Application of Z-9-methyl octadecenoate as repellent in prevention and control of spodoptera frugiperda

Abstract

The invention discloses an application of Z-9-methyl octadecenoate as a repellent in prevention and control of spodoptera frugiperda. According to the invention, through an indoor oviposition selection test and separation, purification and identification by means of microchemical analysis, a substance Z-9-octadecenoic acid methyl ester with oviposition repelling activity on female spodoptera frugiperda is extracted from feces of larvae of spodoptera frugiperda, and experiments find that the substance Z-9-octadecenoic acid methyl ester has the activity of repelling oviposition of female spodoptera frugiperda. The Z-9-methyl octadecenoate can significantly reduce the number of spawns of spodoptera frugiperda on crop leaves sprayed with the Z-9-methyl octadecenoate, effectively prevents female spodoptera frugiperda from laying spawns, does not generate drug resistance, and is safe to the environment. Therefore, the Z-9-methyl octadecenoate can be used as an oviposition repellent for preventing and controlling the spodoptera frugiperda, and a new technology is provided for field comprehensive treatment of the spodoptera frugiperda.

Description

Technical Field

[0001] The present invention relates to the technical field of Spodoptera frugiperda control, and particularly to the application of methyl Z-9-octadecenoate as a repellent in the control of Spodoptera frugiperda. Background Art

[0002] Spodoptera frugiperda is an important agricultural pest globally. This species can feed on more than 80 plants, mainly damaging the important food crop maize. Spodoptera frugiperda mainly damages maize by the larvae feeding on leaves. Spodoptera frugiperda has extremely strong reproductive ability, and the egg production of a single female can be as high as 2,000 eggs. Although the current chemical control methods are effective quickly, they are often highly toxic, pose potential threats to non-target organisms and the environment, and long-term use will also cause Spodoptera frugiperda to develop drug resistance.

[0003] Spraying an oviposition repellent on the crops to be controlled can prevent female adult Spodoptera frugiperda from laying eggs on the leaves, thereby regulating the oviposition behavior of Spodoptera frugiperda, which is a potential control means.

[0004] Applying oviposition repellent substances to control Lepidoptera pests is a common means. Currently, there are studies on using oviposition repellent substances to control Spodoptera frugiperda at home and abroad. For example, the patent with the application number CN202411070873.X discloses a variety of isothiocyanate compounds, which have a repellent effect on Spodoptera frugiperda; the patent application with the application number CN202410646802.3 discloses an octanal screened through the Spodoptera frugiperda SfruORV gene, which can produce a repellent effect on Spodoptera frugiperda. However, the substances disclosed in the above two patent documents are irritating to the human respiratory tract and are not conducive to practical application. The patent with the application number CN202310773179.3 discloses a repellent pheromone derived from the surface of Spodoptera frugiperda eggs, and the active ingredients include dibutyl phthalate, palmitic acid, hexadecenoic acid, stearic acid, oleic acid, linoleic acid, and linolenic acid, and their mass concentrations are 1.8 mg / L - 2.2 mg / L, 32 mg / L - 36 mg / L, 0.6 mg / L - 1.2 mg / L, 23 mg / L - 29 mg / L, 28 mg / L - 32 mg / L, 86 mg / L - 94 mg / L, 45 mg / L - 52 mg / L respectively; the patent application with the application number CN202411256815.6 discloses a bacterium isolated from the surface of Spodoptera frugiperda eggs, which has oviposition repellent activity against female insects. However, the substances that produce oviposition repellent activity are not clearly defined in the above two patents. And there is no report on the use of methyl Z-9-octadecenoate in the control of Spodoptera frugiperda at home and abroad. Summary of the Invention

[0005] The primary object of the present invention is to overcome the disadvantages and deficiencies of the prior art and provide the use of methyl Z-9-octadecenoate in the preparation of a repellent for Spodoptera frugiperda.

[0006] Another object of the present invention is to provide the use of methyl Z-9-octadecenoate as a repellent in the control of Spodoptera frugiperda.

[0007] The object of the present invention is achieved by the following technical solutions:

[0008] Use of methyl Z-9-octadecenoate in the preparation of a repellent for Spodoptera frugiperda.

[0009] The chemical structural formula of the methyl Z-9-octadecenoate is shown in Formula I:

[0010]

[0011] The repellent is an oviposition repellent, and the main active ingredient for oviposition repellency is methyl Z-9-octadecenoate.

[0012] Use of methyl Z-9-octadecenoate as a repellent in the control of Spodoptera frugiperda.

[0013] The repellent is an oviposition repellent, with methyl Z-9-octadecenoate as the active ingredient.

