Insecticide based on soybean trypsin inhibitor and application

Abstract

The invention belongs to the technical field of pesticide compounding, and particularly relates to a pesticide based on a soybean trypsin inhibitor and application, the effective components of the pesticide comprise a pesticide and a synergist, the pesticide is azadirachtin, and the synergist is the soybean trypsin inhibitor. In addition, the invention also provides an agricultural composition for preventing and treating lepidoptera pests, the effective components comprise azadirachtin and a soybean trypsin inhibitor, the effective concentration of the azadirachtin is 0.1289-0.3585 mg / L, and the effective concentration of the soybean trypsin inhibitor is 0.1 mM. The soybean trypsin inhibitor has the advantages that the insecticidal effect is obviously enhanced, the pesticide dosage is effectively reduced, the fatality rate of spodoptera frugiperda is effectively increased, the high insecticidal effect can be achieved under the condition of low-concentration azadirachtin, the chemical pesticide dosage is reduced, the drug resistance risk is relieved, and the pesticide reducing and synergistic requirements of green agriculture are met.

Description

Technical Field

[0001] The invention belongs to the technical field of pesticide compounding, and particularly relates to an insecticide based on soybean trypsin inhibitor and its application. Background Art

[0002] Soybean trypsin inhibitor is the main anti-nutritional factor in soybean. Two types have been isolated so far: Kunitz trypsin inhibitor (KTI) and Bowman-Birk trypsin inhibitor (BBI). KTI specifically inhibits the activity of trypsin, while BBI can inhibit the activity of trypsin and chymotrypsin.

[0003] Azadirachtin (Aza) is a limonoid active substance and a tetracyclic triterpenoid compound. It is primarily isolated and extracted from the neem plant Azadirachta indica A. Juss. It has excellent control effects on Hemiptera, Lepidoptera, and Coleoptera pests. It acts on the insect endocrine system, inhibiting brain neurosecretory cells to influence the synthesis and release of prothoracicotropic hormone (PTTH), and the synthesis and release of ecdysone (20-Hydroxyecdysone, 20E) by the prothoracic gland, thereby inhibiting larval molting and causing death.

[0004] The fall armyworm Spodoptera frugiperda, also known as the fall armyworm, belongs to the genus Spodoptera of the family Noctuidae in the order Lepidoptera. It originated in tropical and subtropical regions of the Americas and is one of the global migratory pests warned by the Food and Agriculture Organization of the United Nations. Summary of the Invention

[0005] The present invention aims to achieve "low-toxic, high-efficiency, and sustainable" pest control through the combination of natural ingredients and multi-target synergistic mechanisms, taking into account both ecological and economic benefits.

[0006] To achieve the above-mentioned object, the present invention provides an insecticide based on soybean trypsin inhibitor, the active ingredients of which include pesticide and synergist;

[0007] The synergist is soybean trypsin inhibitor, and the pesticide is azadirachtin.

[0008] In the above pesticides, the soybean trypsin inhibitor inhibits the activity of midgut proteases.

[0009] On the other hand, the present invention also provides use of the above insecticide in preventing and controlling lepidopteran pests.

[0010] In the above application, the lepidopteran pests include fall armyworm.

[0011] In addition, the present invention also provides an agricultural composition for controlling lepidopteran pests, wherein the active ingredients include pesticides and synergists;

[0012] The pesticide is azadirachtin, and the synergist is soybean trypsin inhibitor;

[0013] The effective concentration of the azadirachtin is 0.1289-0.3585 mg / L, and the effective concentration of the soybean trypsin inhibitor is 0.1 mM.

[0014] In the above agricultural composition, the soybean trypsin inhibitor inhibits the activity of midgut proteases.

[0015] In the above-mentioned agricultural composition, the lepidopteran pests include Spodoptera frugiperda.

[0016] In another aspect, the present invention also provides use of the agricultural composition in controlling lepidopteran pests.

[0017] In the above application, the lepidopteran pests include fall armyworm.

[0018] Compared with the prior art, the technical solution provided by the present invention has at least the following beneficial effects or advantages:

[0019] 1. The synergist provided by the present invention significantly enhances insecticidal efficacy and effectively reduces pesticide usage. By combining soybean trypsin inhibitor (SBTI) with azadirachtin, a synergistic effect is achieved. The combination of 1 mM SBTI with varying doses of azadirachtin significantly exceeded theoretical expectations for insecticidal efficacy, effectively increasing mortality against the fall armyworm (Spodoptera frugiperda). The combination achieves high insecticidal efficacy at low azadirachtin concentrations, reducing chemical pesticide usage and mitigating the risk of pesticide resistance, meeting the green agriculture demand for reduced pesticide use and increased synergy.

[0020] 2. The soybean trypsin inhibitor provided by the present invention targets and inhibits the digestive system of pests; the soybean trypsin inhibitor inhibits the activity of trypsin, chymotrypsin and total protease in the midgut of the fall armyworm, thereby destroying its nutrient absorption function, forming a dual-target synergy with the neuroendocrine disrupting effect of azadirachtin, thereby enhancing the persistence of insecticidal effects.

