KR102356059B1 - Culture method for enhancing content of Cordyceps militaris mycelium using Light-Emitting Diode - Google Patents

Abstract

The present invention relates to a culturing method for increasing the content of Militaris Cordyceps mycelium using an LED, comprising: an inoculation step of inoculating a medium with Militaris Cordyceps mycelium; and culturing the inoculated medium under light conditions to obtain Militaris Cordyceps mycelium; It was confirmed that the maximum growth amount of the Militaris Cordyceps mycelium was performed according to the medium and light conditions, and the optimal light amount, the glucose content of the medium, and the By deriving a culture time condition and using it as a culture condition, it is possible to efficiently obtain the maximum growth amount of the mycelium of Cordyceps Cordyceps Militaris.

Description

Culture method for enhancing content of Cordyceps militaris mycelium using Light-Emitting Diode}

The present invention relates to a culturing method for increasing the content of Militaris Cordyceps mycelium using an LED.

Dongchunghacho (冬蟲夏草) got its name from the meaning that it comes out as an insect in winter and a mushroom in summer. About 800 species are known, most of which are entomopathogenic fungi, and a major feature is that they are infected with host insects and mycelium grows inside to produce fruiting bodies. Cordyceps cordyceps has been used as a medicinal herb in Asia (China, Korea, Japan, etc.) since ancient times. For its efficacy, it has been used as a good medicine for strengthening immunity, preventing adult diseases, asthma, and anemia.

In addition, Cordyceps Cordyceps belongs to the group with the highest protein content (more than 28% of dry content) among crops, and contains a large amount of substances that enhance immunity. In particular, among the immunoactive substances of Cordyceps Cordyceps, 'Cordycepin (3'-deoxy-adenosine)', which was found to be an isomer of quinic acid, is included. Cordycepin is a nucleic acid substance that prevents normal cells from being converted into cancer cells by activating the reduced immune function while participating in the genetic information of cells. In clinical trials with Cordyceps Cordyceps, it was confirmed that the content of NK cells (natural killer cells) and cytokines secreted from immune cells, which are immune cells that kill cancer cells, increased by 18% to 25%, so it is used for blood cancer treatment. It is used as a health functional food and is receiving a lot of attention.

Based on these results, Cordyceps Cordyceps containing cordycepin has been certified by the Ministry of Food and Drug Safety as a health functional food in Korea.

Due to the above functions, Cordyceps is consumed a lot among mushrooms, but the amount collected from nature is very small and cannot meet the demand. However, Cordyceps is a plant that cannot be sprayed with pesticides due to its slow growth rate and sensitivity to the external environment, so it is very difficult to cultivate. In addition, although mass cultivation was successful, it is known that the yield of cordyceps cultivation is low even in farms that have been cultivating it for a long time. That is, if 10 are cultivated and harvested, about 1 or 2 are contaminated or abnormal strains grow, which lowers the yield.

Recently, research on the production of mycelium rather than the production of fruiting bodies of Cordyceps is attracting attention. In addition, Feng et al . (2018) and Tang et al . (2018) reported that there was almost no difference in the composition between the fruiting body and mycelium of Cordyceps. Therefore, it is judged that it is economically advantageous to cultivate mycelium that can be mass-produced in a shorter period of time than the fruiting body. However, to date, studies on optimal conditions suitable for increasing the production of Cordyceps mycelium are scarce.

Therefore, there is a need for research on the optimal culture method for increasing the growth amount of mycelia of Cordyceps Militaris containing cordycepin.

1. Republic of Korea Patent Registration No. 10-0465283 (Aug. 14, 2003)

It is an object of the present invention to provide an optimal method for culturing Militaris Cordyceps mycelium for the maximum growth amount of Cordyceps Militaris.

In addition, another object of the present invention is to provide a mycelium of Cordyceps Cordyceps Militaris according to the culturing method.

In addition, another object of the present invention is to provide a method for predicting the maximum growth amount of the mycelium of Cordyceps Cordyceps Militaris.

In order to achieve the above object, the present invention is an inoculation step of inoculating the medium with Militaris Cordyceps mycelium; and culturing the inoculated medium under light conditions to obtain Militaris Cordyceps mycelium; It provides a culturing method for increasing the content of the mycelium of Cordyceps Cordyceps Militaris comprising a.

