CN108901587B - Solid culture method of cordyceps sobolifera - Google Patents
Solid culture method of cordyceps sobolifera Download PDFInfo
- Publication number
- CN108901587B CN108901587B CN201710252724.9A CN201710252724A CN108901587B CN 108901587 B CN108901587 B CN 108901587B CN 201710252724 A CN201710252724 A CN 201710252724A CN 108901587 B CN108901587 B CN 108901587B
- Authority
- CN
- China
- Prior art keywords
- culture
- medium
- light
- solid
- cordyceps sobolifera
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 241000357408 Ophiocordyceps sobolifera Species 0.000 title claims abstract description 33
- 239000007787 solid Substances 0.000 title claims abstract description 28
- 238000012136 culture method Methods 0.000 title claims abstract description 15
- 239000001963 growth medium Substances 0.000 claims abstract description 30
- 238000005286 illumination Methods 0.000 claims abstract description 30
- 238000000034 method Methods 0.000 claims abstract description 10
- 238000003306 harvesting Methods 0.000 claims abstract description 6
- 239000002609 medium Substances 0.000 claims description 14
- 241000233866 Fungi Species 0.000 claims description 11
- 241000931705 Cicada Species 0.000 claims description 10
- 238000011081 inoculation Methods 0.000 claims description 8
- 241000209140 Triticum Species 0.000 claims description 7
- 235000021307 Triticum Nutrition 0.000 claims description 7
- 239000010902 straw Substances 0.000 claims description 5
- 240000007594 Oryza sativa Species 0.000 claims description 4
- 235000007164 Oryza sativa Nutrition 0.000 claims description 4
- 240000008042 Zea mays Species 0.000 claims description 4
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 claims description 4
- 235000002017 Zea mays subsp mays Nutrition 0.000 claims description 4
- 235000013339 cereals Nutrition 0.000 claims description 4
- 235000005822 corn Nutrition 0.000 claims description 4
- 235000009566 rice Nutrition 0.000 claims description 4
- 238000013341 scale-up Methods 0.000 claims description 3
- 235000007319 Avena orientalis Nutrition 0.000 claims description 2
- 241000209763 Avena sativa Species 0.000 claims description 2
- 235000007558 Avena sp Nutrition 0.000 claims description 2
- 229920000742 Cotton Polymers 0.000 claims description 2
- 240000008620 Fagopyrum esculentum Species 0.000 claims description 2
- 235000009419 Fagopyrum esculentum Nutrition 0.000 claims description 2
- 244000068988 Glycine max Species 0.000 claims description 2
- 235000010469 Glycine max Nutrition 0.000 claims description 2
- 240000005979 Hordeum vulgare Species 0.000 claims description 2
- 235000007340 Hordeum vulgare Nutrition 0.000 claims description 2
- 244000046052 Phaseolus vulgaris Species 0.000 claims description 2
- 235000010627 Phaseolus vulgaris Nutrition 0.000 claims description 2
- 244000000231 Sesamum indicum Species 0.000 claims description 2
- 235000003434 Sesamum indicum Nutrition 0.000 claims description 2
- 244000062793 Sorghum vulgare Species 0.000 claims description 2
- 235000019764 Soybean Meal Nutrition 0.000 claims description 2
- 235000021329 brown rice Nutrition 0.000 claims description 2
- 235000012343 cottonseed oil Nutrition 0.000 claims description 2
- 239000002054 inoculum Substances 0.000 claims description 2
- 235000019713 millet Nutrition 0.000 claims description 2
- 239000004455 soybean meal Substances 0.000 claims description 2
- 239000010903 husk Substances 0.000 claims 2
- 239000000243 solution Substances 0.000 description 11
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 9
- 230000000694 effects Effects 0.000 description 8
- 241001625026 Cordyceps cicadae Species 0.000 description 7
- 238000001035 drying Methods 0.000 description 7
- 239000000523 sample Substances 0.000 description 7
- 238000012258 culturing Methods 0.000 description 6
- 239000000843 powder Substances 0.000 description 6
- 230000001954 sterilising effect Effects 0.000 description 6
- 239000002126 C01EB10 - Adenosine Substances 0.000 description 5
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 5
- 229930195725 Mannitol Natural products 0.000 description 5
- 229960005305 adenosine Drugs 0.000 description 5
- 239000000594 mannitol Substances 0.000 description 5
- 235000010355 mannitol Nutrition 0.000 description 5
- 238000002156 mixing Methods 0.000 description 5
- 238000001816 cooling Methods 0.000 description 4
- 238000001514 detection method Methods 0.000 description 4
- 235000013399 edible fruits Nutrition 0.000 description 4
- 150000004676 glycans Chemical class 0.000 description 4
