CN113956212B - Thermolides compounds with nematodc resistance activity and preparation method and application thereof - Google Patents
Thermolides compounds with nematodc resistance activity and preparation method and application thereof Download PDFInfo
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- 150000001875 compounds Chemical class 0.000 title claims abstract description 77
- 230000000694 effects Effects 0.000 title claims abstract description 15
- 238000002360 preparation method Methods 0.000 title abstract description 16
- 239000000575 pesticide Substances 0.000 claims abstract description 10
- 201000010099 disease Diseases 0.000 claims description 7
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 7
- 241000244206 Nematoda Species 0.000 abstract description 23
- 241000351920 Aspergillus nidulans Species 0.000 abstract description 7
- 230000001069 nematicidal effect Effects 0.000 abstract description 7
- 230000015572 biosynthetic process Effects 0.000 abstract description 4
- 239000005645 nematicide Substances 0.000 abstract description 3
- 108091008053 gene clusters Proteins 0.000 abstract 1
- 239000003120 macrolide antibiotic agent Substances 0.000 abstract 1
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 18
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 18
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 15
- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical group O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 description 11
- 108090000623 proteins and genes Proteins 0.000 description 7
- 101001074427 Bacillus subtilis (strain 168) Polyketide synthase PksJ Proteins 0.000 description 6
- 101001074418 Bacillus subtilis (strain 168) Polyketide synthase PksL Proteins 0.000 description 6
- 101000979117 Curvularia clavata Nonribosomal peptide synthetase Proteins 0.000 description 6
- 101000803786 Emericella nidulans (strain FGSC A4 / ATCC 38163 / CBS 112.46 / NRRL 194 / M139) Conidial yellow pigment biosynthesis polyketide synthase Proteins 0.000 description 6
- 101000701349 Emericella nidulans (strain FGSC A4 / ATCC 38163 / CBS 112.46 / NRRL 194 / M139) Putative sterigmatocystin biosynthesis polyketide synthase Proteins 0.000 description 6
- 101000730455 Serratia sp. (strain ATCC 39006) Beta-ketoacyl synthase PigJ Proteins 0.000 description 6
- 101000691656 Streptomyces venezuelae Narbonolide/10-deoxymethynolide synthase PikA1, modules 1 and 2 Proteins 0.000 description 6
- 101000691655 Streptomyces venezuelae Narbonolide/10-deoxymethynolide synthase PikA2, modules 3 and 4 Proteins 0.000 description 6
- 101000691658 Streptomyces venezuelae Narbonolide/10-deoxymethynolide synthase PikA3, module 5 Proteins 0.000 description 6
- 101001125873 Streptomyces venezuelae Narbonolide/10-deoxymethynolide synthase PikA4, module 6 Proteins 0.000 description 6
- 230000001679 anti-nematodal effect Effects 0.000 description 6
- CSCPPACGZOOCGX-WFGJKAKNSA-N deuterated acetone Substances [2H]C([2H])([2H])C(=O)C([2H])([2H])[2H] CSCPPACGZOOCGX-WFGJKAKNSA-N 0.000 description 6
- 239000007791 liquid phase Substances 0.000 description 6
- 239000002904 solvent Substances 0.000 description 6
- HTSGKJQDMSTCGS-UHFFFAOYSA-N 1,4-bis(4-chlorophenyl)-2-(4-methylphenyl)sulfonylbutane-1,4-dione Chemical compound C1=CC(C)=CC=C1S(=O)(=O)C(C(=O)C=1C=CC(Cl)=CC=1)CC(=O)C1=CC=C(Cl)C=C1 HTSGKJQDMSTCGS-UHFFFAOYSA-N 0.000 description 5
- FKLJPTJMIBLJAV-UHFFFAOYSA-N Compound IV Chemical compound O1N=C(C)C=C1CCCCCCCOC1=CC=C(C=2OCCN=2)C=C1 FKLJPTJMIBLJAV-UHFFFAOYSA-N 0.000 description 5
- NLFBCYMMUAKCPC-KQQUZDAGSA-N ethyl (e)-3-[3-amino-2-cyano-1-[(e)-3-ethoxy-3-oxoprop-1-enyl]sulfanyl-3-oxoprop-1-enyl]sulfanylprop-2-enoate Chemical compound CCOC(=O)\C=C\SC(=C(C#N)C(N)=O)S\C=C\C(=O)OCC NLFBCYMMUAKCPC-KQQUZDAGSA-N 0.000 description 5
- 238000000855 fermentation Methods 0.000 description 5
- 230000004151 fermentation Effects 0.000 description 5
- 238000000926 separation method Methods 0.000 description 5
- 238000001269 time-of-flight mass spectrometry Methods 0.000 description 5
- FCSKOFQQCWLGMV-UHFFFAOYSA-N 5-{5-[2-chloro-4-(4,5-dihydro-1,3-oxazol-2-yl)phenoxy]pentyl}-3-methylisoxazole Chemical compound O1N=C(C)C=C1CCCCCOC1=CC=C(C=2OCCN=2)C=C1Cl FCSKOFQQCWLGMV-UHFFFAOYSA-N 0.000 description 4
- 241000196324 Embryophyta Species 0.000 description 4
- 241001136490 Thermomyces dupontii Species 0.000 description 4
- 108020002494 acetyltransferase Proteins 0.000 description 4
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- 238000000034 method Methods 0.000 description 4
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- 101000869050 Homo sapiens Caveolae-associated protein 2 Proteins 0.000 description 3