In addition, the present invention provides a Militaris Cordyceps mycelium cultured according to the method for culturing the Militaris Cordyceps mycelium.

In addition, the present invention is a method for predicting the dry weight of the Militaris Cordyceps mycelium, (a) the amount of light (X ₁ ) in the box-Behnken design (Box-Behnken design), the glucose content of the medium (X ₂ ) , for the incubation time (X ₃ ), designing the experimental range by coding -1, 0, and 1; (b) obtaining experimental values for the amount of light, the glucose content of the medium, and the incubation time in the designed experimental range of step (a); (c) deriving a quadratic regression model expressed by Equation 1 below using the experimental values of step (b); And (d) predicting the dry weight of the Militaris Cordyceps mycelium by performing ANOVA on the quadratic regression model represented by Equation 1 derived in step (c); It provides a method for predicting the dry weight of the mycelium of Cordyceps Cordyceps Militaris comprising a.

[Equation 1]

Y ₁ =13.18-1.22X ₁ -1.08X ₂ +0.4438X ₃

-1.00X ₁ X ₂ +0.7050X ₁ X ₃ +0.1975X ₂ X ₃ +2.60X ₁ ² +2.56X ₂ ² +2.55X ₃ ²

(In Equation 1, Y ₁ is the dry weight (g/L) of Militaris Cordyceps mycelium (g/L), X ₁ is the amount of light (code unit), X ₂ is the glucose content of the medium (code unit), and X ₃ is the incubation time (code unit).

According to the present invention, Militaris Cordyceps mycelium exhibited the maximum growth when irradiated with green LED light in a PDB (potato dextrose broth) liquid medium and cultured, and the optimal light amount, medium glucose content and culture time through response surface analysis. By deriving the conditions and using them as culture conditions, there is an effect of efficiently obtaining the maximum growth amount of the mycelium of Cordyceps Militaris.

In addition, the Militaris Cordyceps mycelium obtained according to the culture method according to the present invention contains cordycepin, which has excellent immune enhancing activity, anticancer activity, antiviral and anti-inflammatory effects, and thus can be usefully used as a health functional food.

Figure 1 is Militaris Cordyceps militaris ) is a diagram showing the mycelium.
2 is a view showing the mycelium growth according to the type of liquid medium of Militaris Cordyceps Cordyceps (data is the mean ± standard deviation of the results of three experiments).
3 is a diagram showing the mycelium growth according to the incubation period of Militaris Cordyceps Cordyceps (data shows the results of three experiments as mean ± standard deviation).
4 is a view showing a shaking incubator installed with a fluorescent lamp and UV-A.
5 is a view showing a control box for adjusting the amount of fluorescent lamp and UV-A light and on/off.
6 is a view showing changes in the growth amount of mycelia of Cordyceps mycelia of Militaris according to irradiation with fluorescent lamps and UV-A (data shows the results of three experiments as mean ± standard deviation).
7 is a view showing a shaking incubator installed with LED single light (red, green, blue).
8 is a view showing a control box for controlling the amount of single light and on/off of the LED.
9 is a view showing changes in the mycelium growth amount according to LED single light (red, green, blue) irradiation of Militaris Cordyceps Cordyceps (data shows the results of three experiments as mean ± standard deviation).
10 is a view showing the change in the growth amount of mycelia of Cordyceps Cordyceps Militaris according to the irradiation time of LED green light (data shows the results of three experiments as mean ± standard deviation).
11 is a view showing changes in the mycelium growth amount according to the type of LED single light and mixed light of Militaris Cordyceps Cordyceps (data is the mean ± standard deviation of the results of three experiments).
12 is a view showing changes in the mycelium growth amount according to the irradiation time of mixed light red*green in Mylitaris Cordyceps mycelium (data shows the results of three experiments as mean±standard deviation).
13 is a diagram showing the correlation between the culture conditions of Mylitaris Cordyceps mycelium and the mycelium growth amount.
14 is a diagram illustrating a fit model between LED control conditions and the amount of mycelial growth of Militaris Cordyceps cordyceps.
15 is a view showing the optimal LED control conditions for the maximum mycelium amount of Militaris Cordyceps mycelium.
16 is a diagram showing the fit of the optimal LED control condition response model for the maximum mycelium amount of Militaris Cordyceps mycelium.

Hereinafter, the present invention will be described in detail.

