CN104756763A - Method for improving content of cordycepin in cordyceps militaris fruiting bodies - Google Patents
Method for improving content of cordycepin in cordyceps militaris fruiting bodies Download PDFInfo
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Abstract
The invention discloses a method for improving the content of cordycepin in cordyceps militaris fruiting bodies. The method comprises the following steps that monascus and cordyceps militaris are independently cultured, then the two microbial strains are mixed for culturing, the mixed microbial strains are exposed to illumination of 200 Ix scattered light no less than 10 hours each day, the temperature is kept at 18-25 DEG C at daytime, the diurnal temperature difference is controlled to be in the range of 5-8 DEG C, illumination is alternated for 14 hours, the relative humidity is controlled to be between 70% and 80%, and ventilation is conducted every day; after the cordyceps militaris send forth stromata, the illumination intensity is enhanced to 800-1000Ix, the temperature is kept at about 22 DEG C, and the air humidity is kept at about 85%; ventilation and air exchange are conducted every day; during the period of the fifteenth day to the twentieth day, the culturing temperature of the cordyceps militaris rises to 29-34 DEG C; then, the culturing temperature of the mixed culture is regulated to a conventional culture temperature for conventional culturing, and the culturing is continued until the harvest time arrives. By the adoption of the method, the yield of the cordyceps militaris fruiting bodies can be obviously improved, the quality of the cordyceps militaris fruiting bodies can further be improved, and the content of the cordycepin in the fruiting bodies is improved by more than 40%.
Description
Technical field
The present invention relates to microorganism and fermentation technical field thereof, be specifically related to a kind of method improving cordycepin content in fruiting bodies of cordyceps militaris.
Background technology
Cordyceps militaris, has another name called northern Chinese caterpillar Fungus, and fungal taxonomy belongs to Ascomycotina, gang pyrenomycetes, ergot Zoopagales, Clavicipitaceae, and Cordyceps is all the one of Cordyceps sinensis fungus with Cordyceps sinensis.Cordyceps sinensis fungus is the general designation parasitizing the class entomogenous fungi compound that the underground fruit body of insect, spider or agglomerate capsule platymiscium is formed.Cordyceps militaris is the type sepecies of Cordyceps sinensis fungus, and main host plants is on the larva and pupa of Lepidoptera, coleoptera and more dipterous insects in the wild, and effect that the classic of TCM records it be " taste is sweet, property is flat; useful lung kidney; benefit marrow, hemostasis and phlegm ", basically identical with the function of Cordyceps sinensis.In the past, due to Cordyceps militaris, output is rare in the wild, and its exploitation are not valued by the people.But from late nineteen nineties in last century, since its extensive artificial cultivation succeeds, its value of exploiting and utilizing is just more and more familiar with by people, the product developed gets more and more, the market demand is also increasing, and the artificial cultivation of fruiting bodies of cordyceps militaris is just becoming the new industry that has very big economic worth.
Cordycepin is first the ucleosides antibiotic separated from fungi, is one of topmost active component in Cordyceps militaris.As far back as nineteen fifty, Cunningham etc. observe by the insect tissue of Cordyceps militaris Cordycepsmilitaris parasitism not perishable, and then therefrom isolate a kind of antibacterial material, name as cordycepin, and deliver on Nature, many scholars have carried out verifying research to it afterwards.Cordycepin molecular formula is C10H13N503, and molecular weight is 251D, alkalescence, needle-like or flaky crystal.Cordycepin has the multiple pharmacological effect such as antitumor, antibacterial, antiviral, immunological regulation, scavenging free radicals.
Summary of the invention
For solving the problem, the invention provides a kind of method improving cordycepin content in fruiting bodies of cordyceps militaris.
