Matrine bisamide derivatives and their preparation method and application

Abstract

The present invention discloses a class of matrine bisamide derivatives, having a chemical structural formula as shown in Formula I, wherein R1 in Formula I is an Fmoc group or a group having a chemical structural formula as shown in Formula II; and R2 is a hydroxyl group, a group having a chemical structural formula as shown in Formula III, a group having a chemical structural formula as shown in Formula IV, or a group having a chemical structural formula as shown in Formula V. The matrine bisamide derivatives have excellent effects of killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells, and can be used to prepare products for killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells. When the matrine bisamide derivatives are used as active ingredients in preparing products for killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells, the concentration of the matrine bisamide derivatives in the products is 0.01 to 2.72 mmol / L.

Description

Technical Field

[0001] The present invention relates to the technical field of organic synthesis, and in particular to a class of matrine bisamide derivatives and a preparation method and application thereof. Background Art

[0002] Matrine (C 15 H 24 Matrine (NO) is an alkaloid extracted from the dried roots, plants, and fruits of the leguminous plant Sophora flavescens through organic solvents such as ethanol. It exhibits certain anti-tumor, antiviral, antibacterial, and insecticidal activities. However, its relatively low bioactivity, slow efficacy, and poor persistence limit its application. Therefore, current research focuses on introducing various structural groups into matrine through structural splicing in order to enhance its bioactivity.

[0003] At present, the research on the structural modification and activity of matrine is mainly concentrated in the field of traditional Chinese medicine, especially the activity research of matrine derivatives after matrine ring opening is mainly concentrated on anti-tumor and antiviral research, and the value of matrine as a plant-based insecticide in pesticides has not been fully utilized.

[0004] Amide insecticides are blockbuster insecticides launched after neonicotinoids, pyrethroids and organophosphorus insecticides. The structural modification active groups of amide insecticides are flexible and diverse. The introduced groups range from electron-withdrawing groups such as halogen elements fluorine, chlorine, bromine, iodine, cyano, trifluoromethyl, nitro, to electron-donating groups such as methyl and tert-butyl, as well as heterocyclic groups such as thiazole, pyrazole and oxazolidinone.

[0005] As a member of the matrine derivatives, there are few reports on the synthesis and activity research of matrine amide derivatives, and they are mainly monoamide derivatives. Moreover, most of the activity research of these monoamide derivatives focuses on the anti-tumor aspect in the medical field, and there are few studies on insecticide.

[0006] Therefore, the present invention provides a matrine bisamide derivative having insecticidal activity. Summary of the Invention

[0007] The purpose of the present invention is to overcome the above-mentioned deficiencies of the prior art and to provide a class of matrine bisamide derivatives and their preparation methods and applications.

[0008] The first object of the present invention is to provide a class of matrine bisamide derivatives.

[0009] The second object of the present invention is to provide the use of the above-mentioned matrine bisamide derivatives in the preparation of products for killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells.

[0010] The third object of the present invention is to provide a product for killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells.

[0011] In order to achieve the above object, the present invention is implemented through the following scheme:

[0012] A class of matrine bisamide derivatives, the chemical structure of which is shown in Formula I,

[0013]

[0014] R1 is an Fmoc group or a group with a chemical structure as shown in Formula II; R2 is a hydroxyl group, a group with a chemical structure as shown in Formula III, a group with a chemical structure as shown in Formula IV, or a group with a chemical structure as shown in Formula V;

[0015]

[0016] The present invention forms a diamide structure on the 12-position secondary amine and the 11-position side carboxyl group of a matrine derivative (matrine) and introduces the groups shown above, thereby constructing a new matrine diamide derivative; the matrine diamide derivative has excellent insecticidal properties and can effectively kill agricultural pests and / or inhibit the cell proliferation of agricultural pests; and the present invention provides a preparation method of the matrine diamide derivative.

[0017] Preferably, the chemical structural formula of the matrine bisamide derivative is as shown in any one of Formula VI to Formula XII;

[0018]

[0019]

[0020] More preferably, the chemical structural formula of the matrine bisamide derivative is as shown in Formula VIII to Formula XI.

[0021] Further preferably, the chemical structural formula of the matrine bisamide derivative is as shown in Formula VIII or Formula XI.

[0022] Among them, the matrine bisamide derivatives represented by formula VIII or formula XI have more excellent insecticidal activity and can effectively inhibit the proliferation of agricultural pest cells. The half inhibitory concentration (IC50) of the matrine bisamide derivatives represented by formula VIII on agricultural pest cells is 50 ) is 0.02 mM, and the half-inhibitory concentration of the matrine bisamide derivative represented by formula XI on agricultural pest cells is 0.01 mM.

[0023] The present invention also seeks to protect the use of any of the above-mentioned matrine bisamide derivatives in the preparation of products for killing agricultural pests and / or inhibiting the proliferation of agricultural pests.

[0024] Preferably, the product is an insecticide and / or a pesticide.

[0025] Preferably, the agricultural pests are Lepidoptera agricultural pests.

[0026] More preferably, the lepidopteran agricultural pests are fall armyworm and / or armyworm.

[0027] Preferably, the agricultural pest cells are Sf9 cells.

[0028] The present invention also seeks to protect a product for killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells, which uses any of the above-mentioned matrine diamide derivatives as an active ingredient.

[0029] Preferably, the product is an insecticide and / or a pesticide.

[0030] Preferably, the agricultural pests are Lepidoptera agricultural pests.

[0031] More preferably, the lepidopteran agricultural pests are fall armyworm and / or armyworm.

[0032] Preferably, the agricultural pest cells are Sf9 cells.

[0033] Preferably, the matrine bisamide derivative having a chemical structural formula as shown in any one of Formula VIII to Formula XI is used as the active ingredient.

[0034] More preferably, the matrine bisamide derivative having a chemical structural formula such as Formula VIII or Formula XI is used as the active ingredient.

[0035] Further preferably, when the matrine bisamide derivative represented by the chemical structural formula VIII is used as the active ingredient, the concentration of the active ingredient in the product is 0.02 mM;

[0036] When the matrine bisamide derivative with the chemical structure shown in Formula XII is used as the active ingredient, the concentration of the active ingredient in the product is 0.01 mM.

[0037] Compared with the prior art, the present invention has the following beneficial effects:

[0038] The present invention provides a class of matrine bisamide derivatives, having a chemical structural formula as shown in Formula I, wherein R1 in Formula I is an Fmoc group or a group having a chemical structural formula as shown in Formula II; and R2 is a hydroxyl group, a group having a chemical structural formula as shown in Formula III, a group having a chemical structural formula as shown in Formula IV, or a group having a chemical structural formula as shown in Formula V. The matrine bisamide derivatives have excellent effects in killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells, and can be used to prepare products for killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells. When the matrine bisamide derivatives are used as active ingredients in preparing products for killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells, the concentration of the matrine bisamide derivatives in the products is 0.01 to 2.72 mmol / L. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 This is the FTIR detection image of the white fluffy solid in Example 1;

[0040] Figure 2 This is the MS detection image of the white fluffy solid in Example 1;

[0041] Figure 3 This is the FTIR detection image of the yellow solid 1 in Example 1;

[0042] Figure 4 This is the MS detection image of the yellow solid 1 in Example 1;

[0043] Figure 5 This is the FTIR detection image of the yellow solid 2 in Example 2;

[0044] Figure 6 This is the MS detection image of the yellow solid 2 in Example 2;

[0045] Figure 7 This is the FTIR detection image of substance 1 in Example 3;

[0046] Figure 8 This is the MS detection image of substance 1 in Example 3;

[0047] Figure 9 This is the FTIR detection image of substance 2 in Example 4;

[0048] Figure 10 This is the MS detection image of substance 2 in Example 4;

[0049] Figure 11 This is the FTIR detection image of substance 3 in Example 5;

[0050] Figure 12 This is the MS detection image of substance 3 in Example 5;

[0051] Figure 13This is the FTIR detection image of substance 4 in Example 6;

[0052] Figure 14 This is the MS detection image of substance 4 in Example 6;

[0053] Figure 15 This is the FTIR detection image of substance 5 in Example 7;

[0054] Figure 16 This is the MS detection image of substance 5 in Example 7;

[0055] Figure 17 Figures 2 and 3 are cell morphology observations after Sf9 cells were cultured with 1.2 mM Mat-Ac in Example 8; A is a cell morphology observation at 0 h of culture, B is a cell morphology observation at 24 h of culture, C is a cell morphology observation at 48 h of culture, and D is a cell morphology observation at 72 h of culture;

