Culture material and cultivation method of ganoderma lucidum antler

Abstract

The invention provides a culture material and a cultivation method of ganoderma lucidum antler. The culture material and the cultivation method are used for increasing the concentration of ganoderma lucidum alcohol F in a ganoderma lucidum antler extract. The method comprises the following steps: preparing a culture material, uniformly mixing 38-42 parts by weight of corncob, 6.65-7.35 parts by weight of wheat bran, 6.65-7.35 parts by weight of rice bran, 19-21 parts by weight of seaweed meal, 23.75-26.25 parts by weight of sawdust and 0.95-1.05 parts by weight of cane sugar, bagging and sealing to obtain a fungus bag; thirdly, sterilizing the fungus bag, and inoculating the fungus bag with a strain of Ganoderma lucidum antler to generate a strain bag; and then the strain bag is moved into a strain room for spawn running. Then, the strain bag is moved into a closed culture room to be cultured, and sporocarp grows out. And finally, harvesting the ganoderma lucidum antler when the ganoderma lucidum antler grows until the stipe length of the sporocarp is 20-25cm and the spore powder is not released.

Description

Technical Field

[0001] The present invention relates to a fungus culture medium and a cultivation method, and in particular to a culture medium and a cultivation method using antler ganoderma. Background Art

[0002] Most of the common Ganoderma species are red Ganoderma and pine Ganoderma, which are mostly semicircular or fan-shaped. However, the antler-shaped Ganoderma is the most precious and rare. Because of its antler-like appearance, it is named "antler Ganoderma" (Ganoderma Amboinense or Ganoderma Lucidum (antle)). When grown, the color is mostly waxy, tending to red or brown. When it grows to a height of about 20 cm and the top is hazy white, it is called antler velvet. At this time, the vitality is the strongest and the nutritional content is the highest.

[0003] Ganoderiol is one of the active ingredients in Ganoderma lucidum, which has multiple physiological activities such as anti-oxidation, anti-inflammatory, immunomodulatory and anti-tumor. These properties make Ganoderiol the focus of research and application of many health issues. Ganoderiol is believed to have benefits in strengthening the immune system, regulating cholesterol and blood pressure, anti-inflammatory, anti-oxidant, anti-tumor, anti-allergic and promoting cardiovascular health.

[0004] Modern nutritional medicine has attached increasing importance to the study of Ganoderma lucidum alcohol, and many scientific studies have been devoted to exploring its biological activity and health benefits, and looking for potential applications in various diseases.

[0005] Therefore, how to develop a culture medium and a cultivation method for Ganoderma lucidum to increase the amount of Ganoderma lucidum alcohol extracted from Ganoderma lucidum has become a problem to be solved in the relevant technical field. Summary of the invention

[0006] To solve the above problems, according to one embodiment of the present invention, a culture medium of Ganoderma lucidum is provided to increase the concentration of Ganoderma lucidum F in Ganoderma lucidum extract, which comprises 38 to 42 parts by weight of corn cobs, 23.75 to 26.25 parts by weight of sawdust, 19 to 21 parts by weight of seaweed powder, 6.65 to 7.35 parts by weight of wheat bran, 6.65 to 7.35 parts by weight of rice bran, and 0.95 to 1.05 parts by weight of sucrose.

[0007] According to another embodiment of the present invention, a cultivation method of antler ganoderma is provided to increase the concentration of ganoderin F in antler ganoderma extract, which comprises the following steps: a culture medium preparation step, a sterilization step, an inoculation step, a mycelium growth management step, a mycelium production management step and a harvesting step.