For achieving the above object, the technical scheme that the present invention takes is:
Improve a method for cordycepin content in fruiting bodies of cordyceps militaris, comprise the steps:
S1, red colouring agent for food, also used as a Chinese medicine are cultivated:
S11, water-soluble starch 2 ~ 4%, sucrose 3 ~ 5%, peptone 3 ~ 4%, agar 1 ~ 5%, sodium carbonate 0.7 ~ 0.9% mixes with the water of surplus and loads in triangular flask, through sterilizing, cooling, be put into inclined-plane, inoculation red colouring agent for food, also used as a Chinese medicine bacterial classification, in the incubator of 30 DEG C, lucifuge is cultured to mycelia and covers with inclined-plane;
S12, get maltose 20 ~ 24g/L, peptone 30 ~ 34g/L, mannitol 20 ~ 22g/L, sodium nitrate 2 ~ 4g/L, zinc-amino acid chelate 2 ~ 4g/L, potassium dihydrogen phosphate 1 ~ 2g/L mix, and load in triangular flask, with the triangular flask inclined-plane of 40 ~ 60mL sterile water washing step S11 gained, obtaining spore suspension, is the amount of 50 ~ 60% of this seed liquor medium by volume, inoculation, at 30 DEG C, liquid cultivation 1d on the rotary shaker of 180r/min;
S13, get starch 16 ~ 18%, sodium nitrate 0.4 ~ 0.6%, yeast extract 0.6 ~ 0.8%, corn steep liquor 0.8 ~ 1% mix with the water of surplus, pH value is regulated to be after 4.0 with lactic acid, load in triangular flask, be the amount of 50 ~ 60% of this liquid Shake flask medium by volume, the seed liquor of inoculation step S12 gained is cultivated and is planted, at 28 DEG C, liquid Shaking culture 2d under the condition of 180r/min, obtain fermented bacterium
S14, get 25g rice load edible mushroom bag, the material-water ratio by 1: 1.6 adds 40ml distilled water, tightens two ends sack with cord; First temperature is risen to 100 ~ 105 DEG C under normal pressure, insulation 20 ~ 25min; Then temperature is risen to 110 ~ 115 DEG C, insulation 60 ~ 70min; Again temperature is risen to 120 ~ 125 DEG C, 60 ~ 70min is incubated when pressure is 0.13 ~ 0.14MPa, last vexedly again put 30 ~ 35min, stop heating, it is allowed to naturally cool to less than 28 DEG C, take out bottling, the fermented bacterium of inoculation step S13 gained after cooling of having bottled, lucifuge constant temperature culture at putting 30 DEG C;
S2, Cordyceps militaris are cultivated: load corncob 36 ~ 48g, potato residues 8 ~ 10g, sweet potato vine 18 ~ 22g, wheat bran 0.5 ~ 0.9g, calcium carbonate fiber 2 ~ 3g in Cans; Material-water ratio by 1: 1.7 adds 46.3g nutrient solution; With high temperature resistant single-layer polypropylene plastic foil sealing, autoclaving 30min at 120 DEG C, after cooling, the spore suspension of inoculation cordyceps militaris link bacterial strain, dark constant temperature culture at putting 26 DEG C;
S3, the mixing of two bacterium: after the red colouring agent for food, also used as a Chinese medicine cultivated and Cordyceps militaris all back cover, aseptically by the culture of red colouring agent for food, also used as a Chinese medicine and Cordyceps militaris in 1: 6 ~ 1: 10 ratio mix, stir after mixing, lucifuge constant temperature culture at being placed in 26 DEG C, enters fruiting bodies of cordyceps militaris management phase after 5 ~ 7d;
S4, fruit body manage: every day is no less than 10h with the illumination of 200lx scattered light, day temperature remains on 18 ~ 25 DEG C, and day and night temperature controls at 5 ~ 8 DEG C, and 14h illumination alternately, relative moisture controls between 70 ~ 80%, and the 10 ~ 15min that ventilates every day is with supplementary fresh air; After Cordyceps militaris stroma grows, increase intensity of illumination to 800 ~ 1000lx, temperature keeps about 22 DEG C, air humidity about 85%; Ventilation every day 3 times, each 10min; During 15th ~ 20d, Cordyceps militaris cultivation temperature is increased to 29 ~ 34 DEG C; Then the cultivation temperature of culture is adjusted to cellar culture temperature 18 ~ 25 DEG C, carries out cellar culture, until results.
Wherein, described nutrient solution prescription is glucose 20g, peptone 5g, amine sulfate 1.5g, potassium dihydrogen phosphate 1.5g, chelating amino acids manganese 50mg, zinc sulphate 50mg, water 1000ml.
