CN115197243A - Sulfur-containing dibenzofuran type alkaloid and preparation method and application thereof - Google Patents
Sulfur-containing dibenzofuran type alkaloid and preparation method and application thereof Download PDFInfo
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- 238000002360 preparation method Methods 0.000 title claims abstract description 17
- TXCDCPKCNAJMEE-UHFFFAOYSA-N dibenzofuran Chemical compound C1=CC=C2C3=CC=CC=C3OC2=C1 TXCDCPKCNAJMEE-UHFFFAOYSA-N 0.000 title abstract description 10
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 title abstract description 6
- 239000011593 sulfur Substances 0.000 title abstract description 6
- 229910052717 sulfur Inorganic materials 0.000 title abstract description 6
- 229930013930 alkaloid Natural products 0.000 title abstract description 5
- 150000003797 alkaloid derivatives Chemical class 0.000 title abstract description 5
- 150000001875 compounds Chemical class 0.000 claims abstract description 36
- 241000266342 Alternaria longissima Species 0.000 claims abstract description 13
- 239000000284 extract Substances 0.000 claims abstract description 11
- 239000000725 suspension Substances 0.000 claims abstract description 11
- 244000052616 bacterial pathogen Species 0.000 claims abstract description 8
- 238000004519 manufacturing process Methods 0.000 claims abstract description 5
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 81
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 45
- 239000012046 mixed solvent Substances 0.000 claims description 23
- 239000003242 anti bacterial agent Substances 0.000 claims description 16
- 238000010898 silica gel chromatography Methods 0.000 claims description 15
- 238000010828 elution Methods 0.000 claims description 14
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 12
- 150000003839 salts Chemical class 0.000 claims description 11
- 229960001701 chloroform Drugs 0.000 claims description 8
- 238000000034 method Methods 0.000 claims description 8
- 241000894006 Bacteria Species 0.000 claims description 7
- 241001655175 Sorbus pohuashanensis Species 0.000 claims description 7
- 235000012072 hua qui shu Nutrition 0.000 claims description 7
- XELZGAJCZANUQH-UHFFFAOYSA-N methyl 1-acetylthieno[3,2-c]pyrazole-5-carboxylate Chemical compound CC(=O)N1N=CC2=C1C=C(C(=O)OC)S2 XELZGAJCZANUQH-UHFFFAOYSA-N 0.000 claims description 7
- 238000002791 soaking Methods 0.000 claims description 7
- 238000004440 column chromatography Methods 0.000 claims description 6
- 238000000926 separation method Methods 0.000 claims description 5
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 4
- 230000001717 pathogenic effect Effects 0.000 claims description 3
- 239000002552 dosage form Substances 0.000 claims description 2
- 208000015181 infectious disease Diseases 0.000 claims description 2
- 239000000375 suspending agent Substances 0.000 claims description 2
- 238000011282 treatment Methods 0.000 claims description 2
- 239000004562 water dispersible granule Substances 0.000 claims description 2
- 239000004563 wettable powder Substances 0.000 claims description 2
- 239000004599 antimicrobial Substances 0.000 claims 2
- 238000011321 prophylaxis Methods 0.000 claims 1
- 241000223602 Alternaria alternata Species 0.000 abstract description 10
- 230000002401 inhibitory effect Effects 0.000 abstract description 8
- 240000004808 Saccharomyces cerevisiae Species 0.000 abstract description 5
- 239000004480 active ingredient Substances 0.000 abstract description 3
- 230000009286 beneficial effect Effects 0.000 abstract description 3
- 241001092391 Sorbus Species 0.000 abstract description 2
- 239000000575 pesticide Substances 0.000 abstract description 2
- 235000008345 mountainash Nutrition 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 24
- 241000196324 Embryophyta Species 0.000 description 13
- 239000003814 drug Substances 0.000 description 13
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 12
- 229940079593 drug Drugs 0.000 description 12
- 235000004489 Sorbus commixta Nutrition 0.000 description 11