- 238000009630 liquid culture Methods 0.000 description 4
- 229920001282 polysaccharide Polymers 0.000 description 4
- 239000005017 polysaccharide Substances 0.000 description 4
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 239000004480 active ingredient Substances 0.000 description 3
- 238000000855 fermentation Methods 0.000 description 3
- 230000004151 fermentation Effects 0.000 description 3
- 239000000706 filtrate Substances 0.000 description 3
- 238000010438 heat treatment Methods 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 239000012488 sample solution Substances 0.000 description 3
- 238000005303 weighing Methods 0.000 description 3
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 2
- 229930006000 Sucrose Natural products 0.000 description 2
- 238000005273 aeration Methods 0.000 description 2
- 230000003698 anagen phase Effects 0.000 description 2
- 239000002518 antifoaming agent Substances 0.000 description 2
- 238000009792 diffusion process Methods 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 239000000428 dust Substances 0.000 description 2
- IDGUHHHQCWSQLU-UHFFFAOYSA-N ethanol;hydrate Chemical compound O.CCO IDGUHHHQCWSQLU-UHFFFAOYSA-N 0.000 description 2
- 229910001385 heavy metal Inorganic materials 0.000 description 2
- 239000012088 reference solution Substances 0.000 description 2
- 230000001105 regulatory effect Effects 0.000 description 2
- 238000012807 shake-flask culturing Methods 0.000 description 2
- 238000004659 sterilization and disinfection Methods 0.000 description 2
- 229960004793 sucrose Drugs 0.000 description 2
- 238000000870 ultraviolet spectroscopy Methods 0.000 description 2
- 238000009423 ventilation Methods 0.000 description 2
- MLIWQXBKMZNZNF-KUHOPJCQSA-N (2e)-2,6-bis[(4-azidophenyl)methylidene]-4-methylcyclohexan-1-one Chemical compound O=C1\C(=C\C=2C=CC(=CC=2)N=[N+]=[N-])CC(C)CC1=CC1=CC=C(N=[N+]=[N-])C=C1 MLIWQXBKMZNZNF-KUHOPJCQSA-N 0.000 description 1
- BKCWMRVXLGGGBD-WOUKDFQISA-N (2r,3r,4s,5r)-2-(6-aminopurin-9-yl)-2-(2-hydroxyethyl)-5-(hydroxymethyl)oxolane-3,4-diol Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@]1(CCO)O[C@H](CO)[C@@H](O)[C@H]1O BKCWMRVXLGGGBD-WOUKDFQISA-N 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 102000001189 Cyclic Peptides Human genes 0.000 description 1
- 108010069514 Cyclic Peptides Proteins 0.000 description 1
- SHZGCJCMOBCMKK-UHFFFAOYSA-N D-mannomethylose Natural products CC1OC(O)C(O)C(O)C1O SHZGCJCMOBCMKK-UHFFFAOYSA-N 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- PNNNRSAQSRJVSB-UHFFFAOYSA-N L-rhamnose Natural products CC(O)C(O)C(O)C(O)C=O PNNNRSAQSRJVSB-UHFFFAOYSA-N 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 241001248610 Ophiocordyceps sinensis Species 0.000 description 1
- 244000061456 Solanum tuberosum Species 0.000 description 1
- 235000002595 Solanum tuberosum Nutrition 0.000 description 1
- 108010073771 Soybean Proteins Proteins 0.000 description 1
- 229930182558 Sterol Natural products 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- PNNNRSAQSRJVSB-BXKVDMCESA-N aldehydo-L-rhamnose Chemical compound C[C@H](O)[C@H](O)[C@@H](O)[C@@H](O)C=O PNNNRSAQSRJVSB-BXKVDMCESA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 230000036772 blood pressure Effects 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 238000010411 cooking Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000005265 energy consumption Methods 0.000 description 1
- 235000019441 ethanol Nutrition 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000012467 final product Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 1
- 235000013402 health food Nutrition 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 230000001678 irradiating effect Effects 0.000 description 1
- 230000003907 kidney function Effects 0.000 description 1
- 230000037356 lipid metabolism Effects 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical compound O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 description 1
- 230000002906 microbiologic effect Effects 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 235000019796 monopotassium phosphate Nutrition 0.000 description 1
- 239000002777 nucleoside Substances 0.000 description 1
- 125000003835 nucleoside group Chemical group 0.000 description 1
- 235000021049 nutrient content Nutrition 0.000 description 1
- 230000000050 nutritive effect Effects 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- KHIWWQKSHDUIBK-UHFFFAOYSA-N periodic acid Chemical compound OI(=O)(=O)=O KHIWWQKSHDUIBK-UHFFFAOYSA-N 0.000 description 1
