CN112575039B - Endophytic fungus extract with DPPH free radical scavenging effect, and preparation method and application thereof - Google Patents
Endophytic fungus extract with DPPH free radical scavenging effect, and preparation method and application thereof Download PDFInfo
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- HHEAADYXPMHMCT-UHFFFAOYSA-N dpph Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1[N]N(C=1C=CC=CC=1)C1=CC=CC=C1 HHEAADYXPMHMCT-UHFFFAOYSA-N 0.000 title claims abstract description 25
- 238000002360 preparation method Methods 0.000 title claims abstract description 14
- 241000233866 Fungi Species 0.000 title abstract description 10
- 230000007760 free radical scavenging Effects 0.000 title description 6
- 238000000855 fermentation Methods 0.000 claims abstract description 32
- 230000004151 fermentation Effects 0.000 claims abstract description 32
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims abstract description 12
- 239000007788 liquid Substances 0.000 claims abstract description 8
- 239000002516 radical scavenger Substances 0.000 claims abstract description 7
- 229940123457 Free radical scavenger Drugs 0.000 claims abstract description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 12
- 238000001914 filtration Methods 0.000 claims description 7
- 238000000034 method Methods 0.000 claims description 6
- 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 5
- 235000002595 Solanum tuberosum Nutrition 0.000 claims description 5
- 244000061456 Solanum tuberosum Species 0.000 claims description 5
- 239000012153 distilled water Substances 0.000 claims description 5
- 235000011389 fruit/vegetable juice Nutrition 0.000 claims description 5
- 239000008103 glucose Substances 0.000 claims description 5
- JZRWCGZRTZMZEH-UHFFFAOYSA-N thiamine Chemical compound CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N JZRWCGZRTZMZEH-UHFFFAOYSA-N 0.000 claims description 5
- 241001603662 Diaporthe liquidambaris Species 0.000 abstract description 39
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 abstract description 22
- 150000003254 radicals Chemical class 0.000 abstract description 14
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 abstract description 11
- 229930003268 Vitamin C Natural products 0.000 abstract description 11
- 235000019154 vitamin C Nutrition 0.000 abstract description 11
- 239000011718 vitamin C Substances 0.000 abstract description 11
- -1 DPPH free radical Chemical class 0.000 abstract description 8
- 239000003814 drug Substances 0.000 abstract description 8
- 239000003963 antioxidant agent Substances 0.000 abstract description 7
- 235000006708 antioxidants Nutrition 0.000 abstract description 7
- 229940079593 drug Drugs 0.000 abstract description 7
- 230000000694 effects Effects 0.000 abstract description 7
- 230000003078 antioxidant effect Effects 0.000 abstract description 5
- 238000011161 development Methods 0.000 abstract description 2
- 238000002474 experimental method Methods 0.000 abstract description 2
- 239000001963 growth medium Substances 0.000 description 9
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 6
- 241000196324 Embryophyta Species 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- 239000002609 medium Substances 0.000 description 4
- 239000013641 positive control Substances 0.000 description 4
- 238000006243 chemical reaction Methods 0.000 description 3
- 238000012258 culturing Methods 0.000 description 3
- 238000011218 seed culture Methods 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- 229920001817 Agar Polymers 0.000 description 2
- 108020004414 DNA Proteins 0.000 description 2
- 108091023242 Internal transcribed spacer Proteins 0.000 description 2
- 238000012408 PCR amplification Methods 0.000 description 2
- 239000013543 active substance Substances 0.000 description 2
- 239000008272 agar Substances 0.000 description 2
- 230000032683 aging Effects 0.000 description 2
- MGJZITXUQXWAKY-UHFFFAOYSA-N diphenyl-(2,4,6-trinitrophenyl)iminoazanium Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1N=[N+](C=1C=CC=CC=1)C1=CC=CC=C1 MGJZITXUQXWAKY-UHFFFAOYSA-N 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 230000002538 fungal effect Effects 0.000 description 2
- 230000000670 limiting effect Effects 0.000 description 2
- 239000002207 metabolite Substances 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 238000004321 preservation Methods 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 238000002791 soaking Methods 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 239000008223 sterile water Substances 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 241000984061 Aquilaria Species 0.000 description 1
- 241000271307 Aquilaria malaccensis Species 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 230000002292 Radical scavenging effect Effects 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 238000007605 air drying Methods 0.000 description 1
- 229960000723 ampicillin Drugs 0.000 description 1
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 1
- 230000003064 anti-oxidating effect Effects 0.000 description 1
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- 238000012512 characterization method Methods 0.000 description 1