- 241001115347 Sorbus commixta Species 0.000 description 11
- 239000007788 liquid Substances 0.000 description 10
- 235000002637 Nicotiana tabacum Nutrition 0.000 description 9
- 241000208125 Nicotiana Species 0.000 description 8
- 230000001580 bacterial effect Effects 0.000 description 8
- YZBQHRLRFGPBSL-RXMQYKEDSA-N carbapenem Chemical compound C1C=CN2C(=O)C[C@H]21 YZBQHRLRFGPBSL-RXMQYKEDSA-N 0.000 description 8
- 238000012360 testing method Methods 0.000 description 8
- 201000010099 disease Diseases 0.000 description 6
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 6
- 239000011259 mixed solution Substances 0.000 description 6
- 241000223600 Alternaria Species 0.000 description 5
- 241000323752 Alternaria longipes Species 0.000 description 5
- 241000191967 Staphylococcus aureus Species 0.000 description 5
- 238000004458 analytical method Methods 0.000 description 5
- 230000000844 anti-bacterial effect Effects 0.000 description 5
- 241000233866 Fungi Species 0.000 description 4
- RJQXTJLFIWVMTO-TYNCELHUSA-N Methicillin Chemical compound COC1=CC=CC(OC)=C1C(=O)N[C@@H]1C(=O)N2[C@@H](C(O)=O)C(C)(C)S[C@@H]21 RJQXTJLFIWVMTO-TYNCELHUSA-N 0.000 description 4
- 241000589517 Pseudomonas aeruginosa Species 0.000 description 4
- MYSWGUAQZAJSOK-UHFFFAOYSA-N ciprofloxacin Chemical compound C12=CC(N3CCNCC3)=C(F)C=C2C(=O)C(C(=O)O)=CN1C1CC1 MYSWGUAQZAJSOK-UHFFFAOYSA-N 0.000 description 4
- 238000012258 culturing Methods 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 4
- 229960003085 meticillin Drugs 0.000 description 4
- 239000013641 positive control Substances 0.000 description 4
- NWBWCXBPKTTZNQ-UHFFFAOYSA-N (16S)-4-(N-Acetyl-anthraniloyloxy)-20-aethyl-1alpha,14alpha,16-trimethoxy-aconitan-8,9-diol Natural products C1CC(OC)C2(C3C4)C5CC(C(C6)OC)C(OC)C5(O)C6(O)C4C2N(CC)CC31OC(=O)C1=CC=CC=C1NC(C)=O NWBWCXBPKTTZNQ-UHFFFAOYSA-N 0.000 description 3
- 241000588626 Acinetobacter baumannii Species 0.000 description 3
- 241000194031 Enterococcus faecium Species 0.000 description 3
- 238000005481 NMR spectroscopy Methods 0.000 description 3
- 241000220324 Pyrus Species 0.000 description 3
- 244000061456 Solanum tuberosum Species 0.000 description 3
- 235000002595 Solanum tuberosum Nutrition 0.000 description 3
- 241000191963 Staphylococcus epidermidis Species 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 235000021017 pears Nutrition 0.000 description 3
- 239000000843 powder Substances 0.000 description 3
- 230000002829 reductive effect Effects 0.000 description 3
- NWBWCXBPKTTZNQ-QOQRDJBUSA-N y4m5974f7z Chemical compound O([C@]12CN([C@@H]3[C@H]4[C@]5(O)[C@@]6(O)[C@@H](OC)[C@@H]([C@H](C5)OC)C[C@H]6[C@@]3([C@@H]1C4)[C@@H](OC)CC2)CC)C(=O)C1=CC=CC=C1N=C(C)O NWBWCXBPKTTZNQ-QOQRDJBUSA-N 0.000 description 3
- XMAYWYJOQHXEEK-OZXSUGGESA-N (2R,4S)-ketoconazole Chemical compound C1CN(C(=O)C)CCN1C(C=C1)=CC=C1OC[C@@H]1O[C@@](CN2C=NC=C2)(C=2C(=CC(Cl)=CC=2)Cl)OC1 XMAYWYJOQHXEEK-OZXSUGGESA-N 0.000 description 2
- 241000412366 Alternaria mali Species 0.000 description 2
- 241000194032 Enterococcus faecalis Species 0.000 description 2
- 241000223218 Fusarium Species 0.000 description 2
- IHPVFYLOGNNZLA-UHFFFAOYSA-N Phytoalexin Natural products COC1=CC=CC=C1C1OC(C=C2C(OCO2)=C2OC)=C2C(=O)C1 IHPVFYLOGNNZLA-UHFFFAOYSA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 241001123668 Verticillium dahliae Species 0.000 description 2
- 230000000843 anti-fungal effect Effects 0.000 description 2
- 235000010290 biphenyl Nutrition 0.000 description 2
- 239000004305 biphenyl Substances 0.000 description 2
- -1 biphenyl phytoalexin Chemical class 0.000 description 2
- 238000009835 boiling Methods 0.000 description 2
- 238000004113 cell culture Methods 0.000 description 2
- 229960003405 ciprofloxacin Drugs 0.000 description 2
- 230000003247 decreasing effect Effects 0.000 description 2
- 238000003113 dilution method Methods 0.000 description 2
- 239000012153 distilled water Substances 0.000 description 2
- 229940032049 enterococcus faecalis Drugs 0.000 description 2
- 244000000004 fungal plant pathogen Species 0.000 description 2
- 229960004125 ketoconazole Drugs 0.000 description 2
