Helicoverpa armigera sterol absorption transport pathway inhibitor and application thereof

Abstract

The invention relates to the field of pest control, in particular to a cotton bollworm sterol absorption transport pathway inhibitor and application thereof. The compound is used for inhibiting the activity of cotton bollworm NPC1b protein. The inhibitor can obviously inhibit the growth and development of cotton bollworms, so that the death rate of the cotton bollworms is obviously increased; by influencing the expression of the NPC1b gene and interfering the absorption of sterol nutrition, a new action target and a candidate compound are provided for developing a novel pest control medicament.

Description

Technical Field

[0001] The invention relates to the field of pest control, and in particular to an inhibitor of sterol absorption and transport pathway of cotton bollworm and application thereof. Background Art

[0002] The cotton bollworm is an important pest that seriously harms crops. It has a wide range of hosts for a variety of crops, including food crops such as cotton, corn, and sorghum, and vegetable crops such as peppers, tomatoes, and beans. At present, the pest has developed varying degrees of resistance to a variety of conventional pesticides, which has seriously affected the production of crops.

[0003] In insects, cholesterol is an important component of cell membranes and participates in the regulation and maintenance of cell membrane fluidity. It is also an important precursor for the synthesis of bile acids, vitamin D, and steroid hormones. Insects cannot synthesize sterol nutrients from scratch because they lack key genes in the sterol synthesis pathway and must obtain this nutrition from food. One of the main pathways for insect intestines to absorb sterol nutrients is through transport mediated by the NPC1b (Nieman pick type C1b) protein.

[0004] Current studies have found that inhibitors that inhibit mammalian NPC1 proteins, such as Ezetimibe and U18666A, do not significantly inhibit the NPC1b protein of cotton bollworm. This indicates that there are significant differences in structure and function between insect NPC1b protein and mammalian NPC1 protein, and existing inhibitors against mammalian NPC1 protein cannot be effectively used for pest control.

[0005] Studies have shown that NPC1b protein is important for the normal growth, development and reproduction of cotton bollworm. However, no specific inhibitors for NPC1b protein of cotton bollworm have been developed so far. Existing control methods are difficult to effectively interfere with the sterol absorption pathway of cotton bollworm, resulting in unsatisfactory control effects. At the same time, the lack of inhibitors that specifically act on NPC1b protein of cotton bollworm also limits in-depth research on the application value of this target in pest control. Summary of the invention

[0006] The purpose of the present invention is to provide a compound as a sterol absorption and transport pathway inhibitor and its use, so as to provide a new technical solution for the prevention and control of agricultural pests such as cotton bollworm.

[0007] In order to solve the above technical problems, the present invention provides a compound for use as a sterol absorption and transport pathway inhibitor, wherein the compound is used to inhibit the activity of the NPC1b protein of cotton bollworm, and the chemical structure of the compound comprises:

[0008]

[0009]

[0010] Optionally, the compound is used at a concentration of 0.05-0.5 mg / mL.

[0011] Optionally, the compound is used at a concentration of 0.1 mg / mL.

[0012] The present invention also provides a pest control method, comprising the following steps:

[0013] Adding the above compounds to feed;

[0014] The feed is used to control pests.

[0015] Optionally, the pests include one or more of cotton bollworm, thrips, aphids, red spider moth, flea beetle, cabbage worm, scale insect, whitefly, European corn borer, spring looper, willow moth, diamondback moth, green stink bug, rice leaf roller, rice planthopper, bean pod borer, beet armyworm, Spodoptera litura, codling moth, mole cricket, grub, wireworm, cutworm, root maggot, root bug, root aphid, pseudo-ground beetle, cricket, root scale, root leaf beetle, root longhorn beetle, peach borer, root weevil, American spotted leafminer, watermelon leaf miner or termite.

[0016] The present invention also provides a method for preparing a pest control preparation containing an inhibitor, comprising the step of mixing the above compound with a carrier.

[0017] Optionally, the carrier comprises feed.

[0018] Optionally, the concentration of the compound in the feed is 0.05-0.5 mg / mL.

