A composition for preventing and treating grape downy mildew and a preparation method and application thereof

Abstract

The present application relates to the technical field of biotechnology, and proposes a composition for preventing and treating grape downy mildew, a preparation method and application thereof.The composition for preventing and treating grape downy mildew comprises cyazofamid and flavonoid glycoside extract, and the mass ratio of cyazofamid to flavonoid glycoside extract is 5:150-250.Through the above technical solution, the problem of poor prevention and treatment effect of the existing technology on grape downy mildew is solved.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, specifically to a composition for preventing and controlling grape downy mildew, its preparation method, and its application. Background Technology

[0002] Downy mildew primarily affects young grape tissues, such as shoots, bunches, and leaves. Factors contributing to downy mildew include the pathogen, temperature, and humidity. The most significant factor inducing downy mildew is the fungal oospores. Downy mildew is widespread in grape production, and despite timely control measures during the growing season, it cannot be completely eradicated. During the summer and autumn seasons, rainfall becomes more frequent, leading to higher humidity levels in the fields and increasing the risk of downy mildew. In such cases, chemical pesticides can be used to enhance control and prevent widespread disease spread within the vineyard. If the overall disease incidence exceeds 10%, a combination of pesticides is necessary. Using a single pesticide alone is insufficient to control the disease's spread. Therefore, existing pesticides for controlling grape downy mildew suffer from low application efficiency and poor overall control. Summary of the Invention

[0003] This invention proposes a composition for controlling grape downy mildew, its preparation method, and its application, which solves the problem that the agents used to control grape downy mildew in related technologies have poor control effects on grape downy mildew.

[0004] The technical solution of the present invention is as follows: This invention proposes a composition for controlling grape downy mildew, comprising cyazofamid and flavonoid glycoside extract; wherein the mass ratio of cyazofamid to flavonoid glycoside extract is 5:150~250.

[0005] As a further technical solution, the flavonoid glycoside extract is lotus leaf flavonoid glycoside and / or Solomon's seal leaf flavonoid glycoside.

[0006] As a further technical solution, when the flavonoid glycoside extract is lotus leaf flavonoid glycoside and Solomon's seal leaf flavonoid glycoside, the mass ratio of the lotus leaf flavonoid glycoside and Solomon's seal leaf flavonoid glycoside is 3:7~7:3.

[0007] As a further technical solution, the raw material for the lotus leaf flavonoid glycoside is lotus leaf; the raw material for the Solomon's seal leaf flavonoid glycoside is Solomon's seal leaf.

[0008] As a further technical solution, the preparation method of the flavonoid glycoside extract includes the following steps: S1. Crush lotus leaves or Solomon's seal leaves, defatt them, and obtain the pretreated material; S2. Take half the mass of the pretreated material and add it to hot water for ultrasonic extraction and filtration to obtain extract A and filter residue A. S3. Take the other half of the pretreated material by mass and add it to the first ethanol, perform ultrasonic extraction, filter, and obtain extract B and filter residue B. S4. Combine the filter residues A and B and add them to the second ethanol for ultrasonic extraction. Filter the mixture to obtain extract C. S5. After mixing extract A, extract B and extract C, purify, concentrate and dry to obtain flavonoid glycoside extract.

[0009] As a further technical solution, in step S5, the purification process involves using a macroporous adsorption resin column of model D101 and eluting it with ethanol at a volume fraction of 40%~50%.

[0010] As a further technical solution, in step S2, the material-to-liquid ratio of ultrasonic extraction is 1g:10~20mL, the temperature is 60~80℃, the time is 50~60min, and the ultrasonic power is 600~800W.

[0011] As a further technical solution, in steps S3 and S4, the material-to-liquid ratio of the ultrasonic extraction is independently 1g:10~15mL, the temperature is independently 30~40℃, the time is independently 45~55min, and the ultrasonic power is independently 650~750W.

[0012] As a further technical solution, the volume fraction of the first ethanol is 60%~70%; the volume fraction of the second ethanol is 80%~90%.

[0013] As a further technical solution, the dosage form of the composition includes wettable powder, suspension, and water emulsion.

