CN114438153B - Process for improving extraction rate of cordycepin from Guni cordyceps sinensis - Google Patents
Process for improving extraction rate of cordycepin from Guni cordyceps sinensis Download PDFInfo
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- KQLDDLUWUFBQHP-UHFFFAOYSA-N Cordycepin Natural products C1=NC=2C(N)=NC=NC=2N1C1OCC(CO)C1O KQLDDLUWUFBQHP-UHFFFAOYSA-N 0.000 title claims abstract description 62
- OFEZSBMBBKLLBJ-BAJZRUMYSA-N cordycepin Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)C[C@H]1O OFEZSBMBBKLLBJ-BAJZRUMYSA-N 0.000 title claims abstract description 62
- OFEZSBMBBKLLBJ-UHFFFAOYSA-N cordycepine Natural products C1=NC=2C(N)=NC=NC=2N1C1OC(CO)CC1O OFEZSBMBBKLLBJ-UHFFFAOYSA-N 0.000 title claims abstract description 62
- 238000000034 method Methods 0.000 title claims abstract description 42
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- 238000000605 extraction Methods 0.000 title claims abstract description 35
- 241001248610 Ophiocordyceps sinensis Species 0.000 title claims abstract description 32
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- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims abstract description 34
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims abstract description 30
- 239000007788 liquid Substances 0.000 claims abstract description 30
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- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 claims description 3
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- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 10
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 description 6
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- UHOVQNZJYSORNB-UHFFFAOYSA-N Benzene Chemical compound C1=CC=CC=C1 UHOVQNZJYSORNB-UHFFFAOYSA-N 0.000 description 3
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/26—Preparation of nitrogen-containing carbohydrates
- C12P19/28—N-glycosides
- C12P19/38—Nucleosides
- C12P19/40—Nucleosides having a condensed ring system containing a six-membered ring having two nitrogen atoms in the same ring, e.g. purine nucleosides
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- C07H19/02—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof sharing nitrogen
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Abstract
The invention discloses a process for improving the extraction rate of cordycepin from Guni cordyceps sinensis, which comprises the following steps: mixing apple juice, sweet potato juice, glucose, peptone, potassium dihydrogen phosphate, magnesium sulfate and water uniformly, sterilizing, adding activated Cordyceps sinensis mycelia, culturing at 20-30deg.C for 5-15 days, and centrifuging to obtain precipitate; drying the precipitate with hot air, maintaining ventilation in the drying process, pulverizing, sieving, adding into a reflux device, reflux-extracting with boiling water bath for 4-6 hr, and performing ultrasonic treatment to obtain crude liquid; regulating the pH value of the crude liquid to 5-5.5, cooling to-40 to-60 ℃ from room temperature, preserving heat for 1-2h, heating to 30-50 ℃ again, stirring for 1-2h, extracting by adopting ethyl acetate, then filtering by a 4.5 mu m filter membrane, loading on a macroporous resin D1400 column, eluting with distilled water, balancing, eluting by using 15-20% methanol, detecting by HPLC, collecting a sampling tube with the cordycepin peak area exceeding 85%, concentrating and drying.
Description
Technical Field
The invention relates to the technical field of cordycepin extraction, in particular to a process for improving the cordycepin extraction rate of Guni cordyceps sinensis.
Background
Cordycepin (Cordycepin) is an analog of adenosine, of the formula: c (C) 10 H 13 N 5 O 3 Molecular weight of:251.24 it is alkaline, needle-like or plate-like crystals, melting point 230-231 deg.C, maximum absorption wavelength 259.0nm, and is soluble in water, hot ethanol and methanol, and insoluble in benzene, diethyl ether and chloroform.
Cordycepin is a natural medicament, has various pharmacological effects of resisting tumor, resisting bacteria and virus, regulating immunity, scavenging free radicals and the like, and has good clinical application prospect. At present, the research of cordycepin is becoming an extremely active field in pharmaceutical chemistry. Its unique antibacterial and antiviral activity has attracted great attention in the world of advanced technology. It can inhibit viral RNA synthesis; has inhibiting effect on bacillus subtilis and tubercle bacillus; the compound also has killing effect on HIV-I virus; especially has strong inhibiting effect on various solid malignant tumors. Therefore, it is a necessary trend to develop it as a new drug with wide application.
