic germanium (100 ppm) prepared by high temperature and high pressure sterilization. After incubating for a sufficient period of time to form the fruiting body, the mycelium part and the fruiting body part were separated and washed thoroughly. After drying the mycelium and fruiting body, the result of measuring the content of organic germanium is shown in Table 3.

[Table 3] Organic germanium content of Cordyceps mycelia and fruiting bodies in pupa culture

Inorganic Germanium Concentration (ppm) Content (ppm / g) Fruiting body Mycelium 0 0 0 100 173 284

As shown in Table 3, Cordyceps mycelium and fruiting bodies produced by solid culture (chrysalis medium culture) also contained a high amount of organic germanium, and compared with the previously reported natural products, about 21 times more in fruiting bodies, mycelium. Was found to contain about 35 times more organic germanium.

When explaining the organic germanium production method using the Cordyceps sinensis of the present invention in more detail.

The liquid culture of this strain was inoculated with a Cordyceps in 1000mL culture flask containing 200mL SDAY medium consisting of 10g / L yeast extract, 10g / L peptone, 40g / L glucose and 1.44 ~ 7.2g / L inorganic germanium. The stirring was performed at 170 rpm and a temperature of 25 ° C. for about 5 days, and the change in pH during the culture was ignored.

Example 1

Cicada cordyceps isolated from Korea was inoculated into 200 mL of germanium-SDAY medium containing 10 g / L yeast extract, 10 g / L peptone, 40 g / L glucose, and 0.144 g / L inorganic germanium. Incubate for about 120 (5 days). The culture solution was centrifuged to collect only the cells, washed twice with physiological saline, and washed once with tertiary distilled water, and then dried in a 75 ° C. hot air dryer for 20 hours to prepare dried mycelium samples.

Example 2

SDAY medium (Yeast extract 10g / L, Peptone 10g / L, Glucose 40g) with 0.144g / L (100ppm) of inorganic germanium and upgrading the Cordyceps sinensis strain with germanium adaptation to 500ppm, 1000ppm and 5000ppm / L) under the same culture and washing conditions as in Example 1 to prepare a Cordyceps sinensis dried sample containing a high concentration of germanium.

Example 3

Chrysalis culture of pupa: 5 ml of Cordyceps sinensis culture cultured in a liquid medium containing no organic germanium and 10 ml of inorganic germanium 100ppm (0.144g / L) concentration in pupa 200g and inoculated in pupa medium prepared by autoclaving at 121 ℃ for 1 hour After shaking the pupa medium and mycelium well mixed well. When the white mycelium was spread throughout the pupa at 25 ° C for 5 days, the culture temperature was lowered to 18 ° C to change from nutrient growth to reproductive growth conditions to form the fruiting body of Cordyceps sinensis. When the temperature rises above 20 ℃ during the reproductive growth period, the fruiting body of Cordyceps sinensis is not formed, so special attention is required to change the temperature. After about 20 days of reproductive growth conditions, the orange fruiting body was raised from the pupa, and after about 60 days, the whole fruiting body was formed. The fruiting body part and the pupa part under the fruiting body were separated separately, and the surface of the mycelium of the pupa part was washed twice with third distilled water to thoroughly remove the inorganic germanium remaining on the cell surface. Drying for 20 hours in a 75 ℃ hot air dryer to prepare an organic germanium-containing dried samples (mycelium and fruiting body).

Using the Cordyceps sinensis strain, it was possible to prepare organic germanium-containing Cordyceps with high yield and high productivity by using SDAY medium (yeast extract 10g / L, peptone 1.0g / L, glucose 40g / L) and culture of pupa. In addition, the developed organic germanium-containing Cordyceps sinensis will be spotlighted as a dietary supplement and pharmacologically active product by adding a pharmacologically active substance called organic germanium to the existing Cordyceps sinensis efficacy. Furthermore, it can also be used as a cosmetic raw material and pharmaceutical raw material.

Claims (5)

Aseptic inoculation of mycelia of Cordyceps militaris into a liquid culture medium containing 10 g / L yeast extract, 10 g / L peptone, 40 g / L glucose and 100 ppm of inorganic germanium, Shaking and incubating the mycelium at 170 rpm for about 120 hours at 25 ° C. to make the mycelium adaptable to germanium; and Inducing the organic germanium to be contained in the mycelium and fruiting body at high concentration by repeating the same culture in the same medium gradually increasing the amount of inorganic germanium to 500ppm, 1000ppm and 5000ppm by adjusting to 100ppm concentration Cicada Cordyceps sinensis mycelium (mycelium) containing a high concentration of organic germanium comprising a. Mycelium of Cicada Cordyceps sinensis containing organo germanium produced according to claim 1. delete Aseptic inoculation of mycelia of Cicada cordyceps militaris in a liquid culture medium containing 10 g / L yeast extract, 10 g / L peptone and 40 g / L glucose, Shaking the mycelium for about 120 hours at 25 rpm at 170 rpm Preparing the liquid culture mycelium by autoclaving the pupa at 121 ° C. for 1 hour and inoculating a solid pupa medium containing inorganic germanium (100 ppm, 0.144 g / L), Incubating the medium for 5 days at 25 ° C. until white mycelium spreads throughout the pupa, and Incubate the medium for 60 days until the culture temperature is lowered to 20 ° C. or lower until an orange fruiting body appears. Method for producing a mycelium and fruiting body containing a high concentration of organic germanium comprising a. Of Cicada Cordyceps Sinensis Containing Organic Germanium Produced According to Claim 4 Mycelium and Fruiting Body.