CN110616153A - Application of trichoderma koningii LCCC30119 in production of cellulase and tobacco fermentation - Google Patents
Application of trichoderma koningii LCCC30119 in production of cellulase and tobacco fermentation Download PDFInfo
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- 238000000855 fermentation Methods 0.000 title claims abstract description 70
- 230000004151 fermentation Effects 0.000 title claims abstract description 70
- 241000378866 Trichoderma koningii Species 0.000 title claims abstract description 59
- 241000208125 Nicotiana Species 0.000 title claims abstract description 41
- 235000002637 Nicotiana tabacum Nutrition 0.000 title claims abstract description 41
- 108010059892 Cellulase Proteins 0.000 title claims abstract description 33
- 229940106157 cellulase Drugs 0.000 title claims abstract description 32
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 12
- 108090000790 Enzymes Proteins 0.000 claims abstract description 20
- 102000004190 Enzymes Human genes 0.000 claims abstract description 20
- 229940088598 enzyme Drugs 0.000 claims abstract description 20
- 239000007788 liquid Substances 0.000 claims description 22
- 239000001963 growth medium Substances 0.000 claims description 16
- 239000007787 solid Substances 0.000 claims description 15
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- 235000011130 ammonium sulphate Nutrition 0.000 claims description 9
- 229940041514 candida albicans extract Drugs 0.000 claims description 9
- 235000005822 corn Nutrition 0.000 claims description 9
- 238000011081 inoculation Methods 0.000 claims description 9
- 238000009630 liquid culture Methods 0.000 claims description 9
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 9
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 9
- 239000012138 yeast extract Substances 0.000 claims description 9
- 235000013312 flour Nutrition 0.000 claims description 7
- 239000000725 suspension Substances 0.000 claims description 7
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 6
- 238000005273 aeration Methods 0.000 claims description 5
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 4
- 239000012153 distilled water Substances 0.000 claims description 4
- 239000008103 glucose Substances 0.000 claims description 4
- 240000007594 Oryza sativa Species 0.000 claims description 3
- 235000007164 Oryza sativa Nutrition 0.000 claims description 3
- 238000001914 filtration Methods 0.000 claims description 3
- 239000000203 mixture Substances 0.000 claims description 3
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 claims description 3
- LWIHDJKSTIGBAC-UHFFFAOYSA-K potassium phosphate Substances [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 claims description 3
- 235000009566 rice Nutrition 0.000 claims description 3
- 229920001817 Agar Polymers 0.000 claims description 2
- 244000061456 Solanum tuberosum Species 0.000 claims description 2
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- 238000011068 loading method Methods 0.000 claims description 2
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- 230000000694 effects Effects 0.000 abstract description 10
- 239000001913 cellulose Substances 0.000 abstract description 4
- 229920002678 cellulose Polymers 0.000 abstract description 4
- 230000015556 catabolic process Effects 0.000 abstract description 2
- 238000006731 degradation reaction Methods 0.000 abstract description 2
- 238000011031 large-scale manufacturing process Methods 0.000 abstract description 2
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- 241000228212 Aspergillus Species 0.000 description 1
- PZIRUHCJZBGLDY-UHFFFAOYSA-N Caffeoylquinic acid Natural products CC(CCC(=O)C(C)C1C(=O)CC2C3CC(O)C4CC(O)CCC4(C)C3CCC12C)C(=O)O PZIRUHCJZBGLDY-UHFFFAOYSA-N 0.000 description 1
- 229920000742 Cotton Polymers 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 241000208278 Hyoscyamus Species 0.000 description 1
- CWVRJTMFETXNAD-KLZCAUPSSA-N Neochlorogenin-saeure Natural products O[C@H]1C[C@@](O)(C[C@@H](OC(=O)C=Cc2ccc(O)c(O)c2)[C@@H]1O)C(=O)O CWVRJTMFETXNAD-KLZCAUPSSA-N 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 241000223259 Trichoderma Species 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- CWVRJTMFETXNAD-JUHZACGLSA-N chlorogenic acid Chemical compound O[C@@H]1[C@H](O)C[C@@](O)(C(O)=O)C[C@H]1OC(=O)\C=C\C1=CC=C(O)C(O)=C1 CWVRJTMFETXNAD-JUHZACGLSA-N 0.000 description 1
- 229940074393 chlorogenic acid Drugs 0.000 description 1
- FFQSDFBBSXGVKF-KHSQJDLVSA-N chlorogenic acid Natural products O[C@@H]1C[C@](O)(C[C@@H](CC(=O)C=Cc2ccc(O)c(O)c2)[C@@H]1O)C(=O)O FFQSDFBBSXGVKF-KHSQJDLVSA-N 0.000 description 1
- 235000001368 chlorogenic acid Nutrition 0.000 description 1