- 238000001819 mass spectrum Methods 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 244000052769 pathogen Species 0.000 description 2
- ZUOUZKKEUPVFJK-UHFFFAOYSA-N phenylbenzene Natural products C1=CC=CC=C1C1=CC=CC=C1 ZUOUZKKEUPVFJK-UHFFFAOYSA-N 0.000 description 2
- 239000000280 phytoalexin Substances 0.000 description 2
- 229930000044 secondary metabolite Natural products 0.000 description 2
- 239000011734 sodium Substances 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000012546 transfer Methods 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- PYIXHKGTJKCVBJ-UHFFFAOYSA-N Astraciceran Natural products C1OC2=CC(O)=CC=C2CC1C1=CC(OCO2)=C2C=C1OC PYIXHKGTJKCVBJ-UHFFFAOYSA-N 0.000 description 1
- 241000221377 Auricularia Species 0.000 description 1
- NDVRQFZUJRMKKP-UHFFFAOYSA-N Betavulgarin Natural products O=C1C=2C(OC)=C3OCOC3=CC=2OC=C1C1=CC=CC=C1O NDVRQFZUJRMKKP-UHFFFAOYSA-N 0.000 description 1
- 241000588694 Erwinia amylovora Species 0.000 description 1
- 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 description 1
- 102000001399 Kallikrein Human genes 0.000 description 1
- 108060005987 Kallikrein Proteins 0.000 description 1
- 241000202564 Kalopanax Species 0.000 description 1
- 235000007688 Lycopersicon esculentum Nutrition 0.000 description 1
- 244000141359 Malus pumila Species 0.000 description 1
- 241000221960 Neurospora Species 0.000 description 1
- 244000061176 Nicotiana tabacum Species 0.000 description 1
- 241001329956 Nothopassalora personata Species 0.000 description 1
- 240000003768 Solanum lycopersicum Species 0.000 description 1
- 235000014459 Sorbus Nutrition 0.000 description 1
- 244000019194 Sorbus aucuparia Species 0.000 description 1
- 235000009790 Sorbus aucuparia Nutrition 0.000 description 1
- 241000191940 Staphylococcus Species 0.000 description 1
- 235000021307 Triticum Nutrition 0.000 description 1
- 244000098338 Triticum aestivum Species 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 230000001775 anti-pathogenic effect Effects 0.000 description 1
- 229940121375 antifungal agent Drugs 0.000 description 1
- 235000021016 apples Nutrition 0.000 description 1
- 239000012752 auxiliary agent Substances 0.000 description 1
- 230000003385 bacteriostatic effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 1
- 229960005091 chloramphenicol Drugs 0.000 description 1
- 238000013375 chromatographic separation Methods 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 230000008260 defense mechanism Effects 0.000 description 1
- 230000012969 defense response to bacterium Effects 0.000 description 1
- 150000004826 dibenzofurans Chemical class 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 238000002330 electrospray ionisation mass spectrometry Methods 0.000 description 1
- 239000002702 enteric coating Substances 0.000 description 1
- 238000009505 enteric coating Methods 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 244000053095 fungal pathogen Species 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 238000002114 high-resolution electrospray ionisation mass spectrometry Methods 0.000 description 1
- 238000005470 impregnation Methods 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 239000000411 inducer Substances 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 229930014626 natural product Natural products 0.000 description 1
- 229930000223 plant secondary metabolite Natural products 0.000 description 1
- 235000012015 potatoes Nutrition 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 238000004611 spectroscopical analysis Methods 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 239000003104 tissue culture media Substances 0.000 description 1
- 239000012137 tryptone Substances 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D513/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00
- C07D513/02—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00 in which the condensed system contains two hetero rings
- C07D513/04—Ortho-condensed systems
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N43/00—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds
- A01N43/90—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having two or more relevant hetero rings, condensed among themselves or with a common carbocyclic ring system
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Agronomy & Crop Science (AREA)
- Pest Control & Pesticides (AREA)
- Plant Pathology (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Dentistry (AREA)
- General Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Environmental Sciences (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses sulfur-containing dibenzofuran type alkaloid and a preparation method and application thereof, belonging to the technical field of pesticide preparation. The structural formula of the compound provided by the invention is
Description
Technical Field
The invention relates to the technical field of pesticide preparation, in particular to sulfur-containing dibenzofuran type alkaloid and a preparation method and application thereof.
Background
The alternaria alternate is a main leaf disease of tobacco (Nicotiana tabacum) plants in the later growth stage and occurs in all tobacco production areas in the world. According to the estimation report of the literature, the disease area of the tobacco brown spot in China accounts for 30-50% of the tobacco planting area in China every year, and the disease is a tobacco leaf disease which has the widest occurrence range and the greatest harm in the tobacco planting industry in China. Alternaria alternata is caused by the impregnation of Alternaria (Alternaria) fungi, the main pathogenic bacteria of which are Alternaria (a. Alternata) and Alternaria longissima (a. Longipes). Meanwhile, alternaria fungus is an important plant pathogenic fungus widely distributed in the global scope, and the fungus has more than about 40 species, wherein more than 95 percent of the species can parasitize on plants in a facultative way, so that plant diseases of more than ten important crops, economic crops and fruit trees including wheat, tobacco, potatoes, tomatoes, apples, oranges and pears can be caused. The American quarantine department in 2003 discovers that the pears imported from China are infected by alternaria fungus, so that the pears imported from China are suspended indefinitely, and huge economic loss is caused. Therefore, the development of the high-efficiency low-toxicity natural environment-friendly antibacterial agent has important significance on the production of economic crops and the safety of food.
The plant is often influenced by adverse factors such as physics, chemistry, biology and the like in the environment in the growth process, and can cause the change of plant secondary metabolites and start an in vivo defense mechanism, thereby reducing or avoiding the damage of the plant, leading the plant to present certain stress resistance, and simultaneously causing the increase of the accumulation of active secondary metabolites in the plant. For example, after the European mountain ash (Sorbus auricularia) is infected with the epidemic disease, biphenyl phytoalexin can be generated around the focus to inhibit the further proliferation of the Erwinia amylovora. Recently, we found that the de novo synthesis of biphenyl phytoalexins is induced by stimulating Sorbus Pohuashanensis Suspension Cells (SPSC) for a certain period of time using yeast extract as a biological inducer. Therefore, the plant suspension cells are cultured by a biotechnology means, an antibacterial defense mechanism of the plant cells is started by adding the fungus extract, the plant cells are induced to synthesize secondary metabolites with antibacterial activity, more antibacterial substances with novel structure, remarkable activity, safety and effectiveness are found, and the method has important significance for development of natural antibacterial agents.
Disclosure of Invention
The compound provided by the invention has a novel structure and has an obvious inhibiting effect on Alternaria longissima (Alternaria longissima) which is a pathogenic bacterium of Alternaria alternata of tobacco Alternaria alternata.