[0019] Compared with the prior art, the present invention has at least the following beneficial effects:

[0020] The compound provided by the present invention can specifically inhibit the activity of NPC1b protein of cotton bollworm, and achieve pest control by interfering with the absorption of sterol nutrition in the insect body. Bioassay experiments have shown that at a concentration of 0.1 mg / mL, the mortality rate of cotton bollworm after 14 days of treatment with the compound exceeds 70%, and the sterol content in the cotton bollworm body can be significantly reduced.

[0021] The compound can be applied by mixing with feed, is easy to use, and has a broad-spectrum pest control effect. At the recommended concentration, it shows good control effects on a variety of agricultural pests and can significantly increase the expression level of the target gene NPC1b, verifying its mechanism of action and providing a new solution for pest control. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is the virtual screening process of the lead compound in the embodiment of the present invention;

[0023] Figure 2a This is the survival curve of cotton bollworm larvae after feeding with R17 in the embodiment of the present invention;

[0024] Figure 2b This is the survival curve of cotton bollworm larvae after feeding with R20 and R21 in the embodiment of the present invention;

[0025] Figure 2c This is the survival curve of cotton bollworm larvae after feeding with R25 and R32 in the embodiment of the present invention;

[0026] Figure 3 Schematic diagram showing the effects of different inhibitors on the expression of NPC1b gene in cotton bollworm according to the embodiments of the present invention;

[0027] Figure 4 This is a schematic diagram of the analysis of the relative total sterol content in the intestine of cotton bollworm after being treated with different compounds in the examples of the present invention. DETAILED DESCRIPTION

[0028] The following will be described in more detail with reference to a schematic diagram of a sterol absorption and transport pathway inhibitor of cotton bollworm and its use, wherein preferred embodiments of the present invention are shown, and it should be understood that those skilled in the art can modify the present invention described herein while still achieving the beneficial effects of the present invention. Therefore, the following description should be understood as being widely known to those skilled in the art and not as a limitation of the present invention.

[0029] The present invention is described in more detail in the following paragraphs by way of example with reference to the accompanying drawings. The advantages and features of the present invention will become more apparent from the following description. It should be noted that the accompanying drawings are in very simplified form and are not in exact proportions, and are only used to facilitate and clearly assist in illustrating the purpose of the embodiments of the present invention.

[0030] Embodiment 1

[0031] The embodiment of the present invention describes in detail the process of obtaining NPC1b inhibitor compounds based on virtual screening, molecular docking, and screening and analysis processes.

[0032] S11. Acquisition of NPC1b inhibitor compounds based on virtual screening:

[0033] In a specific example, the sequence of the NPC1b_N domain of cotton bollworm was first predicted using the Pfam online database, and then the predicted NPC1b_N domain sequence of cotton bollworm was submitted to the AlphaFold2 software to simulate its three-dimensional structure.

[0034] The initial three-dimensional structure of the cotton bollworm NPC1b protein constructed by AlphaFold2 software was imported into SYBYL software, and Biopolymer>Prepare Struture>Struture Preparing Tool>Analyze Selected Struture was selected in the M1 area; and relevant parameters were set to perform a series of optimization operations on the protein structure, including repairing the side chain of residues, removing water, adding hydrogen and charge, protonation, and preliminary force field optimization. The grid file generated by the optimized protein three-dimensional structure was saved in a timely manner as a .maps.fld format file at the default pH value (7.0), and the cholesterol molecular structure was downloaded and saved as a .pdbqt format file.

[0035] The small molecule structures used for virtual screening in the embodiments of the present invention are all from the SPECS commercial database, and a total of 185,000 compound molecules are selected for virtual screening. First, the Filter tool in OpenEye is used to desalt and remove pan-assay interference compounds (PAINS) to clean the original database (Baell and Holloway 2010; Xie and Du 2015), and then ChemAxon's JfChem is used to predict the protonation state of small molecule compounds at physiological pH and generate reasonable tautomers. Finally, the Omage program of OpenEye is used to generate a three-dimensional conformation for docking. Through the above preliminary screening, compounds that do not conform to the three-dimensional structure of proteins are removed, and finally 80,000 candidate small molecule compounds for molecular docking are obtained.

[0036] S12. Molecular docking:

[0037] In the embodiment of the present invention, semi-flexible docking technology is used for molecular docking to keep the protein ligand structure stable and allow the orientation and structural changes of small molecule compounds. Under the premise of ensuring docking accuracy, the active box should cover the key residues around the active center as much as possible. The grid x, y, and z coordinate values ​​are set to 127, 127, and 127 respectively. The coordinates of the active center are The coordinates of the center of the active site box are set to The molecular docking module Surflex-Dock in SYBYL software was used to dock the ligand onto the receptor grid.