- 150000001413 amino acids Chemical class 0.000 description 3
- 238000004587 chromatography analysis Methods 0.000 description 3
- 238000004440 column chromatography Methods 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 241000228431 Acremonium chrysogenum Species 0.000 description 2
- 241000588724 Escherichia coli Species 0.000 description 2
- 241000233866 Fungi Species 0.000 description 2
- 238000003820 Medium-pressure liquid chromatography Methods 0.000 description 2
- PBCJIPOGFJYBJE-UHFFFAOYSA-N acetonitrile;hydrate Chemical compound O.CC#N PBCJIPOGFJYBJE-UHFFFAOYSA-N 0.000 description 2
- 239000012043 crude product Substances 0.000 description 2
- 238000010348 incorporation Methods 0.000 description 2
- 125000001449 isopropyl group Chemical group [H]C([H])([H])C([H])(*)C([H])([H])[H] 0.000 description 2
- 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 description 2
- 239000000203 mixture Substances 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 125000000094 2-phenylethyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])C([H])([H])* 0.000 description 1
- UHPMCKVQTMMPCG-UHFFFAOYSA-N 5,8-dihydroxy-2-methoxy-6-methyl-7-(2-oxopropyl)naphthalene-1,4-dione Chemical compound CC1=C(CC(C)=O)C(O)=C2C(=O)C(OC)=CC(=O)C2=C1O UHPMCKVQTMMPCG-UHFFFAOYSA-N 0.000 description 1
- 239000005660 Abamectin Substances 0.000 description 1
- 108700016155 Acyl transferases Proteins 0.000 description 1
- 102000057234 Acyl transferases Human genes 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 241000243770 Bursaphelenchus Species 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 241000223218 Fusarium Species 0.000 description 1
- 241000243786 Meloidogyne incognita Species 0.000 description 1
- 208000031888 Mycoses Diseases 0.000 description 1
- 241000228341 Talaromyces Species 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 125000000218 acetic acid group Chemical group C(C)(=O)* 0.000 description 1
- 230000010933 acylation Effects 0.000 description 1
- 238000005917 acylation reaction Methods 0.000 description 1
- 238000012271 agricultural production Methods 0.000 description 1
- RRZXIRBKKLTSOM-XPNPUAGNSA-N avermectin B1a Chemical compound C1=C[C@H](C)[C@@H]([C@@H](C)CC)O[C@]11O[C@H](C\C=C(C)\[C@@H](O[C@@H]2O[C@@H](C)[C@H](O[C@@H]3O[C@@H](C)[C@H](O)[C@@H](OC)C3)[C@@H](OC)C2)[C@@H](C)\C=C\C=C/2[C@]3([C@H](C(=O)O4)C=C(C)[C@@H](O)[C@H]3OC\2)O)C[C@H]4C1 RRZXIRBKKLTSOM-XPNPUAGNSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 125000004063 butyryl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 229940125904 compound 1 Drugs 0.000 description 1
- 238000009795 derivation Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000013505 freshwater Substances 0.000 description 1
- 150000007931 macrolactones Chemical class 0.000 description 1
- 238000012269 metabolic engineering Methods 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 238000005065 mining Methods 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 230000001717 pathogenic effect Effects 0.000 description 1
- 150000002994 phenylalanines Chemical class 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000013535 sea water Substances 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D273/00—Heterocyclic compounds containing rings having nitrogen and oxygen atoms as the only ring hetero atoms, not provided for by groups C07D261/00 - C07D271/00
- C07D273/01—Heterocyclic compounds containing rings having nitrogen and oxygen atoms as the only ring hetero atoms, not provided for by groups C07D261/00 - C07D271/00 having one nitrogen atom
-
- 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
- A01N43/72—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with nitrogen atoms and oxygen or sulfur atoms as ring hetero atoms
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
- C07B2200/00—Indexing scheme relating to specific properties of organic compounds
- C07B2200/07—Optical isomers
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- 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)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention discloses thermolides compounds with nematocide resisting activity, a preparation method and application thereof, wherein the general formula of the compounds is shown as follows, the compounds are obtained by heterologous expression of a biosynthesis gene cluster of a macrolide compound thermolides in aspergillus nidulans, and the prepared compounds have better nematocide resisting activity, can be used for preparing biological pesticides, and have important significance for preventing and treating nematodes.
