CN112494522B - Application of endophytic fungus A871 extract in preparation of antibacterial drugs - Google Patents
Application of endophytic fungus A871 extract in preparation of antibacterial drugs Download PDFInfo
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- CN112494522B CN112494522B CN202011518314.2A CN202011518314A CN112494522B CN 112494522 B CN112494522 B CN 112494522B CN 202011518314 A CN202011518314 A CN 202011518314A CN 112494522 B CN112494522 B CN 112494522B
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- 229940124350 antibacterial drug Drugs 0.000 title claims abstract description 14
- 241000233866 Fungi Species 0.000 title abstract description 8
- 238000002360 preparation method Methods 0.000 title abstract description 5
- 241000191967 Staphylococcus aureus Species 0.000 claims abstract description 13
- 244000063299 Bacillus subtilis Species 0.000 claims abstract description 12
- 235000014469 Bacillus subtilis Nutrition 0.000 claims abstract description 12
- 239000003814 drug Substances 0.000 claims abstract description 10
- 229940079593 drug Drugs 0.000 claims abstract description 9
- 238000000855 fermentation Methods 0.000 claims description 28
- 230000004151 fermentation Effects 0.000 claims description 28
- 239000001963 growth medium Substances 0.000 claims description 10
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 9
- 239000007788 liquid Substances 0.000 claims description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 5
- 238000001914 filtration 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
- 239000007836 KH2PO4 Substances 0.000 claims description 4
- 244000061456 Solanum tuberosum Species 0.000 claims description 4
- 235000002595 Solanum tuberosum Nutrition 0.000 claims description 4
- 239000012153 distilled water Substances 0.000 claims description 4
- 235000011389 fruit/vegetable juice Nutrition 0.000 claims description 4
- 239000008103 glucose Substances 0.000 claims description 4
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 4
- 238000000034 method Methods 0.000 claims description 4
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 4
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 claims description 4
- 239000011691 vitamin B1 Substances 0.000 claims description 4
- 239000002609 medium Substances 0.000 claims description 3
- DPJRMOMPQZCRJU-UHFFFAOYSA-M thiamine hydrochloride Chemical compound Cl.[Cl-].CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N DPJRMOMPQZCRJU-UHFFFAOYSA-M 0.000 claims description 2
- 241001373667 Peniophora sp. Species 0.000 abstract description 23
- 238000011161 development Methods 0.000 abstract description 5
- 239000013543 active substance Substances 0.000 abstract description 4
- 230000002401 inhibitory effect Effects 0.000 abstract description 3
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 6
- 241000196324 Embryophyta Species 0.000 description 6
- 230000000844 anti-bacterial effect Effects 0.000 description 4
- 238000012258 culturing Methods 0.000 description 4
- 230000018109 developmental process Effects 0.000 description 4
- 230000005764 inhibitory process Effects 0.000 description 4
- 238000011218 seed culture Methods 0.000 description 3
- 230000004913 activation Effects 0.000 description 2
- 230000006978 adaptation Effects 0.000 description 2
- 229960000723 ampicillin Drugs 0.000 description 2
- 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 2
- 239000003242 anti bacterial agent Substances 0.000 description 2
- 230000001580 bacterial effect Effects 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 239000002547 new drug Substances 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 1
- 241000984061 Aquilaria Species 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 230000003698 anagen phase Effects 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000009509 drug development Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 238000000691 measurement method Methods 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 244000000010 microbial pathogen Species 0.000 description 1
- 229930014626 natural product Natural products 0.000 description 1
- 230000010355 oscillation Effects 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 239000012898 sample dilution Substances 0.000 description 1
- 230000001502 supplementing effect Effects 0.000 description 1
Classifications
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- A—HUMAN NECESSITIES
- 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
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
-
- 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
- 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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- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
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- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Mycology (AREA)
- Biotechnology (AREA)
- Wood Science & Technology (AREA)
- General Health & Medical Sciences (AREA)
- Zoology (AREA)
- Medicinal Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- Public Health (AREA)
- Pharmacology & Pharmacy (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Veterinary Medicine (AREA)
- Botany (AREA)
- Animal Behavior & Ethology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Communicable Diseases (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Oncology (AREA)
- Biomedical Technology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Virology (AREA)
- Alternative & Traditional Medicine (AREA)
- Medical Informatics (AREA)
- Epidemiology (AREA)
- Medicines Containing Plant Substances (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses application of endophytic fungus Peniophora sp.A871 in preparation of antibacterial drugs. The invention discovers that the endophytic fungus Peniophora sp.A871 extract has obvious inhibiting effect on staphylococcus aureus and bacillus subtilis, and has good application and development prospect. Therefore, the invention provides a candidate drug for researching and developing a new antibacterial drug and provides a scientific basis for developing and utilizing natural active substances derived from endophytic fungi of plants.
Description
Technical Field
The invention belongs to the technical field of biological medicines, and particularly relates to an application of an aquilaria malabarica endophytic fungus Peniophora sp.A871 extract in preparation of antibacterial drugs.