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 230000002035 prolonged effect Effects 0.000 description 1
- 239000003531 protein hydrolysate Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 239000013558 reference substance Substances 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 238000011218 seed culture Methods 0.000 description 1
- 230000001953 sensory effect Effects 0.000 description 1
- 238000004904 shortening Methods 0.000 description 1
- 235000019710 soybean protein Nutrition 0.000 description 1
- 150000003432 sterols Chemical class 0.000 description 1
- 235000003702 sterols Nutrition 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 239000012085 test solution Substances 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
- 239000003643 water by type Substances 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G7/00—Botany in general
- A01G7/06—Treatment of growing trees or plants, e.g. for preventing decay of wood, for tingeing flowers or wood, for prolonging the life of plants
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Biodiversity & Conservation Biology (AREA)
- Ecology (AREA)
- Forests & Forestry (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Botany (AREA)
- Environmental Sciences (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention provides a solid culture method of cordyceps sobolifera, which is characterized in that the method comprises the steps of inoculating an expanded cordyceps sobolifera strain to a solid culture medium, carrying out dark culture and light culture in sequence, and harvesting sporophores at proper time to obtain the cordyceps sobolifera; wherein, the light culture stage adopts 16-20 hr/day illumination.
Description
Technical Field
The invention relates to a solid culture method of cordyceps sobolifera, in particular to an artificial culture method of cordyceps sobolifera with fixed illumination time in the light culture process.
Background
The Chinese medicine cicada fungus is a fungus and insect complex formed by Isaria cicadae Miquel (Paecilomyces cicadae (Miquel) Samson) parasitizing on cicada nymphs, is one of traditional and famous Chinese medicinal materials in China, and has 800 years earlier medicinal use than cordyceps sinensis. The cicada fungus belongs to fungi used as both medicine and food, and has higher nutritive value and specific efficacy. The cicada fungus can generate a plurality of physiological active substances which have important effects on medical treatment and health care, including nucleosides, cyclic peptides, polysaccharides, alcohols, sterols, organic acids and the like, and has obvious effects on the aspects of regulating immunity, improving renal function, regulating lipid metabolism, resisting tumors, resisting fatigue, easing pain, hypnotizing, reducing blood pressure and blood sugar and the like.
The natural cordyceps sobolifera resource is very limited, and the quality of the wild cordyceps sobolifera is questioned due to the problems that the wild cordyceps sobolifera is different in production place, different in harvesting time, easy to be polluted by mixed bacteria, mildewed, contains heavy metals and the like. The artificially cultured cicada fungus does not contain heavy metal, has no pollution and has stable nutrient content, so that people pay more and more attention to the cordyceps cicadae. At present, the industrial cultivation of cordyceps sobolifera is realized.
The artificial culture of Isaria cicadae Miquel is carried out by inoculating Isaria cicadae Miquel to solid substrate, culturing in dark, culturing under light to obtain branched multi-branch sporophore, collecting, and drying to obtain final product. The yield and active ingredient content of the raw materials are important for the sensory and quality acceptance of consumers, and different illumination time has certain influence on the yield of sporocarp and the content of effective active ingredients.
At present, the illumination time used in the light culture stage of artificially culturing cordyceps sobolifera is mostly 24 hours, which causes unnecessary energy waste.
Disclosure of Invention
In order to solve the problems in the prior art, the invention provides a solid culture method of cordyceps sobolifera, which comprises the steps of inoculating cordyceps sobolifera strains to a solid culture medium, carrying out dark culture and light culture in sequence, and harvesting sporophores at proper time to obtain the cordyceps sobolifera; wherein, the light culture stage adopts 16-20 hr/day illumination.