Description
Technical Field
The invention relates to the field of biological pesticides, in particular to thermolides compounds with nematicide activity, and also relates to a preparation method and application of the compounds.
Background
Nematodes are invertebrates widely distributed in fresh water, seawater, soil and other environments, 23000 kinds of nematodes are found, more than 100 tens of thousands of kinds of nematodes are reported, and plant parasitic nematodes occupy about 10% of the nematodes. Plant parasitic nematodes are an important pathogen, can cause serious harm to crops, trees, edible fungi and the like, and the annual agricultural economic loss caused by the plant parasitic nematodes accounts for 12.3% of the annual loss rate of main economic crops in the world, which exceeds 1000 hundred million dollars. At present, diseases caused by nematodes are more than bacteria and viruses, become the second largest diseases in the world, and are second only to fungal diseases. The method is suitable for the growth and propagation of nematodes in temperate and subtropical climates in China, so that the method is also deeply damaged by the nematodes in China, and worldwide important nematode diseases occur in China. The nematode damage is serious, and the yield of certain crops can be reduced by 10-40% when the nematode damage is common. Can lead to 70-80% yield reduction and even failure in serious diseases. 17 provinces, cities and autonomous regions in China are damaged by pine wood nematodes, so that the national economy in China is affected, and the natural landscape and ecological environment are destroyed. The plant parasitic nematodes have serious harm at home and abroad and seriously affect the increase of economy, so that the effective prevention and treatment of nematode diseases is a serious problem to be solved urgently.
Biological pesticides have become a new trend in recent years for their natural nuisance-free, efficient and environmentally friendly reasons. 2012. The Niu Xuemei subject group was isolated from the thermophilic fungus Talaromyces thermophil μs YM3-4 over the years to thermolides class of compounds. Wherein thermolideA and B have better nematoda resisting activity, the nematoda resisting capability is similar to that of avermectin, and the nematoda resisting capability shows better resistance to nematodes such as M.incognita, xylophil mu s5, P.rediviv mu s and the like. However, the strain has not been used well due to its low yield in the original host and severe culture conditions of the original strain. Zhang Jinmei et cl. discloses Heterologous and engineered biosynthesis of nematocidal polyketide-nonribosomal peptide hybrid macrolactone from extreme thermophilic fungi that high efficiency synthesis of thermolides is achieved by a model microorganism. However, whether the synthesized thermolides compound has biological activity has not been reported.
Disclosure of Invention
Accordingly, it is an object of the present invention to provide thermolides compounds having anti-nematode activity; the second object of the present invention is a pesticide containing the thermolides-class compound having an anti-nematode activity; it is a further object of the present invention to provide a pesticide comprising the thermolides-type compound having an anti-nematode activity.
In order to achieve the above purpose, the present invention provides the following technical solutions:
1. thermolides compounds having anti-nematode activity, said compounds having the general formula:
Wherein r1=acetyl or butyryl, r2=isopropyl or phenethyl, r3=methyl or isopropyl.
Preferably, the compound has the structural formula:
2. The thermolides compounds with nematodc activity are applied to the preparation of pesticides for preventing and treating nematodc diseases.
3. Pesticides containing the thermolides compounds with nematicidal activity.
Preferably, the effective concentration of thermolides compounds in the pesticide is more than 25 mug/ml.