[0014] As a further technical solution, when the dosage form of the composition is a wettable powder, it comprises the following components by mass percentage: 20%~40% cyazofamid and flavonoid glycoside extract, 5%~8% dispersant, 3%~5% wetting agent, and filler to 100%; and / or, When the dosage form of the composition is a suspension, it consists of the following components by mass percentage: Cypermethrin and flavonoid glycoside extract 20%~40%, dispersant 6%~9%, wetting agent 2%~5%, thickener 0.1%~2%, antifreeze 1%~3%, defoamer 0.5%~1%, water to 100%; and / or, When the composition is in the form of an aqueous emulsion, it consists of the following components by mass percentage: 20%~40% cypermethrin and flavonoid glycoside extract, 5%~10% emulsifier, 0.2%~2% antifreeze, 0.5%~2% defoamer, and water to 100%.

[0015] As a further technical solution, the composition is obtained by mixing the components of the composition.

[0016] As a further technical solution, the dispersant is one of lignin sulfonate, alkylphenol polyoxyethylene ether formaldehyde condensate sulfate, and calcium alkylbenzene sulfonate; the wetting agent is one of sodium dodecyl sulfate, sodium dodecylbenzene sulfonate, and alkylphenol polyoxyethylene ether; the thickener is one of xanthan gum, polyvinyl alcohol, and bentonite; the antifreeze is a polyol; the defoamer is one of hexadecyl alcohol and diisobutylmethanol; the filler is one of kaolin and diatomaceous earth; and the emulsifier is one of castor oil polyoxyethylene ether and alkylphenol polyoxyethylene ether.

[0017] The present invention also proposes the application of the composition for controlling grape downy mildew in the control of grape downy mildew.

[0018] The working principle and beneficial effects of this invention are as follows: 1. In this invention, by using cyazofamid and flavonoid glycoside extract as the main components of the composition for controlling grape downy mildew, the combination of cyazofamid and flavonoid glycoside extract has a significant synergistic effect, which significantly improves the control effect on grape downy mildew.

[0019] 2. In this invention, the flavonoid glycoside extract uses lotus leaf flavonoid glycoside and / or Solomon's seal leaf flavonoid glycoside. When the flavonoid glycoside extract is lotus leaf flavonoid glycoside and Solomon's seal leaf flavonoid glycoside, the synergistic effect with cyazofamid is better, further improving the control effect against grape downy mildew.

[0020] 3. In the preparation process of the flavonoid glycoside extract, stepwise ultrasonic extraction is adopted, and flavonoid glycosides are extracted using ethanol of different volume fractions, which increases the content of flavonoid glycosides in the flavonoid glycoside extract and further improves the control effect on grape downy mildew. Detailed Implementation

[0021] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0022] In the following examples and comparative examples, the particle size of kaolin and diatomaceous earth is 500 mesh.

[0023] Example 1 When the composition is in the form of a wettable powder, it consists of the following components by weight percentage: Cyazofamid and lotus leaf flavonoids 20%, sodium lignosulfonate 5%, sodium dodecyl sulfate 3%, kaolin to make up to 100%; By mixing the components of the above composition, a composition for controlling grape downy mildew is obtained. The mass ratio of cypermethrin to lotus leaf flavonoids is 5:150. The preparation method of lotus leaf flavonoid glycosides includes the following steps: S1. Crush the lotus leaves and defatt them to obtain the pretreated material; S2. According to the material-liquid ratio of 1g:10mL, take half the mass of the pretreated material and add it to water. Perform ultrasonic extraction at 600W and 60℃ for 60min. Filter to obtain extract A and filter residue A. S3. With a material-to-liquid ratio of 1g:10mL, take the other half of the pretreated material and add it to 60% ethanol. Perform ultrasonic extraction at 30℃ and 650W for 55min. Filter to obtain extract B and filter residue B. S4. Combine filter residue A and filter residue B and add them to 60% second ethanol by volume. Perform ultrasonic extraction and filter to obtain extract C. The ultrasonic extraction conditions are the same as in step S3. S5. After mixing extract A, extract B and extract C, the mixture is loaded onto a macroporous adsorption resin column of model D101, eluted with 40% ethanol, concentrated and dried to obtain lotus leaf flavonoids. The mass content of lotus leaf flavonoids was determined to be 80.2% by liquid chromatography.