The traditional medicine holds that the cordyceps sinensis is mild in property, warm but not dry, and is not stagnated, so that the cordyceps sinensis can tonify deficiency and strengthen body, and can treat diseases and prolong life, the property of the cordyceps sinensis is more stable than that of ginseng and pilose antler, and the cordyceps sinensis is not easily limited by constitution, symptoms, seasons and ages, and can improve immunity and disease resistance of the patients with strong, weak, healthy and ill diseases. And the modern life has over-fast work rhythm, increased pressure and weakened immune system function, and is very easy to cause diseases such as tumor, hepatitis and the like. Men often have accelerated hair loss and reduced sexual function due to reduced kidney function, and women often have emaciation and insomnia, poor appetite, dysphoria with spleen qi, anxiety, and accelerated arrival of menopause.
The cordycepin has low content in the Guni cordyceps, the selection of a proper enrichment method is a key for extracting the cordycepin, the cordycepin has fat solubility and is easy to damage by heating, the conventional extraction at present is to use 10 times of water for carrying out thermal reflux extraction for 3 times at 80 ℃ for 90 minutes each time, but the extraction efficiency is not high, and the purity of the cordycepin obtained by the method is relatively low, and the cordycepin can realize large-scale production, but the process is complex, so that the production cost is high.
Therefore, the extraction process of the cordyceps polysaccharide and the cordycepin is further researched, so that the extraction rate of the cordyceps polysaccharide and the cordycepin is improved, and the cordyceps polysaccharide and the cordycepin have important economic value and wide market application potential.
Disclosure of Invention
The invention aims to solve the defects in the prior art, and provides a process for improving the extraction rate of cordycepin from Guni cordyceps sinensis.
A process for improving the extraction rate of cordycepin from Guni cordyceps, the method comprises the following steps:
(1) Mixing apple juice, sweet potato juice, glucose, peptone, potassium dihydrogen phosphate, magnesium sulfate and water uniformly, sterilizing, adding activated Cordyceps sinensis mycelia, fermenting at 20-30deg.C for 5-15 days, maintaining the tank pressure at 0.02-0.04MPa during fermentation culture, stirring at 100-150r/min, and centrifuging to obtain precipitate;
(2) Drying the precipitate with hot air, maintaining ventilation in the drying process, pulverizing, sieving, adding into a reflux device, and mixing with 95% ethanol according to a feed-liquid ratio of 1: reflux-extracting with boiling water bath for 4-6 hr, and ultrasonic treatment to obtain coarse liquid;
(3) Regulating the pH value of the crude liquid to be 5-5.5, cooling to-40 to-60 ℃ from room temperature, preserving heat for 1-2h, heating to 30-50 ℃ again, stirring for 1-2h, extracting by adopting ethyl acetate, then passing through a 4.5 mu m filter membrane, loading on a macroporous resin D1400 column, eluting with distilled water, balancing, eluting by using 15-20% methanol, detecting by HPLC, collecting a sampling tube with the cordycepin peak area exceeding 85%, concentrating, and drying to obtain the Gunicordyceps sinensis.
Preferably, in the step (1), the activated cordyceps sinensis mycelia are subjected to sealing and sterilization, the acremonium terrestris is inoculated under aseptic conditions, and the culture is carried out for 5-10 days at the constant temperature of 20-25 ℃; then inoculating into the sterilized second culture medium with an inoculum size of 6-10%, and culturing at 15-25deg.C for 2-4 days while maintaining ventilation of 0.01-0.03vvm to obtain activated Cordyceps sinensis mycelia.
