CN113373059A - Method for increasing content of adenosine and ergosterol in fungus mixed fermentation liquid - Google Patents
Method for increasing content of adenosine and ergosterol in fungus mixed fermentation liquid Download PDFInfo
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Abstract
The invention discloses a method for improving the content of adenosine and ergosterol in fungus mixed fermentation liquid, relates to the technical field of edible fungus mixed fermentation, and particularly relates to a method for improving the content of adenosine and ergosterol in fungus mixed fermentation liquid, which comprises the following steps: (1) firstly, preparing various medical and edible fungus slant mother strains respectively, wherein the culture temperature is 20-25 ℃, the humidity is 45-65%, and the culture time is 7-10 days; (2) and respectively inoculating the mature mother seeds into seed fermentation shake flasks, placing 5-8 glass beads in each flask, controlling the inoculation amount to be 5-8 square centimeters, controlling the culture temperature to be 20-25 ℃, the rotation speed to be 180-240 rpm, and the culture time to be 56-72 h. When the invention is used for researching the mixed culture of the medical and edible fungi, the static culture and the dynamic culture are combined to obviously improve the content of adenosine and sterol, so that the invention further optimizes the method for improving the content of adenosine and sterol.
Description
Technical Field
The invention relates to the technical field of edible fungus mixed fermentation, in particular to a method for improving the content of adenosine and ergosterol in fungus mixed fermentation liquid.
Background
The medicinal edible fungi have no unified and standard definition, and generally refer to microbial fungi which have the functions of preventing, protecting and treating human diseases and can be used daily. Medicinal and edible fungi have a long medicinal and edible history in China, and with the progress of modern biotechnology and medical research, the effects of the medicinal and edible fungi on disease prevention, health care and treatment are widely proved, and therefore, a plurality of well-known clinical medicines and health care products are generated. Modern researches show that the main physiologically active substances contained in the edible fungi comprise polysaccharides, terpenoids, nucleosides, sterols, glycoprotein and the like. Except polysaccharides and terpenoids, adenosine and ergosterol are bioactive substances which are rich in almost all medicinal and edible fungi. Wherein adenosine has vasodilating, blood pressure lowering, heart rate slowing, sedative and anticonvulsant effects; ergosterol has anti-tumor and anti-inflammatory activities, and can inhibit the expression of vascular endothelial cell growth factor A in tumor tissues, thereby inhibiting angiogenesis and promoting apoptosis of tumor cells. Therefore, the method has important pharmacological value for improving the content of adenosine and ergosterol in the medicinal and edible fungi fermentation hyphae.
At present, only one invention patent CN106190865A relates to a liquid fermentation culture medium formula for improving the biomass and the active ingredient content of cordyceps sobolifera fungus, and the invention greatly improves the biomass and the active ingredient content of cordyceps sobolifera fungus by optimizing the culture medium. However, the invention only relates to single edible fungus fermentation, and does not relate to mixed fermentation of multiple edible fungi.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides a method for improving the content of adenosine and ergosterol in fungus mixed fermentation liquor, and solves the problems in the background technology.
In order to achieve the purpose, the invention is realized by the following technical scheme: a method for increasing the content of adenosine and ergosterol in fungus mixed fermentation liquor comprises the following steps:
(1) firstly, preparing various medical and edible fungus slant mother strains respectively, wherein the culture temperature is 20-25 ℃, the humidity is 45-65%, and the culture time is 7-10 days;
(2) respectively inoculating 5-8 glass beads in each bottle after the mother seeds are mature, fermenting and shaking the bottles, controlling the inoculation amount to be 5-8 square centimeters, controlling the culture temperature to be 20-25 ℃, the rotation speed to be 180-240 rpm, and culturing for 56-72 hours;
(3) according to the change rule of each medicinal and edible fungus growth curve, seed liquids with different proportions are mixed, inoculated and subjected to fermentation shake flask culture, the total time of the fermentation shake flask culture is 144-168 hours, static culture and dynamic culture are alternately carried out in the culture process, the rotating speed is controlled to be 200-240 rpm during the dynamic culture, and the temperature is controlled to be 20-25 ℃;
(4) during dynamic culture, the fermentation shake flask is irradiated by a long-wavelength visible light fluorescent lamp, the irradiation is carried out intermittently, and the total irradiation time is controlled;
(5) and after the fermentation is finished, centrifugally separating to obtain a fermentation liquid supernatant, filtering by using a microporous filter membrane, and then carrying out quantitative determination on adenosine and ergosterol by using a high performance liquid chromatograph.