Background
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 endophytic fungi of plants 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.
At present, the infection caused by the increasingly strong pathogenic microorganisms, especially drug-resistant strains, is found to be more serious in clinical treatment, and many traditional antibiotics basically lose the treatment efficacy, so that the search for novel antibacterial drugs is a key field of new drug development in the world. The microbial resources are rich and various, and various microbial fermentation products are always important medical resources. The development of new drugs or health products by utilizing active natural products of various microorganisms has very important significance and development prospect.
Disclosure of Invention
The invention aims to provide application of an endophytic fungus Peniophora sp.A871 extract in antibacterial drugs.
The invention discovers that when the content of endophytic fungi Peniophora sp.A871 extract in each filter paper sheet is 10 mug, the diameters of inhibition zones of Staphylococcus aureus (Staphylococcus aureus) and Bacillus subtilis (Bacillus subtilis) are 15mm and 13mm respectively through activity tests of staphylococcus aureus (Staphylococcus aureus) and Bacillus subtilis, and the inhibition zones of positive control medicament ampicillin for the 2 test strains are 18mm and 20mm respectively under the same concentration; the experimental results show that: the endophytic fungus Peniophora sp.A871 extract has significant antibacterial activity. Therefore, the endophytic fungus Peniophora sp.A871 extract can be used for preparing antibacterial drugs.
The antibacterial drug is preferably a drug for resisting staphylococcus aureus or bacillus subtilis.
A second object of the present invention is to provide an antibacterial agent comprising an extract of endophytic fungus Peniophora sp.a871 as an active ingredient.
The antibacterial drug is preferably a drug for resisting staphylococcus aureus and bacillus subtilis.
Preferably, the endophytic fungus Peniophora sp.A871 extract is prepared by taking endophytic fungus Peniophora sp.A871 as a fermentation strain, performing liquid fermentation to obtain a fermentation culture, separating mycelium and fermentation liquor, extracting the fermentation liquor with ethyl acetate, and concentrating the extract to obtain the endophytic fungus Peniophora sp.A871 extract.
Preferably, the liquid fermentation is to inoculate endophytic fungus Peniophora sp.A871 into a fermentation culture medium, and perform liquid fermentation culture at 28 ℃ and 120r/min to obtain a fermentation culture, wherein each liter of the fermentation culture medium is prepared by the following method: decocting potato 200g in distilled water for 20min, filtering to obtain juice, and adding glucose 20g and KH2PO4 3g、MgSO40.75g, vitamin B110mg, adding water to 1L, and naturally adjusting pH to obtain fermentation medium.
The invention discovers that the endophytic fungus Peniophora sp.A871 extract has strong inhibiting effect on staphylococcus aureus and bacillus subtilis, and shows that the endophytic fungus Peniophora sp.A871 extract has good antibacterial activity, can be used for preparing antibacterial drugs and has good application and development prospects. Therefore, the invention provides a candidate drug for researching and developing a new antibacterial drug and provides a scientific basis for developing and utilizing natural active substances derived from endophytic fungi of plants.
Endophytic fungus Peniophora sp.A871, deposited at 5.12.2019 in the Guangdong province collection of microorganisms (GDMCC) at the address: the Guangzhou city first furious Zhonglu No. 100 large yard No. 59 building No. 5 building, the preservation number is: GDMCC NO: 60911.
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.
Activation, fermentation culture and extract preparation of strain A871(Peniophora sp.)
(1) Inoculating endophytic fungus Peniophora sp.A871 to a slant culture medium, and culturing at 28 deg.C for 5d to obtain 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 KH2PO4 3g、MgSO40.75g, vitamin B1 10mg，20g of agar, supplementing water to 1L, naturally adjusting the pH value, and sterilizing for later use.
(2) Inoculating the strain A871 subjected to activation culture in the step (1) into a seed culture medium, and culturing for 7d under the conditions of 28 ℃ and 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 KH2PO4 3g、MgSO40.75g, vitamin B110mg, adding water to 1L, naturally adjusting pH, and sterilizing.
(3) Inoculating the seed liquid 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 mycelium, extracting the filtrate for 3 times by using ethyl acetate, and concentrating the extract under reduced pressure to obtain the endophytic fungus Peniophora sp.A871 extract.
Example 1: determination of activity of endophytic fungus Peniophora sp.A871 extract against staphylococcus aureus and bacillus subtilis
The endophytic fungus Peniophora sp.A871 extract was dissolved in DMSO and diluted to a concentration of 2mg/mL to obtain a sample dilution.
Blending Staphylococcus aureus or Bacillus subtilis in logarithmic growth phase with normal saline to give a solution with a concentration of 106Absorbing 1mL of prepared bacterial liquid into a culture dish, pouring LB culture medium cooled to a proper temperature, uniformly mixing, clamping a filter paper sheet with the thickness of 1.5mm and the diameter of 6mm by using a sterile forceps, placing the filter paper sheet on a bacterial-containing flat plate, respectively dropwise adding 5 mu L of the sample diluent on the filter paper sheet to serve as a sample group, replacing the sample diluent with DMSO to serve as a blank control group, using ampicillin as a positive control group, placing the flat plate in a 37 ℃ incubator for culturing for 24h, and measuring the diameter of an antibacterial ring by using a cross measurement method. Repeating the steps for three times, and calculating the average value of the diameters of the inhibition zones.