[0038] Specifically, software settings: Click Applications>Docking Suite>Dock Ligands in the toolbar, open the Docking dialog box and set the Docking Mode to Surflex-Dock Screen. The candidate small molecule compounds are docked to the active pocket center of the protein ligand molecule in sequence, and the docking results of the target protein and the small molecule compound are scored based on the search engine of molecular similarity and its unique empirical scoring function. At the same time, the software scoring results are sorted according to the scores. According to the binding mode and docking score of the docked ligand, it is analyzed and screened to obtain the compound small molecules with a total score greater than 6 with good energy and structure for molecular dynamics simulation, and its binding stability is analyzed. In the embodiment of the invention, Pymol software is used for visual analysis.

[0039] Import the 27,000 compounds processed by the SYBYL software screening mode into Autodock Vina, set the parameters of the active center site coordinates and the size of the active box in the toolbar grid-grid box, and execute the docking command with the relevant parameters such as the maximum energy value different from the optimal binding model. After the molecular docking is completed, use Pymol software to view the .pdbqt file of the docking results. 2 The small molecular compounds were filtered and the scores were less than -9 kcal / mol. 2 There are 1500 compounds with a total score greater than or equal to 8. 2 After molecular cluster analysis and visualization of 1024 small molecule compounds that meet the five docking models, 53 small molecule compounds were finally screened, and 5 compounds with better effects were obtained from the 53 small molecule compounds through biological screening. The specific screening process is as follows Figure 1 shown.

[0040] In this example, through molecular docking and screening analysis, the following five compounds with potential inhibitory activity were finally obtained. Specifically, the compounds include:

[0041] (1) 2-{[4-(4-methoxyphenyl)-6-trifluoromethyl-2-pyrimidinyl]thio}-N-(3-pyridylmethyl)acetamide; its structural formula is shown in Formula I:

[0042]

[0043] (2) N-{2-[4-(4-methoxyphenyl)-1-piperazinyl]-2-oxoethyl}-N-(2-phenylethyl)methanesulfonamide; its structural formula is shown in Formula II.

[0044]

[0045] (3) N-{2-[4-(1,3-benzodioxol-5-ylmethyl)-1-piperazinyl]-2-oxoethyl}-N-(3-fluorophenyl)methanesulfonamide; its structural formula is shown in Formula III.

[0046]

[0047] (4) N-[4-(1,1-dioxo-2-isothiazolidinyl)phenyl]-2-{[1-(2-methoxyethyl)-1H-indol-4-yl]oxy}acetamide; its structural formula is shown in Formula IV.

[0048]

[0049] (5) {2-[1-(4-methylphenyl)-5-oxo-3-pyrrolidinyl]-1H-benzimidazol-1-yl}acetic acid methyl ester, the structural formula of which is shown in Formula V.

[0050]

[0051]

[0052] It should be noted that the above five compounds are respectively referred to as R17, R20, R21, R25 and R32 below for the convenience of description and understanding.

[0053] In a specific example, the structures of the five compounds were confirmed by nuclear magnetic resonance hydrogen spectrum, carbon spectrum and mass spectrum, and the purity was greater than 95%.

[0054] Embodiment 2

[0055] The present invention describes in detail a method for preparing a feed containing an inhibitor, which comprises the following steps:

[0056] S21: Prepare the compound into a 0.1 mg / mL working solution. Specifically, use an appropriate amount of solvent to dissolve the compound to prepare a mother solution with a higher concentration, and then dilute it with deionized water to the desired concentration. In a specific example, the concentration of the working solution can be adjusted in the range of 0.05-0.5 mg / mL.

[0057] S22: Fully mix the prepared working solution with the cotton bollworm wheat bran feed. Specifically, spray the working solution evenly on the feed, stir evenly, and dry naturally. In a specific example, the feed can also be other conventional insect feeds, such as artificial feed.

[0058] Embodiment three:

[0059] The embodiment of the present invention provides a pest control method, comprising the following steps:

[0060] S31: adding the compound described in Example 1 to the feed and stirring evenly;

[0061] S32: feeding the feed to which the compound is added to the pests.

