CN109280058B - Plant virus resisting medicament - Google Patents
Plant virus resisting medicament Download PDFInfo
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- CN109280058B CN109280058B CN201710597980.1A CN201710597980A CN109280058B CN 109280058 B CN109280058 B CN 109280058B CN 201710597980 A CN201710597980 A CN 201710597980A CN 109280058 B CN109280058 B CN 109280058B
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D493/00—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system
- C07D493/02—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system in which the condensed system contains two hetero rings
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- 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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Abstract
The invention relates to the field of pesticides, and particularly discloses an anti-plant virus medicament which comprises a micromolecule medicament shown in a formula (I) or a pharmaceutically acceptable salt containing a chemical structure shown in the formula (I), wherein R is1One selected from hydrogen, nitro, nitroso, sulfonic acid group, carboxyl, amino or azo, halogen atom, C1-10 alkyl, C2-10 alkenyl or C2-10 alkynyl. The small molecule medicament has good inhibition effect on virus silencing inhibitors, and the small molecule medicament has high-efficiency antiviral effect through the proof of pharmacodynamic experiments.
Description
Technical Field
The invention relates to the field of pesticides, and particularly relates to a plant virus resistant pesticide.
Background
Plant virus diseases are one of the important diseases of crops, are called plant cancers, and cause losses of billions of dollars to global agricultural production every year. In recent years, particularly for commercial crops, the damage of plant virus diseases is increasingly serious. If the tobacco virus disease is caused, once the quality of the ill tobacco leaves is reduced by at least 2-3 grades, the economic value is even completely lost when the quality is serious, and the economic loss caused by the disease is far more than the tobacco fungal disease due to the extremely difficult control, so that the disease becomes a disease which threatens the maximum in tobacco production.
Since the plant virus is propagated by totally depending on the metabolism of the host after infecting the plant, the virus is difficult to be specifically killed under the condition of not influencing the normal metabolism function of the host, so that the prevention and the treatment of the virus disease are extremely difficult. At home and abroad, existing drugs for preventing and treating diseases, such as ningnanmycin, S-methylbenzo [1,2,3] thiadiazole-7-thiocarboxylate (BTH, syn. acibenzolar-S-methyl) and the like, mainly play a role in obtaining resistance (System acquirescence) by inducing a plant generating System, have poor effects and only have a prevention effect. In fact, there are few drugs that have therapeutic effects against plant viruses at present.
The RNA silencing mechanism (RNA silencing) discovered in recent years has now been considered as the most important defense mechanism of plants against viruses. In short, after the virus is infected, the nucleic acid of the virus can cause the plant to generate specific degradation aiming at the nucleic acid, thereby achieving the aim of eliminating the virus. However, similar to military competition, viruses have evolved a silencing suppressor (Viral RNA silencing suppressor) to counteract the defense of plant RNA silencing. Such as the Hc-Pro encoded by potyvirus, plum pox virus, 2b encoded by cucumovirus and P19 encoded by solanum bushy stunt virus. The mechanism of action is mainly to interfere with the overall silencing mechanism by binding to small RNAs that play an important role in RNA silencing. Therefore, if a medicament capable of effectively inhibiting the virus silencing inhibitor is found, the medicament can help plants win the competition of military preparedness, and the aim of effectively treating virus diseases is fulfilled.
With the more and more clear concept of environmental protection, the problem of high toxicity and high residue of traditional chemical pesticides is also increasingly paid attention. Many components in the natural product have the characteristics of high efficiency, low toxicity, good environmental compatibility, small toxicity to human bodies and the environment and the like. Therefore, the method has important significance and value for screening the high-efficiency novel antiviral pesticide with the accurate molecular target from the natural product library.
Disclosure of Invention
In order to solve the problems in the prior art, the invention aims to provide a novel efficient antiviral agent with an accurate molecular target according to the latest virus interaction molecular mechanism.
In order to realize the purpose of the invention, the technical scheme of the invention is as follows:
the invention takes a virus silencing inhibitor as a target, screens a small molecular medicament (ZINC compound database, ID: ZINC79192430) with good inhibition effect on the target protein from a natural product library, and proves that the small molecular medicament has high-efficiency antiviral effect through pharmacodynamic experiments.
On the basis, the invention provides an anti-plant virus medicament, which comprises a small molecule medicament shown in a formula (I) or a pharmaceutically acceptable salt containing a chemical structure shown in the formula (I).
The small molecule medicament has a structure shown in a formula (I):
wherein R is1One selected from hydrogen, nitro, nitroso, sulfonic acid group, carboxyl, amino or azo, halogen atom, C1-10 alkyl, C2-10 alkenyl or C2-10 alkynyl.