[0056] Figure 18 Cell morphology observation images after culturing Sf9 cells with different concentrations of A1 in Example 8; A to D are cell morphology observation images of cells cultured with 1.2 mM A1 at 0 h, 24 h, 48 h, and 72 h, respectively; E to H are cell morphology observation images of cells cultured with 1.6 mM A1 at 0 h, 24 h, 48 h, and 72 h, respectively; I to L are cell morphology observation images of cells cultured with 2.0 mM A1 at 0 h, 24 h, 48 h, and 72 h, respectively;

[0057] Figure 19 These are cell morphology observation pictures after culturing Sf9 cells with different concentrations of B1 in Example 8; A to D are cell morphology observation pictures of cells cultured with 1.2 mM B1 at 0 h, 24 h, 48 h, and 72 h, respectively; E to H are cell morphology observation pictures of cells cultured with 1.6 mM B1 at 0 h, 24 h, 48 h, and 72 h, respectively; I to L are cell morphology observation pictures of cells cultured with 2.0 mM B1 at 0 h, 24 h, 48 h, and 72 h, respectively;

[0058] Figure 20 These are cell morphology observation images after Sf9 cells were cultured with different concentrations of A2 in Example 8; A to D are cell morphology observation images of cells cultured with 0.20 mM A2 at 0 h, 24 h, 48 h, and 72 h, respectively; E to H are cell morphology observation images of cells cultured with 0.25 mM A2 at 0 h, 24 h, 48 h, and 72 h, respectively; I to L are cell morphology observation images of cells cultured with 0.30 mM A2 at 0 h, 24 h, 48 h, and 72 h, respectively;

[0059] Figure 21These are cell morphology observation images after Sf9 cells were cultured with different concentrations of A3 in Example 8; A to D are cell morphology observation images of cells cultured with 0.10 mM A3 at 0 h, 24 h, 48 h, and 72 h, respectively; E to H are cell morphology observation images of cells cultured with 0.30 mM A3 at 0 h, 24 h, 48 h, and 72 h, respectively; I to L are cell morphology observation images of cells cultured with 0.50 mM A3 at 0 h, 24 h, 48 h, and 72 h, respectively;

[0060] Figure 22 These are cell morphology observation images after culturing Sf9 cells with different concentrations of B2 in Example 8; A to D are cell morphology observation images of cells cultured with 0.20 mM B2 at 0 h, 24 h, 48 h, and 72 h, respectively; E to H are cell morphology observation images of cells cultured with 0.60 mM B2 at 0 h, 24 h, 48 h, and 72 h, respectively; I to L are cell morphology observation images of cells cultured with 0.80 mM B2 at 0 h, 24 h, 48 h, and 72 h, respectively;

[0061] Figure 23 These are cell morphology observation images after Sf9 cells were cultured with different concentrations of B3 in Example 8; A to D are cell morphology observation images of cells cultured with 0.20 mM B3 at 0 h, 24 h, 48 h, and 72 h, respectively; E to H are cell morphology observation images of cells cultured with 0.25 mM B3 at 0 h, 24 h, 48 h, and 72 h, respectively; I to L are cell morphology observation images of cells cultured with 0.30 mM B3 at 0 h, 24 h, 48 h, and 72 h, respectively;

[0062] Figure 24 These are cell morphology observation pictures after Sf9 cells were cultured with different concentrations of B4 in Example 8; A to D are cell morphology observation pictures of cells cultured with 1.00 mM B4 at 0 h, 24 h, 48 h, and 72 h, respectively; E to H are cell morphology observation pictures of cells cultured with 2.00 mM B4 at 0 h, 24 h, 48 h, and 72 h, respectively; I to L are cell morphology observation pictures of cells cultured with 3.00 mM B4 at 0 h, 24 h, 48 h, and 72 h, respectively. DETAILED DESCRIPTION

[0063] The present invention is further described in detail below with reference to the accompanying drawings and specific examples. The examples are intended only to illustrate the present invention and are not intended to limit the scope of the present invention. The experimental methods used in the following examples are conventional methods unless otherwise specified; the materials and reagents used are commercially available unless otherwise specified.

[0064] Example 1 Synthesis of 12N-(9-fluorenylmethoxy)matrine

[0065] 1. Synthesis of Mat-Ac

[0066] 1. Experimental methods

[0067] The synthesis reaction equation of matrine is shown in Equation 1, which is specifically as follows:

[0068] Equation I:

[0069] 2.48 g of matrine (0.01 mol) was placed in a 250 mL round-bottom flask, and 40 mL of 10% sodium hydroxide (4 g of sodium hydroxide dissolved in 36 mL of water) (0.1 mol) was added. The mixture was refluxed under stirring, and the reaction progress was monitored by thin-layer chromatography (TLC). After the reaction was complete, the mixture was extracted with ethyl acetate, the aqueous phase was retained, and the pH was adjusted to 7-8 with a 20% by volume sulfuric acid solution in an ice bath. The filtrate was filtered and retained, and extracted three times with ethyl acetate. The aqueous phase was retained and dried to obtain 2.23 g of a white fluffy solid; the yield was 89.9%.

[0070] The white fluffy solid was subjected to infrared spectroscopy (FTIR), mass spectrometry (MS) and nuclear magnetic resonance (NMR) detection, and the corresponding test results were recorded. The NMR detection included 1 H NMR detection and 13 C nuclear magnetic resonance detection.

[0071] 2. Experimental results

[0072] The FTIR detection image of white fluffy solid is as follows Figure 1 As shown, the results show: characteristic peak 3435cm -1 The broad peak at 2927 cm is the stretching vibration peak of hydroxyl -OH. -1 The antisymmetric stretching vibration and symmetric stretching vibration peaks of methylene are 1632 cm -1 The peak is the stretching vibration peak of C=O in carboxylic acid, 1553 cm -1 The peak is the bending vibration peak of NH on the secondary amine, 1439cm -1 and 1410cm -1 The peak is the bending vibration peak of CH in the alkane group, 1170 cm -1 The peak is the stretching vibration peak of CN, 1129cm -1 is the stretching vibration peak of CO; and based on 1553cm -1 NH bending vibration at 3435 cm -1 The stretching vibration peak of the hydroxyl-OH at the position can be used to preliminarily determine that matrine is hydrolyzed into matrine acid.

[0073] The MS detection image of the white fluffy solid is as follows Figure 2 As shown, the results showed that matrine (C15 H 26 The theoretical m / z of N2O2) is 266.38, while a molecular ion peak appears at 267.7 in the MS detection graph of the white fluffy solid, which is consistent with the theoretical m / z of matrine.

[0074] White fluffy solid 1 The results of H NMR examination are: 1 H NMR (700MHz, D2O) δ3.34 (dt, J=12.2, 5.8Hz, 1H), 3.21 (t, J=13.1Hz, 1H), 3. 05(dd, J=12.8, 4.5Hz, 1H), 2.82(dd, J=22.7, 11.9Hz, 2H), 2.55-2.52(m, 2H ), 2.20 (dt, J=13.4, 5.7Hz, 4H), 2.10-2.05 (m, 1H), 1.93 (d, J=13.2Hz, 1H), 1.87-1.78 (m, 2H), 1.69 (ddt, J=14.4, 11.1, 6.0Hz, 2H), 1.65-1.48 (m, 8H).

[0075] White fluffy solid 13 The results of C nuclear magnetic resonance imaging are as follows: 13 C NMR (176MHz, D2O) δ182.67, 61.63, 56.11, 56.03, 52.67, 43.54, 37.66, 36.99, 32.72, 29.78, 25.78, 24.74, 20.45, 19.67, 19.42.

[0076] The results showed that the white fluffy solid was matrine (Mat-Ac), with the molecular formula C 15 H 26 N2O2.

[0077] 2. Synthesis of 12N-(9-fluorenylmethoxy)matrine

[0078] 1. Experimental methods

[0079] The synthetic chemical formula of 12N-(9-fluorenylmethoxy)matrine is shown in Formula II, which is as follows:

[0080] Equation II:

[0081] 0.2664 g of berberine acid (1 mmol) was weighed and placed in a 50 mL flask, 12 mL of 10% (w / w) Na2CO3 solution was added and stirred to dissolve to obtain a mixed solution; then 0.5174 g (2 eq) of Fmoc-Cl was weighed, dissolved in 12 mL of 1,4-dioxane, and added dropwise to the mixed solution. The mixture was stirred thoroughly at 25°C and the reaction progress was monitored by thin layer chromatography (TLC). After the reaction was completed, 20 mL of water was added and the pH value was adjusted to 6-7 with 3 mol / L HCl solution. The mixture was extracted twice with petroleum ether to remove oil-soluble impurities, and the aqueous phase was collected. The aqueous phase was then extracted three times with dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate, and distilled under reduced pressure. The organic phase was concentrated to obtain a yellow solid 1.

