Norcantharidin derivative as well as preparation method and application thereof

Abstract

The invention discloses a norcantharidin derivative as well as a preparation method and application thereof, and belongs to the field of organic synthesis. Chiral modification is performed on the oxygen ring on the left side, so that the bactericidal performance of the derivative is improved. Experiments show that part of the series of norcantharidin derivatives have better bactericidal activity on sclerotinia sclerotiorum and botrytis cinerea, and the activity is remarkably improved compared with that of parent norcantharidin, so that the norcantharidin derivatives are expected to be used for preparing efficient, environment-friendly and low-toxicity bactericides.

Description

Technical Field

[0001] The invention belongs to the field of organic synthesis, and specifically relates to a norcantharidin derivative and a preparation method and application thereof. Background Art

[0002] Cantharidin (CTD) is a compound with a six-membered oxygen bridge ring structure extracted from the traditional Chinese medicine Mylabris. The earliest existing monograph on Chinese medicine, Shennong's Herbal Classic, has recorded Mylabris. It is cold in nature, pungent in taste, and highly toxic. Its effects are mainly to dissipate lumps and reduce swelling, break blood and remove blood stasis, and attack toxins and ulcers. In terms of pesticides, it has insecticidal, bactericidal and algaecidal activities. It is of great significance to modify cantharidin with high enantioselectivity. Summary of the invention

[0003] The present invention aims to provide a norcantharidin derivative and a preparation method and application thereof. Experiments show that some of the norcantharidin derivatives of the present invention have good fungicidal activity against Brassica sclerotiorum and Botrytis cinerea, and the activity is significantly improved compared with the parent norcantharidin, so it is expected to be used to prepare a highly efficient, environmentally friendly, low-toxic fungicide.

[0004] The norcantharidin derivatives of the present invention have a general structural formula as shown in the following formula I:

[0005]

[0006] in:

[0007] R1 is selected from one of alkyl, alkanoyl, aryl-substituted alkyl, heterocycle-substituted alkyl and halogen-substituted alkyl; R2 is selected from phenyl or substituted phenyl, and the substituent of the substituted phenyl is halogen.

[0008] Further, R1 and R2 are selected from the following groups:

[0009]

[0010] Furthermore, compounds 6, 7, 8, and 13 are preferred, and compound 7 is most preferred.

[0011] The preparation method of the norcantharidin derivatives of the present invention comprises the following steps:

[0012] Step 1: reacting norcantharidin with substituted aniline to obtain intermediate a;

[0013]

[0014] R represents monohalogen or dihalogen substitution.

[0015] Step 2: The intermediate a is subjected to a metal-catalyzed hydroalkylation reaction to obtain the target product represented by formula I.

[0016] Furthermore, in step 1, in the presence of a catalyst, norcantharidin and substituted aniline are subjected to an imidization reaction in DCM, and after the reaction is completed, the intermediate a is separated by column chromatography.

[0017] In step 1, the catalyst is one or more of DCC and EDCI.

[0018] In step 1, the reaction temperature is 0-5°C.

[0019] In step 1, the substituted aniline is selected from aromatic amine structures substituted with monohalogen or dihalogen at positions 2, 3, 4, and 5, and its structure is shown below:

[0020] .

[0021] Further, in step 2, the intermediate a, alkyl iodide, catalyst, ligand and base obtained in step 1 are placed in a Shrek tube, and the inert gas is exchanged three times. A solvent and a hydrogen source are added thereto under the protection of inert gas, and the reaction is carried out at a certain temperature for 1-12 hours.

[0022] The alkyl iodide is selected from one of methyl iodide, deuterated methyl iodide, ethyl iodide, n-propane iodide and n-butane iodide.

[0023] The solvent is selected from one of ethylene glycol dimethyl ether, ethyl acetate and tetrahydrofuran.

[0024] In step 2, the catalyst is selected from one of CoCl2, CoBr2, CoBr2(DME), CoI2, Co(acac)2 or CoF3.

[0025] In step 2, the ligand is selected from one of the following ligands L1-L18:

[0026] .