[0062] In the specific implementation process, the pests can be selected from a variety of agricultural pests, including but not limited to: cotton bollworm, thrips, aphids, etc. The compound is added to the feed at a concentration of 0.05-0.5 mg / mL, preferably 0.1 mg / mL.

[0063] The present invention also provides a method for preparing a pest control formulation containing the inhibitor, which specifically comprises:

[0064] The compound described in Example 1 is mixed with a carrier, wherein the carrier comprises feed, and the concentration of the compound in the feed is 0.05-0.5 mg / mL.

[0065] In a specific example, the following steps may be used for preparation:

[0066] Weigh an appropriate amount of compound;

[0067] Dissolve the compound in a suitable amount of solvent;

[0068] Add the dissolved compound solution evenly to the feed;

[0069] Mix thoroughly to prepare the pest control preparation of desired concentration.

[0070] The pest control method provided in this embodiment has the characteristics of simple operation and significant effect, and can effectively control the harm of agricultural pests; in addition, the preparation method provided in this embodiment is simple and feasible, and the obtained preparation has good practicality and insecticidal effect.

[0071] Embodiment 4

[0072] The examples of the present invention provide a specific method for conducting biological activity assays and specific experimental data of the five compounds provided in Example 1 to verify the effects of the five compounds on the growth and development of cotton bollworm.

[0073] S41. Test insect sources and breeding methods.

[0074] The wild-type cotton bollworm eggs used in the examples of the present invention were purchased from Jiyuan Baiyun Industrial Co., Ltd., Henan Province. All feeding experiments were conducted in an intelligent light incubator (14L: 10D) at a temperature of 25-28°C and a humidity of 30-40%. The hatched larvae were transferred to a 1.5mL centrifuge tube and fed with a freshly prepared normal cotton bollworm population feed and a compound-containing inhibitor feed, respectively, and a feed without an inhibitor was used as a control. When the larvae grow to the third instar larvae, they need to be transferred to a jelly box in the later stage to continue feeding with the inhibitor feed until pupation due to the enlargement of the insect body.

[0075] S42. Feed preparation method.

[0076] The small molecule compounds in the examples of the present invention are derived from the SPECS commercial database (Zoetermeer, Netherlands). The five compounds provided in Example 1 were added to the wheat bran feed of cotton bollworm at a concentration of 0.1 mg / mL and listed as experimental groups.

[0077] S43. Feed different inhibitors to observe the growth and development of cotton bollworm.

[0078] The control group's feed had no inhibitors, and other components and preparation methods were the same as those of the experimental group.

[0079] The bioassay experiment in the embodiment of the present invention was carried out in 7 batches, and each batch of feeding experimental control and each inhibitor treatment was set with 24 repetitions, and 1 worm was 1 repetition.

[0080] When feeding, use tweezers to stick a small amount of feed to the wall of a 1.5mL centrifuge tube (the tube cover is pierced with an insect pin to keep it breathable), then use a brush to gently pick up a single-headed cotton bollworm newly hatched larvae onto the feed and place it in an intelligent light incubator for cultivation.

[0081] Before the cotton bollworm grows to the third instar, observe it once a day; from the third instar to before pupation, observe it once in the morning and evening every day. Pay attention to adding feed and controlling humidity during the entire growth and development stage, and record its age and survival. Statistics and collation of data such as survival rate, larval development period, pupation rate, etc., weigh the pupae 24 hours after pupation and record the pupal weight data. The male and female adults after eclosion are placed in a 10×12cm round plastic bottle for single pair pairing, and the adults are fed with 10% honey water. During the egg-laying period, the egg-laying cloth is used to seal the bottle and the egg-laying situation is observed every day. The development period, eclosion rate, egg laying amount, and egg hatching rate of the pupae are recorded, and the egg-laying period and adult life span and other growth and development indicators are recorded at the end of egg-laying of female adults and when the adults die.

[0082] S44. RT-qPCR detection of NPC1b gene expression in cotton bollworm after treatment with different inhibitors.

[0083] The five compounds screened out by the bioassay in Example 1 were fed and then sampled to determine the effect of each group of compounds on the expression of the NPC1b gene of cotton bollworm.