The present inventors confirmed that when irradiated with green LED light in a PDB (potato dextrose broth) liquid medium for 4 to 5 days a day for 12 hours a day and cultured, it exhibited the maximum growth amount of the mycelia of Cordyceps Cordyceps Militaris, and the response surface analysis was performed. The present invention was completed by deriving the optimal light amount, glucose content of the medium, and culture time conditions through this, and using these as culture conditions to efficiently obtain the maximum growth amount of the mycelia of Cordyceps Cordyceps Militaris.

The present invention is an inoculation step of inoculating the medium with Militaris Cordyceps mycelium; and culturing the inoculated medium under light conditions to obtain Militaris Cordyceps mycelium; It provides a culturing method for increasing the content of the mycelium of Cordyceps Cordyceps Militaris comprising a.

In this case, the medium may be a potato dextrose broth medium, and the culture may be performed for 3 to 6 days, preferably 4 to 5 days.

In addition, the light conditions include culturing the mycelium by irradiating the medium inoculated with LED single light or mixed light, the LED single light is green light having a wavelength of 500 nm to 550 nm, and the LED mixed light is green light having a wavelength of 500 nm to 550 nm and It is characterized in that the light is a mixture of red light having a wavelength of 620 nm to 780 nm, but preferably, LED green light having a wavelength of 500 nm to 550 nm can be used, and in this case, the maximum growth amount of the mycelium of Cordyceps Cordyceps mycelium can be shown.

In addition, it is characterized in that the mycelium is cultured by irradiating the LED single light or mixed light to the inoculated medium for 5 to 15 hours a day, and preferably irradiated for 12 hours a day, but is not limited thereto. .

At this time, if the culture conditions and light conditions are deviated as described above, the Militaris Cordyceps mycelium according to the present invention may not be properly obtained or the yield of Militaris Cordyceps cordyceps may be significantly lowered, which may cause uneconomical problems.

In addition, the dry weight of the Militaris Cordyceps mycelium may be characterized in that it has a correlation as shown in Equation 1 below with the amount of light, the glucose content of the medium, and the culture time.

[Equation 1]

Y ₁ =13.18-1.22X ₁ -1.08X ₂ +0.4438X ₃

-1.00X ₁ X ₂ +0.7050X ₁ X ₃ +0.1975X ₂ X ₃ +2.60X ₁ ² +2.56X ₂ ² +2.55X ₃ ²

(In Equation 1, Y ₁ is the dry weight (g/L) of Militaris Cordyceps mycelium, X ₁ is the amount of light, X ₂ is the glucose content of the medium, and X ₃ is the culture time.)

At this time, the culture conditions of the light amount, the glucose content of the medium, and the culture time to obtain the maximum dry weight of the Militaris Cordyceps mycelium are 1000 to 1500 lux, 1 to 10 g/50mL, 10 to 50 hours, respectively, characterized in that, Preferably, the light amount, the glucose content of the medium, and the culture conditions of the culture time may be 1273.75 lux, 5.00853 g/50mL, and 26.5176 hours, respectively, but is not limited thereto.

In addition, the present invention provides a Militaris Cordyceps mycelium cultured according to the method for culturing the Militaris Cordyceps mycelium.

Since the cultured Militaris Cordyceps mycelium contains cordycepin, it may be used as a health functional food for immune enhancing activity and anticancer activity.

In addition, the present invention is a method for predicting the dry weight of the Militaris Cordyceps mycelium, (a) the amount of light (X ₁ ) in the box-Behnken design (Box-Behnken design), the glucose content of the medium (X ₂ ) , for the incubation time (X ₃ ), designing the experimental range by coding -1, 0, and 1; (b) obtaining experimental values for the amount of light, glucose content of the medium and culture time in the designed experimental range of step (a); (c) deriving a quadratic regression model expressed by Equation 1 below using the experimental values of step (b); And (d) predicting the dry weight of the Militaris Cordyceps mycelium by performing ANOVA on the quadratic regression model represented by Equation 1 derived in step (c); It provides a method for predicting the dry weight of the mycelium of Cordyceps Cordyceps Militaris comprising a.