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- 150000001875 compounds Chemical class 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- IDGUHHHQCWSQLU-UHFFFAOYSA-N ethanol;hydrate Chemical compound O.CCO IDGUHHHQCWSQLU-UHFFFAOYSA-N 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- OOYGSFOGFJDDHP-KMCOLRRFSA-N kanamycin A sulfate Chemical compound OS(O)(=O)=O.O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N OOYGSFOGFJDDHP-KMCOLRRFSA-N 0.000 description 1
- 229960002064 kanamycin sulfate Drugs 0.000 description 1
- 229960002523 mercuric chloride Drugs 0.000 description 1
- LWJROJCJINYWOX-UHFFFAOYSA-L mercury dichloride Chemical compound Cl[Hg]Cl LWJROJCJINYWOX-UHFFFAOYSA-L 0.000 description 1
- 238000012543 microbiological analysis Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
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- 230000004048 modification Effects 0.000 description 1
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- 238000007254 oxidation reaction Methods 0.000 description 1
- 238000013081 phylogenetic analysis Methods 0.000 description 1
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- 230000005855 radiation Effects 0.000 description 1
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- 238000004153 renaturation Methods 0.000 description 1
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- 230000002441 reversible effect Effects 0.000 description 1
- 230000002000 scavenging effect Effects 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 238000010008 shearing Methods 0.000 description 1
- 230000009759 skin aging Effects 0.000 description 1
- 239000008399 tap water Substances 0.000 description 1
- 235000020679 tap water Nutrition 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
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- 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
- C12P1/00—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes
- C12P1/02—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes by using fungi
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/06—Fungi, e.g. yeasts
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
- A61K8/97—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
- A61K8/9728—Fungi, e.g. yeasts
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P39/00—General protective or antinoxious agents
- A61P39/06—Free radical scavengers or antioxidants
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/08—Anti-ageing preparations
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Abstract
The invention discloses an endophytic fungus extract with a DPPH free radical removing effect, and a preparation method and application thereof. The endophytic fungus Phomopsis liquidambaris A814 is used as a fermentation strain, liquid fermentation is carried out to obtain a fermentation culture, mycelium and fermentation liquid are filtered and separated, the fermentation liquid is extracted by ethyl acetate, and the extract is concentrated to obtain the endophytic fungus Phomopsis liquidambaris A814 extract. Experiments show that the extract obtained by separating from the fermentation culture of Phomopsis liquidambaris A814 has obvious activity of removing DPPH free radicals, the DPPH free radical removing capacity of the Phomopsis liquidambaris A814 extract is equivalent to that of vitamin C, and the extract can be used for preparing DPPH free radical scavengers, so that the Phomopsis liquidambaris A814 extract has wide application and development prospects in preparation of antioxidant medicines or skin care products.
Description
Technical Field
The invention belongs to the technical field of biological medicines, and particularly relates to an extract of Aquilaria malaccensis endophytic fungus Phomopsis liquidambaris A814 with a DPPH free radical removing effect, a preparation method thereof and application thereof in preparation of DPPH free radical removing agents.
Background
Antioxidation refers to the abbreviation of antioxidant free radicals, which have close relationship with various human diseases. Free radicals are continuously generated in human bodies due to continuous contact of human bodies with the outside, including respiration (oxidation reaction), external pollution, radiation irradiation and the like, and cancer, aging or other diseases are mostly related to the generation of excessive free radicals. Reducing the generation of free radicals and eliminating aging metabolites have become effective methods for delaying human and skin aging, and since synthetic antioxidants used in pharmaceuticals and skin care products generally have toxic and side effects, there is a certain trend to find natural, highly effective, low-toxic antioxidants.
The plant endophytic fungi has rich species diversity and genetic diversity, so that a plurality of new or rare fungal species are discovered from the plant endophytic fungi so far, and the plant endophytic fungi become research hotspots at home and abroad. Researches show that the active compounds produced by the plant endophytic fungi far exceed the range of plant metabolites, can produce various active substances with unique structure types and potential application prospects, and become important resources for finding new natural active substances.