[0082] The collected yellow solid 1 was subjected to FTIR, MS and NMR detection respectively, and the corresponding test results were recorded. The NMR detection included 1 H NMR detection and 13 C nuclear magnetic resonance detection.

[0083] 2. Experimental Results

[0084] The FTIR detection image of yellow solid 1 is as follows Figure 3 As shown, the results show that the wave number is 3063cm -1 and 755cm -1 The peaks at 1651 cm-1 are the stretching vibration peaks of the unsaturated CH of the benzene ring in Fmoc and the characteristic peaks of ortho-substitution; the wave number is 1651 cm-1 -1 The stretching vibration peak of tertiary amide C=O is at the bottom; and the FTIR detection diagram of matrine ( Figure 1 ) compared to 1553cm -1 The bending vibration peak at 37° disappeared, indicating that the Fmoc group was attached to matrine.

[0085] MS detection of yellow solid 1 is shown in Figure 2 Figure 4 As shown, the results showed that: 12N-(9-fluorenylmethoxy)matrine (C 30 H 36 The theoretical m / z value of N2O4) is 488.63, while the MS detection graph of the yellow solid 1 shows a molecular ion peak at 489.5, which is consistent with the theoretical m / z of 12N-(9-fluorenylmethoxy)matrine.

[0086] Yellow solid 1 1 The results of H NMR examination are: 1H NMR (500MHz, MeOD) δ7.79 (d, J=7.5Hz, 2H), 7.62 (t, J=6.7Hz, 2H), 7.38 (t, J=7.5Hz, 2H), 7.31 (tdd, J=7.5, 4.0, 1.2Hz, 2H), 4.75-4.61 (m, 2H), 4.20 (t, J=4.5Hz, 1H), 3.47 (s, 1H), 3.28-3.23 (m, 1H), 3.18 (dd, J=13.8 , 7.0Hz, 1H), 2.81-2.74(m, 3H), 2.19-1.98(m, 6H), 1.69-1.09(m, 12H).

[0087] Yellow solid 1 13 The results of C nuclear magnetic resonance imaging are as follows: 13 C NMR (126MHz, MeOD) δ181.72, 157.77, 145.62, 145.52, 142.89, 142.86, 128.70, 128.70, 128.21, 128.15, 125.75, 125.72, 120.94, 120.91, 67.02, 64.68, 57.37, 55.99, 45.48-45.30(m), 38.12, 35.11, 32.87, 28.76, 28.66, 23.98, 21.51, 21.41.

[0088] The results showed that the yellow solid 1 (A1) was 12N-(9-fluorenylmethoxy)matrine, with the molecular formula C 30 H 36 N2O4, the chemical structure is shown in Formula VI;

[0089]

[0090] Example 2 Synthesis of 12N-(3-fluorobenzoyl)matrine

[0091] 1. Experimental Methods

[0092] The synthesis reaction equation of 12N-(3-fluorobenzoyl)matrine is shown in Equation III, which is specifically shown below:

[0093] Equation III:

[0094] Weigh 0.2664 g (0.4 mmol) of the matrine acid in Example 1 and 0.318 g (3 eq) of Na2CO3 into a 50 mL flask, add 3 mL of water and stir to dissolve, then add 8 mL of 1,4-dioxane, and finally add 243 μL of 3-fluorobenzoyl chloride (2 eq, density 1.304 g / mL), stir thoroughly at 25 ° C, and monitor the reaction progress by TLC. After the reaction is completed, the liquid is spin-dried, 20 mL of water is added, and the pH value is adjusted to 6-7 with HCl solution, followed by extraction three times with dichloromethane. The organic phases are combined, dried over anhydrous sodium sulfate, and distilled under reduced pressure. The organic phase is concentrated to obtain a yellow solid 2.

[0095] The collected yellow solid 2 was subjected to FTIR, MS and NMR detection respectively, and the corresponding test results were recorded. The NMR detection included 1 H NMR detection and 13 C nuclear magnetic resonance detection.

[0096] 2. Experimental Results

[0097] The FTIR detection image of yellow solid 2 is as follows Figure 5 As shown, the results show that the wave number is 3423cm -1 The peaks are hydroxyl-OH stretching vibration peaks, 2937 and 2748 cm -1 The peaks are attributed to the antisymmetric stretching vibration and symmetric stretching vibration of CH in methylene, 1712 cm -1 The peak is C=O stretching vibration peak, 1621cm -1 The peak is the amide C=O stretching vibration peak, 1449 cm -1 The peak is the skeleton vibration of the benzene ring, 1366cm -1 The peak is the hydrocarbon CH bending vibration peak, 1216 cm -1 The peak is the secondary amine CN stretching vibration peak, 909cm -1 and 773cm -1 The peak at is the out-of-plane bending vibration peak of the disubstituted CH on the benzene ring; it can be judged that berberine acid has been successfully connected to the 3-fluorobenzoyl group.

[0098] The MS detection image of yellow solid 2 is as follows Figure 6 As shown, the results showed that: 12N-(3-fluorobenzoyl)matrine (C 22 H 29 The theoretical m / z value of FN2O3) is 388.48, while the MS detection pattern of the yellow solid 2 shows a molecular ion peak at 389.8, which is consistent with the theoretical m / z of 12N-(3-fluorobenzoyl)matrine.

[0099] Yellow solid 2 1 The results of H NMR examination are: 1H NMR (500MHz, MeOD) δ7.46 (td, J=8.0, 5.6Hz, 1H), 7.26 (d, J=7.6Hz, 1H), 7.24-7.18 (m, 1H), 7.17-7.09 (m, 1H), 2.82-2.71 (m, 3H), 2.19 (s, 3H), 2.08-1.79 (m, 6H), 1.76-1.20 (m, 12H).

[0100] Yellow solid 2 13 The results of C nuclear magnetic resonance imaging are as follows: 13 C NMR (126MHz, MeOD) δ180.05, 172.03, 163.85 (d, J=246.4Hz), 140.87 (d, J=7.0Hz), 131.64 (d, J=8.1Hz), 123.70 (d, J=3.2Hz), 117.2 9 (d, J=21.2Hz), 114.74 (d, J=23.2Hz), 63.74, 57.49, 57.35, 41.30, 36.76, 35.30, 34.22, 30.76, 29.72, 23.88, 23.72, 22.06, 21.66.

[0101] The results showed that the yellow solid 2 (B1) was 12N-(3-fluorobenzoyl)matrine, with the molecular formula C 22 H 29 FN2O3, the chemical structure is shown in Formula VII;

[0102]

[0103] Example 3 Synthesis of 12N-(9-fluorenylmethoxy)-18N-(5-indole)matrine butyramide

[0104] 1. Experimental Methods

[0105] The synthetic reaction equation of 12N-(9-fluorenylmethoxy)-18N-(5-indole)matrine butyramide is shown in Equation IV, which is specifically shown below:

[0106] Formula IV:

[0107] 0.2443 g (0.5 mmol) of 12N-(9-fluorenylmethoxy)matrine obtained in Example 1 was weighed and dissolved in 5 mL of DMF. Ultrasonication formed a suspension, followed by the addition of 0.1013 g (1.5 eq) of 1-hydroxybenzotriazole (HOBT) and 0.1150 g (1.2 eq) of N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDCI). After stirring in an ice bath under nitrogen for 1 hour, 0.0793 g (1.2 eq) of 5-aminoindole and 169 μL (3 eq) of N-methylmorpholine were added, and stirring was continued. The reaction was carried out under nitrogen protection throughout the preparation process, and the reaction progress was monitored by TLC. After the reaction was complete, substance 1 was obtained.

[0108] Perform FTIR, MS and NMR tests on substance 1 and record the corresponding test results. NMR test includes 1 H NMR detection and 13 C nuclear magnetic resonance detection.