[0008] The above-mentioned culture medium preparation step is to mix the culture medium of the above-mentioned Ganoderma lucidum evenly, and the culture medium of the above-mentioned Ganoderma lucidum is bagged and sealed to obtain a bacterial bag. The above-mentioned sterilization step is to place the above-mentioned bacterial bag in a sterilization chamber at a sterilization temperature for sterilization, and then cool the above-mentioned bacterial bag to room temperature. The above-mentioned inoculation step is to inoculate the strain of Ganoderma lucidum on the culture medium of Ganoderma lucidum in the above-mentioned bacterial bag in a sterile inoculation room to produce a bacterial bag. The above-mentioned mycelium growth management step is to move the above-mentioned bacterial bag into a dry, ventilated, and light-free bacterial room for cultivation, so that the above-mentioned bacterial bag can be fermented. The above-mentioned mycelium production management step is to move the above-mentioned bacterial bag that has been fermented into a closed culture room, open the bacterial bag for cultivation, and grow the fruiting body of Ganoderma lucidum. The above-mentioned harvesting step is to harvest the above-mentioned Ganoderma lucidum when the stipe length of the above-mentioned fruiting body is 20-25cm and the spore powder has not yet been released.

[0009] According to another embodiment, the sterilization temperature is 121-123°C.

[0010] According to another embodiment, the mycelium growth management step lasts for 22 days, the temperature of the mushroom room is controlled at 23-28°C from day 1 to day 10, and the temperature of the mushroom room is controlled at 25-28°C from day 11 to day 22.

[0011] According to another embodiment, the temperature of the culture room in the above-mentioned mushroom management step is controlled at 25-28°C, and the relative humidity is maintained at around 90% by spraying water or watering on the ground. The light is illuminated for 10 hours a day, and the light intensity is 2000-5000 lux (LX).

[0012] According to another embodiment, spraying and watering are stopped 7 days before the antler ganoderma matures.

[0013] According to another embodiment, the illumination is provided by a fluorescent lamp as a light source.

[0014] According to another embodiment, the above-mentioned culture chamber is kept ventilated for a period of time every day.

[0015] Thus, the cultivation method of the antler ganoderma lucidum of the present invention increases the content of the Ganoderiol F component that can be extracted by the same extraction method in the subsequent step through the new formula of the antler ganoderma lucidum culture medium. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] In order to make the above-mentioned technology and other purposes, features, advantages and embodiments of the present invention more clearly understood, the attached drawings are described as follows:

[0017] Figure 1 Depicted is a tandem mass spectrum (MS / MS) of an 8 ppm ganoderin F standard sample according to an embodiment of the present invention.

[0018] Figure 2Plotted is a concentration analysis table of Ganoderin F according to Example 1 of the present invention. DETAILED DESCRIPTION

[0019] The following will illustrate specific embodiments of the present invention with reference to the drawings, so that those skilled in the art can fully utilize and practice the present invention without excessive interpretation and experimentation. However, readers should understand that these practical details should not be used to limit the present invention. In other words, in some embodiments of the present invention, these practical details are not necessary, but are used to illustrate how to implement the materials and methods of the present invention.

[0020] As described above, in order to extract more ganoderiol components, especially the content of ganoderiol F (structural formula is reproduced below) from antler ganoderma, the present invention provides a culture medium and cultivation method of antler ganoderma. The details are as follows.

[0021]

[0022] Culture medium of Ganoderma lucidum

[0023] According to some embodiments of the present invention, the ingredients of the culture medium of antler ganoderma lucidum include 38 to 42 parts by weight of corn cobs, 23.75 to 26.25 parts by weight of sawdust, 19 to 21 parts by weight of seaweed powder, 6.65 to 7.35 parts by weight of wheat bran, 6.65 to 7.35 parts by weight of rice bran, and 0.95 to 1.05 parts by weight of sucrose. Among them, the seaweed powder is a specially added ingredient to increase the content of ganoderin F that can be extracted from antler ganoderma lucidum.

[0024] Cultivation method of antler ganoderma

[0025] According to other embodiments of the present invention, a method for cultivating Ganoderma lucidum is provided, which comprises a culture medium preparation step, a sterilization step, an inoculation step, a mycelium growth management step, a Ganoderma lucidum production management step and a harvesting step. The details are as follows.

[0026] 1. Preparation of culture medium: Mix the culture medium of the above-mentioned Ganoderma lucidum evenly, and put the mixed culture medium of Ganoderma lucidum into a bag and seal it (for example, put a ring on it and then cover it with a lid) to obtain a fungus bag.