Disclosure of Invention
The invention aims to provide an endophytic fungus Phomopsis liquidambaris A814 extract with the effect of scavenging DPPH free radicals, and a preparation method and application thereof.
The endophytic fungus Phomopsis liquidambaris A814 extract with the effect of removing DPPH free radicals is prepared by the following preparation method, wherein the endophytic fungus Phomopsis liquidambaris A814 is used as a fermentation strain, liquid fermentation is carried out to obtain a fermentation culture, the fermentation culture is filtered, fermentation liquor is collected and extracted by ethyl acetate, and the extract is concentrated to obtain the endophytic fungus Phomopsis liquidambaris A814 extract.
Preferably, the liquid fermentation is to inoculate the endophytic fungus Phomopsis liquidambaris A814 into a fermentation medium, and to perform liquid fermentation culture at 28 ℃ and 120r/min to obtain a fermentation culture, wherein each liter of the fermentation medium is prepared by the following method: decocting potato 200g in distilled water for 20min, filtering to obtain juice, adding glucose 20g and KH 2 PO 4 3g、MgSO 4 0.75g, vitamin B 1 10mg, adding water to 1L, and naturally adjusting pH to obtain fermentation medium.
The second object of the present invention is to provide an extract of the endophytic fungus Phomopsis liquidambaris A814 prepared by the above preparation method.
The third purpose of the invention is to provide the application of the endophytic fungus Phomopsis liquidambaris A814 extract in the preparation of DPPH free radical scavenger.
A DPPH free radical scavenger contains extract of the endophytic fungus Phomopsis Liquidambaris A814 as active ingredient.
An antioxidant pharmaceutical or skin care product comprises the above DPPH free radical scavenger.
According to the invention, experiments show that the endophytic fungus Phomopsis liquidambaris A814 extract has a DPPH free radical clearance rate of 93.68% at a concentration of 500 mu g/mL, and the positive control drug vitamin C has a DPPH free radical clearance rate of 97.91% at the same concentration. The IC of the endophytic fungus Phomopsis liquidambaris A814 extract for eliminating DPPH free radicals is determined by selecting the DPPH free radicals to be subjected to the minimum inhibitory concentration determination under the 7 action concentrations of 200mg/mL, 100mg/mL, 50mg/mL, 25mg/mL, 12.5mg/mL, 6.25mg/mL and 3.15mg/mL 50 IC of positive control drug vitamin C for DPPH free radical scavenging with a value of 29.06 50 The value was 15.70, the clearance was comparable to vitamin C. The experimental results show that: the extract of endophytic fungus Phomopsis liquidambaris A814 has the effect of remarkably eliminating DPPH free radicals and can be used for preparing DPPH free scavenging agents. Therefore, the endophytic fungus Phomopsis liquidambaris A814 extract has wide application prospect in preparing antioxidant drugs or skin care products.
Phomopsis Liquidambaris A814 of the present invention was deposited at the Guangdong province culture Collection (GDMCC) at 12/9/2020 with addresses: the Guangzhou city first furious Zhonglu No. 100 large yard No. 59 building No. 5 building, the preservation number is: GDMCC No: 3.691, which were available to the depository at the filing date of this application.
Detailed Description
The present invention will be better understood by those skilled in the art from the following examples. The examples are described only to illustrate the invention and should not be construed as limiting the invention as detailed in the claims.
Example 1: isolation, purification and characterization of Strain A814(Phomopsis Liquidambaris)
(1) And (3) separating and purifying strains:
washing leaves of Aquilaria malabarica with tap water, and air drying. Soaking in 0.1% (m/v g/ml) mercuric chloride solution for 5-7 min, rinsing with sterile water for 4 times, soaking in 75% ethanol water solution for 3-5 min, and rinsing with sterile water for 3 times. Shearing into 1cm with scissors 2 Left and right. Placing the strain in a separation culture medium (containing 20 mug/mL kanamycin sulfate and 20 mug/mL ampicillin), culturing for 7 days at a constant temperature box at 28 ℃, then picking marginal hyphae and transferring the marginal hyphae to a fresh separation culture medium, and continuing purifying until obtaining a pure strain, namely the strain A814.
The separation culture medium is obtained by the following method: decocting potato 200g in distilled water for 20min, filtering to obtain juice, adding glucose 20g and KH 2 PO 4 3g、MgSO 4 0.75g, vitamin B 1 10mg, 20g of agar, water to 1L, natural pH, and sterilizing for later use.