[0014] The use of methyl Z-9-octadecenoate as a repellent in the control of Spodoptera frugiperda is to dissolve methyl Z-9-octadecenoate in a solvent and then apply it by smearing and / or spraying onto crops for repelling Spodoptera frugiperda from laying eggs.

[0015] The dosage of the methyl Z-9-octadecenoate can be adjusted according to the crop plants. When in use, the methyl Z-9-octadecenoate solution is evenly smeared and / or sprayed on the front and back of the crop leaves; preferably, the spraying dosage of methyl Z-9-octadecenoate is more than 11.4 mg of methyl Z-9-octadecenoate smeared and / or sprayed per square meter of crop leaves (i.e., ≥11.4 mg / m 2 ); more preferably, the spraying dosage of methyl Z-9-octadecenoate is 11.4 - 114 mg of methyl Z-9-octadecenoate smeared and / or sprayed per square meter of crop leaves.

[0016] The solvent is at least one of dichloromethane or n-hexane.

[0017] The crops include corn.

[0018] The present invention has the following advantages and effects compared with the prior art:

[0019] Through indoor oviposition selection experiments, separation, purification, and identification using microchemical analysis methods, a substance, methyl Z-9-octadecenoate, with oviposition deterrent activity against female insects was extracted from the feces of Spodoptera frugiperda larvae, and it was sprayed on the surface of corn plants for oviposition selection experiments. The experimental results showed that methyl Z-9-octadecenoate could significantly reduce the number of eggs on the sprayed leaves, effectively deter the oviposition of female Spodoptera frugiperda, not produce drug resistance, be environmentally safe, and have a relatively low cost, providing a new technology for the integrated field management of Spodoptera frugiperda. Description of the Drawings

[0020] Figure 1 It is a graph showing the results of the oviposition effects of different compounds on female Spodoptera frugiperda under indoor conditions (in the graph, ns indicates that there is no significant difference in the oviposition amount between the control and treatment areas, and "*" indicates that there is a significant difference in the oviposition amount between the control and treatment areas (p < 0.05)).

[0021] Figure 2 It is a graph showing the results of the oviposition effects of different doses of methyl Z-9-octadecenoate on female Spodoptera frugiperda under indoor conditions (in the graph, ns indicates that there is no significant difference in the oviposition amount between the control and treatment areas, and "*" indicates that there is a significant difference in the oviposition amount between the control and treatment areas (p < 0.05)).

[0022] Figure 3 It is a graph showing the oviposition selection results of Spodoptera frugiperda on corn plants treated with different doses of methyl Z-9-octadecenoate under greenhouse conditions (in the graph, ns indicates that there is no significant difference in the oviposition amount between the control and treatment areas, and "**" indicates that there is a significant difference in the oviposition amount between the control and treatment areas (p < 0.01)). Detailed Embodiments

[0023] The present invention will be further described in detail below in conjunction with embodiments, but the embodiments of the present invention are not limited thereto. Unless otherwise specified, the reagents, methods, and equipment used in the present invention are conventional reagents, methods, and equipment in the technical field. The test methods without specific experimental conditions specified in the following embodiments usually follow conventional experimental conditions. Unless otherwise specified, the reagents and raw materials used in the present invention can be obtained commercially.

[0024] Example 1 Screening of the extraction solvent for the oviposition deterrent active substance in the feces of Spodoptera frugiperda

[0025] The feeding and oviposition selection experiments of Spodoptera frugiperda were both conducted in an incubator at 26±1°C, with a relative humidity of 60±5%, and a light-dark cycle of 16h:8h. The feces of Spodoptera frugiperda larvae were collected daily and stored in a -20°C refrigerator for later testing. Feces were weighed on an analytical balance and placed in a headspace bottle, and solvents with different polarities, n-hexane, dichloromethane, and acetone, were added respectively. They were extracted at 26°C for 24h and filtered twice with filter paper to finally form a fecal extract of 10g / L. Spodoptera frugiperda was reared with artificial feed until the pupal stage. Newly emerged Spodoptera frugiperda were placed in a black plastic insect rearing box (16 cm in diameter and 9 cm in height), and allowed to freely mate for 48h. A filter paper with a diameter of 15 cm was cut along the diameter. Half was soaked in the solvent, and the other half was soaked in the fecal extract of different solvents. It was quickly taken out, and after the solvent had evaporated, it was pasted onto the lid of the insect rearing box with double-sided tape. A plastic petri dish with a diameter of 6 cm was placed in the center of the bottom of the insect rearing box, and a degreased cotton ball with 20% (w / v) sucrose solution was placed inside for the adults to supplement nutrition. Subsequently, 3 pairs of Spodoptera frugiperda adults were placed in each insect rearing box. Each insect rearing box was a replicate, and there were 6 replicates for one treatment. The number of eggs laid by female insects in different areas was recorded after 48h. The number of eggs laid in different areas was statistically analyzed using a paired t-test, and the oviposition deterrence index (ODI) was calculated as (the number of eggs laid in the control area - the number of eggs laid in the treatment area) / the number of eggs laid in the control area).