[0109] 2. Experimental Results

[0110] The FTIR detection image of substance 1 is as follows Figure 7 As shown, the results show: 1651cm -1 The peak is the stretching vibration peak of amide C=O, 1555cm -1 The peak is the bending vibration peak of amide NH, 1108 cm -1 The peak is the stretching vibration peak of amide CN. Compared with the FTIR detection chart of 12N-(9-fluorenylmethoxy)matrine in Example 1, it can be proved that 12N-(9-fluorenylmethoxy)matrine is connected to a 5-aminoindole active group.

[0111] The MS detection image of substance 1 is as follows Figure 8 As shown, the results showed that: 12N-(9-fluorenylmethoxy)-18N-(5-indole)matrine butyramide (C 38 H 32 The theoretical m / z value of N4O3) is 603.3, and a molecular ion peak appears at 603.3 in the MS detection graph of substance 1, which is consistent with the theoretical m / z of 12N-(9-fluorenylmethoxy)-18N-(5-indole)matrine butanamide.

[0112] Substance 1 1 The results of H NMR examination are: 1H NMR (700MHz, Methanol-d4) δ7.80 (d, J=7.5Hz, 2H), 7.78 (d, J=2.0Hz, 1H), 7.61 (t, J=7.0Hz, 2H), 7.38 ( td, J=7.4, 2.6Hz, 2H), 7.34-7.29 (m, 3H), 7.23 (d, J=3.1Hz, 1H), 7.19 (dd, J=8.6, 2.0Hz, 1H), 6.40 (d, J= 3.1Hz, 1H), 4.82-4.67 (m, 2H), 4.23 (t, J=4.1Hz, 1H), 3.51-3.38 (m, 1H), 3.27-3.19 (m, 2H), 3.09-2.98 ( m, 2H), 2.89-2.73 (m, 1H), 2.66-2.48 (m, 2H), 2.29-2.18 (m, 2H), 1.78-1.63 (m, 2H), 1.61-1.31 (m, 12H).

[0113] Substance 1 13 The results of C nuclear magnetic resonance imaging are as follows: 13 C NMR (176MHz, Methanol-d4) δ173.97, 157.70, 145.58, 145.51, 142.95, 142.93, 13 5.11, 131.43, 129.36, 128.80, 128.31, 128.27, 128.22, 126.59, 125.76, 125.72, 1 20.99, 120.96, 117.10, 113.71, 112.08, 102.47, 67.14, 64.72, 56.78, 56.51, 55.51, 44.74, 39.35, 37.01, 34.05, 32.22, 30.75, 27.61, 27.27, 23.19, 20.47, 20.36.

[0114] The results showed that substance 1 (A2) was 12N-(9-fluorenylmethoxy)-18N-(5-indole)matrine butyramide, with the molecular formula C 38 H 32 N4O3, the chemical structure is shown in Formula VIII;

[0115]

[0116] Example 4 Synthesis of 12N-(9-fluorenylmethoxy)-18N-(4-fluorophenyl)matrine butyramide

[0117] 1. Experimental Methods

[0118] The synthetic reaction equation of 12N-(9-fluorenylmethoxy)-18N-(4-fluorophenyl)matrine butanamide is shown in Equation V, which is specifically shown below:

[0119] Formula V:

[0120] 0.2025 g (0.5 mmol) of 12N-(9-fluorenylmethoxy)-matrine obtained in Example 1 and 0.0778 g (1.4 eq) of 4-fluoroaniline were dissolved in 3.9 mL of acetonitrile, followed by the addition of 0.1683 g (1.2 eq) of N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (TCFH, molecular weight 280.58) and 140 μL (3.5 eq) of N-methylimidazole (NMI, molecular weight 82.1, density 1.03 g / mL). The mixture was stirred at 25 ° C for 21 hours. After the reaction, the mixture was extracted three times with ethyl acetate, and the organic phase was collected and separated and purified to obtain substance 2.

[0121] Substance 2 was tested by FTIR, MS and NMR respectively, and the corresponding test results were recorded. NMR test included 1 H NMR detection and 13 C nuclear magnetic resonance detection.

[0122] 2. Experimental Results

[0123] The FTIR detection image of substance 2 is as follows Figure 9 As shown, the result shows: 3439cm -1 The peak is the stretching vibration peak of NH on the amide bond, 2929cm -1 、2862cm -1 and 2763cm -1 The peak is attributed to the antisymmetric stretching vibration and symmetric stretching vibration of CH in methylene, 1664 cm -1 The peak is the stretching vibration peak of the amide bond C=O, 1555cm -1 and 1509cm -1 The peak is the bending vibration peak of amide NH, 1447 cm -1 and 1402cm -1 The peak is the bending vibration peak of CH in the hydrocarbon group, 1216 cm -1 The peak is the stretching vibration peak of CN, 1072cm -1 The peak is attributed to the stretching vibration peak of amide CN, 732 cm -1 The peak is the out-of-plane swinging vibration of CH in the long-chain hydrocarbon group; compared with the FTIR detection chart of 12N-(9-fluorenylmethoxy)matrine in Example 1, it can be proved that 12N-(9-fluorenylmethoxy)matrine is connected to the 4-fluoroaniline active group.

[0124] The MS detection chart of substance 2 is as follows Figure 10 As shown, the results showed that: 12N-(9-fluorenylmethoxy)-18N-(4-fluorophenyl) matrine butyramide (C 36 H 40 The theoretical m / z value of FN3O3) is 581.73, while the molecular ion peak appears at 582.4 in the MS detection graph of substance 2, which is consistent with the theoretical m / z of 12N-(9-fluorenylmethoxy)-18N-(4-fluorophenyl)matrine butanamide.

[0125] Substance 2 1 The results of H NMR examination are: 1 H NMR (500MHz, MeOD) δ7.78 (d, J=7.5Hz, 2H), 7.60 (d, J=7.4Hz, 2H), 7.56-7.51 (m, 2H), 7.36 (td, J=7.4, 4.1Hz, 2H), 7.28 (tt, J=7.6, 1.6Hz, 2H), 7.09-6.98 (m, 2H), 4.67 (dd, J=10.7, 4.3Hz, 1H) , 4.20 (t, J=4.4Hz, 1H), 3.67-3.46 (m, 1H), 3.35 (s, 1H), 3.26 (dd, J=9.3, 3.9Hz, 1H), 3.19-3.12 (m, 1H), 2.61 (td, J=8.5, 4.5Hz, 4H), 2.31-2.12 (m, 1H), 1.83-1.72 (m, 3H), 1.66-0.75 (m, 12H).

[0126] Substance 2 13 The results of C nuclear magnetic resonance imaging are as follows: 13 C NMR (126MHz, MeOD) δ174.30, 161.54, 159.62, 145.63, 145.53, 142.93, 136. 16, 129.87, 128.66, 128.63, 128.16, 128.11, 125.68, 125.63, 123.05, 122. 99, 120.94, 120.90, 116.28, 116.09, 66.85, 64.79, 57.87, 57.82, 56.15, 46.31, 41.11, 37.38, 35.98, 31.99, 30.74, 29.50, 28.97, 23.49, 22.06, 22.02.

[0127] The results showed that substance 2 (A3) was 12N-(9-fluorenylmethoxy)-18N-(4-fluorophenyl)matrine butyramide, with the molecular formula C 36 H 40FN3O3, the chemical structure is shown in IX;

[0128]

[0129] Example 5 Synthesis of 12N-(3-fluorobenzoyl)-18N-(5-indole)matrine butyramide

[0130] 1. Experimental Methods

[0131] The synthetic reaction equation of 12N-(3-fluorobenzoyl)-18N-(5-indole)matrine butanamide is shown in Equation VI, which is specifically shown below:

[0132] Equation VI:

[0133] 0.2025 g (0.5 mmol) of 12N-(3-fluorobenzoyl)-matrine obtained in Example 2 and 0.0858 g (1.3 eq) of 5-aminoindole were dissolved in 3.9 mL of acetonitrile, and 0.1683 g (1.2 eq) of N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (TCFH, molecular weight 280.58) and 140 μL (3.5 eq) of N-methylimidazole (NMI, molecular weight 82.1, density 1.03 g / mL) were added in sequence. The mixture was stirred at 25 ° C. for 21 hours. After the reaction, it was extracted three times with ethyl acetate, and the organic phase was collected and separated and purified to obtain substance 3.

[0134] Substance 3 was tested by FTIR, MS and NMR respectively, and the corresponding test results were recorded. NMR test included 1 H NMR detection and 13 C nuclear magnetic resonance detection.