(2) Identification of the strains:
and extracting the genome DNA of the strain A814 by using a fungus genome DNA extraction kit as a PCR amplification template. PCR amplification the rDNA ITS region of the isolated strain was amplified using the fungal rDNA internal transcribed spacer (rDNA ITS) universal primers ITS1(5'-TCCGTAGGTGAACCTGCGG-3', forward) and ITS4(5'-TCCTCCGCTTATTGATATGC-3', reverse). PCR was carried out using 20. mu.L of the reaction system by Ex Taq (TaKa-Ra). The reaction conditions are as follows: pre-denaturation at 93 deg.C for 3 min; denaturation at 93 deg.C for 45s, renaturation at 55 deg.C for 45s, and extension at 72 deg.C for 1.5min for 30 cycles; finally, extension is carried out for 10min at 72 ℃. The PCR product was sequenced by Guangzhou division, McJK, and the sequence was shown as SEQ ID NO. 1. Sequence information obtained by sequencing was BLAST, and the BLAST results showed that the strain A814 has 99.63% similarity to Phomopsis liquidambaris (KR 708977). Therefore, the strain was determined to be Phomopsis liquidambaris by molecular phylogenetic analysis and named Phomopsis liquidambaris A814.
The strain Phomopsis liquidambaris A814 was deposited at the Guangdong province culture Collection (GDMCC) at 12/9/2020, address: guangzhou city, Xielizhonglu No. 100 college No. 59 building No. 5, with the preservation number GDMCC NO: 3.691, which were available to the depository at the filing date of this application.
Example 2: activation, fermentation culture and extract preparation of strain A814(Phomopsis liquidambaris)
(1) The pure strain A14 obtained in example 1 was inoculated into a slant culture medium and cultured at 28 ℃ for 5 days to obtain an activated strain. The slant culture medium is obtained by the following method: decocting potato 200g in distilled water for 20min, filtering to obtain juice, and adding glucose 20g and KH 2 PO 4 3g、MgSO 4 0.75g, vitamin B 1 10mg of agar and 20g of water, the solution is made up to 1L with natural pH and sterilized for later use.
(2) Inoculating the activated and cultured strain A814 in the step (1) into a seed culture medium, and culturing for 7d at 28 ℃ under the condition of 120r/min oscillation to obtain a seed solution, wherein the seed culture medium is obtained by the following method: decocting potato 200g in distilled water for 20min, filtering to obtain juice, and adding glucose 20g and KH 2 PO 4 3g、MgSO 4 0.75g, vitamin B 1 10mg, adding water to 1L, naturally adjusting pH, and sterilizing.
(3) Inoculating the seed solution obtained in the step (2) into a fermentation culture medium in an inoculation amount of 10% (v/v), and culturing at 28 ℃ and 120r/min for 7d to obtain a fermentation culture. The fermentation medium is the same as the seed culture medium.
(4) Preparing an extract: and (3) filtering the fermentation culture obtained in the step (3) to remove mycelia, extracting the filtrate for 3 times by using ethyl acetate, and concentrating the extract under reduced pressure to obtain the endophytic fungus Phomopsis liquidambaris A814 extract.
Example 3: determination of the DPPH radical scavenging Capacity of an extract of the endophytic fungus Phomopsis Liquidambaris A814
The extract of Phomopsis Liquidambaris A814 obtained in example 2 and vitamin C were each mixed with absolute ethanol to give 500. mu.g/mL solutions, and DPPH was mixed with absolute ethanol to give 0.2mmoL/L solution. Sucking 100 mu L of Phomopsis liquidambaris A814 extract solution into a 96-well plate, adding 100 mu L of DPPH solution as a sample group, using absolute ethyl alcohol as a blank control group, using absolute ethyl alcohol to replace DPPH solution as a sample background group, using absolute ethyl alcohol to replace Phomopsis liquidambaris A814 extract solution as a negative control group, using vitamin C to replace Phomopsis liquidambaris A814 extract solution as a positive control group, placing the mixture into an incubator at 30 ℃ for reaction for 30min, measuring absorbance (A) at 517nm of each well by using a microplate reader, and calculating the clearance rate% of DPPH free radicals according to the following formula:
clearance% sample/Positive -A Sample background )/(A Negative control -A Blank control )]×100％
3 replicates per sample. The experimental results show that: under the same concentration condition, the DPPH free radical clearance rate of the Phomopsis liquidambaris A814 extract is 93.68%, and the DPPH free radical clearance rate of the vitamin C is 97.91%.