Preferably, in step (1), the first medium comprises, in parts by weight: 1-4 parts of maltose, 5-10 parts of egg albumen powder and 0.1-0.2 part of KH 2 PO 4 0.1-0.2 part of magnesium sulfate and 80-100 parts of water;
the second culture medium comprises the following components in parts by weight: 15-25 parts of potato powder, 1-5 parts of glucose and 2 parts of8 parts of whey protein concentrate, 0.1-0.2 parts of KH 2 PO 4 0.1-0.2 part of magnesium sulfate and 60-100 parts of water.
Preferably, in the step (1), the mass ratio of apple juice, sweet potato juice, glucose, peptone, potassium dihydrogen phosphate, magnesium sulfate and activated cordyceps sinensis mycelia is 1-5:1-5:1-3:1-2:0.01-0.05:0.01-0.05:5-10.
Preferably, in the step (1), the sterilization temperature is 115-125 ℃ and the sterilization time is 10-20min.
Preferably, in the fermentation culture process of the step (1), continuously introducing sterile air, and introducing air volume of 0.02-0.03vvm.
Preferably, in the step (2), the hot air drying temperature is 40-50 ℃ and the hot air drying time is 2-6h.
Preferably, in the step (2), the ultrasonic frequency is 20-30kHz, and the ultrasonic power is 300-350W.
Preferably, in the step (3), the temperature is reduced from room temperature to-40 to-60 ℃ at a temperature reducing speed of 5-10 ℃/min.
Preferably, in the step (3), the temperature is raised from-40 to-60 ℃ to 30-50 ℃ at a temperature raising speed of 1-3 ℃/min.
The technical effects of the invention are as follows:
(1) In the step (2), after the precipitate is dried by hot air at the temperature of 40-50 ℃, the precipitate has no damage to active ingredients, and is further matched with ethanol hot water bath reflux, and ultrasonic treatment is assisted in the reflux extraction process, so that the use amount of ethanol can be effectively reduced, and the extraction efficiency can be improved.
(2) The invention further adjusts the pH value of the crude liquid to 5-5.5, then carries out quick freezing and cooling, does not cause any damage to components, and also cooperates with the quick cooling and slow heating processes, thereby effectively causing rupture of hydrophobic bond structures of cell membranes, further causing rupture of cell membranes and cell walls, and effectively enhancing the permeability of the cell membranes and the cell walls in an acidic environment, and effectively promoting quick diffusion of active components in the rupture process, and accelerating precipitation of cordycepin components.
(3) Cordycepin has fat solubility, is easy to damage by heating, adopts thermal reflux extraction in conventional extraction, and has low extraction efficiency. Aiming at the technical problems to be solved, after fermentation solid-liquid separation, the invention adopts improved low-temperature and other modes to extract the cordyceps sinensis mycelia, so that the components are not damaged, the extraction rate of cordycepin is improved, the invention greatly saves resources, and the content and efficacy of active ingredients are improved.
Detailed Description
The invention is further illustrated below in connection with specific embodiments.
Example 1
A process for improving the extraction rate of cordycepin from Guni Cordyceps comprises the following steps:
(1) Uniformly mixing 1kg of apple juice, 1kg of sweet potato juice, 1kg of glucose, 1kg of peptone, 0.01kg of monopotassium phosphate, 0.01kg of magnesium sulfate and 500kg of water, sterilizing at 125 ℃ for 20min, adding 5kg of activated cordyceps sinensis mycelia, fermenting and culturing at 30 ℃ for 15 days, keeping the tank pressure at 0.04MPa in the fermentation and culturing process, stirring at 150r/min, introducing sterile air at 0.03vvm, and centrifuging to obtain a precipitate;
(2) Drying the precipitate with 40 ℃ hot air for 2 hours, maintaining ventilation in the drying process, crushing, sieving with a 60-mesh sieve, adding into a reflux device, and adopting 95% ethanol according to a feed-liquid ratio of 1: reflux-extracting in boiling water bath for 4h, wherein ultrasonic treatment is assisted in the reflux-extracting process, the ultrasonic frequency is 20kHz, and the ultrasonic power is 300W, so as to obtain crude liquid;
(3) Regulating the pH value of the crude liquid to 5-5.5 by adopting hydrochloric acid with the concentration of 0.1mol/L, cooling to-40 ℃ at the speed of 5 ℃/min from room temperature, preserving heat for 1h, heating to 30 ℃ at the speed of 1 ℃/min, stirring for 1h at the speed of 1000r/min, extracting by adopting ethyl acetate, filtering by a 4.5 mu m filter membrane, loading a sample macroporous resin D1400 column, eluting with distilled water for balancing, eluting by using 15-20% methanol, detecting by HPLC, collecting a sampling tube with the cordycepin peak area exceeding 85%, concentrating and drying to obtain the cordycepin.