- BMRSEYFENKXDIS-KLZCAUPSSA-N cis-3-O-p-coumaroylquinic acid Natural products O[C@H]1C[C@@](O)(C[C@@H](OC(=O)C=Cc2ccc(O)cc2)[C@@H]1O)C(=O)O BMRSEYFENKXDIS-KLZCAUPSSA-N 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 229920005610 lignin Polymers 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 239000001814 pectin Substances 0.000 description 1
- 229920001277 pectin Polymers 0.000 description 1
- 235000010987 pectin Nutrition 0.000 description 1
- 239000008104 plant cellulose Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000000391 smoking effect Effects 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000008223 sterile water Substances 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 239000002023 wood Substances 0.000 description 1
Classifications
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- A—HUMAN NECESSITIES
- A24—TOBACCO; CIGARS; CIGARETTES; SIMULATED SMOKING DEVICES; SMOKERS' REQUISITES
- A24B—MANUFACTURE OR PREPARATION OF TOBACCO FOR SMOKING OR CHEWING; TOBACCO; SNUFF
- A24B15/00—Chemical features or treatment of tobacco; Tobacco substitutes, e.g. in liquid form
- A24B15/18—Treatment of tobacco products or tobacco substitutes
- A24B15/20—Biochemical treatment
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
- C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
- C12N9/2405—Glucanases
- C12N9/2434—Glucanases acting on beta-1,4-glucosidic bonds
- C12N9/2437—Cellulases (3.2.1.4; 3.2.1.74; 3.2.1.91; 3.2.1.150)
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- Organic Chemistry (AREA)
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- Wood Science & Technology (AREA)
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- Microbiology (AREA)
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Abstract
The invention provides an application of Trichoderma koningii (LCCC 30119) in cellulase production and tobacco fermentation, wherein Trichoderma koningii (LCCC 30119) can produce cellulase with high enzyme activity, and Trichoderma koningii (LCCC 30119) is used for producing the cellulase, so that a new way is provided for the production source of the cellulase, and the large-scale production of the cellulase can be realized; the trichoderma koningii with cellulase capable of producing high enzyme activity is applied to tobacco leaf fermentation, so that the degradation of cellulose in tobacco leaves can be accelerated, the fermentation period of the tobacco leaves is obviously shortened, and the quality of the tobacco leaves is improved.
Description
Technical Field
The invention relates to application of Trichoderma koningii (Trichoderma koningi) LCCC30119 in tobacco fermentation for producing cellulase, and belongs to the technical field of microbial application.
Background
Cellulase is not a single enzyme, but refers to a group of three enzymes which have synergistic action and can biodegrade plant cellulose to generate glucose, and only under the synergistic action of the three enzymes, cellulose molecules can be finally decomposed into glucose. The main enzymes of the cellulase in the tobacco mellowing process can promote the change of the physical and chemical properties of the tobacco.
At present, after tobacco leaves are picked and subjected to threshing and redrying, industrial usability cannot be realized, the tobacco leaves also need to be naturally alcoholized and artificially alcoholized to cause backlog of time and funds, great loss can be brought to tobacco enterprises, meanwhile, industrial unavailability can be caused by some tobacco leaves along with the lapse of time, and greater loss can be brought, and the significance of the biological enzyme is that the usability of the tobacco leaves can be improved.
Research shows that many microorganisms can decompose cellulose, and the current cellulase-producing fungi mainly comprise Trichoderma and Aspergillus. Trichoderma koningii is the most commonly used strain for producing cellulase, but the report of producing high-activity cellulase and fermenting tobacco leaves by utilizing the existing Trichoderma koningii strain is not seen.
Disclosure of Invention
The invention aims to provide application of Trichoderma koningii (LCCC 30119) in cellulase production and tobacco fermentation, and the Trichoderma koningii (LCCC 30119) can produce cellulase in high yield and is very suitable for cellulase production and tobacco fermentation.
On one hand, the invention provides application of Trichoderma koningii (Trichoderma koningii) LCCC30119 in production of cellulase, and the Trichoderma koningii LCCC30119 can produce cellulase with high enzyme activity, so that a new source is provided for a production source of the cellulase.
On the other hand, the invention also provides a method for producing cellulase by using Trichoderma koningii (LCCC 30119), which comprises the steps of fermenting and culturing Trichoderma koningii (LCCC 30119), and centrifuging the fermentation liquor to obtain crude enzyme liquid.