[0024] Example 2 When the composition is in the form of a wettable powder, it consists of the following components by weight percentage: Cyazofamid and lotus leaf flavonoid glycosides 40%, calcium dodecylbenzenesulfonate 8%, sodium dodecylbenzenesulfonate 5%, diatomaceous earth to make up to 100%; By mixing the components of the above composition, a composition for controlling grape downy mildew is obtained. The mass ratio of cypermethrin to lotus leaf flavonoid glycosides is 5:170. The preparation method of lotus leaf flavonoid glycosides includes the following steps: S1. Crush the lotus leaves and defatt them to obtain the pretreated material; S2. According to the material-liquid ratio of 1g:15mL, take half the mass of the pretreated material and add it to water. Perform ultrasonic extraction at 700W and 70℃ for 55min. Filter to obtain extract A and filter residue A. S3. With a material-to-liquid ratio of 1g:13mL, take the other half of the pretreated material and add it to 65% ethanol. Perform ultrasonic extraction at 700W and 35℃ for 50min. Filter to obtain extract B and filter residue B. S4. Combine filter residue A and filter residue B and add them to 65% second ethanol by volume. Perform ultrasonic extraction and filter to obtain extract C. The ultrasonic extraction conditions are the same as in step S3. S5. After mixing extract A, extract B and extract C, the mixture is loaded onto a macroporous adsorption resin column of model D101, eluted with 50% ethanol, concentrated and dried to obtain lotus leaf flavonoids. The mass content of lotus leaf flavonoids was determined to be 80.6% by liquid chromatography.

[0025] Example 3 When the composition is in the form of a suspension, it consists of the following components by mass percentage: Cyazofamid and lotus leaf flavonoid glycosides 20%, calcium dodecylbenzenesulfonate 6%, sodium dodecylbenzenesulfonate 2%, xanthan gum 0.1%, glycerol 1%, diisobutylmethanol 0.5%, water to 100%; By mixing the components of the above composition, a composition for controlling grape downy mildew is obtained. The mass ratio of cypermethrin to lotus leaf flavonoids is 5:190. The preparation method of lotus leaf flavonoid glycosides includes the following steps: S1. Crush the lotus leaves and defatt them to obtain the pretreated material; S2. According to the material-liquid ratio of 1g:20mL, take half the mass of the pretreated material and add it to water. Perform ultrasonic extraction at 800W and 80℃ for 50min. Filter to obtain extract A and filter residue A. S3. With a material-to-liquid ratio of 1g:15mL, take the other half of the pretreated material and add it to 70% ethanol. Perform ultrasonic extraction at 750W and 40℃ for 45min. Filter to obtain extract B and filter residue B. S4. Combine filter residue A and filter residue B and add them to 70% second ethanol by volume. Perform ultrasonic extraction and filter to obtain extract C. The ultrasonic extraction conditions are the same as in step S3. S5. After mixing extract A, extract B and extract C, the mixture is loaded onto a macroporous adsorption resin column of model D101, eluted with 40% ethanol, concentrated and dried to obtain lotus leaf flavonoids. The mass content of lotus leaf flavonoids was determined to be 81.0% by liquid chromatography.

[0026] Example 4 When the composition is in the form of a suspension, it consists of the following components by mass percentage: Cyazofamid and lotus leaf flavonoid glycosides 40%, calcium dodecylbenzenesulfonate 9%, sodium dodecylbenzenesulfonate 5%, xanthan gum 2%, glycerol 3%, diisobutylmethanol 1%, water to make up to 100%; By mixing the components of the above composition, a composition for controlling grape downy mildew is obtained. The mass ratio of cypermethrin to lotus leaf flavonoids is 5:210. The preparation method of lotus leaf flavonoid glycosides is the same as in Example 3.

[0027] Example 5 When the composition is in the form of an aqueous emulsion, it consists of the following components by mass percentage: Cyazofamid and lotus leaf flavonoid glycosides 20%, dodecylphenol polyoxyethylene ether 5%, glycerol 0.2%, hexadecyl alcohol 0.5%, water to 100%; By mixing the components of the above composition, a composition for controlling grape downy mildew is obtained. The mass ratio of cypermethrin to lotus leaf flavonoids is 5:230. The preparation method of lotus leaf flavonoid glycosides is the same as in Example 3.