[0027] In step 2, the reaction temperature is -40 to 40°C, preferably -20 to 0°C.

[0028] In step 2, the base is a combination of one or more of CsF, KF, K3PO4, NaHCO3, KH2PO4, Cs2CO3, Na2CO4, KHCO3, and K2CO3, and more preferably CsF and K3PO4.

[0029] In step 2, the hydrogen source is a combination of one or more of DEMS, (MeO)3SiH, (MeO)2MeSiH, and PHMS.

[0030] The invention discloses an application of the norcantharidin derivatives in preparing a preparation for inhibiting plant pathogenic bacteria.

[0031] The plant pathogens include common rapeseed sclerotinia, tomato gray mold, etc. The demethylcantharidin derivatives of the present invention have good fungicidal activity against rapeseed sclerotinia, tomato gray mold, etc.

[0032] It should be noted that the modification of cantharidin and norcantharidin in the past was on the anhydride ring on the right side of the norcantharidin structure. The present invention is different from the above-mentioned prior art, but improves the bactericidal properties of the derivatives by chiral modification of the oxygen ring on the left side. Among them, compound 7 obtained by methylation and deuterated methylation has excellent antibacterial activity. Its antibacterial effect is 2 times higher than that of the commercially available drug procymidone and about 40 times higher than that of its skeleton structure.

[0033] In addition, the methylation of existing structures is not a simple matter. For example, there is no precedent for methylation of the six-membered oxygen-containing ring in the structure of norcantharidin. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 This is the H NMR spectrum of compound 6.

[0035] Figure 2 This is the H NMR spectrum of compound 7.

[0036] Figure 3 This is the H NMR spectrum of compound 8.

[0037] Figure 4 Pymol2.1 was used to visualize the complex of protein and small molecule. Among them: Figure A is the molecular diagram of compound 7, Figure B is the spatial 3D structure of HK protein, Figure C is the electron cloud density simulation diagram after compound 7 is embedded in HK protein, and Figure D is a schematic diagram of the interaction between compound 7 and different amino acid residues in HK protein.

[0038] Figure 5 The mycelial morphology of S. sclerotiorum was observed by scanning electron microscopy after culturing on PDA medium containing 1.0 mg / L compound 7 for 48 hours. a and b are the SEM morphologies of mycelium at different magnifications of the blank group; c and d are the SEM morphologies of mycelium at different magnifications after treatment with 1 ppm of the drug.

[0039] Figure 6 This is the in vitro activity experiment of compound 7 against Sclerotinia sclerotiorum of rapeseed (100 mg / L and 200 mg / L).

[0040] The present invention is further described in detail below through the accompanying drawings and embodiments given by the inventor. DETAILED DESCRIPTION

[0041] Unless otherwise specified, the terms used herein are understood according to those of ordinary skill in the relevant art.

[0042] The synthetic route of the derivatives of the present invention is as follows:

[0043] Embodiment 1:

[0044]

[0045] 1. Weigh 10 mmol of nordehydrocantharidin and 7.5 mmol of aniline, dissolve in 100 mL of DCM, stir for 30 min in an ice bath, add 10 mmol of EDCI, stir at room temperature for 12 hours, and detect by TLC; after the reaction is complete, remove DCM, dry, concentrate, and separate by column chromatography. The eluent is PE:EA=2:1 (V / V) to obtain intermediate a1 with a yield of 60%.

[0046] 2. Weigh intermediate a1 (0.1 mmol), iodomethane (0.5 mmol), CoBr2(DME) 0.01mmol, ligand L10.012mmol, CsF 0.1mmol, K3PO4 0.2mmol, DME 0.8mL, DEMS 0.3mmol, react at -20℃ for 12h, and separate by thin layer chromatography to obtain compound 1 as a white solid with a yield of 60% and an ee value of 94%.

[0047] Embodiment 2:

[0048]

[0049] 1. For the preparation of intermediate a1, see Example 1.