[Equation 1]

Y ₁ =13.18-1.22X ₁ -1.08X ₂ +0.4438X ₃

-1.00X ₁ X ₂ +0.7050X ₁ X ₃ +0.1975X ₂ X ₃ +2.60X ₁ ² +2.56X ₂ ² +2.55X ₃ ²

(In Equation 1, Y ₁ is the dry weight (g/L) of Militaris Cordyceps mycelium (g/L), X ₁ is the amount of light (code unit), X ₂ is the glucose content of the medium (code unit), and X ₃ is the incubation time (code unit).

At this time, the culture conditions of the light amount, the glucose content of the medium, and the culture time to obtain the maximum dry weight of the Militaris Cordyceps mycelium are 1000 to 1500 lux, 1 to 10 g/50mL, 10 to 50 hours, respectively, characterized in that, Preferably, the light amount, the glucose content of the medium, and the culture conditions of the culture time may be 1273.75 lux, 5.00853 g/50mL, and 26.5176 hours, respectively, but is not limited thereto.

Hereinafter, the present invention will be described in more detail through examples. These examples are only for explaining the present invention in more detail, and it is to those of ordinary skill in the art to which the present invention pertains that the scope of the present invention is not limited by these examples according to the gist of the present invention. it will be self-evident

< Example 1> Establishment of liquid culture technology of Cordyceps mycelium

1. Disclosure material

Cordyceps militaris (KCCM 60304) was distributed from Korea Center for Microorganisms Conservation (KCCM) in order to evaluate the growth amount of Cordyceps mycelium (Fig. 1).

2. Militaris Cordyceps ( Cordyceps militaris ) of liquid culture and growth analysis

Militaris Cordyceps militaris ) of three kinds (YMG (yeast extract, malt extract, glucose medium), PDB (potato dextrose broth), SDB (Sabouraud dextrose broth) having the culture composition shown in Table 1 below to establish the culture environment conditions for increasing the growth amount A liquid culture medium was used.

badge person Medium pH Medium composition (g/L) yeast extract malt extract Glucose Potato extract Peptone YMG 6.0 4 10 4 - - PDB 5.2 - - 20 4 - SDB 4.5 - - 20 - 10

The other pre-sale from the received millimeter-less KCCM Cordyceps (Cordyceps militaris ) mycelia were cultured for 7 days at 25° C. in an incubator using PDA (Potato Dextrose Agar) plate medium. After that, the edge portion of the cultured mycelium was inoculated into a liquid medium (YMG, PDB, SDB) by 3 cores using a cork borer with a diameter of 5 mm, and cultured with shaking (100 rpm) at 25 ° C for 7 days to produce a liquid spawn, one day The mycelium growth amount of the liquid seed culture cultured at intervals was measured.

Each of the YMG, PDB or SDB medium was put into a 100 mL Erlenmeyer flask by 50 mL, sterilized at 121° C. for 15 minutes, and then cooled before use.

In addition, the growth amount of the cultured Militaris Cordyceps mycelium was determined by Carvajal et al . (2012) was measured by the following method by the dry weight measurement method.

① Transfer the cultured mycelium to the falcon tube.

② Centrifuge (4000 rpm, 10 min, 2 times, Union 32R Plus, HANIL, Korea) to separate the culture medium and mycelium.

③ Dry the falcon tube contained in the mycelium at 60℃ for 24 hours.

④ Derive the mycelium weight using the pre-measured total dry weight of the falcon tube.

3. Militaris Cordyceps ( Cordyceps militaris ) of growth according to the liquid culture medium

The amount of mycelium growth according to the type of liquid medium of Mylitaris Cordyceps mycelium is shown in FIG. 2 .

As a result, it was confirmed that the growth amount of the mycelium of Cordyceps Cordyceps Militaris showed the highest growth rate of 12.0 g/L when cultured in PDB (Potato Dextros Broth) medium.

In addition, as a result of analyzing the growth amount of Militaris Cordyceps according to the culture period, the mycelium growth period of Militaris Cordyceps mycelium was fixed to 5 days as shown in FIG.

< Example 2> Evaluation of the effect of fluorescent lamps and UV-A on the growth of Cordyceps mycelium

1. Test material and method for culturing Militaris Cordyceps mycelium

Fluorescent lamp and UV-A (356 nm) were installed in the shaking incubator as shown in FIG. 4 in order to evaluate the change in the growth amount of the Mylitaris Cordyceps mycelium sold in <Example 1-1> according to the irradiation with fluorescent lamps and UV-A. did The light source was installed at a height of about 30 cm of the culture medium, and the light intensity was set to ^{100 pmol·m -2} ·s ^-1.