Example 2
A process for improving the extraction rate of cordycepin from Guni Cordyceps comprises the following steps:
(1) Uniformly mixing 5kg of apple juice, 5kg of sweet potato juice, 3kg of glucose, 2kg of peptone, 0.05kg of monopotassium phosphate, 0.05kg of magnesium sulfate and 500kg of water, sterilizing at 125 ℃ for 20min, adding 10kg of activated cordyceps sinensis mycelia, fermenting and culturing at 30 ℃ for 15 days, keeping the tank pressure at 0.04MPa in the fermentation and culturing process, stirring at 150r/min, introducing sterile air at 0.03vvm, and centrifuging to obtain a precipitate;
(2) Drying the precipitate with 50 ℃ hot air for 6 hours, maintaining ventilation in the drying process, crushing, sieving with a 60-mesh sieve, adding into a reflux device, and adopting 95% ethanol according to a feed-liquid ratio of 1: reflux-extracting in boiling water bath for 6h, wherein ultrasonic treatment is assisted in the reflux-extracting process, the ultrasonic frequency is 30kHz, and the ultrasonic power is 350W, so as to obtain crude liquid;
(3) Regulating the pH value of the crude liquid to 5-5.5 by adopting hydrochloric acid with the concentration of 1mol/L, cooling to-60 ℃ at the speed of 10 ℃/min from room temperature, preserving heat for 2 hours, heating to 50 ℃ at the speed of 3 ℃/min, stirring for 2 hours at the speed of 2000r/min, extracting by adopting ethyl acetate, filtering by a 4.5 mu m filter membrane, loading a macroporous resin D1400 column, eluting with distilled water for balancing, eluting by using 15-20% methanol, detecting by HPLC, collecting a sampling tube with the cordycepin peak area exceeding 85%, concentrating and drying to obtain the cordycepin.
Example 3
A process for improving the extraction rate of cordycepin from Guni Cordyceps comprises the following steps:
(1) Uniformly mixing 2kg of apple juice, 4kg of sweet potato juice, 1.5kg of glucose, 1.7kg of peptone, 0.02kg of monopotassium phosphate, 0.04kg of magnesium sulfate and 300kg of water, sterilizing at 122 ℃ for 13min, adding 8kg of activated cordyceps sinensis mycelia, fermenting and culturing at 22 ℃ for 13 days, keeping the tank pressure at 0.025MPa in the fermentation and culturing process, stirring at 140r/min, and centrifuging to obtain a precipitate, wherein the ventilation rate of sterile air is 0.022 vvm;
(2) Drying the precipitate with 47 ℃ hot air for 3 hours, maintaining ventilation in the drying process, crushing, sieving with a 60-mesh sieve, adding into a reflux device, and adopting 95% ethanol according to a feed-liquid ratio of 1:7, reflux-extracting for 4.5h in boiling water bath, wherein ultrasonic treatment is assisted in the reflux-extracting process, the ultrasonic frequency is 27kHz, and the ultrasonic power is 320W, so as to obtain crude liquid;
(3) Regulating the pH value of the crude liquid to 5-5.5 by adopting hydrochloric acid with the concentration of 0.6mol/L, cooling to-55 ℃ at the speed of 6 ℃/min from room temperature, preserving heat for 1.3h, heating to 35 ℃ at the speed of 2.5 ℃/min, stirring for 1.3h at the speed of 1800r/min, extracting by adopting ethyl acetate, then filtering by a 4.5 mu m filter membrane, loading on a macroporous resin D1400 column, eluting with distilled water for balancing, eluting by using 15-20% methanol, detecting by HPLC, collecting a sampling tube with the cordycepin peak area exceeding 85%, concentrating and drying to obtain the cordycepin.