[0050] 2. Weigh intermediate a1 (0.1 mmol), deuterated iodomethane (0.5 mmol), CoBr2(DME) 0.01mmol, ligand L1 0.012mmol, CsF 0.1mmol, K3PO4 0.2mmol, DME 0.8mL, DEMS 0.3mmol, react at -20℃ for 12h, and separate by thin layer chromatography to obtain compound 2 as a white solid with a yield of 62% and an ee value of 94%.

[0051] Embodiment 3:

[0052]

[0053] 1. For the preparation of intermediate a1, see Example 1.

[0054] 2. Intermediate a1 (0.1 mmol), iodoethane (0.5 mmol), CoBr2(DME) 0.01mmol, ligand L10.012mmol, CsF 0.1mmol, K3PO4 0.2mmol, DME 0.8mL, DEMS 0.3mmol, react at -20℃ for 12h, and separate by thin layer chromatography to obtain compound 3 as a white solid with a yield of 61% and an ee value of 92%.

[0055] Embodiment 4:

[0056]

[0057] 1. For the preparation of intermediate a1, see Example 1.

[0058] 2. Intermediate a1 (0.1 mmol), iodinated n-propane (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 4 as a white solid with a yield of 65% and an ee value of 92%.

[0059] Embodiment 5:

[0060]

[0061] 1. For the preparation of intermediate a1, see Example 1.

[0062] 2. Intermediate a1 (0.1 mmol), n-butyl iodide (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 5 as a white solid with a yield of 60% and an ee value of 90%.

[0063] Embodiment 6:

[0064]

[0065] 1. Weigh 10 mmol of nordehydrocantharidin and 7.5 mmol of 3-F-aniline and dissolve them in 100 mL of DCM. Stir for 30 min in an ice bath. Add 10 mmol of EDCI and stir at room temperature for 12 hours. Detect by TLC. After the reaction is complete, remove DCM, dry, concentrate, and separate by column chromatography to obtain intermediate a2 with a yield of 65%.

[0066] 2. Weigh intermediate a2 (0.1 mmol), iodomethane (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 6 as a white solid with a yield of 66% and an ee value of 95%. 1 H NMR (400 MHz, Chloroform-d) δ7.61-7.32 (m, 1H), 7.16-6.90 (m, 3H), 4.97 (d, J = 5.3 Hz, 1H), 4.58 (s, 1H),3.35-2.78 (m, 2H), 2.13-1.97 (m, 1H), 1.98-1.79 (m, 1H), 1.48-1.38 (m, 1H), 1.07 (d, J = 6.9 Hz, 3H).

[0067] Embodiment 7:

[0068]

[0069] 1. For the preparation of intermediate a2, see Example 6.

[0070] 2. Weigh intermediate a2 (0.1 mmol), deuterated iodomethane (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 7 as a white solid with a yield of 50% and an ee value of 95%. 1 H NMR (400 MHz, Chloroform-d) δ7.50-7.36 (m, 1H), 7.16-6.95 (m, 3H), 4.97 (d, J = 5.3 Hz, 1H), 4.58 (s, 1H),3.27-2.60 (m, 2H), 2.11-1.97 (m, 1H), 1.95-1.79 (m, 1H), 1.46-1.37 (m, 1H).

[0071] Embodiment 8:

[0072]

[0073] 1. For the preparation of intermediate a2, see Example 6.

[0074] 2. Intermediate a2 (0.1 mmol), iodoethane (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 8 as a white solid with a yield of 45% and an ee value of 90%. 1 H NMR (400 MHz, Chloroform-d) δ7.55-7.35 (m, 1H), 7.15-6.94 (m, 3H), 4.95 (d, J = 5.3 Hz, 1H), 4.69 (s, 1H),3.03 (d, J = 1.7 Hz, 2H), 1.94-1.74 (m, 2H), 1.58-1.41 (m, 2H), 1.37-1.16 (m,1H), 0.93 (t, J = 7.4 Hz, 3H).

[0075] Embodiment 9:

[0076]

[0077] 1. For the preparation of intermediate a2, see Example 6.

[0078] 2. Intermediate a2 (0.1 mmol), iodinated n-propane (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 9 as a white solid with a yield of 45% and an ee value of 88%.