[0028] Example 6 When the composition is in the form of an aqueous emulsion, it consists of the following components by mass percentage: Cyazofamid and lotus leaf flavonoid glycosides 40%, dodecylphenol polyoxyethylene ether 10%, glycerol 2%, hexadecyl alcohol 2%, water to make up to 100%; By mixing the components of the above composition, a composition for controlling grape downy mildew is obtained. The mass ratio of cypermethrin to lotus leaf flavonoids is 5:250. The preparation method of lotus leaf flavonoid glycosides is the same as in Example 3.

[0029] Example 7 The only difference between this embodiment and Example 4 is the preparation method of lotus leaf flavonoids; in step S4, the volume fraction of the second ethanol is 80%. The mass content of lotus leaf flavonoids was determined to be 84.6% by liquid chromatography.

[0030] Example 8 The only difference between this embodiment and Example 7 is the preparation method of lotus leaf flavonoids; in step S4, the volume fraction of the second ethanol is 90%. The mass content of lotus leaf flavonoids was determined to be 83.9% by liquid chromatography.

[0031] Example 9 The only difference between this embodiment and embodiment 7 is that lotus leaf flavonoids are replaced with Solomon's seal leaf flavonoids, and lotus leaves are replaced with Solomon's seal leaves. The content of flavonoid glycosides in Polygonatum odoratum leaves was determined to be 84.8% by liquid chromatography.

[0032] Example 10 The only difference between this embodiment and Example 9 is that the flavonoid glycosides of Solomon's seal leaf are replaced with a mixture of lotus leaf flavonoid glycosides and Solomon's seal leaf flavonoid glycosides; the mass ratio of lotus leaf flavonoid glycosides to Solomon's seal leaf flavonoid glycosides is 3:7; the preparation method of lotus leaf flavonoid glycosides is the same as in Example 7, and the preparation method of Solomon's seal leaf flavonoid glycosides is the same as in Example 9.

[0033] Example 11 The only difference between this embodiment and Example 10 is that the flavonoid glycosides of Solomon's seal leaf are replaced with a mixture of lotus leaf flavonoid glycosides and Solomon's seal leaf flavonoid glycosides; the mass ratio of lotus leaf flavonoid glycosides to Solomon's seal leaf flavonoid glycosides is 7:3.

[0034] Example 12 The only difference between this embodiment and Example 3 is the preparation method of lotus leaf flavonoid glycosides, which includes the following steps: S1. Crush the lotus leaves, defatt them, and obtain the pretreated material; S2. According to the material-liquid ratio of 1g:20mL, the pretreated material was added to 70% ethanol by volume, and ultrasonic extraction was performed at 800W and 80℃ for 100min. After filtration, the filtrate was concentrated to obtain lotus leaf flavonoids. The mass content of lotus leaf flavonoids was determined to be 70.5% by liquid chromatography.

[0035] Comparative Example 1 The only difference between this comparative example and Example 3 is that when the dosage form of the composition is a suspension, it consists of the following components by mass percentage: 20% cyazofamid, 6% dispersant, 2% wetting agent, 0.1% thickener, 1% antifreeze, 0.5% defoamer, and water to bring the total to 100%.

[0036] Comparative Example 2 Lotus leaf flavonoid glycosides 20%, dispersant 6%, wetting agent 2%, thickener 0.1%, antifreeze 1%, defoamer 0.5%, water to 100%.