[0135] 2. Experimental Results

[0136] The FTIR detection image of substance 3 is as follows Figure 11 As shown, the results show: 3286cm -1 The peak is the stretching vibration peak of NH, 2927cm -1 and 2780cm -1 The peaks are attributed to the antisymmetric stretching vibration and symmetric stretching vibration of CH in methylene, 1606 cm -1 The peak is the stretching vibration peak of amide C=O, 1451 cm -1 and 1352cm -1 The peak is the bending vibration peak of CH in the hydrocarbon group, 1260 cm -1 and 1226cm -1 The peak is the stretching vibration peak of CN, 1058cm -1 The peak is attributed to the stretching vibration peak of amide CN, 780 cm-1 The peak is the out-of-plane swing vibration of CH in the long-chain hydrocarbon group; compared with the FTIR detection chart of 12N-(3-fluorobenzoyl)matrine in Example 2, it can be determined that 12N-(3-fluorobenzoyl)matrine is connected to a 5-aminoindole active group.

[0137] The MS detection chart of substance 3 is as follows Figure 12 As shown, the results showed that: 12N-(3-fluorobenzoyl)-18N-(5-indole)matrine butyramide (C 30 H 35 The theoretical m / z value of FN4O2) is 502.63, while the molecular ion peak appears at 503.5 in the MS detection graph of substance 3, which is consistent with the theoretical m / z of 12N-(3-fluorobenzoyl)-18N-(5-indole)matrine butanamide.

[0138] Substance 3 1 The results of H NMR examination are: 1 H NMR (500MHz, MeOD) δ7.76 (d, J=2.0Hz, 1H), 7.45 (td, J=7.9, 5.6Hz, 1H), 7.32 (d, J=8.7Hz, 1H), 7.28-7.25 (m, 1H), 7.24 (d, J=2.4Hz, 1H), 7.21 (d, J=4.1Hz, 1H), 7.19 (t, J=2.2Hz, 1H) , 7.17 (d, J=2.0Hz, 1H), 6.40 (d, J=3.1Hz, 1H), 3.60 (s, 1H), 2.92-2.79 (m, 2H), 2.42 (q, J=7 .2Hz, 2H), 2.22 (d, J=4.3Hz, 1H), 2.07-1.87 (m, 6H), 1.82-1.62 (m, 5H), 1.55-1.25 (m, 7H).

[0139] Substance 3 13 The results of C nuclear magnetic resonance imaging are as follows: 13C NMR (126MHz, MeOD) δ174.08, 172.33, 163.89 (d, J=246.6Hz), 140.68 (d, J=7.0Hz), 1 35.12, 131.72 (d, J=8.0Hz), 131.40, 129.36, 126.55, 123.76 (d, J=3.1Hz), 117.48 (d , J=21.6Hz), 117.14, 114.82 (d, J=23.1Hz), 113.76, 112.07, 102.46, 64.31, 63.82, 5 7.42, 57.30, 41.00, 37.26, 35.46, 33.61, 30.73, 30.32, 29.42, 23.90, 21.93, 21.54.

[0140] The results showed that substance 3 (B2) was 12N-(3-fluorobenzoyl)-18N-(5-indole)matrine butyramide, with the molecular formula C 30 H 35 FN4O2, the chemical structure is shown in Formula X;

[0141]

[0142] Example 6 Synthesis of 12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl)matrine butyramide

[0143] 1. Experimental Methods

[0144] The synthesis reaction equation of 12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl)matrine butanamide is shown in Equation VII, which is specifically shown below:

[0145] Formula VII:

[0146] 0.2025 g (0.5 mmol) of 12N-(3-fluorobenzoyl)-matrine obtained in Example 2 and 0.0722 g (1.3 eq) of 4-fluoroaniline were dissolved in 3.9 mL of acetonitrile, and 0.1683 g (1.2 eq) of N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (TCFH, molecular weight 280.58) and 140 μL (3.5 eq) of N-methylimidazole (NMI, molecular weight 82.1, density 1.03 g / mL) were added in sequence. The mixture was stirred at 25 ° C. for 21 hours. After the reaction, the mixture was extracted three times with ethyl acetate, and the organic phase was collected and separated and purified to obtain substance 4.

[0147] Substance 4 was tested by FTIR, MS and NMR respectively, and the corresponding test results were recorded. NMR test included 1 H NMR detection and13 C nuclear magnetic resonance detection.

[0148] 2. Experimental Results

[0149] The FTIR detection image of substance 4 is as follows Figure 13 As shown, the results show: 3385cm -1 and 3294cm -1 The peak is the stretching vibration peak of NH on the amide bond, 2930 cm -1 The peak is attributed to the antisymmetric stretching vibration and symmetric stretching vibration of CH in methylene, 1682 cm -1 The peak is the stretching vibration peak of the amide bond C=O, 1428 cm -1 The peak is the bending vibration peak of CH in the hydrocarbon group, 1204 cm -1 The peak is the stretching vibration peak of CN, 1062cm -1 The peak is attributed to the stretching vibration peak of amide CN; compared with the FTIR detection chart of 12N-(3-fluorobenzoyl)matrine in Example 2, 1712cm -1 The stretching vibration peak of C=O of the carboxylic acid disappears, which can confirm that 12N-(3-fluorobenzoyl)matrine is connected to the 4-fluoroaniline active group.

[0150] The MS detection image of substance 4 is as follows Figure 14 As shown, the results showed that: 12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl)matrine butyramide (C 28 H 33 The theoretical m / z value of F2N3O2) is 481.59, while a molecular ion peak appears at 482.6 in the MS detection graph of substance 4, which is consistent with the theoretical m / z of 12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl)matrine butanamide.

[0151] Substance 4 1 The results of H NMR examination are: 1 H NMR (500MHz, MeOD) δ7.59-7.52 (m, 2H), 7.45 (td, J=7.9, 5.6Hz, 1H), 7.25 (d, J=7.6Hz, 1H), 7.18 (ddd, J=11.1, 8.6, 4.3Hz, 2H), 7.03 (t, J=8.8Hz, 2H), 3.48-3.33 (m, 2H), 2.74 (dd, J=27.2, 10.3Hz, 2H), 2.40 (s, 2H), 1.75-1.56 (m, 6H), 1.97-1.80 (m, 6H), 1.54-1.24 (m, 6H).

[0152] Substance 4 13 The results of C nuclear magnetic resonance imaging are as follows:13 C NMR (126MHz, MeOD) δ174.10, 172.04, 163.81 (d, J=246.4Hz), 160.56 (d, J=2 41.7Hz), 140.88 (d, J=7.1Hz), 136.09, 131.64 (d, J=8.1Hz), 123.61 (d, J=3 .1Hz), 123.04, 122.98, 117.36, 117.19, 116.28, 116.10, 114.67 (d, J=23.1 Hz), 63.58, 57.58, 57.45, 41.50, 37.24, 35.45, 30.02, 23.69, 22.26, 21.84.

[0153] The results showed that substance 4 (B3) was 12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl)matrine butyramide, with the molecular formula C 28 H 33 F2N3O2, chemical structure is shown in XI;

[0154]

[0155] Example 7 Synthesis of 12N-(3-fluorobenzoyl)-18N-(2-fluoroethyl)matrine butyramide

[0156] 1. Experimental Methods

[0157] The synthetic reaction equation of 12N-(3-fluorobenzoyl)-18N-(2-fluoroethyl)matrine butanamide is shown in Equation VIII, which is specifically shown below:

[0158] Formula VIII:

[0159] 0.2025 g (0.5 mmol) of 12N-(3-fluorobenzoyl)-matrine obtained in Example 2 and 0.0410 g of 2-fluoroethylamine (1.3 eq) were dissolved in 3.9 mL of acetonitrile, and 0.1683 g (1.2 eq) of N,N,N',N'-tetramethylchloroformamidine hexafluorophosphate (TCFH, molecular weight 280.58) and 140 μL (3.5 eq) of N-methylimidazole (NMI, molecular weight 82.1, density 1.03 g / mL) were added in sequence. The mixture was stirred at 25° C. for 21 hours. After the reaction, the mixture was extracted three times with ethyl acetate, and the organic phase was collected for separation and purification to obtain substance 5.

[0160] Substance 5 was subjected to FTIR, MS and NMR tests respectively, and the corresponding test results were recorded. NMR test included 1 H NMR detection and 13C nuclear magnetic resonance detection.