[0079] Embodiment 10:

[0080]

[0081] 1. For the preparation of intermediate a2, see Example 6.

[0082] 2. Intermediate a2 (0.1 mmol), n-butyl iodide (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 10 as a white solid with a yield of 42% and an ee value of 87%.

[0083] Embodiment 11:

[0084]

[0085] 1. Weigh 10 mmol of nordehydrocantharidin and 7.5 mmol of 3,4-difluoroaniline and dissolve them in 100 mL of DCM. Stir for 30 min in an ice bath. Add 10 mmol of EDCI and stir at room temperature for 12 hours. Detect by TLC. After the reaction is complete, remove DCM, dry, concentrate, and separate by column chromatography to obtain intermediate a3 with a yield of 61%.

[0086] 2. Intermediate a3 (0.1 mmol), iodomethane (0.5 mmol), CoBr2(DME) 0.01mmol, ligand L10.012mmol, CsF 0.1mmol, K3PO4 0.2mmol, DME 0.8mL, DEMS 0.3mmol, react at -20℃ for 12h, and separate by thin layer chromatography to obtain compound 11 as a white solid with a yield of 56% and an ee value of 90%.

[0087] Embodiment 12:

[0088]

[0089] 1. For the preparation of intermediate a3, see Example 11.

[0090] 2. Intermediate a3 (0.1 mmol), deuterated iodomethane (0.5 mmol), CoBr2(DME) 0.01mmol, ligand L10.012mmol, CsF 0.1mmol, K3PO4 0.2mmol, DME 0.8mL, DEMS 0.3mmol, react at -20℃ for 12h, and separate by thin layer chromatography to obtain compound 12 as a white solid with a yield of 54% and an ee value of 90%.

[0091] Embodiment 13:

[0092]

[0093] 1. For the preparation of intermediate a3, see Example 11.

[0094] 2. Intermediate a3 (0.1 mmol), iodoethane (0.5 mmol), CoBr2(DME) 0.01mmol, ligand L10.012mmol, CsF 0.1mmol, K3PO4 0.2mmol, DME 0.8mL, DEMS 0.3mmol, react at -20℃ for 12h, and separate by thin layer chromatography to obtain compound 13 as a white solid with a yield of 58% and an ee value of 91%.

[0095] Embodiment 14:

[0096]

[0097] 1. For the preparation of intermediate a3, see Example 11.

[0098] 2. Intermediate a3 (0.1 mmol), iodinated n-propane (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 14 as a white solid with a yield of 58% and an ee value of 86%.

[0099] Embodiment 15:

[0100]

[0101] 1. For the preparation of intermediate a3, see Example 11.

[0102] 2. Intermediate a3 (0.1 mmol), n-butyl iodide (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 15 as a white solid with a yield of 55% and an ee value of 85%.

[0103] Embodiment 16:

[0104]

[0105] 1. Weigh 10 mmol of nordehydrocantharidin and 7.5 mmol of 3-chloroaniline and dissolve them in 100 mL of DCM. Stir for 30 min in an ice bath. Add 10 mmol of EDCI and stir at room temperature for 12 hours. Detect by TLC. After the reaction is complete, remove DCM, dry, concentrate, and separate by column chromatography to obtain intermediate a4 with a yield of 63%.

[0106] 2. Intermediate a4 (0.1 mmol), iodomethane (0.5 mmol), CoBr2(DME) 0.01mmol, ligand L10.012mmol, CsF 0.1mmol, K3PO4 0.2mmol, DME 0.8mL, DEMS 0.3mmol, react at -20℃ for 12h, and separate by thin layer chromatography to obtain compound 16 as a white solid with a yield of 60% and an ee value of 92%.

[0107] Embodiment 17:

[0108]

[0109] 1. For the preparation of intermediate a4, see Example 16.

[0110] 2. Intermediate a4 (0.1 mmol), deuterated iodomethane (0.5 mmol), CoBr2(DME) 0.01mmol, ligand L10.012mmol, CsF 0.1mmol, K3PO4 0.2mmol, DME 0.8mL, DEMS 0.3mmol, react at -20℃ for 12h, and separate by thin layer chromatography to obtain compound 17 as a white solid with a yield of 55% and an ee value of 92%.