Preferably, the fermentation culture is a liquid fermentation culture or a solid fermentation culture.
Preferably, the conditions of the liquid fermentation culture are as follows: the inoculation amount of Trichoderma koningii (Trichoderma koningii) LCCC30119 is 105-107cfu/ml, the fermentation temperature is 30-60 ℃, the fermentation time is 2-4 d, the fermentation culture is aeration culture, and the ventilation quantity is 1: 0.8-1.2V/V.min;
preferably, the inoculum size is 106cfu/ml, fermentation temperature of 50 ℃, time of 3d, and ventilation volume of 1.0V/V.min.
Preferably, the liquid culture medium used for liquid fermentation culture contains 4g/L of bran powder, 0.5g/L of yeast extract, 2g/L of corn flour, 0.1g/L of ammonium sulfate and 0.2g/L of potassium dihydrogen phosphate, the pH value is 5-8, and the loading amount of the liquid culture medium is 60-80%;
preferably, the pH is 7.6 and the liquid medium is 75% in volume.
Preferably, the solid fermentation culture adopts a three-stage fermentation process, and specifically comprises the following steps:
(1) culturing Trichoderma koningii LCCC30119 strain on a slant PDA culture medium to obtain a strain suspension;
(2) inoculating the strain suspension on a liquid culture medium for amplification culture to obtain a seed solution;
(3) mixing the seed liquid with the mixture at a ratio of 105-107Inoculating cfu/g into a solid culture medium, culturing at 30-60 ℃ in a ventilating way, then drying in vacuum, adding distilled water and filtering to obtain fermentation liquor.
Preferably, the PDA medium contains: 200g/L of potato, 20g/L of glucose and 15g/L of agar;
and/or the liquid medium contains: 4g/L of bran powder, 0.5g/L of yeast extract, 2g/L of corn powder, 0.1g/L of ammonium sulfate and 0.2g/L of potassium dihydrogen phosphate;
and/or the solid medium contains: 55% of rice hull powder, 30% of bran, 10% of corn flour, 3% of yeast extract, 1.5% of ammonium sulfate and 0.5% of monopotassium phosphate;
preferably, the ventilation rate of the ventilation culture is 0.8-1.2V/V.min;
preferably, the ventilation rate is 1.0V/V.min.
Preferably, the temperature of the fermentation liquor centrifugation is 0-4 ℃, the rotating speed is 5000-8000 rpm, and the time is 10-20 min; preferably, the temperature is 4 ℃, the rotation speed is 6000rpm, and the time is 15 min.
On the other hand, the invention also provides application of Trichoderma koningii (Trichoderma koningi) LCCC30119 or the method in tobacco leaf fermentation.
On the other hand, the invention also provides a tobacco leaf fermentation method, and Trichoderma koningii LCCC30119 is inoculated in tobacco leaves for fermentation.
Preferably, the inoculation amount of Trichoderma koningii (Trichoderma koningi) LCCC30119 is 105～107cfu/g, the fermentation temperature is 30-60 ℃, the humidity is 40-70%, and the time is 5-10 d.
The invention has the beneficial effects that:
the invention discloses that Trichoderma koningii (Trichoderma koningii) LCCC30119 can produce cellulase with high enzyme activity, provides a new way for the production source of the cellulase, and can realize the large-scale production of the cellulase; the trichoderma koningii with cellulase capable of producing high enzyme activity is applied to tobacco leaf fermentation, so that the degradation of cellulose in tobacco leaves can be accelerated, the fermentation period of the tobacco leaves is obviously shortened, and the quality of the tobacco leaves is improved.
Detailed Description
The present invention is described in detail with reference to specific examples, which are provided to facilitate the understanding of the technical solutions of the present invention by those skilled in the art, and the implementation or use of the present invention is not limited by the description of the present invention.
In the present invention, the raw materials and equipment used are commercially available or commonly used in the art, if not specified. Trichoderma koningii (Trichoderma koningi) used in the invention is purchased from Liaoning province microorganism strain preservation center, and the serial numbers are as follows: LCCC 30119.
The methods in the examples are conventional in the art unless otherwise specified.