[0027] 2. Sterilization step: After the above-mentioned bacteria bag is placed in a sterilization chamber at a sterilization temperature for sterilization, the above-mentioned bacteria bag is cooled to room temperature. The above-mentioned sterilization temperature is 121-123°C, the sterilization time can be 3 hours, and the pH value of the bacteria bag after sterilization is 6.8-7.5.

[0028] 3. Inoculation step: After the temperature of the fungus bag is cooled to room temperature (about 26-28°C), the fungus species of Ganoderma lucidum are inoculated onto the Ganoderma lucidum culture medium in the fungus bag in a sterile inoculation room to obtain a fungus species bag.

[0029] 4. Mycelium growth management step: Move the above-mentioned spawn package into a dry, ventilated, light-free spawn room to allow the spawn to germinate. The above-mentioned mycelium growth management step takes 22 days, and the temperature of the spawn room is controlled at 23-28°C from the 1st to the 10th day, and the temperature of the spawn room is controlled at 25-28°C from the 11th to the 22nd day.

[0030] 5. Ganoderma management steps: After the fermented strain package is moved into a closed culture room, the strain package is opened for cultivation to grow the fruiting body of Deer Horn Ganoderma. The temperature of the culture room in the above-mentioned Ganoderma management steps is controlled at 25-28°C through the air conditioner outside the culture room. The humidity of the culture room in the above-mentioned Ganoderma management steps is adjusted by spraying water and watering the ground through a humidifier in the culture room to control the relative humidity at around 90%. Stop spraying and watering 7 days before the Deer Horn Ganoderma matures in the above-mentioned Ganoderma management steps. The above-mentioned Ganoderma management steps use fluorescent lamps as the light source for 10 hours a day, and the light intensity is 2000-5000 lux (LX).

[0031] The culture room in the above-mentioned Ganoderma lucidum management steps needs to be ventilated for a period of time every day. This is because Ganoderma lucidum is an aerobic fungus that requires sufficient oxygen to promote the healthy growth of mycelium and avoid the formation of deformed Ganoderma lucidum. Therefore, as the mycelium of Ganoderma lucidum grows, the ventilation time of the culture room needs to be increased in time to meet the oxygen demand of Ganoderma lucidum. The ventilation of the culture room can be achieved by adjusting the opening and closing degree of the vents of the culture room in the above-mentioned Ganoderma lucidum management steps. The ventilation time of the culture room in the above-mentioned Ganoderma lucidum management steps should be adjusted according to the temperature in the shed to ensure sufficient oxygen supply.

[0032] 6. Harvesting steps: Harvest when the stipe of the fruiting body of Ganoderma lucidum grows to a length of 20-25 cm and has not yet released spores.

[0033] Experimental Examples and Comparative Examples

[0034] Here, different culture media of antler ganoderma lucidum are used to cultivate antler ganoderma lucidum, and the concentration of ganoderin F detected in the samples after extraction of antler ganoderma lucidum is observed. The cultivation method and extraction steps of antler ganoderma lucidum in Experimental Example 1 and Comparative Example 1 are the same, and the difference lies in the composition of the culture media of antler ganoderma lucidum. The culture media of antler ganoderma lucidum in Experimental Example 1 contains 20 parts by weight of seaweed powder, while the culture media of antler ganoderma lucidum in Comparative Example 1 uses a conventional culture media without any seaweed powder added. The detailed composition of the culture media of antler ganoderma lucidum in Experimental Example 1 and Comparative Example 1 is shown in Table 1.

[0035] Table 1: Composition of the culture medium of Ganoderma lucidum in Experimental Example 1 and Comparative Example 1

[0036] Element Experimental Example 1 (parts by weight) Comparative Example 1 (parts by weight) Corn cob 40 50 Wood chips 25 35 Seaweed Powder 20 -- Rice bran 7 7 Wheat bran 7 7 sucrose 1 1

[0037] From Table 1, it can be seen that the difference between the culture medium of the experimental example and the comparative example is that the experimental example 1 reduces the corn cob and the sawdust by about 10 parts by weight, and adds 20 parts by weight of seaweed powder. The main purpose is to increase the content of ganoderin F in the extract under the same extraction method in the future.