Optionally, the plurality of edible fungi in step (1) may be any two or more of Ganoderma lucidum, Cordyceps sinensis, Tremella Aurantialba, Grifola frondosa, Hericium erinaceus, Lentinus Edodes, Armillariella mellea, and Phellinus linteus.
Optionally, the time ratio of static culture to dynamic culture in the step (3) is 1: 2-1: 6, preferably 1: 3-1: 4.5.
Optionally, in the step (4), the wavelength range of the long-wavelength visible light is 600-700 nm, and preferably 650-700 nm.
Optionally, in the step (4), the irradiation time of the long-wavelength visible light is 24-72 hours, and preferably 48-60 hours.
The invention provides a method for improving the content of adenosine and ergosterol in fungus mixed fermentation liquor, which has the following beneficial effects:
when the applicant researches mixed culture of medicinal and edible fungi, the invention unexpectedly discovers that the content of adenosine and sterol can be obviously improved by combining static culture and dynamic culture, and the method for improving the content of adenosine and sterol is further optimized in the invention.
Detailed Description
In the following, technical solutions in the embodiments of the present invention are clearly and completely described, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments.
Embodiment 1
The invention provides a technical scheme that: a method for improving the content of adenosine and ergosterol in fungus mixed fermentation liquor specifically comprises the following steps:
(1) firstly, respectively preparing various medical and edible fungi PDA slant mother strains, wherein the medical and edible fungi can be any two or more of lucid ganoderma, cordyceps sinensis, tremella aurantialba, grifola frondosa, hericium erinaceus, lentinus edodes, armillaria mellea and phellinus igniarius, culturing at the temperature of 20-25 ℃ and the humidity of 45-65%, and culturing for 7-10 days;
(2) respectively inoculating seed fermentation shake flasks after the mother seeds are mature, wherein the liquid loading amount of the seed flasks is 50mL, the seed culture medium is a liquid PDA culture medium, 5-8 glass beads are placed in each flask, the inoculation amount is controlled to be 5-8 square centimeters, the culture temperature is 20-25 ℃, the rotation speed is 180-240 rpm, and the culture time is 56-72 hours;
(3) according to the change rule of each medicine and food fungus growth curve, seed liquids with different inoculation amounts in proportion are mixed and inoculated into a fermentation shake flask, the volume of fermentation liquid is 150mL, the total time of the fermentation shake flask culture is 144-168h, static culture and dynamic culture are alternately carried out in the culture process, the rotating speed is controlled to be 200-240 rpm during the dynamic culture, the temperature is controlled to be 20-25 ℃, the time proportion of the static culture to the dynamic culture is 1: 2-1: 6, and the optimal proportion is 1: 3-1: 4.5;
(4) during dynamic culture, the fermentation shake flask is irradiated by a long-wavelength visible light fluorescent lamp, the irradiation is carried out intermittently, and the total irradiation time is controlled. The wavelength range of the long-wavelength visible light is 600-700 nm, and the preferable wavelength range is 650-700 nm; the long-wavelength visible light irradiation time is 24-72 h, preferably 48-60 h;
(5) and after the fermentation is finished, performing centrifugal separation to obtain mycelia, and performing quantitative determination on adenosine and ergosterol by using a high performance liquid chromatograph after leaching treatment and microfiltration membrane filtration.
Example II
A method for increasing the content of adenosine and ergosterol in fungus mixed fermentation liquor further comprises the following steps:
(1) firstly, preparing various medical and edible fungi PDA slant mother seeds respectively, wherein the medical and edible fungi are two combinations of lucid ganoderma and cordyceps, the culture temperature is 23 ℃, the humidity is 50%, and the culture time is 7 days;
(2) respectively inoculating seed fermentation shake flasks after the mother seeds are mature, wherein the liquid loading amount of the seed flasks is 50mL, the seed culture medium is a liquid PDA culture medium, 5 glass beads are placed in each flask, the inoculation amount is controlled to be 5 square centimeters, the culture temperature is 23 ℃, the rotation speed is 180rpm, and the culture time is 72 hours;
(3) according to the change rule of each medicine and food fungus growth curve, seed liquids with different inoculation amounts in proportion are mixed and inoculated into a fermentation shake flask, the volume of fermentation liquid is 150mL, the total time of the fermentation shake flask culture is 168h, static culture and dynamic culture are alternately carried out in the culture process, the rotating speed is controlled at 200rpm during the dynamic culture, the temperature is controlled at 23 ℃, and the time proportion of the static culture to the dynamic culture is 1: 3;
(4) during dynamic culture, the fermentation shake flask is irradiated by a long-wavelength visible light fluorescent lamp, the irradiation is carried out intermittently, and the total irradiation time is controlled. The wavelength of the long-wavelength visible light is 680nm, and the irradiation time of the long-wavelength visible light is 60 h;
(5) after fermentation, mycelium is obtained by centrifugal separation, and quantitative determination of adenosine and ergosterol is carried out by a high performance liquid chromatograph after leaching treatment and microfiltration membrane filtration, wherein the quantitative determination is respectively 2.5mg/g and 6.9 mg/g.