Preferably, the cordyceps sobolifera species is selected from the group consisting of scale-up cultured cordyceps sobolifera species; the expanding culture comprises any one or more of slant culture, shake flask culture, seed tank culture, fermenter culture and the like.
Preferably, the light culture period is 18 hours/day light.
In some embodiments, the solid medium is a conventional solid medium used in cicada fungus culture process, such as a grain culture medium, and may be selected from any one or more of wheat, corn, rice, millet, soybean meal, buckwheat, barley, oat, brown rice, and polished round-grained rice; crop straw culture medium, such as one or more of wheat straw, corn stalk, cotton stalk, bean stalk, and sesame stalk; culture medium for crop hulls, such as bran, soybean hull, and cottonseed hull. Wheat medium is preferred.
The inoculation amount is 5-15%; preferably, the inoculation amount is 7 to 12 percent; more preferably, the inoculation amount is 10%.
The culture conditions are as follows: the culture temperature in the dark culture stage is 22-24 ℃, and the relative humidity is 70-85%; the culture temperature in the light culture stage is 20-22 deg.C, and the relative humidity is 70-85%.
The light culture stage adopts composite white light, and the illumination intensity is 150-250 Lx.
The invention has the beneficial effects that: according to the invention, LED white light is adopted in the cicada fungus light culture stage and irradiation is carried out for 18 h/day, and compared with the irradiation of 24h all day illumination in the prior art, the following remarkable effects are achieved: the energy consumption is obviously reduced, and the service lives of the lamp and the control equipment thereof are prolonged; secondly, the illumination time is shortened, the appearance characters and the content of effective components of the sporocarp are not influenced, and the yield is slightly increased; thirdly, the spore yield is greatly reduced, and the trouble caused by dust diffusion in the culture process can be obviously reduced.
Detailed Description
The cordyceps sobolifera strain used in example 1 below was deposited in the general microbiological center of the China Committee for culture Collection of microorganisms with the accession number of CGMCC No. 3453 (the strain is a known strain disclosed in the invention patent with the application number of 201110120603.1). The research of the invention shows that the type of the cordyceps sobolifera strain does not form a factor influencing the effect of the invention, and the drying method of the invention can realize the effect on different cordyceps sobolifera strains.
Example 1 expanded culture of Cordyceps sobolifera
Slant culture: inoculating strains into a slant test tube, and placing the slant test tube inoculated with the strains into an incubator at 22 ℃ for 7 days until hyphae grow over the test tube;
and (3) shake flask culture: placing 200m1 liquid culture medium into a 500m1 triangular flask, sterilizing for 30 minutes under 0.11Mpa, cooling to room temperature, inoculating strains of 1 inclined test tube to the culture medium of the 500m1 triangular flask, and culturing in a constant temperature shaking incubator at 22 +/-1 ℃ and 150 rpm for 3 days;
seed tank culture: charging 20L liquid culture medium into 50L airlift fermentation tank, wherein the temperature of the culture medium is higher than 95 deg.C, adding edible defoaming agent in an amount of 0.03 wt% of the culture medium, introducing steam under 0.11Mpa, heating to 121 deg.C, and maintaining the pressure for 30-45 min; sterilizing, cooling to 20 deg.CWhen in use, 4 bottles of the cultured 500m1 shake flask strains are inoculated into a culture medium, the culture temperature is 22 +/-1 ℃, and the ventilation volume is 1: 0.5V/V min, and maintaining the pressure of 4.903-7.0845 × 104Pa, after 3 days aeration culture, the culture volume is 20L after reaching the logarithmic growth phase.
Culturing in a fermentation tank: charging 200L liquid culture medium into 500L airlift fermentation tank, wherein the temperature of the culture medium is higher than 95 deg.C, adding edible defoaming agent in an amount of 0.03 wt% of the culture medium, introducing steam under 0.11Mpa, heating to 121 deg.C, and maintaining the pressure for 30-45 min; after sterilization, when the seed is cooled to 20 ℃, the seed in the 200L seed tank cultured as above is inoculated and cultured, the culture temperature is 22 +/-1 ℃, and the ventilation volume is 1: 0.5V/V min, and maintaining the pressure of 4.903-7.0845 × 104Pa, after 3 days aeration culture, the logarithmic growth phase is reached. The final culture volume was 200L.