In a specific embodiment of the invention, the agent comprises a small molecule agent represented by formula (II):
which is based on a small molecular medicament shown as a formula (I), R1In the case of hydrogen, the name is (8S, E) -5-hydroxy-8-methyl-4- (2-oxo-1, 2-dihydroquinolin-3-yl) -3,4,8,9,10,11,14, 15-octahydro- [1]Oxetamido [3,4-g ]]Chromene-2, 6,12(13H) -trione. Through experimental research, the small molecule medicament shown in the formula (II) has obvious inhibition effect on virus silencing inhibitors.
Furthermore, the invention also provides application of the small molecule medicament shown in the formula (I)/formula (II) or pharmaceutically acceptable salt thereof in inhibiting virus silencing inhibitor (Viral RNA sitting inhibitor). Such viral silencing inhibitors include, but are not limited to, P19, 2b, Hc-Pro, and the like.
Since most viruses have a suppressor of silencing, the target virus to which the agent for resisting plant virus is directed is not particularly limited.
Optionally and preferably, e.g., a Hc-Pro inhibitor-containing potyvirus, a plum pox virus, etc.; tomato sterility virus containing 2b inhibitor, cucumber mosaic virus, etc.; p19 inhibitor-containing carnation Italian ringspot virus, tomato bushy stunt virus, etc.
The medicament is also understood to be a pharmaceutical composition comprising an effective amount of a compound of formula (I)/formula (II) or a pharmaceutically acceptable salt thereof. The same applies to the control of diseases caused by plant viruses.
The invention has the beneficial effects that:
the invention provides a new application of a compound shown as a formula (I)/a formula (II), finds a new application in inhibiting a virus silencing inhibitor, and provides an anti-plant virus medicament based on the application.
Drawings
FIG. 1 is a diagram of EMSA assay for detecting the activity of small molecule agents in inhibiting the silencing inhibitor P19 in example 1 of the present invention.
FIG. 2 is a diagram showing the application of small molecule agents to the EMSA assay for the inhibition of the activity of the suppressor 2b in example 2 of the present invention.
Detailed Description
Preferred embodiments of the present invention will be described in detail with reference to the following examples. It is to be understood that the following examples are given for illustrative purposes only and are not intended to limit the scope of the present invention. Various modifications and alterations of this invention will become apparent to those skilled in the art without departing from the spirit and scope of this invention.
The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.
Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
Example 1
The micromolecule medicament has a structural formula shown as follows:
the results of applying the small molecule agent to an activity detection gel electrophoresis migration assay (EMSA) for inhibiting the suppressor of silencing P19 are shown in fig. 1. P19 and the small molecule agent are mixed in binding buffer and reacted fully for 25 minutes at room temperature. Wherein the concentration of P19 is 2 μ M, and the concentration of the small molecule agent is 2-200 ppm. Subsequently, siRNA was added to a final concentration of 50nM, and after thorough mixing, the reaction was continued at room temperature for 25 minutes. And then, observing the result by electrophoresis and color development after film transfer. In the color results, only siRNA showed a band and was in the lower position of the figure (as indicated by the UnboundsiRNA arrow). The silencing inhibitor protein combined with siRNA will make the electrophoresis rate of siRNA slow and the electrophoresis distance shorter, so it is in the upper position of the figure (shown by the arrow P19-siRNA complex). While when the compound is added and the protein is inhibited from being combined with the siRNA, the siRNA restores the original electrophoresis rate, so that the strip signal is enhanced at the lower part of the figure.
In FIG. 1, lane 1 is a negative control containing siRNA only. Lane 2 is a positive control containing siRNA and P19. Lanes 3-9 are experimental groups, each containing siRNA and P19, and containing the small molecule agent at a concentration gradient of 2, 5, 10, 20, 40, 100, 200 ppm. As can be seen, the bands of the negative control group all appear at the unbounded siRNA positions at the lower part of the figure. After the addition of the inhibitor protein to the positive control, a portion of the siRNA bound to the protein, as shown in the upper band of the figure for P19-siRNA complex. When small molecule agents with increasing concentrations are added, the bands at the upper part of the graph gradually weaken, indicating that the small molecule agents can effectively prevent the binding reaction of the suppressor protein and siRNA. As shown in the figure, the small molecule agent with the concentration of 20ppm can well inhibit the binding reaction of P19 with siRNA with the concentration of 2 μ M.