The invention firstly provides a compound shown as a formula I or an agriculturally and pharmaceutically acceptable salt thereof;
the invention also provides a preparation method of the compound shown in the formula I, which comprises the following steps:
s1: soaking and extracting the dried Sorbus pohuashanensis suspension cells with methanol or ethanol, and concentrating the extract to obtain an extract;
s2: performing silica gel column chromatography gradient elution on the extract obtained in the step S1 by using a dichloromethane/methanol mixed solvent or a trichloromethane/methanol mixed solvent to obtain 9 components, and collecting the 4 th component;
s3: performing silica gel column chromatography gradient elution on the collected component of S2 by using a dichloromethane/methanol mixed solvent or a trichloromethane/methanol mixed solvent, and collecting the 4 th component;
s4: and (3) carrying out Sephadex LH-20 column chromatography separation on the 4 th component obtained from the S3 by using a dichloromethane/methanol mixed solvent to obtain the compound shown in the formula I.
In the above preparation method, in S2, the silica gel column chromatography gradient elution is performed by sequentially performing silica gel column chromatography gradient elution with a dichloromethane/methanol mixed solvent or a chloroform/methanol mixed solvent at a volume ratio of 200, 99.5;
in S3, the silica gel column chromatography gradient elution is silica gel column chromatography gradient elution sequentially by using a dichloromethane/methanol mixed solvent or a chloroform/methanol mixed solvent with a volume ratio of 100;
in S4, the volume ratio of the dichloromethane/methanol mixed solvent is 1:1.
In the preparation method, in the step S1, the temperature for soaking and extracting is room temperature; the times of soaking and extracting are 1 to 3 times;
in S3, the number of gradient elution of the silica gel column chromatography is 1;
in S4, the chromatographic separation times of the Sephadex LH-20 column is 2 times; the Sephadex LH-20 column chromatography separation also comprises a step of concentrating the chromatographic solution.
The room temperature is well known to those skilled in the art and is generally from 15 to 40 ℃.
In the above preparation method, the concentration is a 40 ℃ reduced pressure concentration.
The use of the compounds of formula I or their agriculturally acceptable salts for the preparation of antibacterial agents also falls within the scope of the present invention.
Specifically, the bacteria controlled by the antibacterial agent are Alternaria longissima longipes.
The application of the compound shown in the formula I or the agriculturally and pharmaceutically acceptable salt thereof in preventing and treating pathogenic bacteria infection also belongs to the protection scope of the invention.
Specifically, the pathogenic bacteria are Alternaria longissima longipes.
The invention further provides an antibacterial agent which comprises the compound shown in the formula I or the agriculturally and pharmaceutically acceptable salt thereof.
The antibacterial agent also comprises an agriculturally and pharmaceutically acceptable auxiliary agent.
The dosage form of the antibacterial agent is wettable powder, water dispersible granules, a suspending agent or missible oil.
In the antibacterial agent, the content of the compound shown in the formula I or the agriculturally and pharmaceutically acceptable salt thereof is 1-99 wt%; specifically, the content of the compound shown in the formula I or the agriculturally and pharmaceutically acceptable salt thereof is 10-90 wt%.
The bacteria for preventing and treating the antibacterial agent are Alternaria longissima longipes.
Experiments show that the compound shown in the formula I has a strong inhibiting effect on the growth of Alternaria alternate (Alternaria longipes) which is a pathogenic bacterium of Alternaria alternata.
The invention has the following beneficial effects:
(1) The invention provides a method for preparing sulfur-containing dibenzofuran type alkaloid Sorbus commixta A from yeast-induced Sorbus commixta suspension cells, and the prepared compound has a novel structure and has an obvious inhibiting effect on Alternaria longissima (Alternaria longipes) which is a pathogen of Alternaria alternata.
(2) The traditional method of extracting active ingredients from plant planting bodies is generally affected by the disadvantages of long growth cycle of plants, unstable yield and quality, and the like. The invention uses the yeast-induced plant suspension cell culture technology to produce and extract active ingredients, has the characteristics of high production speed, easily controlled growth conditions and stable yield and quality, and is beneficial to industrial operation; moreover, the yield can be continuously improved by optimizing the cell culture conditions.
Detailed Description
The present invention is described in further detail below with reference to specific embodiments, which are given for the purpose of illustration only and are not intended to limit the scope of the invention.
The experimental procedures in the following examples are conventional unless otherwise specified.
Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
The method for preparing the suspension cells of the dried Sorbus commixta used in the following examples is identical to the method for preparing suspension cells of the dried Sorbus commixta in the following documents, except that the Sorbus commixta is replaced by Sorbus commixta (Li Jiaxing, li Huiliang, zhou Liangyun, etc. [ J ] Yeast induces chemical components and bacteriostatic activity of Sorbus commixta suspension cells [ J ] Natural products research and development, 2019,031 (012): 2071-2076 ].
The silica gel column chromatography filler is 200-300 meshes.
Methicillin-resistant Staphylococcus aureus (methicillin-resistant Staphylococcus aureus), carbapenem-resistant Pseudomonas aeruginosa (carbapenem-resistant Pseudomonas aeruginosa), carbapenem-resistant Acinetobacter baumannii (carbapenem-resistant Enterococcus faecalis), multi-drug-resistant Enterococcus faecium (multi-drug-resistant Enterococcus faecalis) and multi-drug-resistant Staphylococcus epidermidis (multi-drug-resistant Staphylococcus epidermidis) in the following examples are described in the literature (±) -preissoide: a new enteric resistant a raw enteric resistant Staphylococcus aureus-2H-3236 zx3236-oxcarbazn enteric coating tissue culture medium, which is obtained from university of military medical science 3763, or other medical science 3763-resistant Staphylococcus aureus (which is a Chinese medicine university), namely, a research university 3763, a medical university 3763, a research university of military science 3762, or a research university of traditional Chinese medicine 3763.
Example 1 preparation of Sorbus commixta
1. Preparation method
S1: soaking and extracting 20kg of dried Sorbus commixta suspension cells with methanol at room temperature for 3 times, mixing extractive solutions, and concentrating under reduced pressure at 40 deg.C to obtain 6kg of extract;
s2: performing silica gel column chromatography gradient elution on the extractum obtained in the step S1 by using a dichloromethane/methanol mixed solvent with the volume ratio of 200, 99.5, 99.1, 98;
s3: sequentially carrying out silica gel column chromatography gradient elution on the collected components of the S2 by using a dichloromethane/methanol mixed solvent with the volume ratio of 100, 99 and 98, wherein the total time is 1 time, and collecting the 4 th component;
s4: subjecting the 4 th component obtained by S3 treatment to Sephadex LH-20 column chromatography with dichloromethane/methanol mixed solvent of 1:1 for 2 times, and concentrating the chromatography solution under reduced pressure at 40 deg.C to obtain 5mg of lappaconitine.
2. Analysis of results
The obtained kalopanax pohuashanjian white powder finished product is subjected to mass spectrum, ultraviolet and infrared analysis, and the analysis result is as follows:
sorbus pohuashanensis A molecular formula C 14 H 9 NO 3 S；ESIMS(pos.)：m/z 294[M+Na] + ，565[2M+Na] + ；HRESIMS(neg.)：m/z 270.0231[M-1] - (270.0230calcd for C 14 H 8 NO 3 S)；UV(MeOH)λ max ：230(sh)，305，330(sh)nm；IR(KBr)ν max ：3235，2956，2922，2852，1735，1623，1587，1494，1454，1385，1313，1251，1212，1029，969，939cm -1 .
The nuclear magnetic resonance analysis of the lappaconitine was performed, and the results are shown in table 1.
TABLE 1 NMR spectroscopic data for Sorbus pohuashanensis A (delta in ppm, J in Hz)
According to the results of the mass spectrum, ultraviolet, infrared and nuclear magnetic resonance analysis, the kallikrein A prepared by the invention has the following structure:
example 2
1. In order to examine the anti-pathogenic fungus effect of the prepared compound, five pathogenic fungi of potato Verticillium dahliae (Verticillium dahliae) ATCC 42216, alternaria mali (Alternaria mali) ATCC 6663, gibberella saubenii (Gibberella saubenecii) ATCC 20193, alternaria longissima (Alternaria longipes) ATCC 26293 and Neurospora brassicae (Cercospora personata) ATCC 18592 are selected, and the antibacterial activity of the kalopanaemonium majus is determined by a double dilution method.
