Application of a nitrogen-containing heterocyclic compound in preventing and treating golden apple snail

Abstract

The application belongs to the technical field of pest control, and discloses application of a nitrogen-containing heterocyclic compound in the prevention and treatment of apple snails. The application takes the arginine kinase of the apple snail as an action target, and screens out a nitrogen-containing heterocyclic compound with the activity of killing the apple snail, which is N-[4-(3,5-diphenyl-4,5-dihydro-1H-pyrazol-1-yl)benzyl]-N-(1-hexyl-1H-benzimidazol-2-yl)amine. The nitrogen-containing heterocyclic compound is prepared into a suspension agent or a granule as an active ingredient, and both of them have excellent killing effects on the apple snails. Compared with the existing arginine kinase inhibitor quercetin, the nitrogen-containing heterocyclic compound has higher activity, better selectivity, lower toxicity to vertebrates, and great application potential.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of pest control, and particularly relates to application of a nitrogen-containing heterocyclic compound in Pomacea canaliculata control. BACKGROUND

[0002] Pomacea canaliculata belongs to amphibious freshwater mollusks, and has very strong environmental adaptability. Different water environments such as rivers, ponds and paddy fields can be survived. In addition to threatening the ecological system of the invaded area, Pomacea canaliculata is also the intermediate host of Angiostrongylus cantonensis, which is easy to cause diseases due to the infection of Angiostrongylus cantonensis caused by eating uncooked and unripe Pomacea canaliculata. The influence of Pomacea canaliculata on agriculture is increasingly serious. At present, the management of Pomacea canaliculata still mainly relies on chemical agents. The long-term use of traditional chemical agents not only pollutes the environment, but also causes the continuous enhancement of the drug resistance of Pomacea canaliculata, which in turn promotes the continuous increase of the drug dosage, and enters a vicious circle. Therefore, it is particularly important to develop new Pomacea canaliculata killing agents from the target of the drug.

[0003] Studies have shown that by inhibiting or interfering with arginine kinase (AK), and then affecting energy metabolism, the purpose of killing or killing pests can be achieved. Arginine kinase is a phosphoarginine kinase widely distributed in invertebrates, which is directly related to energy transport, muscle contraction and ATP regeneration in insects. In addition to the above functions, arginine kinase also has the functions of participating in stress, maintaining immunity and paralyzing host insects. Since arginine kinase only exists in invertebrates, it is used as a target molecule in the control of agricultural pests. The lethal study of arginine kinase RNA interference has been verified in many insects such as wood lice, houseflies and red flour beetles.

[0004] Computer aided drug design (CADD) is a method of drug design based on computational chemistry, which simulates and predicts the interaction between small molecule compounds and target receptors to design and optimize lead compounds. With the continuous progress of computer science and the mutual penetration of quantum mechanics, molecular mechanics, molecular dynamics and pharmaceutical science, CADD technology has developed from basic theory research to a practical discipline. In recent years, CADD technology has made some progress in insecticides, molluscicides, bactericidal compounds, herbicides and other aspects. For example, Yang et al. took the PcRoo protein of apple snail as the receptor, and screened in the DrugBank database to obtain two target active compounds of baclofen and acetaminophen with molluscicidal activity. Chen et al. selected PcAdv and PcnWAS of apple snail as target proteins, and screened in the DrugBank database to finally obtain a compound potassium alginate with molluscicidal activity.

[0005] However, as an important enzyme existing in the body of apple snail, arginine kinase is rarely used as a target of killing compounds. Therefore, it is necessary to screen killing compounds with arginine kinase as the target of apple snail, so as to further realize the effective control of apple snail. SUMMARY

[0006] In order to further enrich the existing molluscicidal ingredients and realize the effective control of apple snail, the present application screens a nitrogen-containing heterocyclic compound with molluscicidal activity by using CADD technology. The present application specifically provides the following technical solutions.

[0007] Firstly, the present application provides an application of a nitrogen-containing heterocyclic compound in preventing and treating apple snail, wherein the nitrogen-containing heterocyclic compound is N-[4-(3,5-diphenyl-4,5-dihydro-1H-pyrazol-1-yl)benzyl]-N-(1-hexyl-1H-benzimidazol-2-yl)amine.

[0008] Further, in the application, the nitrogen-containing heterocyclic compound is used as an active ingredient for apple snail.

[0009] Further, in the application, the nitrogen-containing heterocyclic compound is prepared into a suspension or a granule.