Example three
A method for increasing the content of adenosine and ergosterol in fungus mixed fermentation liquor further comprises the following steps:
(1) firstly, preparing various medical and edible fungi PDA slant mother strains respectively, wherein the medical and edible fungi are three combinations of cordyceps, tremella aurantialba and grifola frondosa, the culture temperature is 25 ℃, the humidity is 65%, and the culture time is 7 days;
(2) respectively inoculating seed fermentation shake flasks after the mother seeds are mature, wherein the liquid loading amount of the seed flasks is 50mL, the seed culture medium is a liquid PDA culture medium, 8 glass beads are placed in each flask, the inoculation amount is controlled to be 8 square centimeters, the culture temperature is 25 ℃, the rotation speed is 200rpm, and the culture time is 72 hours;
(3) according to the change rule of each medicine and food fungus growth curve, seed liquids with different inoculation amounts in proportion are mixed and inoculated into a fermentation shake flask, the volume of fermentation liquid is 150mL, the total time of the fermentation shake flask culture is 168h, static culture and dynamic culture are alternately carried out in the culture process, the rotating speed is controlled at 220rpm during the dynamic culture, the temperature is controlled at 25 ℃, and the time proportion of the static culture to the dynamic culture is 1: 4.5;
(4) during dynamic culture, the fermentation shake flask is irradiated by a long-wavelength visible light fluorescent lamp, the irradiation is carried out intermittently, and the total irradiation time is controlled. The wavelength of the long wavelength visible light is 700 nm; the irradiation time of the long-wavelength visible light is 60 h;
(5) after fermentation, mycelium is obtained by centrifugal separation, and quantitative determination of adenosine and ergosterol is carried out by a high performance liquid chromatograph after leaching treatment and microfiltration membrane filtration, wherein the quantitative determination is respectively 3.6mg/g and 8.2 mg/g.
Example four
A method for increasing the content of adenosine and ergosterol in fungus mixed fermentation liquor further comprises the following steps:
(1) firstly, preparing various medical and edible fungi PDA inclined plane mother seeds respectively, wherein the medical and edible fungi are five combinations of lucid ganoderma, cordyceps sinensis, tremella aurantialba, grifola frondosa and hericium erinaceus, the culture temperature is 25 ℃, the humidity is 65%, and the culture time is 7 days;
(2) respectively inoculating seed fermentation shake flasks after the mother seeds are mature, wherein the liquid loading of the seed flasks is 50mL, the seed culture medium is a liquid PDA culture medium, 8 glass beads are placed in each flask, the inoculation amount is controlled to be 5 square centimeters, the culture temperature is 25 ℃, the rotation speed is 220rpm, and the culture time is 72 hours;
(3) according to the change rule of each medicine and food fungus growth curve, seed liquids with different inoculation amounts in proportion are mixed and inoculated into a fermentation shake flask, the volume of fermentation liquid is 150mL, the total time of the fermentation shake flask culture is 168h, static culture and dynamic culture are alternately carried out in the culture process, the rotating speed is controlled at 220rpm during the dynamic culture, the temperature is controlled at 25 ℃, and the time proportion of the static culture to the dynamic culture is 1: 4;
(4) during dynamic culture, the fermentation shake flask is irradiated by a long-wavelength visible light fluorescent lamp, the irradiation is carried out intermittently, and the total irradiation time is controlled; the wavelength of the long-wavelength visible light is 660 nm; the irradiation time of the long-wavelength visible light is 50 h;
(5) after fermentation, mycelium is obtained by centrifugal separation, and quantitative determination of adenosine and ergosterol is carried out by a high performance liquid chromatograph after leaching treatment and microfiltration membrane filtration, wherein the quantitative determination is respectively 2.7mg/g and 5.1 mg/g.