In addition, the control box for on/off of the light source was purchased and installed from Bitsol LED, an LED lighting brand for plant growth of Esrez, as shown in FIG. 5, and the culture was performed as in <Example 1-2>. Liquid spawn was produced by shaking culture (100 rpm) in PDB medium for 7 days at 25 ° C., and irradiated with fluorescent lamps and UV-A at 12 h/day for 5 days, and the growth amount was measured by dry weight measurement.

As a control, Militaris Cordyceps mycelium cultured under cancer culture conditions was used.

2. Growth amount of Militaris Cordyceps according to fluorescent lamp and UV-A light source

Fig. 6 shows the growth amount of mycelium according to fluorescent lamp and UV-A irradiation of Militaris Cordyceps, and it was confirmed that Militaris Cordyceps growth showed the highest amount of mycelium when cultured under a fluorescent light source than in dark culture or UV-A. , it was confirmed that about 2 times more mycelium was produced than that of cancer culture.

< Example 3> LED single light Evaluation of the growth rate of Cordyceps mycelium according to control conditions (wavelength type, irradiation time)

1. Test material and method for culturing Militaris Cordyceps mycelium

In order to evaluate the change in the amount of mycelial growth of Militaris Cordyceps mycelium according to the irradiation of LED single light, LED single light was installed in a shaking incubator as shown in FIG. 7 . A total of three colors were installed for the single light, red, green, and blue, and 120 pieces were installed for each color (Table 2). The light source was installed at a height of about 30 cm of the culture medium, and the light intensity was set to 64.9-108.0 pmol·m ^-2 ·s ^-1 .

In addition, as shown in FIG. 8, the LED and the control box were installed by purchasing and installing Bitsol LED, an LED lighting brand for plant growth of Esrez, at 25° C. in PDB medium under the same conditions as in <Example 1-2>. Shaking culture (100 rpm) was performed for 7 days to produce liquid spawn, and the three types of single light were irradiated for 5 days at 12 h/day, and the growth amount was measured by dry weight measurement.

As a control, Cordyceps mycelium cultured under dark culture, fluorescent light and UV-A conditions (Example 2) was used.

colour Wavelength (nm) LED quantity PCB quantity LED quantity per PCB Red 650 120 10 12 Green 525 120 10 12 Blue 450 120 10 12

2. LED of Militaris Cordyceps single light Growth according to conditions

9 shows the growth amount of mycelium according to the single light irradiation of LED Cordyceps Militaris, and when irradiated with green light, it was confirmed that Militaris Cordyceps showed the highest amount of mycelium growth at 25.3 g/L, which is a cancer Mycelium growth rate was 3.88 times higher than cultured, 1.71 times higher than fluorescent lamps, and 1.74 times higher than UV-A.

Therefore, it is judged that the LED green light is effective in increasing the growth amount of the Mylitaris Cordyceps mycelium. It was confirmed that the highest growth rate was exhibited.

< Example 4> LED mixed light control condition ( mixed light Evaluation of mycelium growth of Cordyceps by type, irradiation time)

1. Test material and method for culturing Militaris Cordyceps mycelium

Red*green (650+525nm), red*blue (650+450nm), green*blue (525+450nm) were used as the mixed light to evaluate the change in the growth amount of Militaris Cordyceps mycelium according to the irradiation of LED mixed light. brightness of the light source is ^{64.9 ~ 108.0 pmol · m -2 ·} s - was set to ^one.

In addition, the culture was performed under the same conditions as in <Example 1-2> in PDB medium at 25° C. for 7 days with shaking (100 rpm) to produce liquid spawns, and the three types of mixed light were irradiated for 5 days at 12 h/day. Thus, the amount of growth was measured by a dry weight measurement method.

2. LED of Militaris Cordyceps mixed light Growth according to conditions

11 shows the growth amount of mycelium according to the LED mixed light irradiation of Militaris Cordyceps, when irradiated with red*green, red*blue, green*blue mixed light, 13.9 g/L, 8.3 g/L, respectively; 13.7 g/L.

Therefore, when red*green of the mixed light was irradiated on the mycelium of Cordyceps, it was confirmed that it was most effective for increasing the mycelium growth.