[0161] 2. Experimental Results

[0162] The FTIR detection image of substance 5 is as follows Figure 15 As shown, the results show that compared with the FTIR detection chart of 12N-(3-fluorobenzoyl)matrine in Example 2, 3072 cm -1 The peak at 2930 cm is the stretching vibration peak of the methylene group at the newly added fluorine atom, and -1 The stretching vibration peak of alkane -CH2- was enhanced, which confirmed that 12N-(3-fluorobenzoyl)matrine was connected to the 2-fluoroethylamine active group.

[0163] The MS detection image of substance 5 is as follows Figure 16 As shown, the results showed that: 12N-(3-fluorobenzoyl)-18N-(2-fluoroethyl) matrine butyramide (C 24 H 33 The theoretical m / z value of F2N3O2) is 433.54, while a molecular ion peak appears at 434.7 in the MS detection graph of substance 5, which is consistent with the theoretical m / z of 12N-(3-fluorobenzoyl)-18N-(2-fluoroethyl)matrine butanamide.

[0164] Substance 5 1 The results of H NMR examination are: 1 H NMR (500MHz, MeOD) δ7.48 (td, J=7.9, 5.6Hz, 1H), 7.30 (d, J=7.6Hz, 1H), 7.23 (qd, J=8.7, 2. 2Hz, 3H), 4.49 (t, J=5.0Hz, 1H), 4.40 (t, J=5.0Hz, 1H), 4.08 (s, 1H), 3.69-3.54 (m, 1H), 3.4 9 (t, J=5.0Hz, 1H), 3.45 (q, J=5.6, 5.0Hz, 1H), 3.07 (dd, J=26.7, 11.6Hz, 2H), 2.71 (s, 1H), 2.36 (d, J=12.3Hz, 2H), 2.27 (q, J=7.0Hz, 2H), 2.03 (t, J=11.8Hz, 3H), 1.95-1.19 (m, 12H).

[0165] Substance 5 13 The results of C nuclear magnetic resonance imaging are as follows: 13C NMR (126MHz, MeOD) δ176.16, 172.85, 163.97 (dd, J=245.6, 8.7Hz), 140.32 (d, J= 7.1Hz), 131.34 (dd, J=115.8, 7.9Hz), 123.93 (d, J=3.1Hz), 117.81 (d, J=21.3Hz ), 115.03 (d, J = 23.1Hz), 83.18 (d, J = 167.1Hz), 64.23, 57.08 (d, J = 9.8Hz), 55.4 4, 41.09, 40.92, 40.15, 36.37, 35.73, 32.69, 30.74, 28.30, 23.70, 21.31, 21.05.

[0166] The results showed that substance 5 (B4) was 12N-(3-fluorobenzoyl)-18N-(2-fluoroethyl)matrine butyramide, with the molecular formula C 24 H 33 F2N3O2, the chemical structure of which is shown in Formula XII;

[0167]

[0168] Example 8 Effects of Matrine Diamide Derivatives on Spodoptera Frugiperda Sf9 Cells

[0169] 1. Experimental Methods

[0170] 1. Inhibition rate test

[0171] The effects of the matrine obtained in Example 1, A1 prepared in Example 1, B1 prepared in Example 2, A2 prepared in Example 3, A3 prepared in Example 4, B2 prepared in Example 5, B3 prepared in Example 6, and B4 prepared in Example 7 on Spodoptera frugiperda Sf9 cells were detected respectively. Taking the detection of the matrine obtained in Example 1 as an example, the specific results are as follows:

[0172] Spodoptera frugiperda Sf9 cells with good growth conditions were selected and cultured in insect cell culture medium {10% FBS and 90% Grace's Insect Medium, with 1% antibiotics (penicillin-streptomycin solution, w / w) to a cell density of 1×10 5 / mL cell suspension, the cell suspension was inoculated into each cell well of a 96-well plate at 100 μL / well, and placed in a 27 ° C cell constant temperature incubator for pre-cultured for 24 h, and then the matrine (Mat-Ac) obtained in Example 1 was added to the 96-well plate to obtain cell wells with Mat-Ac concentrations of 0.1 mM, 0.4 mM, 0.8 mM and 1.2 mM, respectively. Three experimental time points (24 h, 48 h and 72 h) were set for each cell well of each Mat-Ac concentration, and each cell well of each Mat-Ac concentration was repeated 4 wells at each experimental time point.

[0173] After adding the matrine obtained in Example 1 to the 96-well plate, after the cell wells reached the corresponding experimental time point, methyl thiazolyl blue (MTT) was added to the cell wells at 10 μL / well, sealed and placed in a 27°C constant temperature incubator in the dark for 4 hours, after the treatment, the liquid in the cell wells was discarded, and then dimethyl sulfoxide (DMSO) was added to the cell wells at 150 μL / well, placed in a shaker at 60 rpm / min for 15 minutes, and then placed in a microplate reader to measure the OD value of the cell wells at 429 nm to obtain the OD value of Sf9 cells treated with different concentrations of Mat-Ac for different treatment times (OD 实验组 ).

[0174] Blank control wells: The cell suspension was inoculated into a 96-well plate at 100 μL / well, and 1% (w / w) DMSO was added for culture.

[0175] The OD values ​​of the blank control wells at 24h, 48h and 72h at 429nm were measured using an enzyme marker and recorded as OD 空白组 ; And the inhibition rate of different concentrations of Mat-Ac on Spodoptera frugiperda Sf9 cells was calculated according to formula 1;

[0176] Formula I: Inhibition rate (%) = {1-(OD 实验组 / OD 空白组 )}×100%;

[0177] OD 实验组 and OD 空白组 The OD value is the OD value at the same experimental time (i.e., the same 24 h, the same 48 h, or the same 72 h); the inhibition rate results of each replicate well are expressed as (mean ± standard deviation), and the Duncan multiple comparison method in SPSS27.0 is used to test and analyze the differences, with the test level being P = 0.05.

[0178] The matrine (Mat-Ac) obtained in Example 1 was replaced by A1 prepared in Example 1, and the concentrations of A1 were set to 0.1 mM, 0.4 mM, 0.8 mM, 1.2 mM, 1.6 mM and 2.0 mM, respectively. The inhibition rates of different concentrations of A1 on Sf9 cells at different treatment times were calculated.

[0179] The matrine (Mat-Ac) obtained in Example 1 was replaced by B1 prepared in Example 1, and the concentrations of B1 were set to 0.1 mM, 0.4 mM, 0.8 mM, 1.2 mM, 1.6 mM and 2.0 mM, respectively. The inhibition rates of different concentrations of B1 on Sf9 cells at different treatment times were calculated.

[0180] The matrine (Mat-Ac) obtained in Example 1 was replaced by A2 prepared in Example 1, and the concentrations of A2 were set to 0.05 mM, 0.10 mM, 0.15 mM, 0.20 mM, 0.25 mM and 0.30 mM, respectively, and the inhibition rates of different concentrations of A2 on Sf9 cells at different treatment times were calculated.

[0181] The matrine (Mat-Ac) obtained in Example 1 was replaced by A3 prepared in Example 1, and the concentrations of A3 were set to 0.1 mM, 0.2 mM, 0.3 mM, 0.4 mM and 0.5 mM, respectively. The inhibition rates of different concentrations of A3 on Sf9 cells at different treatment times were calculated.

[0182] The matrine (Mat-Ac) obtained in Example 1 was replaced by B2 prepared in Example 1, and the concentrations of B2 were set to 0.2M, 0.4mM, 0.6mM and 0.8mM, respectively. The inhibition rates of different concentrations of B2 on Sf9 cells at different treatment times were calculated.

[0183] The matrine (Mat-Ac) obtained in Example 1 was replaced by B3 prepared in Example 1, and the concentrations of B3 were set to 0.05 mM, 0.10 mM, 0.15 mM, 0.20 mM, 0.25 mM and 0.30 mM, respectively, and the inhibition rates of different concentrations of B3 on Sf9 cells at different treatment times were calculated.

[0184] The matrine (Mat-Ac) obtained in Example 1 was replaced by B4 prepared in Example 1, and the concentrations of B4 were set to 1.0 mM, 1.5 mM, 2.0 mM, 2.5 mM and 3.0 mM, respectively. The inhibition rates of different concentrations of B4 on Sf9 cells at different treatment times were calculated.