[0010] Further, in the application, the concentration of the nitrogen-containing heterocyclic compound in the suspension is ≥3.8999 mg / L, and the content of the nitrogen-containing heterocyclic compound in the granule is ≥1 wt%.

[0011] Based on the above use, another aspect of the present application provides a method for controlling apple snail, which comprises: using the above-mentioned nitrogen-containing heterocyclic compound as an active ingredient for apple snail.

[0012] Further, in the method, the nitrogen-containing heterocyclic compound is prepared into a suspension agent or a granule.

[0013] Further, in the method, the concentration of the nitrogen-containing heterocyclic compound in the suspension agent is ≥ 3.8999 mg / L, and the content in the granule is ≥ 1 wt%.

[0014] Compared with the prior art, the application of the nitrogen-containing heterocyclic compound in the prevention and treatment of golden apple snails has the following beneficial effects:

[0015] The application takes the arginine kinase of the golden apple snail as a target, and uses the CADD technology to screen a nitrogen-containing heterocyclic compound with activity of killing the golden apple snail. After 24 h, 48 h and 72 h of immersion and killing treatment of the golden apple snail, the killing rate of the nitrogen-containing heterocyclic compound on the golden apple snail is 30%, 70% and 80% respectively, the corrected mortality is 30.00%, 66.67% and 77.78% respectively, and the LC 50 value of 72 h is 3.8999 mg / L, which has a significant effect.

[0016] Compared with the existing arginine kinase inhibitor quercetin, the nitrogen-containing heterocyclic compound provided by the application has higher activity, is low in toxicity to vertebrates, is safe, and has great potential for agricultural application. Specifically, after 24 h, 48 h and 72 h of treatment of the golden apple snail, the LC 50 value of the nitrogen-containing heterocyclic compound is 7.9919 mg / L, 5.3477 mg / L and 3.8999 mg / L respectively, which is lower than 319.2637 mg / L, 53.6744 mg / L and 30.4203 mg / L of quercetin respectively, and the killing effect is better.

[0017] The application has been proved by experiments that the nitrogen-containing heterocyclic compound has good effect on killing the golden apple snail in a field environment. Specifically, the average control effect of 1 wt% and 3 wt% nitrogen-containing heterocyclic compound granules in 14 d is 87% and 95% respectively, indicating that the field application prospect of the nitrogen-containing heterocyclic compound is good.

[0018] The application is of great significance for reducing the biting harm of the golden apple snail to aquatic crops such as rice, lotus, water bamboo, taro, water chestnut, water caltrop, gourd, and swamp cabbage, and avoiding the yield reduction of the aquatic crops caused by the harm of the golden apple snail. BRIEF DESCRIPTION OF DRAWINGS

[0019] Figure 1 The chemical structural formula of the nitrogen-containing heterocyclic compound is shown. DETAILED DESCRIPTION

[0020] The present application will be described below in connection with embodiments, and the technical solutions in the embodiments of the present application will be clearly and completely described. Obviously, the described embodiments are only some of the embodiments of the present application, but not all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative work fall within the scope of the present application.

[0021] In the following examples, the experimental methods are conventional methods, and are performed according to the techniques or conditions described in the literature in the art or according to the product instructions, unless otherwise specified. The materials, reagents and the like used in the following examples can be obtained from commercial channels, unless otherwise specified.

[0022] In the following examples, the nitrogen-containing heterocyclic compound is N-[4-(3,5-diphenyl-4,5-dihydro-1H-pyrazol-1-yl)benzyl]-N-(1-hexyl-1H-benzimidazol-2-yl)amine, which is purchased from specs company, with a purity of 99%, and its chemical structural formula is shown in Figure 1 .

[0023] Example 1

[0024] This example describes the screening and snail-killing activity of the nitrogen-containing heterocyclic compound.

[0025] 1. Test snails

[0026] The snails were artificially bred by the Pesticide Research Laboratory of Sichuan Agricultural University, and snails with a shell diameter of 5±3 mm were selected for the test.

[0027] 2. Screening method

[0028] Computer-aided drug design (CADD) technology was used for design: the tertiary structure of arginine kinase was constructed through the I-TASSER server; rigid docking was performed on the target protein by using Discovery Studio; and flexible docking was performed on the target protein by using Autodock4.0 to simulate and screen arginine kinase inhibitors of snails.