Compound solution preparation: dissolving the kalopanain prepared in example 1 and the ketoconazole serving as a positive control in DMSO to prepare a solution of 10 mg/mL;
preparing a PDB culture solution: adding 1000mL of distilled water into 25g of PDB (potato extract powder 5.0g/L, glucose 20.0g/L, chloramphenicol 0.1g/L, shanghai Bo microbial science and technology Co., ltd.), boiling for dissolving, and sterilizing at 121 deg.C for 20 min;
preparing test bacterial liquid: inoculating plant pathogenic fungi stored in a 4 ℃ inclined plane into PDB culture solution, and culturing at 28 ℃ and 160r/min for 3-4d;
the experimental method comprises the following steps: the test cell liquid was diluted 100 times with the PDB culture solution. In the 96-well plate, 198. Mu.L of diluted test bacterial solution was added to each well of the first row. Adding 100 mu L of diluted test bacterial liquid into each hole of the other rows; adding 2 mu L of compound solution into the first row, sucking 100 mu L of mixed solution to the second row after the compound solution is uniformly sucked and mixed by a liquid transfer gun, sucking 100 mu L of mixed solution to the third row after the compound solution is uniformly mixed, operating the last rows in the same way, and discarding the mixed solution sucked from the last row; the concentration of the compounds in each row is 0.1mg/mL,0.05mg/mL,0.025mg/mL and 0.0125mg/mL in turn, and is decreased progressively. One positive and negative control per plate (2. Mu.L of ketoconazole solution and 2. Mu.L of DMSO solution, respectively).
And (5) judging a result: culturing at 28 deg.C, and observing after 72 hr. The minimal compound concentration at which no fungal growth was observed was the Minimal Inhibitory Concentration (MIC), and the results are shown in table 2.
TABLE 2 antifungal Activity of Sorbus pohuashanensis A (MIC, μ g/mL)
As can be seen from Table 2, sorbus pohuashanensis A has a significant inhibitory activity against Alternaria longissima (Alternaria longipes), which is a pathogen of Alternaria alternata.
2. In order to examine the effect of the prepared compound on drug-resistant pathogenic bacteria, the invention selects 5 drug-resistant bacteria (provided by army medical university) of methicillin-resistant Staphylococcus aureus (methicillin-resistant Staphylococcus aureus), carbapenem-resistant Pseudomonas aeruginosa (carbapenem-resistant Pseudomonas aeruginosa), carbapenem-resistant Acinetobacter baumannii (carbapenem-resistant Acinetobacter baumannii), multi-drug-resistant Enterococcus faecium (multi-drug-resistant Enterococcus faecium) and multi-drug-resistant Staphylococcus epidermidis (provided by army medical university), and the antibacterial activity of the compound anthocarcinonide is determined by a double dilution method.
Compound solution preparation: the kalopanain compound prepared in example 1 and ciprofloxacin as a positive control were dissolved in DMSO and prepared into a solution of 10 mg/mL;
preparing an LB culture solution: taking 25g of LB (the components are tryptone 10.0g/L, yeast powder 5.0g/L, sodium chloride 10.0g/L, shanghai Bo microbial science and technology Co., ltd.), adding 1000mL of distilled water, heating, boiling, dissolving, and sterilizing at 121 ℃ for 15 minutes for later use;
preparing test bacterial liquid: inoculating the drug-resistant bacteria to LB culture solution from a freezing tube, and culturing at the constant temperature of 37 ℃ and 160r/min for 10h until the culture solution becomes turbid.
The experimental method comprises the following steps: diluting the test bacterial liquid by 1000 times with LB culture liquid; in a 96-well plate, 198. Mu.L of diluted test bacterial liquid was added to each well of the first row. Adding 100 mu L of diluted test bacterial liquid into each hole of the other rows; adding 2 mu L of compound solution into the first row, sucking 100 mu L of mixed solution to the second row after the compound solution is uniformly sucked and mixed by a liquid transfer gun, sucking 100 mu L of mixed solution to the third row after the compound solution is uniformly mixed, operating the last row, and discarding the mixed solution sucked out; the concentration of the compounds in each row is 0.1mg/mL,0.05mg/mL,0.025mg/mL and 0.0125mg/mL in turn, and is decreased progressively. One positive and negative control per plate (2. Mu.L ciprofloxacin solution and 2. Mu.L DMSO solution, respectively).