[0111] Embodiment 18:

[0112]

[0113] 1. For the preparation of intermediate a4, see Example 16.

[0114] 2. Intermediate a4 (0.1 mmol), iodoethane (0.5 mmol), CoBr2(DME) 0.01mmol, ligand L10.012mmol, CsF 0.1mmol, K3PO4 0.2mmol, DME 0.8mL, DEMS 0.3mmol, react at -20℃ for 12h, and separate by thin layer chromatography to obtain compound 18 as a white solid with a yield of 58% and an ee value of 90%.

[0115] Embodiment 19:

[0116]

[0117] 1. For the preparation of intermediate a4, see Example 16.

[0118] 2. Intermediate a4 (0.1 mmol), iodinated n-propane (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 19 as a white solid with a yield of 55% and an ee value of 88%.

[0119] Embodiment 20:

[0120] 1. For the preparation of intermediate a4, see Example 16.

[0121] 2. Intermediate a4 (0.1 mmol), n-iodobutane (0.5 mmol), CoBr2 0.01 mmol, ligand L1 0.012 mmol, CsF 0.1 mmol, K3PO4 0.2 mmol, DME 0.8 mL, DEMS 0.3 mmol, react at -20 °C for 12 h, and separate by thin layer chromatography to obtain compound 20 as a white solid with a yield of 56% and an ee value of 86%.

[0122] Embodiment 21:

[0123] Test strains: There are two test strains, Sclerotinia scleotiorum and Botrytis cinerea.

[0124] Test compounds: compounds 1-20 and positive control commercial drugs sclerotinol and fucymidone, and control natural products norcantharidin, olefin skeleton, and nor skeleton.

[0125] Others: glucose, agar powder, cutting board, kitchen knife, gauze, potato, cotton wool, sealing film, marker pen, etc.

[0126] Antibacterial activity determination method:

[0127] Growth rate method - refer to "Research Methods of Plant Chemical Protection" edited by Mu Liyi. The growth rate method is also called the inhibition zone method or agar plate method. It is a method of mixing different concentrations of liquid medicine with melted culture medium to make a toxic culture medium, inoculating pathogens on the culture medium plane, and judging the toxicity of the agent by the growth rate of the pathogens. It is expressed by the size of the colony diameter within a certain period of time.

[0128] The aseptic culture of potato dextrose agar (PDA) is generally prepared by the following method: 200 g peeled potatoes are cut into dices, added to 1000 mL distilled water, boiled for 30 minutes, filtered through 4 layers of gauze, and the filtrate is poured into a clean pan, and an appropriate amount of water is added, followed by 20 g glucose and 18 g agar powder, heated and stirred with a glass rod to dissolve them all. After all the dissolution is completed, the volume is fixed to 1000 mL with distilled water, and finally the calculated amount is divided into 300 mL conical flasks.

[0129] First, a dimethyl sulfoxide (DMSO) solution containing the test compound was added to a sterile Petri dish containing nearly 10 mL of potato dextrose agar (PDA). Then, a mycelial cake with a diameter of 6 mm was cut from the fungal colony and placed in the center of the PDA Petri dish at 28 °C for 3-7 days. For each compound, three antifungal experiments were performed. In addition, pure DMSO and commercial fungicides, sclerotinol and fucymidone, were used as negative and positive controls, respectively. The inhibition rate (I) of the test compound was calculated according to the following formula:

[0130]

[0131] In this formula, C represents the average hyphal diameter of the negative control, and T represents the average hyphal diameter of the compound-treated PDA. Finally, some selected compounds were further tested at different concentrations to calculate their EC50 (half-maximal effective concentration) values ​​using probabilistic analysis based on SPSS 23.0 software package.