The present invention has following beneficial effect:
Fruiting bodies of cordyceps militaris output can be significantly improved, improve and reach more than 35%; And the quality of fruiting bodies of cordyceps militaris can also be improved, cordycepin content in fruit body is made to improve 40%, this is to the output and the efficiency that improve Cordyceps militaris solid fermentation, and the content improving active ingredient in medicine cordyceps powder and capsule is significant, has good application and development prospect.
Embodiment
In order to make objects and advantages of the present invention clearly understand, below in conjunction with embodiment, the present invention is further elaborated.Should be appreciated that specific embodiment described herein only in order to explain the present invention, be not intended to limit the present invention.
In below implementing, nutrient solution prescription is glucose 20g, peptone 5g, amine sulfate 1.5g, potassium dihydrogen phosphate 1.5g, chelating amino acids manganese 50mg, zinc sulphate 50mg, water 1000ml.
Embodiment 1
S1, red colouring agent for food, also used as a Chinese medicine are cultivated:
S11, water-soluble starch 4%, sucrose 5%, peptone 4%, agar 5%, sodium carbonate 0.9% mixes with the water of surplus and loads in triangular flask, through sterilizing, cooling, is put into inclined-plane, inoculation red colouring agent for food, also used as a Chinese medicine bacterial classification, in the incubator of 30 DEG C, lucifuge is cultured to mycelia and covers with inclined-plane;
S12, get maltose 24g/L, peptone 34g/L, mannitol 22g/L, sodium nitrate 4g/L, zinc-amino acid chelate 4g/L, potassium dihydrogen phosphate 2g/L mix, and load in triangular flask, with the triangular flask inclined-plane of 60mL sterile water washing step S11 gained, obtaining spore suspension, is the amount of 60% of this seed liquor medium by volume, inoculation, at 30 DEG C, liquid cultivation 1d on the rotary shaker of 180r/min;
S13, get starch 18%, sodium nitrate 0.6%, yeast extract 0.8%, corn steep liquor 1% mix with the water of surplus, pH value is regulated to be after 4.0 with lactic acid, load in triangular flask, be the amount of 60% of this liquid Shake flask medium by volume, the seed liquor of inoculation step S12 gained is cultivated and is planted, at 28 DEG C, liquid Shaking culture 2d under the condition of 180r/min, obtain fermented bacterium
S14, get 25g rice load edible mushroom bag, the material-water ratio by 1: 1.6 adds 40ml distilled water, tightens two ends sack with cord; First temperature is risen to 105 DEG C under normal pressure, insulation 25min; Then temperature is risen to 115 DEG C.Insulation 70min; Again temperature is risen to 125 DEG C, be incubated 70min when pressure is 0.14MPa, finally vexedly again put 35min, stop heating, allow it naturally cool to less than 28 DEG C, take out bottling, the fermented bacterium of the rear inoculation step S13 gained of cooling of having bottled, lucifuge constant temperature culture at putting 30 DEG C;
S2, Cordyceps militaris are cultivated: load corncob 48g, potato residues 10g, sweet potato vine 22g, wheat bran 0.9g, calcium carbonate fiber 3g in Cans; Material-water ratio by 1: 1.7 adds 46.3g nutrient solution; With high temperature resistant single-layer polypropylene plastic foil sealing, autoclaving 30min at 120 DEG C, after cooling, the spore suspension of inoculation cordyceps militaris link bacterial strain, dark constant temperature culture at putting 26 DEG C;
S3, the mixing of two bacterium: after the red colouring agent for food, also used as a Chinese medicine cultivated and Cordyceps militaris all back cover, aseptically by the culture of red colouring agent for food, also used as a Chinese medicine and Cordyceps militaris in 1: 10 ratio mix, stir after mixing, lucifuge constant temperature culture at being placed in 26 DEG C, enters fruiting bodies of cordyceps militaris management phase after 7d;
S4, fruit body manage: every day is no less than 10h with the illumination of 200lx scattered light, and day temperature remains on 25 DEG C, and day and night temperature controls at 8 DEG C, and alternately, relative moisture controls between 80%, and the 10 ~ 15min that ventilates every day is with supplementary fresh air in 14h illumination; After Cordyceps militaris stroma grows, increase intensity of illumination to 1000lx, temperature keeps about 22 DEG C, air humidity about 85%; Ventilation every day 3 times, each 10min; During 20d, Cordyceps militaris cultivation temperature is increased to 34 DEG C; Then the cultivation temperature of culture is adjusted to cellar culture temperature 25 DEG C, carries out cellar culture, until results.