Slant tube medium: 200g of potato cooking juice, 20g of cane sugar, 20g of agar and the balance of water to 1000ml, wherein the pH value is 6.5;
liquid culture medium in shake flasks, seed tanks and fermentors: 30g of yeast extract powder, 30g of cane sugar, 5g of soybean protein hydrolysate and the balance of water, wherein the water is supplemented to 1000ml, and the pH value is 6.5.
Example 2 solid culture of Cordyceps sobolifera
Preparation of solid medium: after wheat is cleaned and water is controlled, a proper amount of water is added, and the weight ratio of the wheat: 1 part of water: 1.4, mixing uniformly; mixing, and placing into culture container with thickness of culture medium controlled at 4 cm. Placing the culture container filled with culture medium in a sterilizing pot, and sterilizing at 121 deg.C for 30-50 min; after sterilization, the culture vessel was transferred into a buffer chamber, naturally cooled to 24 ℃ or lower, and transferred into an inoculation chamber.
Inoculation: irradiating the culture container with ultraviolet light in a superclean bench or a hundred-level laminar flow hood (below) for 0.5h before inoculation; the seed culture obtained in the scale-up culture of example 1 was inoculated in 10% of the inoculum size per culture vessel, and the inoculated vessel was placed in a culture chamber for culture.
The culture conditions are as follows: the culture temperature in the dark culture stage is 22-24 ℃, and the relative air humidity is 70-85%; when the mycelium grows over the culture medium, the culture is switched to light culture, the light source adopts LED white light, the illumination intensity is 200Lx, the illumination time is 18 hours/day, the temperature of the culture room is 20-22 ℃, and the relative air humidity is 70-85%. Ventilating the culture room at proper time to keep the air in the spawning room fresh; until the fruit body is mature, a large amount of spores are not produced (about 23-26 days).
Example 3 solid culture of Cordyceps sobolifera
The cultivation process was substantially the same as in example 2 except that the light irradiation time was 16 hours.
Example 4 solid culture of Cordyceps sobolifera
The cultivation process was substantially the same as in example 2 except that the light irradiation time was 20 hours.
Example 5 solid culture of Cordyceps sobolifera
The cultivation was carried out in substantially the same manner as in example 2 except that the light irradiation time was 22 hours.
Example 6 solid culture of Cordyceps sobolifera
The cultivation was carried out in substantially the same manner as in example 2, except that the light irradiation time was 24 hours.
Example 7 harvesting and drying
Harvesting the fruiting body cultured in any one of examples 2 to 6; and directly feeding the harvested sporocarps into drying equipment for drying. The drying temperature is 60 ℃, and the drying time is 4-5 h.
Effect of illumination time on yield and quality of subelements
The fruiting bodies obtained by culturing in examples 2 to 6 were harvested and dried by the method of example 7, and the yield and quality of the fruiting bodies were evaluated as follows.
1. Influence of illumination time on appearance of cordyceps sobolifera fruiting body
The fruit bodies harvested in each example were scored for their appearance color by 10 scorers according to the scoring criteria in table 1, and the average values are reported in table 2.
TABLE 1 appearance scoring criteria
| Colour(s) | Score value |
| Light yellow-dark yellow | 15-11 |
| Dark yellow-dark gray | 10-6 |
| Dark gray-black | 5-0 |
TABLE 2 Effect of illumination time on appearance color
| Examples | Light culture stage with daily illumination time | Average score |
| Example 2 | 18 h/day | 13.8 |
| Example 3 | 16 h/day | 13.0 |
| Example 4 | 20 h/day | 10.4 |
| Examples5 | 22 h/day | 12.7 |
| Example 6 | 24 h/day | 13.4 |
The results show that the fruit body is yellow in color when the light culture is carried out for 18h, and the color is not very different from the light time groups of other types. Therefore, the appearance shape of the cordyceps sobolifera fruiting body is not influenced by shortening the illumination time to 18 h/day.
2. Influence of illumination time on Cordyceps cicadae yield
The fruiting body yield per 1150g solid medium (dry weight) in examples 2-6 was calculated and the results are shown in Table 3.