[0037] A grape experimental field was established with a plant spacing of 300×300cm. The field was divided into 15 zones. After the grape leaves unfolded and before the onset of downy mildew (mid-June), the compositions for controlling grape downy mildew prepared in Examples 1-12 and Comparative Examples 1-2 were prepared as 1 mg / L solutions and sprayed onto the grape leaves in the corresponding zones: Zones 1-12 corresponded to Examples 1-12 respectively; Zones 13-14 corresponded to Comparative Examples 1-2 respectively; Zone 15 (control zone) was sprayed with an equal amount of water; every 7... Spray once in mid-September. In each area, 10 new shoots were investigated, and 10 leaves were investigated from top to bottom on each new shoot. The disease condition of each leaf was investigated and recorded according to the disease assessment and grading method shown in Table 1. The control effect on grape downy mildew was calculated, as shown in Table 2. Control effect = [(disease index of control area - disease index of area sprayed with grape downy mildew control composition) / disease index of control area] × 100%, where, disease index = [Σ(number of diseased leaves at each level × relative level value) / (total number of investigated leaves / × 9)] × 100.

[0038] Table 1 Disease Assessment Grading

[0039] Table 2 Prevention and control effects

[0040] By comparing the data from Examples 1-12 and Comparative Examples 1-2, it was found that the compositions for controlling grape downy mildew prepared in Examples 1-12 had a higher anti-mildew effect than those in Comparative Examples 1-2. This indicates that by combining cyazofamid and flavonoid glycoside extracts, the synergistic effect can be significantly improved in controlling grape downy mildew.

[0041] By comparing the data from Examples 7 and 9-11, it was found that the compositions for controlling grape downy mildew prepared in Examples 10-11 had a higher anti-mildew effect than those in Examples 7 and 9. This indicates that when the flavonoid extracts are made from lotus leaf flavonoids and Solomon's seal leaf flavonoids, the synergistic effect with cyazofamid is better, which can further improve the control effect against grape downy mildew.

[0042] By comparing the data from Examples 4, 7-8, and 12, it was found that the composition for controlling grape downy mildew prepared in Examples 7-8 had a higher anti-mildew effect compared with Examples 4 and 12. This indicates that the use of multiple stepwise ultrasonic extractions and different volume fractions of ethanol during the extraction process of the flavonoid glycoside extract can further improve the control effect against grape downy mildew.

[0043] The above are merely preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A composition for controlling grape downy mildew, characterized in that, It includes cyazofamid and flavonoid glycoside extracts; the mass ratio of cyazofamid and flavonoid glycoside extracts is 5:150~250; The flavonoid extract is lotus leaf flavonoid glycoside and / or Solomon's seal leaf flavonoid glycoside; The raw material for the lotus leaf flavonoid glycosides is lotus leaf; the raw material for the Solomon's seal leaf flavonoid glycosides is Solomon's seal leaf. The preparation method of the flavonoid glycoside extract includes the following steps: S1. Crush lotus leaves or Solomon's seal leaves, defatt them, and obtain the pretreated material; S2. Take half the mass of the pretreated material and add it to water, perform ultrasonic extraction, filter, and obtain extract A and filter residue A; S3. Take the other half of the pretreated material by mass and add it to the first ethanol, perform ultrasonic extraction, filter, and obtain extract B and filter residue B. S4. Combine the filter residues A and B and add them to the second ethanol for ultrasonic extraction. Filter the mixture to obtain extract C. S5. After mixing extract A, extract B and extract C, purify, concentrate and dry to obtain flavonoid glycoside extract; In step S2, the material-to-liquid ratio for ultrasonic extraction is 1g:10~20mL, the temperature is 60~80℃, the time is 50~60min, and the ultrasonic power is 600~800W. In steps S3 and S4, the material-to-liquid ratio of the ultrasonic extraction is 1g:10~15mL, the temperature is 30~40℃, the time is 45~55min, and the ultrasonic power is 650~750W. The volume fraction of the first ethanol is 60% to 70%; the volume fraction of the second ethanol is 80% to 90%.

2. The composition for controlling grape downy mildew according to claim 1, characterized in that, When the flavonoid extract is lotus leaf flavonoid glycoside and Solomon's seal leaf flavonoid glycoside, the mass ratio of the lotus leaf flavonoid glycoside and Solomon's seal leaf flavonoid glycoside is 3:7 to 7:

3.

3. The composition for controlling grape downy mildew according to claim 1, characterized in that, The dosage forms of the composition include wettable powders, suspensions, and emulsions.

4. The application of the composition for controlling grape downy mildew according to any one of claims 1 to 3 in the control of grape downy mildew.

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