[0185] 2. Observation of Sf9 cell morphology

[0186] Spodoptera frugiperda Sf9 cells with good cell morphology were selected and prepared with insect cell culture medium to a cell density of 1×10 6 Cells were plated at a concentration of 1 mL / well in a 24-well plate, and Mat-Ac, 1.2 mM A1, 1.6 mM A1, 2.0 mM A1, 0.2 mM A2, 0.25 mM A2, 0.30 mM A2, 0.10 mM A3, 0.30 mM A3, 0.50 mM A3, 1.2 mM B1, 1.6 mM B1, 2.0 mM A2, were added to the 24 wells. B1, 0.20mM B2, 0.60mM B2, 0.80mM B2, 0.20mM B3, 0.25mM B3, 0.30mM B3, 1.00mM B4, 2.00mM B4 and 3.00mM B4, each substance at each concentration was added to a different well and cultured at 27°C. The cells were observed under an inverted phase contrast microscope at 0h, 24h, 48h and 72h of culture, and photographed to record the cell morphology.

[0187] 2. Experimental Results

[0188] 1. The inhibition rates of different concentrations of matrine (Mat-Ac) on Spodoptera frugiperda Sf9 are shown in Table 1.

[0189] Table 1 Inhibition rate of different concentrations of matrine (Mat-Ac) on Spodoptera frugiperda Sf9

[0190]

[0191] Note: Different letters in the same column of data represent significant differences (n=3, P<0.05).

[0192] The results showed that after 24 hours of treatment with matrine, except for 0.8mM matrine, which had no inhibitory effect on the proliferation of Sf9 cells, other concentrations of matrine had a weak inhibitory effect on Sf9 cells; after 48 hours of treatment, matrine of various concentrations had no obvious inhibitory effect on the proliferation of Sf9 cells, and the inhibition rate was about 15%; after 72 hours of treatment, matrine of various concentrations showed a certain inhibitory effect on the proliferation of Sf9 cells, among which the inhibition rate of Sf9 cells reached 38.85% when the concentration of matrine was 1.2mM; and the inhibition rate of Sf9 cells gradually increased with the increase of treatment time; the IC value of matrine was 0.8mM, and the inhibition rate of Sf9 cells increased with the increase of treatment time. 50 (half inhibitory concentration) was 7.44 mM.

[0193] The cell morphology of Sf9 cells after culturing with 1.2mM Mat-Ac is shown in the figure below. Figure 17As shown, A is the cell morphology observation picture at 0 h of culture, B is the cell morphology observation picture at 24 h of culture, C is the cell morphology observation picture at 48 h of culture, and D is the cell morphology observation picture at 72 h of culture.

[0194] The results showed that after treatment of Sf9 cells with 1.2 mM Mat-Ac for 24 h and 48 h, there was no obvious change in cell morphology and they were in a normal growth state, but the growth at 48 h was lower than that at 24 h; and after treatment for 72 h, the cells had uneven and bumpy cell membranes, uneven cytoplasm, uneven cell brightness, and were transparent, and the cells appeared vacuolated; this indicated that Mat-Ac had a certain inhibitory effect on Sf9 cells, but the inhibition rate was low.

[0195] 2. The inhibition rates of different concentrations of A1 on Spodoptera frugiperda Sf9 are shown in Table 2.

[0196] Table 2 Inhibition rate of different concentrations of A1 on Spodoptera frugiperda Sf9

[0197]

[0198] Note: Different letters in the same column of data represent significant differences (n=3, P<0.05).

[0199] The results showed that when 0.1mM A1 (12N-(9-fluorenylmethoxy)matrine) was used to treat Sf9 cells, no inhibition rate of A1 on Sf9 cell proliferation was observed within 0 to 72 hours. When the concentration of A1 was ≥0.4mM, the proliferation of Sf9 cells was inhibited after treatment, and the inhibition rate increased with time, indicating that the inhibitory effect of A1 on Sf9 cell proliferation was time-dependent. Moreover, after 72 hours of treatment with 0.4mM A1, the inhibition rate reached 60.11%. After 72 hours of treatment with A1, the inhibition rates of the groups with concentrations ≥0.8mM remained at around 65%, with no significant difference. The half inhibitory concentration (IC50) of A1 (12N-(9-fluorenylmethoxy)matrine) was 1.57kJ / mL. 50 ) is 0.61mM.

[0200] The cell morphology of Sf9 cells after being cultured with different concentrations of A1 is shown in the figure below. Figure 18 As shown, A to D are cell morphology observation pictures of cells cultured with 1.2 mM A1 at 0 h, 24 h, 48 h and 72 h, respectively; E to H are cell morphology observation pictures of cells cultured with 1.6 mM A1 at 0 h, 24 h, 48 h and 72 h, respectively; I to L are cell morphology observation pictures of cells cultured with 2.0 mM A1 at 0 h, 24 h, 48 h and 72 h, respectively; Figure 18 The green circles are normal cells, and the red circles are abnormal cells.

[0201] The results showed that after culturing Sf9 cells with A1 at various concentrations for 24 hours, a small number of cells showed uneven brightness; after culturing for 48 hours, many cells in the cell morphology observation diagram became irregular, with uneven cytoplasm, uneven cell membrane, granular matter on the cell surface, and obvious cell shrinkage and vacuolation; after culturing for 72 hours, a large number of Sf9 cells cultured with A1 at various concentrations showed shrinkage or vacuolation, with only a small number of cells remaining with normal morphology; this indicates that A1 has a good inhibitory effect on Sf9 cells.

[0202] 3. The inhibition rates of different concentrations of B1 on Spodoptera frugiperda Sf9 are shown in Table 3.

[0203] Table 3 Inhibition rate of different concentrations of B1 on Spodoptera frugiperda Sf9

[0204]

[0205] Note: Different letters in the same column of data represent significant differences (n=3, P<0.05).

[0206] The results showed that after culturing Sf9 cells with 0.8 mM B1 (12N-(3-fluorobenzoyl)matrine) for 24 hours, no inhibitory effect was observed on the proliferation of Sf9 cells, while other concentrations of B1 had an inhibitory effect on the proliferation of Sf9 cells; after culturing Sf9 cells with B1 at various concentrations for 48 hours and 72 hours, the proliferation of Sf9 cells was inhibited, and the inhibitory effect on the proliferation of Sf9 cells gradually increased with the increase of B1 concentration and culture time, indicating that the inhibitory effect of B1 on the proliferation of Sf9 cells was concentration-dependent and time-dependent; the half inhibitory concentration (IC50) of B1 (12N-(3-fluorobenzoyl)matrine) was 1. 50 ) is 2.72mM.

[0207] The cell morphology of Sf9 cells after being cultured with different concentrations of B1 is shown in the figure below. Figure 19 As shown, A to D are cell morphology observation pictures of cells cultured with 1.2 mM B1 at 0 h, 24 h, 48 h and 72 h, respectively; E to H are cell morphology observation pictures of cells cultured with 1.6 mM B1 at 0 h, 24 h, 48 h and 72 h, respectively; I to L are cell morphology observation pictures of cells cultured with 2.0 mM B1 at 0 h, 24 h, 48 h and 72 h, respectively; Figure 19 The green circles are normal cells, and the red circles are abnormal cells.

[0208] The results showed that after culturing Sf9 cells with various concentrations of B1 for 24 hours, there was no obvious change in the morphology of Sf9 cells; after culturing for 48 hours, Sf9 cells treated with various concentrations of B1 showed a small amount of cell shrinkage, and some cells did not adhere to the wall, some cells had rough cell membranes, and the cytoplasm was uneven and contained small particles; after culturing for 72 hours, the shrinkage of Sf9 cells treated with various concentrations of B1 increased compared with that at 48 hours of culture, and the cell density decreased compared with that at 0 hours of culture; this indicates that B1 has a certain inhibitory effect on Sf9 cells and has a certain influence on the cell morphology of Sf9.

[0209] 4. The inhibition rates of different concentrations of A2 on Spodoptera frugiperda Sf9 are shown in Table 4.

[0210] Table 4 Inhibition rate of different concentrations of A2 on Spodoptera frugiperda Sf9

[0211]

[0212]

[0213] Note: Different letters in the same column of data represent significant differences (n=3, P<0.05).

[0214] The results showed that after culturing Sf9 cells for 24 hours, A2 (12N-(9-fluorenylmethoxy)-18N-(5-indole)matrine butanamide) at various concentrations had a certain inhibitory effect on the proliferation of Sf9 cells, and the inhibition rate gradually increased with the increase of A2 concentration; after culturing for 48 hours, there was no significant difference in the inhibition rate of Sf9 cells cultured with A2 concentrations ≥ 0.10mM, indicating that 0.10mM A2 had reached a saturation concentration, and the inhibition rate was about 60% after 48 hours of culture, and about 70% after 72 hours of culture; the half inhibitory concentration (IC50) of A2 (12N-(9-fluorenylmethoxy)-18N-(5-indole)matrine butanamide) 50 ) is 0.02mM.