Table 3 effect of illumination time on yield results (n ═ 3)
| Examples | Light culture stage with daily illumination time | Yield g/1150g solid medium |
| Example 2 | 18 h/day | 129.75±8.41 |
| Example 3 | 16 h/day | 119.04±6.59 |
| Example 4 | 20 h/day | 121.22±6.03 |
| Example 5 | 22 h/day | 129.18±9.49 |
| Example 6 | 24 h/day | 128.67±9.55 |
As can be seen from Table 3, the yield of the fruiting body is 128.67g/1150g of the solid culture medium when 24 hours of illumination are given each day, the yield of the fruiting body is gradually reduced along with the reduction of the illumination time, and when the illumination time is 20 hours/day, the yield of the fruiting body is only 121.22g/1150g of the solid culture medium, which is reduced by 6%; however, when the light is applied for 18h per day, the yield of fruiting bodies is improved to 129.75g/1150g solid medium, which is equivalent to the 24 h/day light condition. Therefore, the light irradiation time in the light culture stage is shortened to 18 h/day, and the yield of the fruiting body is not reduced.
In addition, the invention unexpectedly discovers that the spore yield can be obviously reduced by adopting illumination for 18 h/day for culture, and the research of the invention shows that the spore yield is 1.2-1.4 g/1150g of solid culture medium by adopting other illumination time for culture, and the spore yield is 0.5-0.8 g/1150g of solid culture medium by adopting illumination for culture, so that the trouble caused by dust diffusion in the culture process can be obviously reduced.
3. Influence of illumination time on effective component content
The fruiting bodies of Cordyceps cicadae cultured in examples 2-6 were collected and the contents of adenosine, HEA (N6- (2-hydroxyethyl adenosine)) and polysaccharide as effective components were measured.
3.1 detection method
3.1.1 determination of adenosine and HEA high performance liquid chromatography was used:
chromatographic conditions and system applicability test: a chromatographic column: symmetry C18(Waters, 4.6mm × 250mm, 5 μm, l.n.0264313291); mobile phase: acetonitrile-0.04 mol/L potassium dihydrogen phosphate (5: 95); flow rate: 1.0 mL/min; detection wavelength: 260 nm; column temperature: 35 ℃; sample introduction amount: 10 μ L. The theoretical plate number is not less than 3000 calculated according to adenosine peak.
Preparation of control solutions: accurately weighing appropriate amount of adenosine and HEA reference substance, and adding 50% methanol water solution to obtain 25 μ g/mL solution.
Preparation of a test solution: precisely weighing 0.2g of cordyceps sobolifera fruiting body powder, placing the powder in a 20mL test tube with a plug, precisely adding 5mL of distilled water, carrying out ultrasonic (40KHz) treatment for 30 minutes, taking out the powder, immediately precisely adding 5mL of methanol, uniformly mixing, passing through a 0.22 mu m microporous filter membrane, and taking out a subsequent filtrate to obtain the cordyceps sobolifera fruiting body powder.
And (3) determination: respectively and precisely sucking 10 μ L of reference solution and sample solution, injecting into high performance liquid chromatograph for 20min, measuring, recording the peak areas corresponding to the reference solution and sample solution at the wavelength, and calculating according to the concentration conversion result.
3.1.2 determination of polysaccharide according to the method of "detection method of functional ingredients of health food" (Baihong Sundao).
3.1.3 detection of mannitol by UV-Vis spectrophotometry
Preparing a sample extracting solution: precisely weighing 0.1g of cordyceps sobolifera fruiting body powder, adding 100mL of 70% ethanol water solution, carrying out reflux extraction in a water bath at 85-90 ℃, taking out after the solution boils for 5min, filtering, taking the filtrate, cooling the filtrate to room temperature, using 70% ethanol water solution to fix the volume to 100mL, and shaking up to obtain the sample solution to be detected.
And (3) measuring the content of mannitol in the sample: precisely transferring 1mL of sample extracting solution, placing the sample extracting solution into a 15mL test tube with a plug, precisely adding 1mL of potassium periodate solution, uniformly mixing, placing the test tube at room temperature for 10min, adding 2mL of 0.1% L-rhamnose solution to remove excessive periodate, shaking and uniformly mixing, finally adding 4mL of freshly prepared Nash reagent, heating the test tube in a water bath at 53 ℃ for 15min to enable the test tube to develop color, and rapidly cooling the test tube to the room temperature. Measuring absorbance at 412nm according to ultraviolet-visible spectrophotometry (appendix VA of pharmacopoeia of the people's republic of China), introducing into a standard curve, determining the concentration (mg/mL) of mannitol in the sample, and calculating the content (mg/g) of mannitol in the sample according to a formula.