The invention has the beneficial effects that: the thermolides compounds with nematodc resistance are screened, and a thermolides product of L-type amino acid is obtained through marfey derivation, which is different from the D-type amino acid product reported in the prior literature; the thermolides analogues of phenylalanine incorporation and unnatural amino acid incorporation are obtained by genome mining and combined biosynthesis; through metabolic engineering, thermolides derivatives with different acylation modifications are obtained; the anti-nematode activity of the obtained compound is higher than that of the original compound, and more efficient and safe candidate medicines are provided for preventing and treating nematodes in the later agricultural production.
Detailed Description
The present invention will be further described with reference to specific examples, which are not intended to be limiting, so that those skilled in the art will better understand the present invention and practice it.
Example 1 preparation of Compounds
Preparation of Compounds II and VII: genes ThmABE (PKS, NRPS and acetyltransferase) from Talaromyces thermophilus NRRL 2155 were expressed heterologously in aspergillus nidulans and the heterologously expressed strain was mass fermented and after fermentation was completed, acetone was used: ethyl acetate (1:3) was repeatedly extracted three times and concentrated to dryness under reduced pressure. Crude separation is carried out by MCI column chromatography, the solvent is methanol-water, the elution program is 40%,50%,60%,70%,80%,90%,100% methanol, two column volumes are eluted at each concentration, and the components containing the compound II and the components containing the compound VII are obtained by combining. Subsequently, the mixture was separated again by Sephadex LH-20 (200 mm. Times.20 mm) with methanol as a solvent. The crude product obtained was again subjected to separation in a preparative liquid phase (column type: YMC ODS-A5 μm 120A (10X 250 mm)), and isocratic elution with 73% methanol water gave 2mg of pure compound II, 14mg of pure compound VII. Deuterated acetone was dissolved and nuclear magnetic identification was performed, and the assignment of the compounds is shown in tables 1 and 2.
Identification of Compound II: colorless oily form; the calculated value of HR TOFMS m/z 566.3299[ M+Na ] +,C28H49NO9 in positive ion mode is 566.3300, indicating that the molecule of the compound is C 28H49NO9, and the structure of the compound is determined by combining the nuclear magnetic data in Table 1, as follows:
TABLE 1 Nuclear magnetic data assignment for Compound II
Identification of Compound VII: colorless oily form; calculated value of HR TOFMS m/z 594.3613[ M+Na ] +,C30H53NO9 under positive ion mode is 594.3613, which shows that the molecular formula of the compound is C 30H53NO9, and the structure of the compound is determined by combining the nuclear magnetic data in Table 2:
TABLE 2 Nuclear magnetic data assignment for Compound VII
Preparation of compound III: the method comprises the steps of (1) carrying out heterologous expression of a gene AcrAB (PKS, NRPS) from Acremonium chrysogenum in Aspergillus nidulans, carrying out mass fermentation to obtain a compound X, feeding the compound X (22 mg) into a ThmE (acetyltransferase) escherichia coli expression strain, repeatedly extracting three times with acetone and ethyl acetate (1:3) after feeding, concentrating under reduced pressure, and preparing a chromatographic MPLC (medium pressure chromatography) by adopting a medium pressureX2 (BUCHI, inc.), column model: 80g reverse C18 column), 45% -80% methanol-water gradient for 40min, collecting and concentrating the fraction containing compound III. Separating by preparative liquid phase (preparative column type: YMC ODS-A5 μm 120A (10X 250 mm)), eluting with 45% acetonitrile-water isocratically to obtain 1.5mg of pure compound III. Deuterated acetone was dissolved and nuclear magnetic identification was performed, and the assignment of the compounds is shown in table 3.
Identification of Compound III: colorless oily form; the structure of this compound was identified as follows based on the assignment of nuclear magnetic data in table 3.
TABLE 3 Nuclear magnetic data assignment for Compound III
Preparation of compound IV: genes AcrABCD (PKS, NRPS, SDR and acetyltransferase) from Acremonium chrysogenum were combined and co-expressed in aspergillus nidulans, the heterologous expression strain was mass fermented and after fermentation was completed, acetone was used: ethyl acetate (1:3) was repeatedly extracted three times and concentrated to dryness under reduced pressure. Adopts medium pressure preparation chromatography MPLCX2 (BUCHI, inc.), column model: 80g reverse C18 column), 45% -90% methanol-water gradient for 40min, collecting and concentrating the fraction containing compound IV. Separating again by preparative liquid phase (preparative column type: YMC ODS-A5 μm 120A (10X 250 mm)), eluting with 41% acetonitrile-water isocratically to obtain 6.5mg of pure compound IV. Deuterated acetone was dissolved and nuclear magnetic identification was performed, and the assignment of the compounds is shown in table 4.