[0132]

[0133] Example 22: Molecular docking simulation

[0134] 1. Preparation of compound structure

[0135] The compound for docking this time was compound 7, which was constructed using ChemDraw software, imported into Chem3D software, optimized using the MM2 module and energy minimized, and saved as an sdf file as the ligand molecule for molecular docking. It was then imported into Maestro 12.8 software, optimized using the LigPrep module, and the force field selected was OPLS3e.

[0136] 2. Preparation of target protein structure

[0137] Since the receptor protein of phthalimide drugs is generally HK protein, HK protein was selected for simulation. The HK (PDB ID: 2C2A) protein structure comes from the RCSB database (https: / / www.rcsb.org / ). The protein structure was processed on the Maestro11.9 platform to remove unreasonable structures, and the protein was processed using Schrodinger's Protein Preparation Wizard to minimize the energy of the protein and optimize the geometric structure.

[0138] 3. Molecular docking

[0139] Molecular docking processing and optimization were performed by the Glide module in Schrödinger Maestro software. Protein processing was performed using the Protein Preparation Wizard module. Receptors were preprocessed, optimized, and minimized (constrained minimization using the OPLS3e force field). Compound structures were prepared using the default settings of the LigPrep module. When screening was performed in the Glide module, the prepared receptors were imported, and the box size was set to 10Å×10Å×10Å based on the protein ligand as the active site center. Finally, molecular docking and screening were performed using the standard precision docking (SP) method.

[0140] 4. Screening and analysis of docking results

[0141] Analyze the action mode of the compound and the target protein to obtain the interaction between the compound and the protein residue, such as the hydrogen bonding, π-π interaction, hydrophobic interaction, etc., and then refer to the docking score of the compound to infer whether the compound to be screened has a certain active effect.

[0142] 5. Docking results

[0143] The molecular docking results are shown in Table 2. In addition, the complex of protein and small molecule was visualized and analyzed using Pymol2.1. Figure 4 .

[0144]

[0145] 6. Protein interaction analysis

[0146] In this experiment, compound 7 was molecularly docked with the HK target protein. The molecular docking results showed that the compound and the target protein had a binding effect and a good match, with a binding energy of -8.545 kcal / mol, which was less than -6.0 kcal / mol. The complex formed by the docked compound and the protein was visualized using Pymol2.1 software to obtain the binding mode of the compound and the protein. According to the binding mode, the amino acid residues that bind to the compound and the protein pocket can be clearly seen, such as: Compound 7 can form strong hydrogen bond interactions with the key residues VAL-431 (valine) and TYR-384 (tyrosine) in the active site of the HK protein, and can also form weak hydrogen bonds with ARG-430 (arginine). It has strong binding ability and plays an important role in anchoring small molecules in the protein pocket. In addition, the compound can also form hydrophobic interactions with TYR-384 (tyrosine) and ILE-416 (isoleucine), especially the benzene ring of the compound forms a π-π conjugated interaction with the amino acid benzene ring of TYR-384 (tyrosine), which makes an important contribution to the stabilization of small molecules. These interactions can promote the formation of stable complexes between the compound and proteins, and are highly correlated with proteins.

[0147] Then, the rapeseed sclerotinia was cultured on a PDA medium containing 1.0 mg / L compound 7 for 48 hours. The mycelial morphology was observed using a scanning electron microscope. Figure 5 As shown in the figure, the mycelium of Sclerotinia sclerotiorum was obviously wrinkled under the scanning electron microscope, which proved that the growth of mycelium was indeed inhibited.

[0148] We then conducted an in vitro activity test on rapeseed leaves for compound 7. The inhibitory effects of 100 ppm and 200 ppm compound 7 on rapeseed sclerotinia were tested, and 100 ppm and 200 ppm sclerotium net were used as the control group. The results showed that the leaves of the experimental group and the blank group showed a large difference. After 72 hours of inoculation, the blank group had been severely infected by the pathogen, with large areas of wilting and a large number of white hyphae. The wilting area of ​​the experimental group and the positive control group using 100 ppm compound 7 and 100 ppm sclerotium net was significantly smaller than that of the blank group. The inhibition rate of compound 7 was calculated to be 44.2%, and the inhibition rate of the positive control group was 31.9%. Subsequently, the concentration was increased to 200 ppm, and there was no disease on the leaves of compound 7 and the positive control group. The inhibition rate of both was 100%. It shows an excellent antibacterial effect.