[0021] 3. The agricultural composition and insecticide provided by the present invention are environmentally friendly and highly safe; azadirachtin and soybean trypsin inhibitor are both natural substances of plant origin, have low toxicity to non-target organisms, reduce chemical pesticide residues, and are suitable for eco-friendly pest control systems; they achieve "low-toxic, high-efficiency, and sustainable" pest control, taking into account both ecological and economic benefits. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0023] Figure 1 This is a bar graph showing the toxicity of agricultural compositions containing different concentrations of azadirachtin and soybean trypsin inhibitor to Spodoptera frugiperda;

[0024] Figure 2 Effects of different concentrations of soybean trypsin inhibitor on the activity of total proteases in the midgut of Spodoptera frugiperda after 24h and 48h.

[0025] Figure 3 Effects of different concentrations of soybean trypsin inhibitor on the enzyme activity of midgut trypsin in Spodoptera frugiperda after 24h and 48h;

[0026] Figure 4 Effects of different concentrations of soybean trypsin inhibitor on the enzyme activity of chymotrypsin in the midgut of Spodoptera frugiperda after 24h and 48h. DETAILED DESCRIPTION

[0027] The technical solutions of the present invention are described below with reference to the following examples. However, the present invention is not limited to the following examples. The experimental and detection methods described in each example are conventional methods unless otherwise specified. The reagents and materials described are commercially available unless otherwise specified. The percentages in the following examples are percentages by mass unless otherwise specified. The ratios in the following examples are ratios by mass unless otherwise specified.

[0028] Example 1

[0029] This example is intended to illustrate the indoor toxicity assay of azadirachtin against Spodoptera frugiperda.

[0030] 0.3% emulsifiable concentrate of azadirachtin was purchased from Shandong Huimin Zhonglian Biotechnology Co., Ltd. and diluted with water as a solvent according to the working concentration and the content of the active ingredient.

[0031] In this embodiment, the feed mixing method is used for bioassay. First, an artificial feed for fall armyworm is prepared. The main ingredients of the artificial feed are corn flour and soybean flour. Before the artificial feed is completely cooled and solidified, the drug is mixed into the artificial feed and stirred evenly. After cooling, the feed is cut into small pieces. After the fall armyworm is starved for 2 hours, the artificial feed is placed in a 30 cm 3 Larvae were reared individually in round plastic boxes with five replicates per concentration, with 20 larvae per replicate. After 7 days, the number of dead S. frugiperda larvae was counted, and the regression equation was calculated using DPS 9.01 software. The results are shown in Table 1. The regression curve for the toxicity of azadirachtin to S. frugiperda larvae obtained through bioassay was: y = 2.8851x + 6.2855, with a correlation coefficient of 0.9391.

[0032] Table 1: Regression equation for the toxicity of azadirachtin to Spodoptera frugiperda

[0033] Pharmacy Regression equation Correlation coefficient <![CDATA[LC 50 (95% confidence limit) (mg / L)]]> P-value Azadirachtin y=2.8851x+6.2855 0.9391 0.3585 0.0017

[0034] Example 2

[0035] This example is intended to illustrate the effect of soybean trypsin inhibitor (SBTI) on the midgut protease activity of Spodoptera frugiperda.

[0036] Soybean trypsin inhibitor was purchased from Shanghai MacLean Biochemical Technology Co., Ltd., catalog number: 9035-81-8.

[0037] Soybean trypsin inhibitor of different concentrations was mixed into artificial feed and stirred evenly. Artificial feed with ddH2O added was used as control (CK group). After the fall armyworm was starved for 2 hours, the artificial feed was placed on a 30 cm 3 The larvae were reared individually in a plastic round box with 3 replicates for each concentration, and 50 larvae in each replicate. The midguts were dissected after 24h and 48h of feeding, and midgut protease extracts were prepared. The protein concentration was determined using a BCA protein quantification kit, and the activities of total protease, trypsin, and chymotrypsin in the midgut were determined using a total protease kit, a trypsin kit, and a chymotrypsin kit, respectively. The test results are shown in the figure. Figure 2 、 Figure 3 and Figure 4 shown.

[0038] Depend on Figure 2 、 Figure 3 and Figure 4 It can be seen that after feeding 0.2mM and 1mM soybean trypsin inhibitor for 48h, the total protease, trypsin and chymotrypsin in the midgut were significantly downregulated, so 0.2mM and 1mM soybean trypsin inhibitor can inhibit the activity of the three midgut proteases.

[0039] Example 3

[0040] This example is intended to illustrate the indoor toxicity test of the agricultural composition provided by the present invention against Spodoptera frugiperda.