And (5) judging a result: culturing at 37 deg.C, and observing after 12 h. The minimal compound concentration at which no bacterial growth was observed was the Minimal Inhibitory Concentration (MIC), and the results are shown in table 3.
TABLE 3 antibacterial Activity of Sorbus commixta (MIC, μ g/mL)
As can be seen from Table 3, the lappaconitine has no significant inhibitory activity against 5 kinds of drug-resistant bacteria.
In conclusion, the results of the present example fully illustrate that the sulfur-containing dibenzofuran alkaloid Sorbus commixta A has potential application value for further development into antifungal drugs.
Claims (9)
1. A compound of formula I or an agriculturally acceptable salt thereof;
2. a process for the preparation of a compound of formula i as claimed in claim 1, comprising the steps of:
s1: soaking and extracting the dried Sorbus pohuashanensis suspension cells with methanol or ethanol, and concentrating the extract to obtain an extract;
s2: performing silica gel column chromatography gradient elution on the extract obtained in the step S1 by using a dichloromethane/methanol mixed solvent or a trichloromethane/methanol mixed solvent to obtain 9 components, and collecting the 4 th component;
s3: performing silica gel column chromatography gradient elution on the collected component of S2 by using a dichloromethane/methanol mixed solvent or a trichloromethane/methanol mixed solvent, and collecting the 4 th component;
s4: and (3) carrying out Sephadex LH-20 column chromatography separation on the 4 th component obtained from the S3 by using a dichloromethane/methanol mixed solvent to obtain the compound shown in the formula I.
3. The method of claim 2, wherein: in S2, the silica gel column chromatography gradient elution is carried out by sequentially using a dichloromethane/methanol mixed solvent or a chloroform/methanol mixed solvent at a volume ratio of 200, 99.5, 99.1, 98, 10, 80, 20, 70, 60, 40, 50;
in S3, the silica gel column chromatography gradient elution is carried out by sequentially using a dichloromethane/methanol mixed solvent or a chloroform/methanol mixed solvent with the volume ratio of 100, 99;
in S4, the volume ratio of the dichloromethane/methanol mixed solvent is 1:1.
4. The production method according to claim 2 or 3, characterized in that: in S1, the temperature for soaking and extracting is room temperature; the soaking and extracting times are 1 to 3;
in S3, the number of gradient elution of the silica gel column chromatography is 1;
in S4, the times of Sephadex LH-20 column chromatography separation are 2 times; the Sephadex LH-20 column chromatography separation also comprises a step of concentrating the chromatographic solution.
5. The use of a compound of formula i as claimed in claim 1 or an agriculturally acceptable salt thereof for the preparation of an antibacterial agent; or
The use of a compound of formula i as claimed in claim 1 or an agriculturally acceptable salt thereof for the prophylaxis or treatment of pathogenic infections;
specifically, the bacteria controlled by the antibacterial agent are Alternaria longissima longipes;
the pathogenic bacteria are Alternaria longissima longissipes.
6. An antimicrobial agent characterized by: comprising a compound of formula I as claimed in claim 1 or an agriculturally pharmaceutically acceptable salt thereof.
7. The antimicrobial agent of claim 6, wherein: the dosage form of the antibacterial agent is wettable powder, water dispersible granules, suspending agent or missible oil.
8. The antibacterial agent according to claim 6 or 7, characterized in that: the antibacterial agent contains the compound shown in the formula I or the agriculturally and pharmaceutically acceptable salt thereof according to claim 1 in an amount of 1-99 wt%.
9. The antibacterial agent according to any one of claims 6 to 8, characterized in that: the bacteria controlled by the antibacterial agent are Alternaria longissima longissipes.
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| CN103183669A (en) * | 2011-12-27 | 2013-07-03 | 湖南化工研究院 | Thiazole methylaminopyridine compounds and preparation method for same |
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| CN103183669A (en) * | 2011-12-27 | 2013-07-03 | 湖南化工研究院 | Thiazole methylaminopyridine compounds and preparation method for same |
| CN103342703A (en) * | 2013-07-22 | 2013-10-09 | 湖南大学 | N-[4-(benzofuran-5-yl)thiazole-2-yl] amide as well as preparation method and application for same |
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