Example 4
A process for improving the extraction rate of cordycepin from Guni Cordyceps comprises the following steps:
(1) Uniformly mixing 4kg of apple juice, 2kg of sweet potato juice, 2.5kg of glucose, 1.3kg of peptone, 0.04kg of monopotassium phosphate, 0.02kg of magnesium sulfate and 400kg of water, sterilizing at 118 ℃ for 17min, adding 6kg of activated cordyceps sinensis mycelia, fermenting and culturing at 28 ℃ for 8 days, keeping the tank pressure at 0.035MPa in the fermentation and culturing process, stirring at 110r/min, introducing sterile air at 0.028vvm, and centrifuging to obtain precipitate;
(2) Drying the precipitate with hot air at 43deg.C for 5 hr, maintaining ventilation in the drying process, pulverizing, sieving with 60 mesh sieve, adding into reflux device, and mixing with 95% ethanol at a feed-liquid ratio of 1: reflux-extracting in boiling water bath for 5.5h, wherein ultrasonic treatment is assisted in the reflux-extracting process, the ultrasonic frequency is 24kHz, and the ultrasonic power is 330W, so as to obtain crude liquid;
(3) Regulating the pH value of the crude liquid to 5-5.5 by adopting hydrochloric acid with the concentration of 0.3mol/L, cooling to-45 ℃ at the speed of 8 ℃/min from room temperature, preserving heat for 1.7h, heating to 45 ℃ at the speed of 1.5 ℃/min, stirring for 1.7h at the speed of 1200r/min, extracting by adopting ethyl acetate, then filtering with 4.5 μm filter membrane, loading onto macroporous resin D1400 column, eluting with distilled water, eluting with 15-20% methanol, detecting by HPLC, collecting sample tube with cordycepin peak area exceeding 85%, concentrating, and drying to obtain cordycepin.
Example 5
A process for improving the extraction rate of cordycepin from Guni Cordyceps comprises the following steps:
(1) Uniformly mixing 3kg of apple juice, 3kg of sweet potato juice, 2kg of glucose, 1.5kg of peptone, 0.03kg of monopotassium phosphate, 0.03kg of magnesium sulfate and 350kg of water, sterilizing at 120 ℃ for 15min, adding 7kg of activated cordyceps sinensis mycelia, fermenting and culturing at 25 ℃ for 10 days, keeping the tank pressure at 0.03MPa in the fermentation and culturing process, stirring at 120r/min, introducing air volume of sterile air at 0.025vvm, and centrifuging to obtain a precipitate;
sealing the first culture medium in activated Cordyceps sinensis mycelia, sterilizing, inoculating Acremonium terrestris under aseptic condition, and culturing at 23deg.C for 6 days; then inoculating the strain into a sterilized second culture medium with an inoculum size of 8%, and culturing at 25 ℃ for 3 days while maintaining ventilation of 0.02vvm to obtain activated cordyceps sinensis mycelia;
the first culture medium comprises the following components in parts by weight: 2 parts of maltose, 6 parts of egg albumen powder and 0.12 part of KH 2 PO 4 0.12 part of magnesium sulfate and 100 parts of water; the second culture medium comprises the following components in parts by weight: 20 parts of potato powder, 2 parts of glucose, 4 parts of whey protein concentrate and 0.12 part of KH 2 PO 4 0.12 part of magnesium sulfate and 100 parts of water;
(2) Drying the precipitate with 45 ℃ hot air for 4 hours, maintaining ventilation in the drying process, crushing, sieving with a 60-mesh sieve, adding into a reflux device, and adopting 95% ethanol according to a feed-liquid ratio of 1:6.5 reflux-extracting for 5h in boiling water bath, wherein ultrasonic treatment is assisted in the reflux-extracting process, the ultrasonic frequency is 26kHz, and the ultrasonic power is 325W, so as to obtain crude liquid;
(3) Regulating the pH value of the crude liquid to 5-5.5 by adopting hydrochloric acid with the concentration of 0.45mol/L, cooling to-50 ℃ at the speed of 7 ℃/min from room temperature, preserving heat for 1.5h, heating to 40 ℃ at the speed of 2 ℃/min, stirring for 1.5h at the speed of 1500r/min, extracting by adopting ethyl acetate, then passing through a 4.5 mu m filter membrane, loading a macroporous resin D1400 column, eluting with distilled water for balancing, eluting by using 15-20% methanol, detecting by HPLC, collecting a sampling tube with the cordycepin peak area exceeding 85%, concentrating and drying to obtain the cordycepin.