Identification of Compound IV: colorless oily form; based on the assignment of nuclear magnetic data in table 4, the structure of this compound was identified as follows:
table 4 Nuclear magnetic data assignment of Compound IV
Preparation of compound V: genes FcaAB (PKS, NRPS) from Fusarium catenulatum cgmcc 3.4704 and genes ThmC and ThmE (SDR and acyltransferase) from Talaromyces thermophilus NRRL 2155 were combined for heterologous expression in aspergillus nidulans and after extensive fermentation, were fermented with acetone: ethyl acetate (1:3) was repeatedly extracted three times and concentrated to dryness under reduced pressure. Adopts medium pressure preparation chromatography MPLCX2 (BUCHI, inc.), column model: 120g reverse C18 column), 40% -90% methanol-water gradient for 50min, collecting and concentrating the fraction containing compound V. Further purification was carried out by preparing a liquid phase (preparation column type: YMC ODS-A5 μm 120A (10X 250 mm)), and isocratic elution with 71% methanol-water gave 3mg of pure compound V. Deuterated acetone was dissolved and nuclear magnetic identification was performed, and the assignment of the compounds is shown in table 5.
Identification of Compound V: colorless oily form; the calculated value of HR TOFMS m/z 644.3779[ M+Na ] +,C34H55NO9 in positive ion mode is 644.3769, indicating the molecular formula of this compound is C 34H55NO9, and the structure of this compound is determined by combining the nuclear magnetic data in Table 5 as follows:
Table 5 Nuclear magnetic data assignment of Compound V
Preparation of Compound VI: gene ThmAB (PKS, NRPS is heterologously expressed in Aspergillus nidulans) from Talaromyces thermophilus NRRL 2155 was prepared by fermenting a large amount to obtain compound IX, structure of IX was as follows, feeding compound IX into ThmC and ThmE E.coli coexpression strains, repeating extraction three times with acetone: ethyl acetate (1:3) after the end of feeding, concentrating under reduced pressure to dryness, carrying out crude separation by MCI column chromatography with methanol-water as solvent, eluting with 40%,50%,60%,70%, 80%,90%,100% methanol as solvent, eluting with two column volumes as each concentration, concentrating compound VI mainly in 80% and 90% of the components, combining concentrating under reduced pressure to dryness, then further purifying with semi-prepared liquid phase (preparation column model: YMC ODS-A5 μm 120A (10×250 mm)), eluting with 80% methanol-water to obtain 3mg pure product of compound VI, dissolving with deuterated acetone, carrying out nuclear magnetic identification, and attaching the compound as shown in Table 6.
Identification of Compound VI: colorless oily form; calculated value of HR TOFMS m/z 624.4085[ M+Na ] +,C32H59NO9 under positive ion mode is 624.4082, which shows that the molecular formula of the compound is C 32H59NO9, and the structure of the compound is determined by combining the nuclear magnetic data in Table 6 as follows:
TABLE 6 Nuclear magnetic data assignment for Compound VI
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Compound I and compound VIII preparation: genes ThmABCE (PKS, NRPS, SDR and acetyltransferase) from Talaromyces thermophilus NRRL 2155 were expressed heterologously in aspergillus nidulans, the heterologously expressed strains were mass fermented and after the fermentation was completed, acetone was used: ethyl acetate (1:3) was repeatedly extracted three times and concentrated to dryness under reduced pressure. Crude separation was performed using MCI column chromatography with methanol-water as solvent and 30%,40%,50%,60%,70%,80%,90%,100% methanol as elution procedure, two column volumes were eluted for each concentration, and the fractions containing compound I and compound VIII were combined separately. Subsequently, the mixture was separated again by Sephadex LH-20 (150 mm. Times.10 mm) with methanol as a solvent. The crude product obtained was again subjected to separation by preparative liquid phase (column type: YMC ODS-A5 μm 120A (10X 250 mm)), isocratic elution with 65% methanol water, to give 2mg of pure compound I, isocratic elution with 75% methanol water, to give 11mg of pure compound VIII. Deuterated acetone was dissolved and nuclear magnetic identification was performed, and the assignment of the compounds is shown in tables 7 and 8.
Identification of Compound I: colorless oily form; from the assignment of the nuclear magnetic data in Table 7, it was compared with the reported thermolide A carbon spectrum data, and the structure of the compound was determined as follows.