Example 1Production of cellulase by fermentation of Trichoderma koningii (Trichoderma koningi) LCCC30119
The liquid fermentation process comprises the following steps: and (3) determining the liquid fermentation process of the Corning wood enzyme LCCC30119 by integrating the results of the single-factor multi-level test and the multi-factor multi-level orthogonal test:
(1) preparing strains: preparing the strain on the slant PDA culture medium into spore suspension with sterile water, wherein the spore suspension has a bacterial count of 107-108cfu/ml。
(2) Liquid fermentation medium: contains bran powder 4g/L, yeast extract 0.5g/L, corn flour 2g/L, ammonium sulfate 0.1g/L, potassium dihydrogen phosphate 0.2g/L, pH 6.5.5, and culture medium in an amount of 75% of the tank volume.
(3) Sterilization of a fermentation tank culture medium: 121 ℃ and 2 h.
(4) Inoculation: the sterile inoculation is carried out by utilizing a flame ring, and the inoculation amount is 5 percent.
(5) Liquid culture: the culture temperature is 34 ℃, the culture medium is inoculated into a fermentation tank with 75% of liquid culture medium, and the culture is carried out for 72 hours under ventilation with the ventilation rate of 1:1 (V/V.min).
The solid fermentation process comprises the following steps: the solid fermentation process of trichoderma koningii LCCC30119 adopts a three-stage fermentation process: slant culture-liquid culture-solid culture.
(1) Slant surface strain: culturing the strain on slant PDA culture medium to obtain the concentration of 107-108cfu/ml of bacterial suspension.
(2) Seed liquid culture: comprises bran powder 4g/L, yeast extract 0.5g/L, corn flour 2g/L, ammonium sulfate 0.1g/L, potassium dihydrogen phosphate 0.2g/L, pH 6.5, ventilating culture for 48 hr, ventilation amount of 1:1(V/V.min), seed liquid concentration of 108cfu/ml。
(3) Solid culture: 55% of rice hull powder, 30% of bran, 10% of corn flour, 3% of yeast extract, 1.5% of ammonium sulfate and 0.5% of monopotassium phosphate, and distilled water is added according to the mass ratio of the materials to the water of 1: 1.
(4) And (3) sterilizing a culture medium: 121 ℃ and 4 h.
(5) Inoculation amount: 5 to 10 percent.
(6) Solid culture: the culture was carried out for 96h under aeration.
(7) Performing aeration culture, vacuum drying, adding distilled water 10 times of the dry weight of the solid koji, shaking at 40 deg.C (200rpm) for 45min, and filtering with absorbent cotton to obtain fermentation broth.
Preparation of crude enzyme solution: and (3) freezing and centrifuging the fermentation liquor obtained by liquid fermentation culture or solid fermentation culture for 15min at the temperature of 4 ℃ and the rotating speed of 6000rpm to obtain supernatant, namely the crude enzyme liquid.
Through the research on the optimum reaction temperature, the optimum pH value, the thermal stability and the pH stability of cellulase produced by Trichoderma koningii (Trichoderma koningi) LCCC30119, the optimum reaction temperature of cellulase produced by Trichoderma koningii LCCC30119 is determined to be 50 ℃, the optimum pH value is determined to be 7.6, and the enzyme activity of fermentation liquor reaches 154U/ml. The enzyme activity is reduced at 30-60 deg.C, and the temperature is higher than 60 deg.C. The enzyme activity is still kept above 50% of the maximum value when the pH is 5.0.
Example 2Application of Trichoderma koningii (Trichoderma koningii) LCCC30119 in tobacco leaf fermentation
Inoculating Trichoderma koningii (Trichoderma koningii) LCCC30119 to tobacco leaves, wherein the inoculation amount of Trichoderma koningii LCCC30119 is 105～107cfu/g, the fermentation temperature is 30-60 ℃, the humidity is 40-70%, and the time is 5-10 d.
Or adding the fermentation supernatant enzyme solution of Trichoderma koningii (Trichoderma koningii) LCCC30119 obtained in example 2 into tobacco leaves for fermentation, wherein the cellulase activity of the added fermentation supernatant enzyme solution in the tobacco leaves is 10-20U/mg, and the fermentation conditions are as follows: the temperature is 30-60 ℃, the humidity is 40-70%, and the time is 5-10 d.
The invention takes 2018 Chongqing B4F raw tobacco as a control group, and 10 is added6The tobacco leaves are fermented by using cfu/g Trichoderma koningii (Trichoderma koningii) LCCC30119 as a treatment group 1 and adding 15U/mg prepared cellulase enzyme powder as a treatment group 2, wherein the fermentation temperature is 50 ℃, the humidity is 55% and the time is 8d, the fermentation period of the tobacco leaves is obviously shortened, and the change of each chemical component in the fermented tobacco leaves is shown in Table 1.