Comparative example
A process for improving the extraction rate of cordycepin from Guni Cordyceps comprises the following steps:
(1) Uniformly mixing 3kg of apple juice, 3kg of sweet potato juice, 2kg of glucose, 1.5kg of peptone, 0.03kg of monopotassium phosphate, 0.03kg of magnesium sulfate and 350kg of water, sterilizing at 120 ℃ for 15min, adding 7kg of activated cordyceps sinensis mycelia, fermenting and culturing at 25 ℃ for 10 days, keeping the tank pressure at 0.03MPa in the fermentation and culturing process, stirring at 120r/min, introducing air volume of sterile air at 0.025vvm, and centrifuging to obtain a precipitate; wherein the activated cordyceps sinensis mycelia are the same as the activation method of example 5;
(2) Drying the precipitate with 45 ℃ hot air for 4 hours, maintaining ventilation in the drying process, crushing, sieving with a 60-mesh sieve, adding into a reflux device, and adopting 95% ethanol according to a feed-liquid ratio of 1:6.5 reflux-extracting for 5h in boiling water bath, wherein ultrasonic treatment is assisted in the reflux-extracting process, the ultrasonic frequency is 26kHz, and the ultrasonic power is 325W, so as to obtain crude liquid;
(3) Extracting the crude liquid by using ethyl acetate, then filtering the crude liquid by a 4.5 mu m filter membrane, loading the crude liquid into a macroporous resin D1400 column, eluting and balancing by distilled water, eluting by using 15-20% methanol, detecting by HPLC, collecting a sampling tube with the cordycepin peak area exceeding 85%, concentrating and drying to obtain the cordycepin of Gunie.
The method of example 5 and comparative example was used to extract the cordycepin of the Guni, and was specifically as follows:
the calculated formula of the extraction rate of the cordycepin of the Guni cordyceps sinensis is as follows:
the purity of the Guni Cordyceps sinensis cordycepin is calculated as follows:
from the above table, it can be seen that: the extraction rate of the cordycepin from the Chinese caterpillar fungus can reach 4.02 per mill, which is far more than that of the comparative example (2.14 per mill), and the purity can reach 99.35 percent.
The foregoing is only a preferred embodiment of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art, who is within the scope of the present invention, should make equivalent substitutions or modifications according to the technical scheme of the present invention and the inventive concept thereof, and should be covered by the scope of the present invention.
Claims (9)
1. A process for improving the extraction rate of cordycepin from Guni cordyceps sinensis is characterized by comprising the following steps:
(1) Mixing apple juice, sweet potato juice, glucose, peptone, potassium dihydrogen phosphate, magnesium sulfate and water uniformly, sterilizing, adding activated Cordyceps sinensis mycelia, fermenting at 20-30deg.C for 5-15 days, maintaining the tank pressure at 0.02-0.04MPa during fermentation culture, stirring at 100-150r/min, and centrifuging to obtain precipitate;
in the step (1), the mass ratio of apple juice, sweet potato juice, glucose, peptone, potassium dihydrogen phosphate, magnesium sulfate and activated cordyceps sinensis mycelia is 1-5:1-5:1-3:1-2:0.01-0.05:0.01-0.05:5-10;
(2) Drying the precipitate with hot air, maintaining ventilation in the drying process, pulverizing, sieving, adding into a reflux device, and mixing with 95% ethanol according to a feed-liquid ratio of 1: reflux-extracting with boiling water bath for 4-6 hr, and ultrasonic treatment to obtain coarse liquid;
(3) Regulating the pH value of the crude liquid to be 5-5.5, cooling to-40 to-60 ℃ from room temperature, preserving heat for 1-2h, heating to 30-50 ℃ again, stirring for 1-2h, extracting by adopting ethyl acetate, then passing through a 4.5 mu m filter membrane, loading on a macroporous resin D1400 column, eluting with distilled water, balancing, eluting by using 15-20% methanol, detecting by HPLC, collecting a sampling tube with the cordycepin peak area exceeding 85%, concentrating, and drying to obtain the Gunicordyceps sinensis.