Table 7 Nuclear magnetic data belonging to Compound 1
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Identification of Compound VIII: colorless oily form; the calculated value of HR TOFMS m/z 596.3773[ M+Na ] +,C30H55NO9 in positive ion mode is 596.3769, indicating the molecular formula of the compound is C 30H55NO9, and the structure of the compound is determined by combining the nuclear magnetic data in Table 8.
Table 8 Nuclear magnetic data assignment of Compound VIII
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EXAMPLE 2 and thermolides nematicidal Effect of Compounds
The compounds I to VIII of example 1 were subjected to an anti-nematode test, in particular as follows: the above-mentioned compounds I to VIII were prepared into solutions having concentrations of 50. Mu.g/mL and 25. Mu.g/mL, respectively, and then 400 nematodes were added, and after 30 hours of treatment, the death of the nematodes was counted, and the statistical results are shown in Table 1.
Nematoda-resisting effect of the compound shown in Table 1 and thermolides
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The results show that the compound II in the compounds I to IV has better nematicidal effect, and the compound V, VI and VII have better effect when treated for 30 hours at 25 mug/ml, so that the nematicidal rate of the compound V, VI and VII reaches 100 percent.
The above-described embodiments are merely preferred embodiments for fully explaining the present invention, and the scope of the present invention is not limited thereto. Equivalent substitutions and modifications will occur to those skilled in the art based on the present invention, and are intended to be within the scope of the present invention. The protection scope of the invention is subject to the claims.
Claims (2)
1. The application of thermolides compounds with nematodc activity in preparing pesticides for preventing and treating nematodc disease is characterized in that: the structural formula of the compound is as follows:
2. the use according to claim 1, characterized in that: the effective concentration of thermolides compounds in the pesticide is 25 mug/ml.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111276470.7A CN113956212B (en) | 2021-10-29 | 2021-10-29 | Thermolides compounds with nematodc resistance activity and preparation method and application thereof |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111276470.7A CN113956212B (en) | 2021-10-29 | 2021-10-29 | Thermolides compounds with nematodc resistance activity and preparation method and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN113956212A CN113956212A (en) | 2022-01-21 |
| CN113956212B true CN113956212B (en) | 2024-04-30 |
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Citations (4)
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|---|---|---|---|---|
| US5510372A (en) * | 1988-02-25 | 1996-04-23 | Pfizer Inc | Antiparasitic macrolide antiobiotics |
| CN1861593A (en) * | 2006-04-26 | 2006-11-15 | 云南大学 | Large ring lactone compound containing meta-phenol ring and its application |
| CN102532026A (en) * | 2011-11-16 | 2012-07-04 | 云南大学 | Talamidolide compounds and application thereof |
| CN111588717A (en) * | 2020-06-17 | 2020-08-28 | 中国人民解放军空军军医大学 | Application of two quadruple lactone antibiotics as MRSA (methicillin resistant Staphylococcus aureus) resisting drugs and extraction and separation method thereof |
-
2021
- 2021-10-29 CN CN202111276470.7A patent/CN113956212B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5510372A (en) * | 1988-02-25 | 1996-04-23 | Pfizer Inc | Antiparasitic macrolide antiobiotics |
| CN1861593A (en) * | 2006-04-26 | 2006-11-15 | 云南大学 | Large ring lactone compound containing meta-phenol ring and its application |
| CN102532026A (en) * | 2011-11-16 | 2012-07-04 | 云南大学 | Talamidolide compounds and application thereof |
| CN111588717A (en) * | 2020-06-17 | 2020-08-28 | 中国人民解放军空军军医大学 | Application of two quadruple lactone antibiotics as MRSA (methicillin resistant Staphylococcus aureus) resisting drugs and extraction and separation method thereof |
Non-Patent Citations (3)
| Title |
|---|
| "Thermolides, Potent Nematocidal PKS-NRPS Hybrid Metabolites from Thermophilic Fungus Talaromyces thermophilus";Ji-Peng Guo等;《J. Am. Chem. Soc.》(第134期);第20306−20309页 * |
| Characterization of Thermolide Biosynthetic Genes and a New Thermolide from Sister Thermophilic Fungi;Xuemei Niu等;《Org. Lett.》(第16期);第3744−3747页 * |
| Metabolites from nematophagous fungi and nematicidal natural products from fungi as alternatives for biological control. Part II: metabolites from nematophagous basidiomycetes and non-nematophagous fungi;Thomas Degenkolb等;《Appl Microbiol Biotechnol》;第3813–3824页 * |
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