[0029] 3. Snail-killing test

[0030] The snail-killing effect of the nitrogen-containing heterocyclic compound on snails under different treatment times was determined by immersion killing, and the specific method was as follows:

[0031] The compound screened in step 2 is weighed out to prepare a 5 mL suspension at a concentration of 1 g / L. The compound content in the suspension is 5 mg, dimethyl sulfoxide accounts for 2% of the volume of the suspension, Tween 80 accounts for 1% of the volume of the suspension, and water is added to 100%. The suspension is diluted to 5 mg / L and added to a twelve-hole plate, 5 mL per hole. The test golden apple snails are then placed in the twelve-hole plate, 5 per hole, and water is used as a blank control instead of the diluent. The remaining treatment steps are the same. Each treatment is repeated 3 times, and the plates are placed in an incubator at a temperature of 25°C and a humidity of 80%. The holes are covered with gauze to prevent the golden apple snails from escaping. The mortality of the golden apple snails is checked at 24 h, 48 h, and 72 h, and the corrected mortality rate is calculated.

[0032] Corrected mortality rate = [(treatment group mortality rate - control group mortality rate) / (1 - control group mortality rate)] x 100%.

[0033] 4. Test results

[0034] The results of the killing effect of the nitrogen-containing heterocyclic compound on golden apple snails at different times are shown in Table 1.

[0035] Table 1. Corrected mortality rate of golden apple snails treated with the nitrogen-containing heterocyclic compound

[0036] Treatment time Treatment group mortality Control group mortality Corrected mortality 24h 30％ 0％ 30.00％ 48h 70％ 10％ 66.67％ 72h 80％ 10％ 77.78％

[0037] As can be seen from Table 1, under the condition of 25°C, the killing rate of the nitrogen-containing heterocyclic compound on the test golden apple snails after 24 h, 48 h, and 72 h of immersion treatment was 30%, 70%, and 80%, respectively, and the corrected mortality rate was 30.00%, 66.67%, and 77.78%, respectively. This shows that the nitrogen-containing heterocyclic compound has a significant killing effect on golden apple snails, and the killing effect after 48 h and 72 h of treatment is significantly better than that after 24 h of treatment.

[0038] Example 2

[0039] This example describes the comparison of the killing effect of the nitrogen-containing heterocyclic compound and quercetin (an existing arginine kinase inhibitor) on golden apple snails.

[0040] 1. The test golden apple snails are the same as in Example 1.

[0041] 2. Test method

[0042] The killing effect of the nitrogen-containing heterocyclic compound and quercetin on golden apple snails at different concentrations is determined by immersion method, and the specific process is as follows:

[0043] The compounds screened in step 2 of example 1 and quercetin were respectively prepared into 5 mL suspending agents with a concentration of 1 g / L, the compound content in the suspending agent was 5 mg, dimethyl sulfoxide accounted for 2% of the volume of the suspending agent, Tween 80 accounted for 1% of the volume of the suspending agent, and water was supplemented to 100%. And respectively diluted to 4 mg / L, 6 mg / L, 8 mg / L, 10 mg / L, 12 mg / L. The diluents were respectively added to twelve-hole plates, 5 mL was added to each hole. Then the test golden apple snails were respectively put into the twelve-hole plates, 5 were put into each hole, and water instead of the diluent was used as a blank control, and the remaining treatment steps were the same, each treatment was repeated 3 times. Placed in a incubator with a temperature of 25°C and a humidity of 80%, the hole opening was covered with gauze to prevent the golden apple snails from escaping, and the death of the golden apple snails was checked at 24 h, 48 h and 72 h. The toxicity regression equation was established, and the LC 50 values were calculated.

[0044] 3. Test results

[0045] The toxicity regression equation and LC 50 values of the nitrogen-containing heterocyclic compound and quercetin in killing golden apple snails are shown in Table 2.

[0046] Table 2. Comparison of the effects of the nitrogen-containing heterocyclic compound and quercetin in killing golden apple snails

[0047]

[0048]

[0049] From Table 2, it can be seen that after 24 h, 48 h and 72 h of treatment, the LC 50 values of quercetin on the test golden apple snails were 319.2637 mg / L, 53.6744 mg / L and 30.4203 mg / L, respectively, and the LC 50 values of the nitrogen-containing heterocyclic compound on the test golden apple snails were 7.9919 mg / L, 5.3477 mg / L and 3.8999 mg / L, respectively, which were all lower than that of quercetin. And the LC 50 values of the nitrogen-containing heterocyclic compound after 48 h and 72 h of treatment were significantly lower than the LC 50 values after 24 h of treatment. This result shows that the nitrogen-containing heterocyclic compound has higher activity and better killing effect on golden apple snails than quercetin, and the killing effect is significantly better after 48 h of treatment.

[0050] Example 3

[0051] This example describes the field test effect of the nitrogen-containing heterocyclic compound in killing golden apple snails.