Embodiment 2
S1, red colouring agent for food, also used as a Chinese medicine are cultivated:
S11, water-soluble starch 2%, sucrose 3%, peptone 3%, agar 1%, sodium carbonate 0.7 mixes with the water of surplus and loads in triangular flask, through sterilizing, cooling, is put into inclined-plane, inoculation red colouring agent for food, also used as a Chinese medicine bacterial classification, in the incubator of 30 DEG C, lucifuge is cultured to mycelia and covers with inclined-plane;
S12, get maltose 20g/L, peptone 30g/L, mannitol 20g/L, sodium nitrate 2g/L, zinc-amino acid chelate 2g/L, potassium dihydrogen phosphate 1g/L mix, and load in triangular flask, with the triangular flask inclined-plane of 40mL sterile water washing step S11 gained, obtaining spore suspension, is the amount of 50% of this seed liquor medium by volume, inoculation, at 30 DEG C, liquid cultivation 1d on the rotary shaker of 180r/min;
S13, get starch 16%, sodium nitrate 0.4%, yeast extract 0.6%, corn steep liquor 0.8% mix with the water of surplus, pH value is regulated to be after 4.0 with lactic acid, load in triangular flask, be the amount of 50% of this liquid Shake flask medium by volume, the seed liquor of inoculation step S12 gained is cultivated and is planted, at 28 DEG C, liquid Shaking culture 2d under the condition of 180r/min, obtain fermented bacterium
S14, get 25g rice load edible mushroom bag, the material-water ratio by 1: 1.6 adds 40ml distilled water, tightens two ends sack with cord; First temperature is risen to 100 DEG C under normal pressure, insulation 20min; Then temperature is risen to 110 DEG C, insulation 60min; Again temperature is risen to 120 DEG C, be incubated 60min when pressure is 0.13MPa, finally vexedly again put 30min, stop heating, allow it naturally cool to less than 28 DEG C, take out bottling, the fermented bacterium of the rear inoculation step S13 gained of cooling of having bottled, lucifuge constant temperature culture at putting 30 DEG C;
S2, Cordyceps militaris are cultivated: load corncob 36g, potato residues 8g, sweet potato vine 18g, wheat bran 0.5g, calcium carbonate fiber 2g in Cans; By 1: 1, the material-water ratio of 7 adds 46.3g nutrient solution; With high temperature resistant single-layer polypropylene plastic foil sealing, autoclaving 30min at 120 DEG C, after cooling, the spore suspension of inoculation cordyceps militaris link bacterial strain, dark constant temperature culture at putting 26 DEG C;
S3, the mixing of two bacterium: after the red colouring agent for food, also used as a Chinese medicine cultivated and Cordyceps militaris all back cover, aseptically by the culture of red colouring agent for food, also used as a Chinese medicine and Cordyceps militaris in 1: 6 ~ 1: 10 ratio mix, stir after mixing, lucifuge constant temperature culture at being placed in 26 DEG C, enters fruiting bodies of cordyceps militaris management phase after 5d;
S4, fruit body manage: every day is no less than 10h with the illumination of 200lx scattered light, and day temperature remains on 18 DEG C, and day and night temperature controls at 5 DEG C, and alternately, relative moisture controls between 70%, and the 10min that ventilates every day is with supplementary fresh air in 14h illumination; After Cordyceps militaris stroma grows, increase intensity of illumination to 800lx, temperature keeps about 22 DEG C, air humidity about 85%; Ventilation every day 3 times, each 10min; During 15d, Cordyceps militaris cultivation temperature is increased to 29 DEG C; Then the cultivation temperature of culture is adjusted to cellar culture temperature 18 DEG C, carries out cellar culture, until results.