Example five
A method for increasing the content of adenosine and ergosterol in fungus mixed fermentation liquor further comprises the following steps:
(1) firstly, preparing various medical and edible fungi PDA inclined plane mother strains respectively, wherein the medical and edible fungi are three combinations of hericium erinaceus, shiitake mushrooms and armillaria mellea, the culture temperature is 22 ℃, the humidity is 50%, and the culture time is 7 days;
(2) respectively inoculating seed fermentation shake flasks after the mother seeds are mature, wherein the liquid loading amount of the seed flasks is 50mL, the seed culture medium is a liquid PDA culture medium, 8 glass beads are placed in each flask, the inoculation amount is controlled to be 8 square centimeters, the culture temperature is 22 ℃, the rotating speed is 240rpm, and the culture time is 72 hours;
(3) according to the change rule of each medicine and food fungus growth curve, seed liquids with different inoculation amounts in proportion are mixed and inoculated into a fermentation shake flask, the volume of fermentation liquid is 150mL, the total time of the fermentation shake flask culture is 168h, static culture and dynamic culture are alternately carried out in the culture process, the rotating speed is controlled at 220rpm during the dynamic culture, the temperature is controlled at 22 ℃, and the time proportion of the static culture to the dynamic culture is 1: 3;
(4) during dynamic culture, the fermentation shake flask is irradiated by a long-wavelength visible light fluorescent lamp, the irradiation is carried out intermittently, and the total irradiation time is controlled; the wavelength of the long wavelength visible light is 680 nm; the irradiation time of the long-wavelength visible light is 72 h;
(5) and after the fermentation is finished, performing centrifugal separation to obtain mycelium, performing extraction treatment and microfiltration membrane filtration, and performing quantitative determination on adenosine and ergosterol by using a high performance liquid chromatograph, wherein the content of adenosine and ergosterol is 2.8mg/g and 7.6mg/g respectively.
The above description is only for the preferred embodiment of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art should be considered to be within the technical scope of the present invention, and the technical solutions and the inventive concepts thereof according to the present invention should be equivalent or changed within the scope of the present invention.
Claims (5)
1. A method for increasing the content of adenosine and ergosterol in fungus mixed fermentation liquor comprises the following steps:
(1) firstly, preparing various medical and edible fungus slant mother strains respectively, wherein the culture temperature is 20-25 ℃, the humidity is 45-65%, and the culture time is 7-10 days;
(2) respectively inoculating 5-8 glass beads in each bottle after the mother seeds are mature, fermenting and shaking the bottles, controlling the inoculation amount to be 5-8 square centimeters, controlling the culture temperature to be 20-25 ℃, the rotation speed to be 180-240 rpm, and culturing for 56-72 hours;
(3) according to the change rule of each medicine and food fungus growth curve, seed liquids with different proportions are mixed, inoculated into a fermentation shake flask, the total time of the fermentation shake flask culture is 144-168h, static culture and dynamic culture are alternately carried out in the culture process, the rotating speed is controlled to be 200-240 rpm during the dynamic culture, and the temperature is controlled to be 20-25 ℃;
(4) during dynamic culture, the fermentation shake flask is irradiated by a long-wavelength visible light fluorescent lamp, the irradiation is carried out intermittently, and the total irradiation time is controlled;
(5) and after the fermentation is finished, centrifugally separating to obtain a fermentation liquid supernatant, filtering by using a microporous filter membrane, and then carrying out quantitative determination on adenosine and ergosterol by using a high performance liquid chromatograph.
2. The method of claim 1, wherein the content of adenosine and ergosterol in the fungal mixed fermentation broth is increased by: the various fungi used as medicine and food in the step (1) can be any two or more of lucid ganoderma, cordyceps sinensis, tremella aurantialba, grifola frondosa, hericium erinaceus, shiitake mushroom, armillaria mellea and phellinus igniarius.
3. The method of claim 1, wherein the content of adenosine and ergosterol in the fungal mixed fermentation broth is increased by: the time ratio of static culture to dynamic culture in the step (3) is 1: 2-1: 6, and preferably the ratio is 1: 3-1: 4.5.
4. The method of claim 1, wherein the content of adenosine and ergosterol in the fungal mixed fermentation broth is increased by: the wavelength range of the long-wavelength visible light in the step (4) is 600-700 nm, and the preferable wavelength range is 650-700 nm.
5. The method of claim 1, wherein the content of adenosine and ergosterol in the fungal mixed fermentation broth is increased by: the irradiation time of the long-wavelength visible light in the step (4) is 24-72 hours, and preferably 48-60 hours.
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| CN113616681A (en) * | 2021-09-17 | 2021-11-09 | 江西国药有限责任公司 | Fermented cordyceps sinensis powder with high adenosine and high ergosterol and production method thereof |
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