[0084] It should be noted that when the feed contains R17, R20, R21, R25 and R32 compounds, the first-instar larvae of cotton bollworm cannot grow to the third instar. Therefore, this experiment used cotton bollworm larvae that had just molted to the second instar, fed on the feed containing the compound until the second day of the fourth instar, and dissected the intestines and sampled. Every three intestines were used as a replicate, and a total of 6 biological replicates were taken. The control group was sampled in the same way. RNA was extracted and reversed to cDNA.

[0085] The expression of candidate genes was detected by fluorescence quantitative PCR using the Beijing Polymer Company kit Realtime PCR Super mix SYBR green with anti-Taq (Cat: MF013-01). (-ΔΔCt) The relative gene expression was calculated.

[0086] S45. Determine the changes in sterol content in cotton bollworms fed with 5 highly active inhibitors.

[0087] The five bioactive compounds provided in Example 1 were added to cotton bollworm feed, and the feed was used for cotton bollworm larvae that had just molted to the second instar, so as to determine the effects of these compounds on the sterol content in the cotton bollworm.

[0088] The larvae of cotton bollworm that had just molted to the second instar were fed with feed containing inhibitors until the second day of the fourth instar, and then starved for 2 hours. After feeding with feed without sterols and inhibitors for 6 hours, the weight of the larvae was weighed, and the intestines of cotton bollworm larvae were dissected in pre-cooled PBS buffer for sampling. Every 5 intestines were used as a replicate, and a total of 5 biological replicates were taken, and the same sampling treatment was performed on the control. After the dissected intestinal samples were placed in a vacuum freeze dryer for freeze drying for 2 days, the dry weight of the samples was weighed with a one-millionth balance for extraction and detection of the sterol content in the samples. The entire sterol extraction process was carried out in a biosafety fume hood. Data acquisition was performed using a gas chromatograph-mass spectrometer (GC-MS, Thermo Scientific, USA).

[0089] S46. Data processing.

[0090] For the treatments that failed to develop into pupae after feeding with the compound, the Log-rank test method was used for survival curve analysis. The survival rate and growth and development of other compound treatments were analyzed for significant differences using independent sample t-tests and plotted using GraphPad Prism 9.3.0 (GraphPad Software_USA).

[0091] The data of NPC1b gene expression and sterol content in cotton bollworm after compound feeding treatment were analyzed for significant differences using independent sample t test, and the relative total sterol content was analyzed by LSD and Duncan of one-way analysis of variance (ANOVA). All data are shown as Mean ± SEM and analyzed and plotted using GraphPad Prism 9.3.0 (GraphPad Software_USA). The letters above the columns indicate the differences between different treatments. Different letters indicate differences between treatments, and the same letters indicate no significant differences. Asterisks represent significant differences between different treatments (*p<0.05, **p<0.01, ***p<0.001), and ns indicates no statistical difference between two treatments.

[0092] S47. Experimental results.

[0093] Please refer to the growth and development of cotton bollworm fed with different inhibitors Figure 2a-2c , Figure 2a The survival rate of the R17-treated group dropped to about 25% (i.e., mortality rate of 75%) at about 14 days. Figure 2b : The survival rate of the R20-treated group dropped to about 10% (i.e., mortality rate of 90%) at about 14 days; Figure 2b : The survival rate of the R21-treated group dropped to about 20% (i.e., mortality rate 80%) at about 14 days; Figure 2c : The survival rate of the R25 and R32 treatment groups dropped to about 20% (ie, mortality rate 80%) at about 14 days.

[0094] Among them, the mortality rate of the R20 treatment group was as high as 90% at 14 days, and that of the R21, R25, and R32 treatment groups was 80%.

[0095] The RT-qPCR method was used to detect the changes in the expression level of the NPC1b gene. The relative expression levels of the NPC1b gene in cotton bollworm after treatment with different high-efficiency inhibitors detected by RT-qPCR are shown in Table 1 below:

[0096] Table 1: Relative expression levels of NPC1b gene in cotton bollworm after treatment with five different inhibitors

[0097]

[0098] Please refer to the effects of 5 different inhibitors on the expression of NPC1b gene in cotton bollworm Figure 3 The results showed that compared with the control group, the NPC1b gene expression levels in the five compound treatment groups provided in Example 1 were significantly upregulated, with the maximum upregulation reaching 129%. Specifically:

[0099] The 6 replicate values ​​of the Control group (CK group) are:

[0100] 1.951898454, 0.500513308, 1.445105012, 0.967543882, 0.769714857 and 0.951102236.