Example 2
The micromolecule medicament has a structural formula shown as follows:
the results of applying the small molecule agent to the EMSA test for inhibiting the activity of the suppressor 2b are shown in fig. 2. The 2b and the small molecule agent are mixed in the binding buffer solution and fully reacted for 25 minutes at room temperature. Wherein the concentration of 2b is 2 μ M, and the concentration of the small molecule agent is 20-200 ppm. Subsequently, siRNA was added to a final concentration of 50nM, and after thorough mixing, the reaction was continued at room temperature for 25 minutes. And then, observing the result by electrophoresis and color development after film transfer. In the color results, only siRNA showed a band and was in the lower position of the figure (indicated by Unbound siRNA arrow). The silencing inhibitor protein combined with siRNA will make the electrophoresis rate of siRNA slow and the electrophoresis distance shorter, so it is in the upper position of the figure (shown by the arrow P19-siRNA complex). While when the compound is added and the protein is inhibited from being combined with the siRNA, the siRNA restores the original electrophoresis rate, so that the strip signal is enhanced at the lower part of the figure.
In FIG. 2, lane 1 is a negative control containing siRNA only. Lane 2 is a positive control containing siRNA and 2 b. Lanes 3-9 are experimental groups, each containing siRNA and 2b, and containing the small molecule agent at a concentration gradient of 20, 40, 80, 100, 150, 200 ppm. As can be seen, the bands of the negative control group all appear at the unbounded siRNA positions at the lower part of the figure. After the addition of the inhibitor protein to the positive control, a portion of the siRNA binds to the protein, shown in the upper 2b-siRNA complete band of the figure. When small molecule agents with increasing concentrations are added, the bands at the upper part of the graph gradually weaken, indicating that the small molecule agents can effectively prevent the binding reaction of the suppressor protein and siRNA. As shown in the figure, the small molecule agent with 100ppm concentration can well inhibit the binding reaction of 2b with siRNA with 2 μ M concentration.
Example 3
The compound shown in the formula (II) has the effect of preventing and treating the tomato bushy stunt virus.
The experimental method comprises the following steps:
selecting Benziyan tobacco with similar growth vigor at the 4-5 leaf stage, fully grinding 0.3g of fresh diseased leaves with obvious symptoms by using 30m L double distilled water, carrying out friction inoculation on 1% virus inoculation liquid through diatomite, inoculating 2 leaves on each strain, spraying 150ppm of medicament aqueous solution 2 hours, 1 day and 3 days after inoculation, and setting 20 strains in each treatment group.
Disease indices were calculated at 8, 11, and 14 days after inoculation and the results are shown in table 1.
a. Grading the disease condition standard:
grade 0-no symptoms.
Grade 1-mild symptoms appeared in inoculated leaves.
2-stage-one to two systematic leaves are clear and deformed.
Grade 3-most upper leaves are deformed or main side veins are necrotic, and diseased plants are dwarfed.
Grade 4-the whole plant is severely deformed or necrotic.
b. Index of disease condition
Disease index is [ ∑ (number of diseased leaves at each stage × relative stage)/(total leaf number investigated × 9) ] × 100%
c. Controlling effect
The control effect (%) is (control disease index-treatment disease index)/control disease index) × 100%
TABLE 1
Example 4
The compound of formula (II) has the effect of preventing and treating cucumber mosaic virus.
Selecting Benziyan tobacco with similar growth vigor at the 4-5 leaf stage, fully grinding 0.3g of fresh diseased leaves with obvious symptoms by using 30m L double distilled water, carrying out friction inoculation on 1% virus inoculation liquid through diatomite, inoculating 2 leaves for each strain, spraying 150ppm of medicament aqueous solution 2 hours after inoculation, 1 day and 3 days after inoculation, setting 20 strains for each treatment group, and calculating disease indexes from 9 th, 15 th and 20 th after inoculation, wherein the results are shown in a table 2.
a. Grading the disease condition standard:
grade 0-no symptoms.
Grade 1-mild symptoms appeared in inoculated leaves.
2-stage-one to two systematic leaves are clear and deformed.
Grade 3-most upper leaves are leafy, chlorosis or deformed.
Grade 4-leaf of the whole plant, severe deformation or necrosis, and severe dwarfing of diseased plants.
b. Index of disease condition
Disease index is [ ∑ (number of diseased leaves at each stage × relative stage)/(total leaf number investigated × 9) ] × 100%
c. Controlling effect
The control effect (%) is (control disease index-treatment disease index)/control disease index) × 100%
TABLE 2
Example 5
The compound of the formula (II) has the effect of preventing and treating turnip mosaic virus.