Embodiment 3
S1, red colouring agent for food, also used as a Chinese medicine are cultivated:
S11, water-soluble starch 3%, sucrose 4%, peptone 3.5%, agar 3%, sodium carbonate 0.8% mixes with the water of surplus and loads in triangular flask, through sterilizing, cooling, is put into inclined-plane, inoculation red colouring agent for food, also used as a Chinese medicine bacterial classification, in the incubator of 30 DEG C, lucifuge is cultured to mycelia and covers with inclined-plane;
S12, get maltose 22g/L, peptone 32g/L, mannitol 21g/L, sodium nitrate 3g/L, zinc-amino acid chelate 3g/L, potassium dihydrogen phosphate 1.5g/L mix, and load in triangular flask, with the triangular flask inclined-plane of 50mL sterile water washing step S11 gained, obtaining spore suspension, is the amount of 55% of this seed liquor medium by volume, inoculation, at 30 DEG C, liquid cultivation 1d on the rotary shaker of 180r/min;
S13, get starch 17%, sodium nitrate 0.5%, yeast extract 0.7%, corn steep liquor 0.9% mix with the water of surplus, pH value is regulated to be after 4.0 with lactic acid, load in triangular flask, be the amount of 55% of this liquid Shake flask medium by volume, the seed liquor of inoculation step S12 gained is cultivated and is planted, at 28 DEG C, liquid Shaking culture 2d under the condition of 180r/min, obtain fermented bacterium
S14, get 25g rice load edible mushroom bag, the material-water ratio by 1: 1.6 adds 40ml distilled water, tightens two ends sack with cord; First temperature is risen to 102.5 DEG C under normal pressure, insulation 22.5min; Then temperature is risen to 112.5 DEG C, insulation 65min; Again temperature is risen to 122.5 DEG C, be incubated 65min when pressure is 0.135MPa, finally vexedly again put 32.5min, stop heating, allow it naturally cool to less than 28 DEG C, take out bottling, the fermented bacterium of the rear inoculation step S13 gained of cooling of having bottled, lucifuge constant temperature culture at putting 30 DEG C;
S2, Cordyceps militaris are cultivated: load corncob 42g, potato residues 9g, sweet potato vine 20g, wheat bran 0.7g, calcium carbonate fiber 2.5g in Cans; Material-water ratio by 1: 1.7 adds 46.3g nutrient solution; With high temperature resistant single-layer polypropylene plastic foil sealing, autoclaving 30min at 120 DEG C, after cooling, the spore suspension of inoculation cordyceps militaris link bacterial strain, dark constant temperature culture at putting 26 DEG C;
S3, the mixing of two bacterium: after the red colouring agent for food, also used as a Chinese medicine cultivated and Cordyceps militaris all back cover, aseptically by the culture of red colouring agent for food, also used as a Chinese medicine and Cordyceps militaris in 1: 8 ratio mix, stir after mixing, lucifuge constant temperature culture at being placed in 26 DEG C, enters fruiting bodies of cordyceps militaris management phase after 6d;
S4, fruit body manage: every day is no less than 10h with the illumination of 200lx scattered light, and day temperature remains on 21.5 DEG C, and day and night temperature controls at 6.5 DEG C, and alternately, relative moisture controls between 75%, and the 10 ~ 15min that ventilates every day is with supplementary fresh air in 14h illumination; After Cordyceps militaris stroma grows, increase intensity of illumination to 800 ~ 1000lx, temperature keeps about 22 DEG C, air humidity about 85%; Ventilation every day 3 times, each 10min; During 18d, Cordyceps militaris cultivation temperature is increased to 31 DEG C; Then the cultivation temperature of culture is adjusted to cellar culture temperature 21.5 DEG C, carries out cellar culture, until results.
The above is only the preferred embodiment of the present invention; it should be pointed out that for those skilled in the art, under the premise without departing from the principles of the invention; can also make some improvements and modifications, these improvements and modifications also should be considered as protection scope of the present invention.