[0052] 1. Test agent

[0053] The nitrogen-containing heterocyclic compound is prepared into granules, and the granule components are as follows: 1-6 wt% of the nitrogen-containing heterocyclic compound, 1 wt% of rapeseed oil, 2 wt% of sodium carboxymethyl cellulose (binder), 5 wt% of white sugar, 5% of butter flavor, and corn flour to make up to 100 wt%. The granules are prepared and provided by the Pesticide Research Laboratory of Sichuan Agricultural University.

[0054] 2. Test method

[0055] The test site is selected as clay with medium soil fertility, and the plot cultivation conditions are consistent. The plot area is 5 m 2 Each plot is transplanted with 100 clusters of seedlings, and each cluster has 5 plants. The four plots are isolated by plastic film, and the water depth is maintained at 5±2 cm. 7 days after transplantation, 100 apple snails with the same size are artificially raised in each plot. The granules containing 1 wt%, 3 wt%, and 6 wt% of the nitrogen-containing heterocyclic compound are uniformly applied in the water, and the control group is not treated. Each treatment has 3 replicates. 1 day, 3 days, 7 days, and 14 days after application, the mortality of apple snails in the plot is investigated, and the corrected mortality is calculated.

[0056] Corrected mortality = [(treatment mortality-control mortality) / (1-control mortality)] x 100%.

[0057] 3. Test results

[0058] As shown in Table 3, the 1 wt% nitrogen-containing heterocyclic compound granules and the 3 wt% nitrogen-containing heterocyclic compound granules have good field control effects on apple snails, and the average control effects are 87% and 95% respectively after 14 days. However, the mortality of apple snails is reduced when the 6 wt% nitrogen-containing heterocyclic compound granules are used, and the main reason is that apple snails refuse to eat.

[0059] Table 3. Field control effect of nitrogen-containing heterocyclic compound granules (%)

[0060]

[0061] In summary, the present application takes apple snail arginine kinase as the target, and screens a nitrogen-containing heterocyclic compound with apple snail killing activity, which is N-[4-(3,5-diphenyl-4,5-dihydro-1H-pyrazol-1-yl)benzyl]-N-(1-hexyl-1H-benzimidazol-2-yl)amine. The nitrogen-containing heterocyclic compound is prepared into a suspension or granules as an active ingredient, which has excellent killing effect on apple snails, and has higher activity, good selectivity, and low toxicity to vertebrates compared with the existing arginine kinase inhibitor quercetin, and has great application potential.

[0062] The above-described embodiments are merely some of the embodiments of the present application, but not all the embodiments. The detailed description of the embodiments of the present application is not intended to limit the scope of the claimed application, but merely represents selected embodiments of the application. All other embodiments obtained by persons of ordinary skill in the art based on the concept of the present application, without making creative labor, are within the scope of the present application.

Claims

1. The application of a nitrogen-containing heterocyclic compound in the control of golden apple snails, characterized in that, The nitrogen-containing heterocyclic compound is N-[4-(3,5-diphenyl-4,5-dihydro-1H-pyrazol-1-yl)benzyl]-N-(1-hexyl-1H-benzimidazol-2-yl)amine.

2. The application according to claim 1, characterized in that, The nitrogen-containing heterocyclic compound was used as an active ingredient in the golden apple snail.

3. The application according to claim 2, characterized in that, The nitrogen-containing heterocyclic compound is prepared into a suspension or granules.

4. The application according to claim 3, characterized in that, The concentration of the nitrogen-containing heterocyclic compound in the suspension is ≥3.8999 mg / L.

5. The application according to claim 3, characterized in that, The nitrogen-containing heterocyclic compound has a content of ≥1 wt% in the granules.

6. A method for controlling golden apple snails, characterized in that, The method includes applying the nitrogen-containing heterocyclic compound N-[4-(3,5-diphenyl-4,5-dihydro-1H-pyrazol-1-yl)benzyl]-N-(1-hexyl-1H-benzimidazol-2-yl)amine as an active ingredient to *Pomacea canaliculata*.

7. The method for controlling golden apple snails according to claim 6, characterized in that, The nitrogen-containing heterocyclic compound is prepared into a suspension or granules.

8. The method for controlling golden apple snails according to claim 7, characterized in that, The concentration of the nitrogen-containing heterocyclic compound in the suspension is ≥3.8999 mg / L.

9. The method for controlling golden apple snails according to claim 7, characterized in that, The nitrogen-containing heterocyclic compound has a content of ≥1 wt% in the granules.

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