[0026] In addition, since a small amount of feed might be collected during the feces collection process, the oviposition selection of female Spodoptera frugiperda adults for the feed extract was also tested. The artificial feed eaten by Spodoptera frugiperda larvae was weighed in the same way and extracted with n-hexane, dichloromethane, and acetone. The remaining steps of the oviposition selection experiment and environmental conditions were as described above.

[0027] The experimental results showed (Table 1) that the fecal extracts of n-hexane and dichloromethane both showed significant oviposition deterrence activity against female adults, while the fecal extract of acetone had no significant oviposition deterrence activity against female adults. Among them, the fecal extract of dichloromethane had the highest oviposition deterrence activity, which was 64.29%. Table 2 showed that female adults had no significant oviposition preference for the feed extracts of the tested solvents. Therefore, dichloromethane was selected as the subsequent extraction solvent in this example.

[0028] Table 1 Oviposition deterrence activity of fecal extracts of different solvents against female Spodoptera frugiperda

[0029]

[0030] Table 2 Oviposition selection of female Spodoptera frugiperda adults for feed extracts of different solvents

[0031]

[0032] Example 2 Identification of Oviposition Deterrent Substances in the Feces of Spodoptera frugiperda

[0033] According to the method in Example 1, the feces of Spodoptera frugiperda larvae collected were extracted with dichloromethane for 24 h to form an extract with a concentration of 200 g / L. After the extract passed through a 0.22 μm PTFE filter membrane, it was subjected to component analysis by gas chromatography-mass spectrometry. The model of the gas chromatography was Agilent 7890B, and it was subsequently coupled with a mass spectrometer of model Agilent 7200. Column model: HP-5MS, detector was an ion detector (70 eV, 180 °C), and the carrier gas was helium. The injection volume was 1 μL, without splitting, and the temperature programming was to hold at 40 °C for 2 min, and then rise to 240 °C at a rate of 20 °C / min.

[0034] Since fatty acids are easily derivatized in the chromatographic column, resulting in inaccurate substance matching results, the extract was derivatized separately. The specific derivatization steps refer to the sample pretreatment method in the national standard "Determination of Fatty Acids in Foods" (GB 5009.168-2016) with slight modifications. Specifically: Weigh 400 mg of the sample into a stoppered test tube, add 4 mL of isooctane, let it stand for 1 h, shake it for 15 s every 15 min, then add 200 μL of 2% (w / v) potassium hydroxide methanol solution, cover the test tube with a glass stopper and shake it vigorously for 30 s, and then let it stand until it becomes clear. Add 1 g of sodium bisulfate and shake it vigorously. After the salt precipitates, the upper layer solution passes through a 0.22 μm PTFE filter membrane and waits to be injected into the instrument. The column model used in the gas chromatography-mass spectrometry is DB-WAX, the injection volume is 1 μL, without splitting, and the temperature programming: the initial temperature is 100 °C, hold for 13 min, rise to 180 °C at a rate of 10 °C / min, hold for 6 min, then rise to 200 °C at a rate of 1 °C / min, hold for 20 min, and finally rise to 230 °C at a rate of 4 °C / min, hold for 10.5 °C.

[0035] The mass-to-charge ratio of the substance was compared through the NIST database to clarify the molecular weight and basic structure of the substance. The analysis results are shown in Table 3.

[0036] Table 3 Identification Results of Compounds in the Feces of Spodoptera frugiperda

[0037]

[0038] Subsequently, standards of hexadecanoic acid, methyl hexadecanoate, Z-9-octadecenoic acid, methyl octadecanoate, (Z,Z)-9,12-octadecadienoic acid, and methyl Z-9-hexadecenoate were purchased from Sigma-aldrich for screening oviposition deterrent substances. Different standards were dissolved in dichloromethane respectively. A filter paper with a diameter of 15 cm was cut along the diameter. 2 mL of the solution was added to one half of the filter paper, and finally 0.1 mg of the test substance was formed on this half of the filter paper. The solvent (dichloromethane) was used as the control. The remaining procedures of the oviposition selection test were as described in Example 1.

[0039] The results are as Figure 1 shown. The results indicate that hexadecanoic acid has a significant effect of attracting the oviposition of Spodoptera frugiperda, while methyl Z-9-octadecenoate has a significant effect of deterring the oviposition of Spodoptera frugiperda. Other substances have no significant effect on the oviposition of female Spodoptera frugiperda.