[0038] The Ganoderma lucidum of Experimental Example 1 and Comparative Example 1 were extracted by the same extraction method. The steps of the extraction method are described in detail as follows.

[0039] 1. Use a grinder to grind the Ganoderma lucidum into powder with a particle size less than 0.25 cm.

[0040] 2. Add the antler ganoderma lucidum powder into the 60% ethanol solvent, and use a stirrer or a stirring rod to slowly stir the mixture of the antler ganoderma lucidum powder and the 60% ethanol solvent for 60 minutes.

[0041] 3. Place the stirred mixture in a constant temperature device and set the appropriate temperature (according to the extraction requirements). Perform constant temperature extraction for 60 minutes. During this process, the active ingredients of Ganoderma lucidum will dissolve into the solvent.

[0042] 4. Prepare a Buchner funnel and filter paper. Place the Buchner funnel on the collection container and put the filter paper in the funnel. Gently pour the extract into the funnel, filter it by vacuum filtration, and collect the filtrate.

[0043] 5. Mix the residue on the filter paper with 60% ethanol solvent and repeat steps 3 and 4.

[0044] 6. Repeat steps 3 to 5 once more.

[0045] 7. Combine the three filtrates, concentrate under vacuum to remove the alcohol, and then freeze the antler ganoderma lucidum extract at -80°C.

[0046] 8. The frozen Ganoderma lucidum extract is placed under vacuum conditions to allow the frozen liquid to sublime directly from the solid state to the gaseous state, so that the frozen water molecules are directly sublimated into water vapor and escape, thereby achieving the purpose of drying. The freeze-drying process may take several hours or days, depending on the nature and quantity of the sample. The results are listed in Table 2.

[0047] Table 2: Yield of the Ganoderma lucidum extract of Experimental Example 1

[0048]

[0049] As shown in Table 2, the antler ganoderma powder of Experimental Example 1 was added to 1000mL of 60% ethanol solvent, and then extracted according to the above steps. The extract obtained in the first extraction was 4.730g, and the yield was 4.73%. In the second extraction, the antler ganoderma powder was added to 1000mL of 60% ethanol solvent for extraction, and the extract of the second extraction weighed 5.630g, and the yield was 5.63%. In the third extraction, the antler ganoderma powder was added to 1000mL of 60% ethanol solvent for extraction, and the extract of the third extraction weighed 6.660g, and the yield was 6.66%.

[0050] like Figure 1 As shown, the molecular weight of ganoderin F is 454.344, and the MS / MS spectrum peak of ganoderin F is m / z 455.35 (M+1 peak). Therefore, the present invention indeed extracts ganoderin F from antler ganoderma.

[0051] Standard samples of ganoderin F were prepared at concentrations of 2, 4, 6, and 8 ppm, and ganoderin F was quantified using an MS / MS mass spectrometer. Linear regression was performed using the sum of the integrated areas of the peaks in the mass spectrum and the concentration of the ganoderin F standard samples (e.g. Figure 2 As shown), we get the regression equation y=4220.4x+6673.6, R 2 =0.9997. The concentration of the standard sample of ganoderin F and the total integrated area of ​​the peaks in the mass spectrum are shown in Table 3.

[0052] Table 3: Concentration of the standard sample of Lingzhi alcohol F and the total integrated area of ​​the peaks in the mass spectrum

[0053]

[0054]

[0055] Next, after harvesting the antler ganoderma lucidum extracts of Experimental Example 1 and Comparative Example 1, the ganoderma lucidum F standard was first tested to produce the calibration curves of Experimental Example 1 and Comparative Example 1. It is known that the peak area of ​​the sample analysis of Experimental Example 1 and Comparative Example 1 is 20334.695, and then the ganoderma lucidum F concentration of the antler ganoderma lucidum extracts of Experimental Example 1 and Comparative Example 1 is obtained by interpolation.

[0056] The concentration of ganoderin F in the antler ganoderma lucidum extract of Experimental Example 1 of the present invention was compared with the concentration of ganoderin F in the antler ganoderma lucidum extract of Comparative Example 1 to illustrate the difference between the concentration of ganoderin F in the antler ganoderma lucidum extract of Experimental Example 1 of the present invention and the concentration of ganoderin F in the antler ganoderma lucidum extract of Comparative Example 1. Please refer to Table 4.