Claims

1. A norcantharidin derivative, characterized in that Its general structural formula is shown in the following formula I: ； in: R1 is selected from one of alkyl, alkanoyl, aryl-substituted alkyl, heterocycle-substituted alkyl and halogen-substituted alkyl; R2 is selected from phenyl or substituted phenyl, and the substituent of the substituted phenyl is halogen.

2. The norcantharidin derivative according to claim 1, characterized in that: R1, R2 are selected from the following groups: Compound 1: R1=CH3, R2=Ph; Compound 2: R1=CD3, R2=Ph; Compound 3: R1 = C2H5, R2 = Ph; Compound 4: R1 = n-C3H7, R2 = Ph; Compound 5: R1 = n-C4H9, R2 = Ph; Compound 6: R1 = CH3, R2 = 3-F-Ph; Compound 7: R1 = CD3, R2 = 3-F-Ph; Compound 8: R1 = C2H5, R2 = 3-F-Ph; Compound 9: R1 = n-C3H7, R2 = 3-F-Ph; Compound 10: R1 = n-C4H9, R2 = 3-F-Ph; Compound 11: R1=CH3, R2=3,4-F-Ph; Compound 12: R1=CD3, R2=3,4-F-Ph; Compound 13: R1 = C2H5, R2 = 3,4-F-Ph; Compound 14: R1 = n-C3H7, R2 = 3,4-F-Ph; Compound 15: R1 = n-C4H9, R2 = 3,4-F-Ph; Compound 16: R1 = CH3, R2 = 3-Cl-Ph; Compound 17: R1 = CD3, R2 = 3-Cl-Ph; Compound 18: R1 = C2H5, R2 = 3-Cl-Ph; Compound 19: R1 = n-C3H7, R2 = 3-Cl-Ph; Compound 20: R1 = n-C4H9, R2 = 3-Cl-Ph.

3. The method for preparing the norcantharidin derivatives according to claim 1, characterized in that The steps include: Step 1: reacting norcantharidin with substituted aniline to obtain intermediate a; ； R represents monohalogen or dihalogen substitution; Step 2: The intermediate a is subjected to a metal-catalyzed hydroalkylation reaction to obtain the target product represented by formula I.

4. The preparation method according to claim 3, characterized in that: In step 1, in the presence of a catalyst, norcantharidin and substituted aniline are subjected to an imidization reaction in DCM, and after the reaction is completed, the intermediate a is separated by column chromatography; The catalyst is one or more of DCC and EDCI.

5. The preparation method according to claim 3, characterized in that: In step 2, the intermediate a obtained in step 1, alkyl iodide, catalyst, ligand and base are placed in a Shrek tube under inert gas protection, and a solvent and a hydrogen source are added to the system to perform a hydroalkylation reaction to obtain the target product shown in formula I; The alkyl iodide is selected from one of methyl iodide, deuterated methyl iodide, ethyl iodide, n-propane iodide and n-butane iodide; The catalyst is selected from one of CoCl2, CoBr2, CoBr2(DME), CoI2, Co(acac)2, and CoF3; The ligand is selected from one of the following ligands L1-L18: 。 6. The preparation method according to claim 5, characterized in that: The base is a combination of one or more of CsF, KF, K3PO4, NaHCO3, KH2PO4, Cs2CO3, Na2CO4, KHCO3, and K2CO3.

7. The preparation method according to claim 5, characterized in that: The hydrogen source is a combination of one or more of DEMS, (MeO)3SiH, (MeO)2MeSiH, and PHMS.

8. The preparation method according to claim 5, characterized in that: The ligand is ligand L1.

9. Use of the norcantharidin derivatives according to claim 1 in the preparation of a preparation for inhibiting plant pathogens.

10. The use according to claim 9, characterized in that: The plant pathogens include rapeseed sclerotinia and tomato gray mold.

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