[0040] Example 3 Oviposition Deterrence Activity Test of Different Doses of Methyl Z-9-octadecenoate

[0041] Methyl Z-9-octadecenoate (purchased from Sigma-aldrich) was dissolved in dichloromethane to prepare solutions with different concentrations. A filter paper with a diameter of 15 cm was cut along the diameter. 2 mL of the solution was added to one half of the filter paper (the area of one half of the filter paper is about 88 cm 2 ), and finally 0.01 mg, 0.1 mg, and 1 mg of the test doses were formed on this half of the filter paper. The solvent (dichloromethane) was used as the control. The remaining procedures and settings of the oviposition selection test were as described in Example 1.

[0042] The results are as Figure 2 shown. Methyl Z-9-hexadecenoate with a dose of 0.01 mg has no effect on the oviposition of female Spodoptera frugiperda, while 0.1 mg and 1 mg of methyl Z-9-octadecenoate have a significant effect of deterring the oviposition of Spodoptera frugiperda.

[0043] Example 4 Pot Experiment of Oviposition Deterrent Substances

[0044] The maize variety used in the experiment was Zhengdan 958. The seeds were first soaked for 8 h, and then placed in a petri dish with moist filter paper for germination. After the seeds germinated, they were sown in flower pots with a diameter of 9 cm. The maize seedlings used in the pot experiment for oviposition selection were 2 weeks old. Methyl Z-9-octadecenoate was dissolved in n-hexane to prepare a stock solution of 100 g / L, and diluted with 0.1% Triton-X aqueous solution (v:v) to the corresponding concentrations. 15 mL of the solution was sprayed on each maize plant, so that the final dose sprayed on each maize plant was 10 mg / plant, 100 mg / plant, and 1000 mg / plant. And 0.1% Triton-X aqueous solution of n-hexane (i.e., without adding methyl Z-9-octadecenoate) was used as a control. After standing for 30 min until the liquid on the maize leaves dried, the plants were placed in a nylon mesh cage (100 mesh) of 60*80*50 cm. One control maize plant and one maize plant treated with different concentrations of methyl Z-9-octadecenoate solution were placed diagonally in each cage. A petri dish containing 20% (w / v) sucrose solution was placed in the center of the bottom of the cage for adult nutrition supplementation. Subsequently, 5 pairs of Spodoptera frugiperda adults were placed in each cage, and the number of egg masses and eggs on each maize plant was counted after 48 h. Each cage was a replicate, and there were 6 replicates in total. The maize planting and pot experiment conditions were carried out in a greenhouse at a temperature of 26±2 °C, a relative humidity of 70%-90%, and a light-dark cycle of 16 h:8 h.

[0045] The results are as Figure 3 shown. The results showed that the oviposition amount on the maize plants sprayed with methyl Z-9-octadecenoate decreased significantly. Methyl Z-9-octadecenoate had a significant effect on repelling oviposition of female Spodoptera frugiperda at the doses of 100 mg / plant and 1000 mg / plant.

[0046] In summary, methyl Z-9-octadecenoate can regulate the oviposition behavior of female Spodoptera frugiperda both indoors and outdoors, and has the activity of repelling oviposition. Therefore, this substance can provide a new means for the integrated control of Spodoptera frugiperda.

[0047] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited by the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications made without departing from the spirit and principle of the present invention shall be equivalent substitution methods and are all included in the protection scope of the present invention.

Claims

1. Application of Z-9-octadecenoic acid methyl ester in the preparation of fall armyworm repellent.

2. The use according to claim 1, characterized in that: The repellent is an egg-laying repellent.

3. Application of Z-9-octadecenoic acid methyl ester as a repellent in controlling fall armyworm.

4. The use according to claim 3, characterized in that: The repellent is an egg-laying repellent.

5. The use according to claim 3 or 4, characterized in that: Z-9-octadecenoic acid methyl ester is dissolved in a solvent and then applied and / or sprayed on crops to repel fall armyworms from laying eggs.

6. The use according to claim 5, characterized in that: The dosage of the Z-9-octadecenoic acid methyl ester is smearing and / or spraying more than 11.4 mg of Z-9-octadecenoic acid methyl ester per square meter of crop leaves.

7. The use according to claim 6, characterized in that: The dosage of the Z-9-octadecenoic acid methyl ester is 11.4-114 mg of Z-9-octadecenoic acid methyl ester applied and / or sprayed per square meter of crop leaves.

8. The use according to claim 5, characterized in that: The solvent is at least one of dichloromethane and n-hexane.

9. The use according to claim 5, characterized in that: The crops include corn.

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