However, as in <Example 3>, when compared with the green light (C. militaris : 25.3 g/L), which is the single most effective light for increasing the growth of mycelium of Cordyceps cordyceps mycelium, the single light was more effective than the mixed light. It was confirmed to be effective in increasing the growth rate.

Therefore, it is judged that single light is more effective than mixed light for the LED application for increasing the growth amount of the Militaris Cordyceps mycelium. When irradiated for 12 h/day, it exhibited the highest mycelium growth.

< Example 5> Derivation of optimal control conditions for LED for maximum growth of mycelium

1. LED control conditions and mycelium between growth Correlation Derivation

Pearson correlation coefficient (Hauke and Kossowski, 2001) and heat map (Wilf et al., 2016) were used to derive the correlation between the mycelium growth condition and the mycelium culture condition of Cordyceps including the LED control condition, and the statistical program was the R program (version 3.4). .3) was used.

Militaris Cordyceps mycelium culture conditions factors were set as LED wavelength, LED light amount, LED irradiation time, glucose content of the medium, and pH of the medium.

It was evaluated that the higher the coefficient in the Pearson correlation coefficient, the higher the correlation between the culture condition factor and the growth amount. model was used.

As a result, the correlation between the Militaris Cordyceps mycelium culture conditions and the mycelium growth amount is shown in FIG. 13, and the culture conditions that have the greatest effect on the Militaris Cordyceps mycelium growth amount are the glucose content and LED wavelength of the medium, In particular, it was confirmed that the LED wavelength showed a positive correlation (0.40) with the mycelium growth.

In addition, the fit model between the LED control condition and the growth amount of the Cordyceps mycelium is shown in FIG. 14, and the LED wavelength with the highest correlation between the LED control condition and the mycelium growth was confirmed to be the ‘Sinusoidal’ model. The regression formula using this is shown in Table 3, and it is judged that the growth amount can be predicted using only the LED control conditions (wavelength, light amount, irradiation time) through the regression formula between the derived LED control conditions and the mycelium growth amount.

No. χ y regression a constant One wavelength Mycelium growth y=a+bcos(cχ+d) a=130.71; b=58.39; c=2.07; d=3.37 2 amount of light Mycelium growth y=a+bcos(cχ+d) a=11.11; b=56.23; c=2.05; d=3.13 3 investigation time Mycelium growth y=a+bχ+cχ ² +d a=-0.388035; b=0.313334; c=-0.001540; d=0.000003

2. Establishment of optimal LED conditions for increasing mycelium growth

The optimal LED control condition for increasing the growth of mycelium of Cordyceps Cordyceps Militaris was the Box-Benkhen design (BBD) experimental design of the Response Surface Model (RSM), and the program was designed in Design-Expert Software Version 10 was used.

The fixing factor was Militaris Cordyceps ( C. militaris ) mycelium, and the variables were set as the light quantity condition, the glucose content of the medium, and the incubation time. At this time, the range of variables was set through preliminary experiments, and the BBD experimental design was set to a total of 17 conditions as shown in Table 4 below.

Run Independent variables
(coded) Independent variables
(actual) Mycelia dry weight
weight), g/L X _One X ₂ X ₃ X _One X ₂ X ₃ Y _One One One 0 -One 1338 7.5 24 16.12 2 0 0 0 669 7.5 72 13.16 3 One 0 One 1338 7.5 120 18.68 4 -One One 0 0 10 72 20.17 5 0 0 0 669 7.5 72 13.55 6 0 -One -One 669 5 24 19.65 7 -One 0 -One 0 7.5 24 19.38 8 One -One 0 1338 5 72 18.51 9 -One 0 One 0 7.5 120 19.12 10 0 -One One 669 5 120 19.88 11 One One 0 1338 10 72 15.14 12 0 One One 669 10 120 17.33 13 0 0 0 669 7.5 72 11.06 14 0 0 0 669 7.5 72 15.02 15 -One -One 0 0 5 72 19.54 16 0 One -One 669 10 24 16.31 17 0 0 0 669 7.5 72 13.11 Independent variables Levels -One 0 One X ₁ : Light quantity, lux 0 669 1338 X ₂ : glucose content, g/50 mL 5 7.5 10 X ₃ : incubation time, h 24 72 120

15 shows a three-dimensional response surface graph showing the correlation between factors as a response surface and a second-order polynomial regression model derived from it. As a result, the optimal culture conditions for the maximum mycelium amount of Militaris Cordyceps mycelium are When 1273.75 lux, the glucose content of the medium was 5.00853 g/50mL, and the incubation period was 26.5176 hours, it was confirmed that the mycelium growth amount was increased to 26.5176 g/L.