Calculating the formula:
3.2 results of measurement
The results are shown in Table 4.
Table 4 influence of illumination time on the content of active ingredient (n ═ 3)
| Solid culture method | Adenosine (%) | HEA(％) | Polysaccharide (%) | Mannitol (%) |
| Example 2 | 0.107±0.004 | 0.088±0.007 | 7.27±0.462 | 5.14±0.387 |
| Example 3 | 0.111±0.007 | 0.103±0.009 | 7.16±0.538 | 5.21±0.297 |
| Example 4 | 0.100±0.005 | 0.094±0.007 | 7.21±0.665 | 5.17±0.384 |
| Example 5 | 0.102±0.008 | 0.096±0.006 | 7.13±0.945 | 5.02±0.257 |
| Example 6 | 0.097±0.004 | 0.093±0.021 | 7.58±0.970 | 5.18±0.265 |
The result shows that the content of the effective components of the fruit body is not obviously changed compared with other illumination time by adopting 18h for light culture, and the inherent quality of the product is not influenced by adopting 18h for light illumination.
Claims (8)
1. A solid culture method of cicada fungus is characterized in that the method comprises the steps of inoculating a cicada fungus strain to a solid culture medium, carrying out dark culture and light culture in sequence, and harvesting sporocarp at proper time to obtain the cicada fungus strain; wherein, the culture temperature in the dark culture stage is 22-24 ℃, and the relative air humidity is 70-85%; when the mycelium grows over the culture medium, the culture is changed into light culture, the light source adopts LED white light, the illumination intensity is 200Lx, the illumination time is 18 hours/day, the temperature of the culture room is 20-22 ℃, and the relative air humidity is 70-85%.
2. The culture method of claim 1, wherein the cordyceps sobolifera species is selected from the group consisting of scale-up culture cordyceps sobolifera species.
3. The culture method according to claim 1 or 2, wherein the solid medium comprises a grain medium, a crop straw medium, a crop husk medium.
4. The culture method according to claim 3, wherein the grain culture medium is a culture medium consisting of one or more of wheat, corn, rice, millet, soybean meal, buckwheat, barley, oat, brown rice and polished round-grained rice; the crop straw culture medium is a culture medium consisting of any one or more of wheat straws, corn stalks, cotton stalks, bean stalks and sesame stalks; the crop husk culture medium is a culture medium consisting of one or more of bran, soybean hull and cottonseed hull.