[0101] The 6 replicate values ​​of the R21 treatment group were: 1.912992654, 3.135169261, 2.058557861, 4.665411492, 2.057519827 and 1.265146102.

[0102] Calculation shows that:

[0103] The average expression level in the CK group was approximately 1.098.

[0104] The average expression level of the R21 group was approximately 2.516.

[0105] Calculation of the increase: (2.516-1.098) / 1.098*100%≈129%.

[0106] The sterol content in the insects was determined by gas chromatography-mass spectrometry (GC-MS). The changes in sterol content in the cotton bollworm fed with 5 highly active inhibitors are shown in Table 2 below:

[0107] Table 2

[0108]

[0109]

[0110] For specific data on the relative total sterol content in the intestine of cotton bollworm after treatment with different compounds, please refer to Figure 4 .

[0111] The results showed that the sterol content in the insects of the five compound treatment groups provided in Example 1 was significantly lower than that of the control group, verifying the mechanism of action of the five compounds in Example 1 to exert insecticidal effects by interfering with the sterol absorption and transport pathway.

[0112] In the examples of the present invention, the mechanism of action of the compound was verified by molecular biology and metabolomics analysis, providing a scientific basis for further development and optimization of this type of compound.

[0113] In summary, the above data show that the five compounds provided in Example 1 have good insecticidal activity:

[0114] 1) The five compounds significantly affect the growth and development of cotton bollworm, preventing it from developing normally to the third instar;

[0115] 2) The mortality rate of cotton bollworm exceeds 70% within 14 days;

[0116] 3) The surviving larvae were significantly smaller than those in the control group and their development period was significantly prolonged.

[0117] Embodiment 5

[0118] Efficacy range test: The five compounds provided in Example 1 were prepared into feed containing inhibitors according to the method of Example 2, and the control effects on other agricultural pests were tested respectively. The results showed that the compounds showed good control effects on various agricultural pests such as thrips and aphids.

[0119] The examples of the present invention prove that the compound has a broad-spectrum insecticidal activity and can be used to control a variety of agricultural pests. In addition, the present invention adopts a feeding mixing application method, which is simple to operate and easy to promote and apply.

[0120] In summary, the five compounds screened by the present invention have a significant growth inhibitory effect on cotton bollworm, which can significantly increase its mortality rate; these compounds can affect the expression level of the NPC1b gene of cotton bollworm, thereby affecting its absorption of sterol nutrition; the preparation and use methods of the compounds are simple and have good practicality; the present invention provides new targets and candidate compounds for the development of new pest control agents.

[0121] Obviously, those skilled in the art can make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. A use of a compound as a sterol absorption and transport pathway inhibitor, characterized in that: The compound is used to inhibit the activity of NPC1b protein of cotton bollworm, and the chemical structure of the compound includes:

2. The use according to claim 1, characterized in that The compound is used at a concentration of 0.05-0.5 mg / mL.

3. The use according to claim 2, characterized in that: The compounds were used at a concentration of 0.1 mg / mL.

4. A method for controlling pests, characterized in that: The following steps are involved: Adding the compound according to any one of claims 1 to 3 to feed; The feed is used to control pests.

5. The method according to claim 4, characterized in that The pests include one or more of cotton bollworm, thrips, aphids, red spider moth, flea beetle, cabbage worm, scale insect, whitefly, European corn borer, spring looper, willow moth, diamondback moth, green stink bug, rice leaf roller, rice planthopper, bean pod borer, beet armyworm, Spodoptera litura, codling moth, mole cricket, grub, wireworm, cutworm, root maggot, root bug, root aphid, pseudo-ground beetle, cricket, root scale, root leaf beetle, root longhorn beetle, peach borer, root weevil, American spotted leafminer, watermelon leaf miner or termite.

6. A method for preparing a pest control preparation containing an inhibitor, characterized in that: The method comprises the step of mixing the compound according to any one of claims 1 to 3 with a carrier.

7. The preparation method according to claim 6, characterized in that: The carrier includes feed.

8. The preparation method according to claim 7, characterized in that: The concentration of the compound in the feed is 0.05-0.5 mg / mL.

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