Example 4: half-scavenging concentration of DPPH free radicals (IC) by extract of the endophytic fungus Phomopsis liquidambaris A814 50 Value) determination
The Phomopsis Liquidambaris A814 extract solution (500. mu.g/mL) and the vitamin C solution (500. mu.g/mL) of example 3 were diluted with absolute ethanol to 200. mu.g/mL, 100. mu.g/mL, 50. mu.g/mL, 25. mu.g/mL, 12.5. mu.g/mL, 6.25. mu.g/mL, and 3.15. mu.g/mL, respectively, and the DPPH radical clearance rates of the Phomopsis Liquidambaris A814 extract solution and the vitamin C solution at each concentration were determined as in example 3, and IC was calculated using SigmaPolot 14.0 software 50 The value is obtained. 3 replicates per sample.
The experimental results show that: DPPH free radical scavenging IC by Phomopsis liquidambaris A814 extract 50 IC of positive control drug vitamin C for DPPH free radical scavenging with a value of 29.06 50 The value was 15.70. The experimental results show that the Phomopsis liquidambaris A814 extract has obvious capacity of eliminating DPPH free radicals, the eliminating capacity of the extract is equivalent to that of vitamin C, and the extract can be used for preparing DPPH free radical scavengers, so that the Phomopsis liquidambaris A814 extract has wide application and development prospects in preparing antioxidant medicines or skin care products.
The above are only preferred embodiments of the present invention, and it should be noted that the above preferred embodiments should not be considered as limiting the present invention, and the protection scope of the present invention should be subject to the scope defined by the claims. It will be apparent to those skilled in the art that various modifications and adaptations can be made without departing from the spirit and scope of the invention, and should be considered to be within the scope of the invention.
Sequence listing
<110> Guangdong province institute for microbiology (Guangdong province center for microbiological analysis and detection)
<120> endophytic fungus extract with DPPH free radical scavenging effect, preparation method and application thereof
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<213> endophytic fungus A814(Phomopsis liquidambaris)
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ccaagttaac tcttgttttt acactgaaac tctgagaaaa aacacaaatg aatcaaaact 180
ttcaacaacg gatctcttgg ttctggcatc gatgaagaac gcagcgaaat gcgataagta 240
atgtgaattg cagaattcag tgaatcatcg aatctttgaa cgcacattgc gccctctggt 300
attccggagg gcatgcctgt tcgagcgtca tttcaaccct caagcattgc ttggtgttgg 360
ggcactgctt ttaacgaagc aggccctgaa atctagtggc gagctcgcca ggaccccgag 420
cgtagtagtt aaaccctcgc tttggaaggc cctggcggtg ccctgccgtt aaacccccaa 480
cttctgaaaa tttgacctcg gatcaggtag gaatacccgc tgaacttaag catatcaata 540
agcggaggaa 550
Claims (2)
1. Endophytic fungiPhomopsis liquidambaris Application of A814 extract in preparation of DPPH free radical scavenger, and endophytic fungiPhomopsis liquidambaris The preparation method of the A814 extract comprises the following steps: with endophytic fungiPhomopsis liquidambaris A814 as fermentation strain, fermenting to obtain fermentation culture, filtering to separate mycelium and fermentation broth, extracting the fermentation broth with ethyl acetate, and concentrating the extractive solution to obtain endophytic fungusPhomopsis liquidambaris A814 extract ofPhomopsis liquidambaris Deposit number of a814 is: GDMCC No: 3.691.
2. the use according to claim 1, wherein the liquid fermentation is of an endophytic fungusPhomopsis liquidambaris A814 is inoculated into a fermentation medium, and liquid fermentation culture is carried out at 28 ℃ and 120r/min to obtain a fermentation culture, wherein each liter of the fermentation medium is prepared by the following method: decocting potato 200g in distilled water for 20min, filtering to obtain juice, and adding glucose 20g and KH 2 PO 4 3 g、MgSO 4 0.75g, vitamin B 1 10mg, adding water to 1L, and naturally adjusting pH to obtain fermentation medium.
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