[0057] Table 4: The concentration of Lingzhi alcohol F and the peak area of ​​sample analysis in Experimental Example 1 and Comparative Example 1

[0058] Experimental Example 1 Comparative Example 1 Peak area 20334.695 21487.204 Lingzhi alcohol F concentration (ppm) 6.40 3.51

[0059] As shown in Table 4, after the extract of the fruiting body of Ganoderma lucidum obtained by the cultivation method of Ganoderma lucidum of the present invention was dissolved in the solution, 10 mg was weighed, 1 ml of methanol was added to dissolve and centrifuged, and then 20 μL of the supernatant + 80 μL of ddH2O / 0.1% FA was taken, and 5 μL was taken for the machine. The concentration of Ganoderma lucidum F obtained was 6.40 ppm, which was significantly improved compared with the concentration of 3.51 ppm in the extract of Ganoderma lucidum obtained by the cultivation method of Ganoderma lucidum of the comparative example.

[0060] The above-mentioned implementation cases are only exemplary and specific descriptions, and are not intended to limit the scope of the present invention. Any equivalent modifications or changes made thereto do not depart from the spirit and scope of the present invention and should be included in the claims of this case.

Claims

1. A culture medium of Ganoderma lucidum, used to increase the concentration of ganoderin F in Ganoderma lucidum extract, characterized in that: 38-42 parts by weight of corn cobs; 23.75-26.25 parts by weight of wood chips; 19-21 parts by weight of seaweed powder; 6.65-7.35 parts by weight of wheat bran; 6.65-7.35 parts by weight of rice bran; and 0.95 to 1.05 parts by weight of sucrose.

2. A method for cultivating Ganoderma lucidum, for increasing the concentration of ganoderin F in Ganoderma lucidum extract, characterized in that: The following steps are involved: A culture medium preparation step, mixing the culture medium of the Ganoderma lucidum according to claim 1 evenly, and bagging and sealing the culture medium of the Ganoderma lucidum to obtain a bacterial bag; Sterilization step, placing the bacteria bag in a sterilization chamber at a sterilization temperature for sterilization, and then cooling the bacteria bag to room temperature; an inoculation step, inoculating the strain of Ganoderma lucidum on the culture medium of Ganoderma lucidum in the strain bag in a sterile inoculation room to produce a strain bag; The mycelium growth management step is to move the bacterial strain package into a dry, ventilated, light-free bacterial room for cultivation to allow the bacterial strain to germinate; The step of managing the production of ganoderma lucidum is to move the fermented spawn package into a closed culture room, open the spawn package for cultivation, and grow the fruiting body of ganoderma lucidum; as well as The step of harvesting is to harvest the antler ganoderma lucidum when the stipe of the fruiting body grows to a length of 20-25 cm and spore powder has not yet been released.

3. The method for cultivating Ganoderma lucidum according to claim 2, characterized in that: The sterilization temperature is 121-123°C.

4. The method for cultivating Ganoderma lucidum according to claim 2, characterized in that: The mycelium growth management step lasts for 22 days, the temperature of the mushroom house is controlled at 23-28°C from the 1st to the 10th day, and the temperature of the mushroom house is controlled at 25-28°C from the 11th to the 22nd day.

5. The method for cultivating Ganoderma lucidum according to claim 2, characterized in that: The temperature of the culture room in the step of managing the growing mushrooms is controlled at 25-28°C, and the relative humidity is maintained at around 90% by spraying water or watering on the ground. The light is illuminated for 10 hours a day, and the light intensity is 2000-5000 lux (LX).

6. The method for cultivating Ganoderma lucidum according to claim 5, characterized in that: Stop spraying and watering 7 days before the antler ganoderma matures.

7. The method for cultivating Ganoderma lucidum according to claim 5, characterized in that: The illumination is provided by fluorescent lamps as light sources.

8. The method for cultivating Ganoderma lucidum according to claim 5, characterized in that: The culture room is kept ventilated for a period of time every day.

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