The results of the experiment are shown in table 1:
table 1: the endophytic fungus Peniophora sp.Inhibition of bacteria by A871 extract
The experimental results show that: the endophytic fungus Peniophora sp.A871 extract has obvious inhibiting effect on staphylococcus aureus and bacillus subtilis, and can be used for preparing medicines for resisting staphylococcus aureus and bacillus subtilis, so that the endophytic fungus Peniophora sp.A871 extract has wide application and development prospects in preparing antibacterial medicines.
The above is only a preferred embodiment of the present invention, and it should be noted that the above preferred embodiment 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 these modifications and adaptations should be considered within the scope of the invention.
Claims (2)
1. Endophytic fungiPeniophorasp. A871 GDMCC NO: 60911 application of the extract in preparing antibacterial drugs; the endophytic fungiPeniophoraThe sp, A871 extract is an endophytic fungusPeniophorasp, A871 as fermentation strain, performing liquid fermentation to obtain fermentation culture, separating mycelium and fermentation broth, extracting the fermentation broth with ethyl acetate, and concentrating the extractive solution to obtain endophytic fungiPeniophorasp, A871 extract, wherein the antibacterial drug is a drug for resisting staphylococcus aureus or bacillus subtilis.
2. Use according to claim 1, wherein the liquid fermentation is of an endophytic fungusPeniophorasp, A871 is inoculated into a fermentation culture medium, liquid fermentation culture is carried out at 28 ℃ and 120r/min to obtain a fermentation culture, and the fermentation cultureThe base is prepared by the following method per liter: decocting potato 200g in distilled water for 20min, filtering to obtain juice, and adding glucose 20g and KH2PO4 3 g、MgSO4 0.75g, vitamin B1 10mg, adding water to 1L, and naturally adjusting pH to obtain fermentation medium.
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| CN202011518314.2A CN112494522B (en) | 2020-12-21 | 2020-12-21 | Application of endophytic fungus A871 extract in preparation of antibacterial drugs |
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| Application Number | Priority Date | Filing Date | Title |
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| CN202011518314.2A CN112494522B (en) | 2020-12-21 | 2020-12-21 | Application of endophytic fungus A871 extract in preparation of antibacterial drugs |
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| CN112494522B true CN112494522B (en) | 2022-02-22 |
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111019838A (en) * | 2019-12-17 | 2020-04-17 | 广东省微生物研究所（广东省微生物分析检测中心） | Endophytic fungus and extract and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| GB0126164D0 (en) * | 2001-10-31 | 2002-01-02 | Danisco | Sequences |
| CN1993379A (en) * | 2004-06-09 | 2007-07-04 | 惠氏公司 | Anti-il-13 antibodies, crystals of anti-il-13 antibodies and complexes comprising the same |
| KR20110137792A (en) * | 2009-03-10 | 2011-12-23 | 팔메드 테바 엘티디 | Novel coprinus comatus and tremella mesenterica muschroom strains, products and extracts thereof and compositions comprising them |
| CN101991715B (en) * | 2010-10-28 | 2012-07-25 | 广东省微生物研究所 | Aquilaria sinensis fruit skin extract and preparation method and application thereof |
| US9125419B2 (en) * | 2012-08-14 | 2015-09-08 | Marrone Bio Innovations, Inc. | Bacillus sp. strain with antifungal, antibacterial and growth promotion activity |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111019838A (en) * | 2019-12-17 | 2020-04-17 | 广东省微生物研究所（广东省微生物分析检测中心） | Endophytic fungus and extract and application thereof |
Non-Patent Citations (3)
| Title |
|---|
| 《一株枝网刺柳珊瑚共附生真菌 Peniophora sp.次级代谢产物研究》;周静;《中国海洋药物杂志》;20120831;第31卷(第4期);第8页左栏第1段 * |
| 《植物内生真菌抗氧化活性成分研究进展》;丁建海;《安徽农业科学》;20141010;第42卷(第29期);第10057页左栏第1段 * |
| 《麦角甾醇过氧化物的抑菌活性研究》;麻兵继;《食品研究与开发》;20120730;第33卷(第7期);第42页摘要 * |
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Address after: 510070 No.56 courtyard, No.100 Xianlie Middle Road, Yuexiu District, Guangzhou City, Guangdong Province Patentee after: Institute of Microbiology, Guangdong Academy of Sciences Address before: 510070 No.56 courtyard, No.100 Xianlie Middle Road, Yuexiu District, Guangzhou City, Guangdong Province Patentee before: GUANGDONG INSTITUTE OF MICROBIOLOGY (GUANGDONG DETECTION CENTER OF MICROBIOLOGY) |
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