5. The culture method according to claim 3, wherein the solid medium is a grain medium.
6. The culture method according to claim 1, wherein the inoculation amount is 5% to 15%.
7. The culture method according to claim 6, wherein the inoculum size is 7% to 12%.
8. The culture method according to claim 7, wherein the inoculation amount is 10%.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710252724.9A CN108901587B (en) | 2017-04-18 | 2017-04-18 | Solid culture method of cordyceps sobolifera |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710252724.9A CN108901587B (en) | 2017-04-18 | 2017-04-18 | Solid culture method of cordyceps sobolifera |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN108901587A CN108901587A (en) | 2018-11-30 |
| CN108901587B true CN108901587B (en) | 2021-02-09 |
Family
ID=64402856
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710252724.9A Active CN108901587B (en) | 2017-04-18 | 2017-04-18 | Solid culture method of cordyceps sobolifera |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108901587B (en) |
Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112005814A (en) * | 2019-05-30 | 2020-12-01 | 大叶大学 | Cordyceps sobolifera, solid and liquid culture method thereof and application of extract thereof |
| CN114214199B (en) * | 2022-01-07 | 2023-08-22 | 沈阳农业大学 | Preservation method of Isaria cicadae spore powder |
| CN114586606A (en) * | 2022-03-17 | 2022-06-07 | 连云港市农业科学院 | Culture method of cordyceps sobolifera |
Family Cites Families (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1283778C (en) * | 2004-10-30 | 2006-11-08 | 浙江省农业科学院 | Culture of Chansi mould strains and extraction and use of anagentic compound therefrom |
| CN103283479B (en) * | 2012-02-24 | 2014-10-15 | 北京市弘科农业科技服务站 | Cordyceps militaris strain and application thereof as well as method for producing cordyceps militaris |
| CN102763560B (en) * | 2012-06-19 | 2014-04-23 | 桐乡市中泰虫草专业合作社 | Cordyceps sobolifera fruiting body with cicada larva as host and culture method thereof |
| CN106318875B (en) * | 2015-06-18 | 2020-02-07 | 浙江泛亚生物医药股份有限公司 | Bidirectional artificial culture method of cordyceps sobolifera |
| CN105543104B (en) * | 2016-01-05 | 2017-08-04 | 江苏农林职业技术学院 | A kind of optimization method of the artificial domesticating cultivation solid medium of wild cicada fungus |
| CN106034738B (en) * | 2016-06-06 | 2017-06-16 | 江苏科技大学 | A kind of method with silkworm pupa as host artificial culture silkworm chrysalis cicada fungus |
| CN106190861A (en) * | 2016-07-19 | 2016-12-07 | 湖州新驰医药科技有限公司 | The bacterial screening of a kind of artificial isaria cicadae miq, preparation and productive culture method |
-
2017
- 2017-04-18 CN CN201710252724.9A patent/CN108901587B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN108901587A (en) | 2018-11-30 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103330258B (en) | Cordyceps militaris health-care beverage prepared by liquid submerged fermentation and preparation method thereof | |
| CN102389139B (en) | Preparation method for edible fungus nutritional health-care functional drink | |
| CN103396956B (en) | A kind of saccharomyces cerevisiae and its screening and culturing method and the method for bean pulp fermentation | |
| CN106544242B (en) | Preparation method of beautiful cordyceps sobolifera wine | |
| CN106318875B (en) | Bidirectional artificial culture method of cordyceps sobolifera | |
| CN108901587B (en) | Solid culture method of cordyceps sobolifera | |
| CN110093283B (en) | Beauveria bassiana strain and culture method thereof | |
| CN105981581A (en) | Artificial culture method of cordyceps sobolifera | |
| CN106212044A (en) | A kind of Antrodia Camphorata ware formula cultural method | |
| KR20020069859A (en) | Method using liquid spawn to reduce growth time for mushroom mycelial grain products | |
| CN108624509B (en) | Recycling method of solid culture medium in artificial culture process of cordyceps sobolifera | |
| CN106922387A (en) | A kind of artificial culture method of cicada fungus | |
| CN106922386A (en) | A kind of artificial culture method of cicada fungus | |
| CN107027516A (en) | A kind of selenium-rich Periostracum cicadae, its cultural method and application | |
| CN115039633B (en) | Artificial culture method for sporophore of Isaria japonica | |
| TWM528053U (en) | Antrodia cinnamomea cultivation jar | |
| CN109762745A (en) | A kind of fermentation culture method of Pleurotus tuber-regium and the method that active constituent is extracted from culture | |
| CN106550763B (en) | Artificial culture method of cordyceps sobolifera spore powder | |
| CN106797801B (en) | Artificial culture method of cordyceps sobolifera | |
| CN106922389A (en) | A kind of artificial culture method of cicada fungus | |
| CN108566982A (en) | A kind of drying means of cicada fungus fructification | |
| CN106912293B (en) | Artificial culture method of cordyceps sobolifera | |
| CN106544243B (en) | Preparation method of cordyceps sobolifera wine | |
| CN106544244B (en) | Preparation method of cordyceps sobolifera wine | |
| CN110236175A (en) | A kind of Semen Coicis rich in Pleurotus eryngii erythrothioneine smears sauce and preparation method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CP03 | Change of name, title or address | ||
| CP03 | Change of name, title or address |
Address after: No.1938, xinqun Road, Pinghu Economic Development Zone, Jiaxing City, Zhejiang Province, 314200 Patentee after: Pan Asia Biopharmaceutical Co.,Ltd. Country or region after: China Address before: No.1938, xinqun Road, Pinghu Economic Development Zone, Jiaxing City, Zhejiang Province, 314200 Patentee before: ZHEJIANG BIOASIA PHARMACEUTICAL Co.,Ltd. Country or region before: China |