In addition, the second-order polynomial equation regression model was analyzed for variance (ANOVA ^{) to obtain R 2} (coefficient of determination) indicating the degree of fit of the response model as shown in FIG. 16 . As a result, R ² (coefficient of determination) was 0.8567, indicating high reliability. Accordingly, the second polynomial equation derived in FIG. 15 is suitable for predicting the response value of optimal culture conditions for the maximum mycelium amount of Militaris Cordyceps. judged

Claims (14)

Inoculation step of inoculating the Militaris Cordyceps mycelium in a potato dextrose broth medium; and
Culturing step of culturing the inoculated medium under LED green light conditions having a wavelength of 525 nm to obtain Militaris Cordyceps mycelium; includes,
The culture method for increasing the content of Militaris Cordyceps mycelium, characterized in that the culture is carried out under the conditions of 1273.75 lux of light, 5.00853 g/50 mL of glucose in the medium, and 26.5176 hours of incubation time. delete delete delete delete delete delete The method of claim 1,
The dry weight of Militaris Cordyceps mycelium is,
A culturing method for increasing the content of Militaris Cordyceps mycelium, characterized in that it has a correlation as shown in Equation 1 below with the amount of light, the glucose content of the medium, and the culture time:
[Equation 1]
Y ₁ =13.18-1.22X ₁ -1.08X ₂ +0.4438X ₃
-1.00X ₁ X ₂ +0.7050X ₁ X ₃ +0.1975X ₂ X ₃ +2.60X ₁ ² +2.56X ₂ ² +2.55X ₃ ²
(In Equation 1, Y ₁ is the dry weight (g/L) of Militaris Cordyceps mycelium, X ₁ is the amount of light, X ₂ is the glucose content of the medium, and X ₃ is the culture time.) 9. The method of claim 8,
A culturing method for increasing the content of Militaris Cordyceps mycelium, characterized in that the culturing conditions for the amount of light, the glucose content of the medium and the incubation time are 1000 to 1500 lux, 1 to 10 g/50 mL, and 10 to 50 hours, respectively. 10. The method of claim 9,
A culturing method for increasing the content of Militaris Cordyceps mycelium, characterized in that the light amount, the glucose content of the medium, and the culture conditions of the culture time are 1273.75 lux, 5.00853 g/50mL, and 26.5176 hours, respectively. delete In the method for predicting the dry weight of Militaris Cordyceps mycelium according to claim 1,
(a) With respect to the amount of light (X ₁ ), the glucose content of the medium (X ₂ ), and the incubation time (X ₃ ) by the Box-Behnken design, -1, 0 and 1 were coded as experiments designing a range;
(b) obtaining experimental values for the amount of light, glucose content of the medium and culture time in the designed experimental range of step (a);
(c) deriving a quadratic regression model expressed by Equation 1 below using the experimental values of step (b); and
(d) predicting the dry weight of the Militaris Cordyceps mycelium by performing ANOVA on the quadratic regression model represented by Equation 1 derived in step (c); Militaris Cordyceps mycelium dry weight prediction method comprising:
[Equation 1]
Y ₁ =13.18-1.22X ₁ -1.08X ₂ +0.4438X ₃
-1.00X ₁ X ₂ +0.7050X ₁ X ₃ +0.1975X ₂ X ₃ +2.60X ₁ ² +2.56X ₂ ² +2.55X ₃ ²
(In Equation 1, Y ₁ is the dry weight (g/L) of Militaris Cordyceps mycelium (g/L), X ₁ is the amount of light (code unit), X ₂ is the glucose content of the medium (code unit), and X ₃ is the incubation time (code unit). 13. The method of claim 12,
Light quantity, glucose content of the medium and culture conditions of culture time are 1000 to 1500 lux, 1 to 10 g/50 mL, and 10 to 50 hours, respectively. Method for predicting dry weight of mycelium of Cordyceps Cordyceps. 14. The method of claim 13,
Light quantity, glucose content of the medium, and culture conditions of the culture time are 1273.75 lux, 5.00853 g/50mL, and 26.5176 hours, respectively. Method for predicting the dry weight of the Militaris Cordyceps mycelium.

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