Selecting tobacco with 4-5 leaf stages and similar growth vigor, fully grinding 0.4g of fresh diseased leaves with obvious symptoms by using 20m L double-distilled water, inoculating 2% of virus inoculation liquid by using diatomite in a rubbing way, washing the leaves after the leaves are dried, applying the liquid 2 hours, 1 and 3 days after inoculation, spraying 150ppm of medicament aqueous solution, and recording the number of viruses with fluorescence signals of a control group and a medicament treatment group 3,4 and 5 days after inoculation respectively.
Each treatment group had 14 plants, 2-3 leaves per plant. Different leaves were taken at each recording time point and 3 replicates were taken. And the inhibition rate of the virus was calculated as follows.
Y＝(C-A)/C*100％
Wherein Y is the inhibition rate of the compound on viruses, C is the virus number of a control group, and A is the virus number of a compound treatment group. The results are shown in Table 3.
TABLE 3
Example 6
The control effect of the compounds of formula (II) of the invention on plum pox virus.
Selecting Benziyan tobacco with similar growth vigor in 4-5 leaf period, fully grinding 0.3g of fresh diseased leaves with obvious symptoms by using 30m L double distilled water, carrying out friction inoculation on 1% virus inoculation liquid through diatomite, inoculating 2 leaves for each strain, spraying 150ppm of medicament aqueous solution 2 hours, 1 day and 3 days after inoculation, setting 15 strains for each treatment group, and calculating disease indexes 8 days, 10 days and 12 days after inoculation.
The results are shown in Table 4.
a. Grading the disease condition standard:
grade 0-no symptoms.
Grade 1-mild symptoms appeared in inoculated leaves.
Stage 2-one to two system blades mottle, deform.
Grade 3-most upper leaves are chlorosis, deformed, and stunted.
Grade 4-the whole plant leaves are chlorosis, severely deformed or necrotic, and the diseased plants are severely dwarfed.
b. Index of disease condition
Disease index is [ ∑ (number of diseased leaves at each stage × relative stage)/(total leaf number investigated × 9) ] × 100%
c. Controlling effect
The control effect (%) is (control disease index-treatment disease index)/control disease index) × 100%
TABLE 4
The use of the small molecule pharmaceutical compounds of the present invention as antiviral agents has been described by way of specific examples, and it will be apparent to those skilled in the art that the present invention can be used for other purposes without departing from the scope of the present invention by appropriately modifying the materials, process conditions, etc. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.
Claims (2)
1. The application of the small molecule compound shown in the formula (II) or the pharmaceutically acceptable salt thereof in inhibiting plant virus silencing suppressors;
2. The use according to claim 1, wherein the plant viral silencing suppressor is P19, 2b or Hc-Pro.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710597980.1A CN109280058B (en) | 2017-07-20 | 2017-07-20 | Plant virus resisting medicament |
| PCT/CN2017/120188 WO2019015264A1 (en) | 2017-07-20 | 2017-12-29 | Agent against plant virus |
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| CN201710597980.1A CN109280058B (en) | 2017-07-20 | 2017-07-20 | Plant virus resisting medicament |
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| CN110669053A (en) * | 2018-07-02 | 2020-01-10 | 北京赫尔默技术有限公司 | Zearalenone derivative and synthesis method thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4331685A (en) * | 1978-05-12 | 1982-05-25 | Kureha Kagaku Kogyo Kabushiki Kaisha | Control of plant virus diseases |
| CN102417505A (en) * | 2011-08-29 | 2012-04-18 | 南开大学 | Tetrazole compounds containing methyl-1,2,3-thiadiazole as well as preparation methods and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DD281111A5 (en) * | 1989-04-05 | 1990-08-01 | Univ Leipzig | AGENTS FOR THE CONTROL OF VEGETABLE VIRUSES |
| MX360866B (en) * | 2012-05-24 | 2018-11-09 | A B Seeds Ltd | Compositions and methods for silencing gene expression. |
| EP2909322A4 (en) * | 2012-10-16 | 2016-03-16 | Monsanto Technology Llc | Methods and compositions for controlling plant viral infection |
| JP2016147836A (en) * | 2015-02-13 | 2016-08-18 | 日本曹達株式会社 | Anti-plant virus composition |
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- 2017-12-29 WO PCT/CN2017/120188 patent/WO2019015264A1/en active Application Filing
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4331685A (en) * | 1978-05-12 | 1982-05-25 | Kureha Kagaku Kogyo Kabushiki Kaisha | Control of plant virus diseases |
| CN102417505A (en) * | 2011-08-29 | 2012-04-18 | 南开大学 | Tetrazole compounds containing methyl-1,2,3-thiadiazole as well as preparation methods and application thereof |
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