2. The process for improving the extraction rate of the cordycepin from the Gunii Cordyceps sinensis according to claim 1, wherein in the step (1), the activated Gunii Cordyceps sinensis mycelia are subjected to sterilization after sealing a first culture medium, the Acremonium terrestris is inoculated under aseptic conditions, and the culture is carried out for 5-10 days at a constant temperature of 20-25 ℃; then inoculating into the sterilized second culture medium with an inoculum size of 6-10%, and culturing at 15-25deg.C for 2-4 days while maintaining ventilation of 0.01-0.03vvm to obtain activated Cordyceps sinensis mycelia.
3. The process for improving the extraction rate of cordycepin of Cordycepin Goodyear of claim 2, wherein in the step (1), the first medium comprises, in parts by weight: 1-4 parts of maltose, 5-10 parts of egg albumen powder and 0.1-0.2 part of KH 2 PO 4 0.1-0.2 part of magnesium sulfate and 80-100 parts of water;
the second culture medium comprises the following components in parts by weight: 15-25 parts of potato powder, 1-5 parts of glucose, 2-8 parts of whey protein concentrate and 0.1-0.2 part of KH 2 PO 4 0.1-0.2 part of magnesium sulfate and 60-100 parts of water.
4. The process for improving the extraction rate of cordycepin from Cordycepin Goodyear of claim 1, wherein in step (1), the sterilization temperature is 115-125 ℃ and the sterilization time is 10-20min.
5. The process for improving the extraction rate of the cordycepin from the Chinese caterpillar fungus, as claimed in claim 1, wherein in the fermentation culture process of the step (1), sterile air is continuously introduced, and the ventilation rate is 0.02-0.03vvm.
6. The process for improving the extraction rate of cordycepin from Cordycepin Goodyear of claim 1, wherein in the step (2), the hot air drying temperature is 40-50deg.C, and the hot air drying time is 2-6h.
7. The process for improving the extraction rate of cordycepin of Cordycepin Goodyear of claim 1, wherein in the step (2), the ultrasonic frequency is 20-30kHz and the ultrasonic power is 300-350W.
8. The process for improving the extraction rate of cordycepin from Cordycepin Goodyear of claim 1, wherein in the step (3), the cooling rate from room temperature to-40 to-60 ℃ is 5-10 ℃/min.
9. The process for improving the extraction rate of cordycepin from Cordycepin Goodyenne according to claim 1, wherein in the step (3), the temperature is raised from-40 to-60 ℃ to 30-50 ℃ at a temperature-raising speed of 1-3 ℃/min.
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| CN109810201A (en) * | 2019-03-01 | 2019-05-28 | 江南大学 | The ultrasonic wave combination of acidic water extracting method of Cordyceps sinensis polysaccharide and cordycepin in a kind of Cordyceps militaris |
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| CN106317148A (en) * | 2016-07-29 | 2017-01-11 | 河北省科学院生物研究所 | Method for extracting cordycepin from cordyceps militaris |
| CN109182152A (en) * | 2018-11-15 | 2019-01-11 | 重庆农笑农业发展有限公司 | For improving the culture medium of aweto mycelium active constituent |
| CN109810201A (en) * | 2019-03-01 | 2019-05-28 | 江南大学 | The ultrasonic wave combination of acidic water extracting method of Cordyceps sinensis polysaccharide and cordycepin in a kind of Cordyceps militaris |
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