[0041] The concentration LC 10 , LC 30 and LC 50 Azadirachtin was mixed with 0.2mM and 1mM soybean trypsin inhibitor, with a total of 6 compound combinations. The agents were mixed into the artificial feed before the artificial feed was completely cooled and solidified, and stirred evenly. After cooling, the feed was cut into small pieces. After the fall armyworm was starved for 2 hours, the artificial feed was placed 30cm 3 Each concentration was repeated 5 times, with 20 larvae in each repeat. After 7 days, the number of dead Spodoptera frugiperda larvae was counted and the adjusted mortality rate was calculated. The theoretical mortality rate and the actual adjusted mortality rate were calculated, and the synergistic toxicity index method was further used to determine the specific combined effect of the two agents. The test results are shown in Table 2 and Figure 1 shown.

[0042] Corrected mortality (%) = (treatment mortality - control mortality) / (1 - control mortality) x 100%.

[0043] Survival rate S (%) = (1-corrected mortality) × 100%.

[0044] Theoretical mortality (%) = (1-S azadirachtin × S soybean trypsin inhibitor) × 100%, S is the survival rate.

[0045] Synergistic toxicity index = (actual corrected mortality rate - theoretical mortality rate) / theoretical mortality rate × 100.

[0046] Criteria for determining synergistic effects: Theoretical and actual mortality rates were subjected to Student's t-test of variance analysis. A significantly higher actual mortality rate than the theoretical mortality rate indicated synergism (p < 0.05), no significant difference indicated no synergism (p ≥ 0.05), and a significantly lower actual mortality rate than the theoretical mortality rate indicated antagonism (p < 0.05). A synergistic toxicity index > 20 indicated synergism, a synergistic toxicity index of -20 ≤ 20 indicated additive effect, and a synergistic toxicity index < -20 indicated antagonism.

[0047] Table 2: Toxicity of different concentrations of azadirachtin and soybean trypsin inhibitor to Spodoptera frugiperda

[0048]

[0049] From Table 2 and Figure 1 It can be found that 0.2mM soybean trypsin inhibitor and azadirachtin LC 10 , LC 30 , LC 50After compounding, there was no significant difference between the theoretical mortality rate and the actual mortality rate of the mixed agent, so 0.2mM soybean trypsin inhibitor and azadirachtin LC 10 , LC 30 , LC 50 There was no synergistic effect (p<0.05). 1mM soybean trypsin inhibitor and azadirachtin LC 10 , LC 30 , LC 50 After compounding, 1mM soybean trypsin inhibitor and azadirachtin LC 10 The actual mortality rate of the mixed agent was significantly higher than the theoretical mortality rate (p < 0.05), 1mM soybean trypsin inhibitor and azadirachtin LC 30 Mixed agent, 1mM soybean trypsin inhibitor and azadirachtin LC 50 The actual mortality rate of the mixed agent was significantly higher than the theoretical mortality rate (p < 0.01), so 0.2mM soybean trypsin inhibitor and azadirachtin LC 10 , LC 30 , LC 50 There is a synergistic effect. 0.2mM soybean trypsin inhibitor and azadirachtin LC 10 , LC 30 , LC 50 The synergistic toxicity index of the combination is greater than 0 and less than 20, so 0.2mM soybean trypsin inhibitor and azadirachtin LC 10 , LC 30 , LC 50 The effect of the mixed agent after compounding is additive; 1mM soybean trypsin inhibitor and azadirachtin LC 10 , LC 30 , LC 50 The effect of the compounded mixture is synergistic. Figure 1 The results are consistent.

[0050] The above results show that 0.1mM soybean trypsin inhibitor can significantly enhance the synergy of azadirachtin in controlling fall armyworm.

[0051] As described above, the basic principles, main features and advantages of the present invention are well described. The above embodiments and descriptions are merely descriptions of preferred embodiments of the present invention, and the present invention is not limited to the above embodiments. Various changes and improvements made to the technical solutions of the present invention by those skilled in the art without departing from the spirit and scope of the present invention should fall within the scope of protection determined by the present invention.

Claims

1. A pesticide based on soybean trypsin inhibitor, characterized in that Active ingredients include pesticides and synergists; The synergist is soybean trypsin inhibitor, and the pesticide is azadirachtin.

2. The insecticide according to claim 1, characterized in that The soybean trypsin inhibitor inhibits the activity of midgut proteases.

3. Use of the insecticide according to claim 1 in controlling lepidopteran pests.

4. The use according to claim 3, characterized in that The lepidopteran pests include fall armyworm.

5. An agricultural composition for controlling lepidopteran pests, characterized in that: Active ingredients include pesticides and synergists; The pesticide is azadirachtin, and the synergist is soybean trypsin inhibitor; The effective concentration of the azadirachtin is 0.1289-0.3585 mg / L, and the effective concentration of the soybean trypsin inhibitor is 0.1 mM.

6. The agricultural composition according to claim 5, characterized in that The soybean trypsin inhibitor inhibits the activity of midgut proteases.

7. The agricultural composition according to claim 5, characterized in that The lepidopteran pests include fall armyworm.

8. Use of the agricultural composition according to claim 5 in controlling lepidopteran pests.

9. The use according to claim 8, characterized in that The lepidopteran pests include fall armyworm.

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