Claims (2)
1. improve a method for cordycepin content in fruiting bodies of cordyceps militaris, it is characterized in that, comprise the steps:
S1, red colouring agent for food, also used as a Chinese medicine are cultivated:
S11, water-soluble starch 2 ~ 4%, sucrose 3 ~ 5%, peptone 3 ~ 4%, agar 1 ~ 5%, sodium carbonate 0.7 ~ 0.9% mixes with the water of surplus and is distributed in triangular flask, through sterilizing, cools, is put into inclined-plane, inoculation red colouring agent for food, also used as a Chinese medicine bacterial classification, in the incubator of 30 DEG C, lucifuge is cultured to mycelia and covers with inclined-plane;
S12, get maltose 20 ~ 24g/L, peptone 30 ~ 34g/L, mannitol 20 ~ 22g/L, sodium nitrate 2 ~ 4g/L, zinc-amino acid chelate 2 ~ 4g/L, potassium dihydrogen phosphate 1 ~ 2g/L mix, and load in triangular flask, with the triangular flask inclined-plane of 40 ~ 60mL aseptic washing bar step S11 gained, obtaining spore suspension, is the amount of 50 ~ 60% of this seed liquor medium by volume, inoculation, at 30 DEG C, liquid cultivation 1d on the rotary shaker of 180r/min;
S13, get starch 16 ~ 18%, sodium nitrate 0.4 ~ 0.6%, yeast extract 0.6 ~ 0.8%, corn steep liquor 0.8 ~ 1% mix with the water of surplus, pH value is regulated to be after 4.0 with lactic acid, load in triangular flask, be the amount of 50 ~ 60% of this liquid Shake flask medium by volume, the seed liquor of inoculation step S12 gained is cultivated and is planted, at 28 DEG C, liquid Shaking culture 2d under the condition of 180r/min, obtain fermented bacterium;
S14, get 25g rice load edible mushroom bag, add 40ml distilled water by 1: 1.6 material-water ratio, tighten two ends sack with cord; First temperature is risen to 100 ~ 105 DEG C under normal pressure, insulation 20 ~ 25min; Then temperature is risen to 110 ~ 115 DEG C, insulation 60 ~ 70min; Again temperature is risen to 120 ~ 125 DEG C, 60 ~ 70min is incubated when pressure is 0.13 ~ 0.14MPa, last vexedly again put 30 ~ 35min, stop heating, it is allowed to naturally cool to less than 28 DEG C, take out bottling, the fermented bacterium of inoculation step S13 gained after cooling of having bottled, lucifuge constant temperature culture at putting 30 DEG C;
S2, Cordyceps militaris are cultivated: load corncob 36 ~ 48g, potato residues 8 ~ 10g, sweet potato vine 18 ~ 22g, wheat bran 0.5 ~ 0.9g, calcium carbonate fiber 2 ~ 3g in Cans; Material-water ratio by 1: 1.7 adds 46.3g nutrient solution; With high temperature resistant single-layer polypropylene plastic foil sealing, autoclaving 30min at 120 DEG C, after cooling, the spore suspension of inoculation cordyceps militaris link bacterial strain, dark constant temperature culture at putting 26 DEG C;
S3, the mixing of two bacterium: after the red colouring agent for food, also used as a Chinese medicine cultivated and Cordyceps militaris all back cover, aseptically by the culture of red colouring agent for food, also used as a Chinese medicine and Cordyceps militaris in 1: 6 ~ 1: 10 ratio mix, stir after mixing, lucifuge constant temperature culture at being placed in 26 DEG C, enters fruiting bodies of cordyceps militaris management phase after 5 ~ 7d;
S4, fruit body manage: every day is no less than 10h with the illumination of 200lx scattered light, day temperature remains on 18 ~ 25 DEG C, and day and night temperature controls at 5 ~ 8 DEG C, and 14h illumination alternately, relative moisture controls between 70 ~ 80%, and the 10 ~ 15min that ventilates every day is with supplementary fresh air; After Cordyceps militaris stroma grows, increase intensity of illumination to 800 ~ 1000lx, temperature keeps about 22 DEG C, air humidity about 85%; Ventilation every day 3 times, each 10min; During 15th ~ 20d, Cordyceps militaris cultivation temperature is increased to 29 ~ 34 DEG C; Then the cultivation temperature of culture is adjusted to cellar culture temperature 18 ~ 25 DEG C, carries out cellar culture, until results.
2. a kind of method improving cordycepin content in fruiting bodies of cordyceps militaris according to claim 1, it is characterized in that, described nutrient solution prescription is glucose 20g, peptone 5g, amine sulfate 1.5g, potassium dihydrogen phosphate 1.5g, chelating amino acids manganese 50mg, zinc sulphate 50mg, water 1000ml.
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