As can be seen from table 1, when Trichoderma koningii (Trichoderma koningii) LCCC30119 or cellulase is added, the contents of chlorogenic acid, starch, pectin and lignin in tobacco leaves are reduced, the contents of total sugar and hyoscyamus are increased, and after smoking evaluation, the tobacco fragrance is coordinated, the smoke is pure and mild, the tobacco fragrance is increased, and the smoothness, comfort and richness of the smoke are improved.
TABLE 1 chemical composition of tobacco leaves in each treatment
The above description is only an example of the present application, and the protection scope of the present application is not limited by these specific examples, but is defined by the claims of the present application. Various modifications and changes may occur to those skilled in the art. Any modification, equivalent replacement, improvement, etc. made within the technical idea and principle of the present application should be included in the protection scope of the present application.
Claims (10)
1. Application of Trichoderma koningii (Trichoderma koningi) LCCC30119 in producing cellulase.
2. The method for producing the cellulase by using the Trichoderma koningi LCCC30119 is characterized by comprising the steps of carrying out fermentation culture on the Trichoderma koningi LCCC30119, and centrifuging the fermentation liquor to obtain a crude enzyme solution.
3. The method of claim 2, wherein the fermentation culture is a liquid fermentation culture or a solid fermentation culture.
4. The method of claim 2, wherein the conditions of the liquid fermentation culture are: the inoculation amount of Trichoderma koningii (Trichoderma koningii) LCCC30119 is 105-107cfu/ml, the fermentation temperature is 30-60 ℃, the fermentation time is 2-4 d, the fermentation culture is aeration culture, and the ventilation quantity is 1: 0.8-1.2V/V.min;
preferably, the inoculum size is 106cfu/ml, fermentation temperature of 50 ℃, time of 3d, ventilation volume of 1: 1.0V/V.min.
5. The method according to claim 2 or 3, wherein the liquid culture medium used for liquid fermentation culture comprises 4g/L of bran powder, 0.5g/L of yeast extract, 2g/L of corn flour, 0.1g/L of ammonium sulfate, 0.2g/L of potassium dihydrogen phosphate, pH of 5-8, and the loading amount of the liquid culture medium is 60-80%;
preferably, the pH is 7.6 and the liquid medium is 75% in volume.
6. The method according to claim 2, wherein the solid fermentation culture adopts a three-stage fermentation process, and specifically comprises the following steps:
(1) cultivation of Trichoderma koningii (Trichoderma koningii) on slant PDA medium
koningii) LCCC30119 strain to obtain a strain suspension;
(2) inoculating the strain suspension on a liquid culture medium for amplification culture to obtain a seed solution;
(3) mixing the seed liquid with the mixture at a ratio of 105-107Inoculating cfu/g into a solid culture medium, performing aeration culture at the temperature of 30-60 ℃, performing vacuum drying, and adding distilled water for filtering to obtain a fermentation liquid;
preferably, the PDA medium contains: 200g/L of potato, 20g/L of glucose and 15g/L of agar;
and/or the liquid medium contains: 4g/L of bran powder, 0.5g/L of yeast extract, 2g/L of corn powder, 0.1g/L of ammonium sulfate and 0.2g/L of potassium dihydrogen phosphate;
and/or the solid medium contains: 55% of rice hull powder, 30% of bran, 10% of corn flour, 3% of yeast extract, 1.5% of ammonium sulfate and 0.5% of monopotassium phosphate;
preferably, the ventilation rate of the ventilation culture is 1: 0.8-1.2V/V.min;
preferably, the ventilation rate is 1: 1.0V/V.min.
7. The method according to claim 2, wherein the fermentation liquid is centrifuged at 0-4 ℃ and 5000-8000 rpm for 10-20 min.
8. Use of Trichoderma koningii (Trichoderma koningii) LCCC30119 or the method of any one of claims 2 to 7 for tobacco leaf fermentation.
9. A tobacco leaf fermentation method is characterized in that Trichoderma koningii (LCCC 30119) is inoculated in tobacco leaves for fermentation.
10. The tobacco leaf fermentation method according to claim 9, wherein the inoculation amount of trichoderma koningii LCCC30119 is 105～107cfu/g, the fermentation temperature is 30-60 ℃, the humidity is 40-70%, and the time is 5-10 d.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201910913660.1A CN110616153A (en) | 2019-09-25 | 2019-09-25 | Application of trichoderma koningii LCCC30119 in production of cellulase and tobacco fermentation |
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| CN111481479A (en) * | 2020-04-20 | 2020-08-04 | 中国农业科学院麻类研究所 | Method for extracting antioxidant component from cannabis sativa leaves and application of antioxidant component |
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