[0215] The cell morphology of Sf9 cells after being cultured with different concentrations of A2 is shown in the figure below. Figure 20 As shown, A to D are cell morphology observation pictures of cells cultured with 0.20 mM A2 at 0 h, 24 h, 48 h and 72 h, respectively; E to H are cell morphology observation pictures of cells cultured with 0.25 mM A2 at 0 h, 24 h, 48 h and 72 h, respectively; I to L are cell morphology observation pictures of cells cultured with 0.30 mM A2 at 0 h, 24 h, 48 h and 72 h, respectively; Figure 20 The green circles are normal cells, and the red circles are abnormal cells.

[0216] The results showed that after culturing Sf9 cells with A2 at various concentrations for 24 hours, a large number of cells showed irregular morphology (indistinct cell membrane, chaotic cytoplasm, and vacuolation); after culturing for 48 hours, the cell vacuolation phenomenon became more serious, the cell morphology became irregular, cell disintegration particles appeared, and the cells did not adhere to the wall; after culturing for 72 hours, the Sf9 cells completely shrank and the cell density decreased significantly, a large number of cells disintegrated, and no normal cells were seen in the cell morphology observation image; this indicates that A2 has significant inhibitory activity on Sf9 cells and can affect cell morphology.

[0217] 5. The inhibition rates of different concentrations of A3 on Spodoptera frugiperda Sf9 are shown in Table 5.

[0218] Table 5 Inhibition rate of different concentrations of A3 on Spodoptera frugiperda Sf9

[0219]

[0220] Note: Different letters in the same column of data represent significant differences (n=3, P<0.05).

[0221] The results showed that after culturing Sf9 cells for 24 hours, A3 (12N-(9-fluorenylmethoxy)-18N-(4-fluorophenyl)matrine butanamide) at various concentrations had a certain inhibitory effect on the proliferation of Sf9 cells, and the inhibition rate gradually increased with the increase of A3 concentration; after culturing for 48 hours, there was no significant difference in the inhibition rate of Sf9 cells cultured with A3 concentration ≥0.20mM, indicating that 0.20mM A3 had reached the saturation concentration, the inhibition rate after 48 hours of culture was about 58%, and the inhibition rate after 72 hours of culture was about 60%; the half-inhibitory concentration of A3 (12N-(9-fluorenylmethoxy)-18N-(4-fluorophenyl)matrine butanamide) was 0.04mM.

[0222] The cell morphology of Sf9 cells after being cultured with different concentrations of A3 is shown in the figure below. Figure 21 As shown, A to D are cell morphology observation pictures of cells cultured with 0.10 mM A3 at 0 h, 24 h, 48 h and 72 h, respectively; E to H are cell morphology observation pictures of cells cultured with 0.30 mM A3 at 0 h, 24 h, 48 h and 72 h, respectively; I to L are cell morphology observation pictures of cells cultured with 0.50 mM A3 at 0 h, 24 h, 48 h and 72 h, respectively; Figure 21 The green circles are normal cells, and the red circles are abnormal cells.

[0223] The results showed that after Sf9 cells were cultured with 0.1mM A3 and 0.3mM A3 for 24h and 48h, Sf9 cells grew normally without obvious changes in cell morphology. After 72h of culture, the cells began to shrink. However, after Sf9 cells were cultured with 0.5mM A3 for 24h, a large number of Sf9 cells showed irregular morphology (unclear cell membrane, chaotic cytoplasm, and vacuolated cells). After 48h of culture, cell vacuolation became more serious, the cells shrank, the cell density decreased significantly, cell disintegration particles appeared, and the cells did not adhere to the wall. After 72h of culture, the Sf9 cells completely shrank and a large number of cells disintegrated.

[0224] This indicates that A3 has a significant inhibitory effect on Sf9 cells, but it only affects cell morphology at high concentrations.

[0225] 6. The inhibition rates of different concentrations of B2 on Spodoptera frugiperda Sf9 are shown in Table 6.

[0226] Table 6 Inhibition rate of different concentrations of B2 on Spodoptera frugiperda Sf9

[0227]

[0228] Note: Different letters in the same column of data represent significant differences (n=3, P<0.05).

[0229] The results showed that after culturing Sf9 cells with various concentrations of B2 (12N-(3-fluorobenzoyl)-18N-(5-indole) matrine butanamide) for 24 hours, the inhibition rate of Sf9 cells was 3% to 25%; after culturing for 48 hours and 72 hours, the inhibition effect increased with the increase of B2 concentration; and for Sf9 cells cultured with the same concentration of B2, the inhibition effect increased with the increase of culture time, indicating that the inhibitory effect of B2 on Sf9 cells was concentration-dependent and time-dependent; after culturing for 72 hours with 0.8mM B2, the inhibition rate of Sf9 cells reached 72.93%; the half inhibitory concentration (IC50) of B2 (12N-(3-fluorobenzoyl)-18N-(5-indole) matrine butanamide) was 1. 50 ) is 0.32mM.

[0230] The cell morphology of Sf9 cells after cultured with different concentrations of B2 is shown in the figure below. Figure 22 As shown, A to D are cell morphology observation pictures of cells cultured with 0.20 mM B2 at 0 h, 24 h, 48 h and 72 h, respectively; E to H are cell morphology observation pictures of cells cultured with 0.60 mM B2 at 0 h, 24 h, 48 h and 72 h, respectively; I to L are cell morphology observation pictures of cells cultured with 0.80 mM B2 at 0 h, 24 h, 48 h and 72 h, respectively; Figure 22 The green circles are normal cells, and the red circles are abnormal cells.

[0231] The results showed that after Sf9 cells were cultured with 0.20mM B2 for 24h and 48h, Sf9 cells grew normally with no obvious changes in cell morphology. After 72h of culture, some Sf9 cells shrank. After 24h of culture with 0.3mM B2, some Sf9 cells shrank. After 48h and 72h of culture, the shrinkage and vacuolation of Sf9 cells increased significantly, and the cell density decreased significantly. After 24h of culture with 0.5mM B2, Sf9 cells showed irregular morphology (unclear cell membrane, chaotic cytoplasm, and vacuolation). After 48h and 72h of culture, cell vacuolation became more serious, the cells shrank, and the cell density decreased significantly. Cell disintegration particles appeared, and the cells did not adhere to the wall.

[0232] 7. The inhibition rates of different concentrations of B3 on Spodoptera frugiperda Sf9 are shown in Table 7.

[0233] Table 7 Inhibition rate of different concentrations of B3 on Spodoptera frugiperda Sf9

[0234]

[0235]

[0236] Note: Different letters in the same column of data represent significant differences (n=3, P<0.05).

[0237] The results showed that after culturing Sf9 cells for 24 hours, the inhibition rate of Sf9 cells with B3 (12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl) matrine butanamide) at various concentrations was about 25%. After culturing for 48 hours and 72 hours, the inhibitory effect on Sf9 cells was significantly enhanced, with the inhibition rate > 60%. This indicated that the inhibitory effect of B3 on the proliferation of Sf9 cells increased with time. B3 had an excellent inhibitory effect on the proliferation of Sf9 cells. After culturing Sf9 cells for 72 hours with 0.3 mM B3, the inhibition rate reached 68.37%. The half inhibitory concentration (IC50) of B3 (12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl) matrine butanamide) was 1.5747 W / cm2. 50 ) is 0.01mM.

[0238] The cell morphology of Sf9 cells after being cultured with different concentrations of B3 is shown in the figure below. Figure 23 As shown, A to D are cell morphology observation pictures of cells cultured with 0.20 mM B3 at 0 h, 24 h, 48 h and 72 h, respectively; E to H are cell morphology observation pictures of cells cultured with 0.25 mM B3 at 0 h, 24 h, 48 h and 72 h, respectively; I to L are cell morphology observation pictures of cells cultured with 0.30 mM B3 at 0 h, 24 h, 48 h and 72 h, respectively; Figure 23 The green circles are normal cells, and the red circles are abnormal cells.

[0239] The results showed that after culturing Sf9 cells with various concentrations of B3 for 24 hours, some Sf9 cells shrank and the cell brightness decreased; after culturing for 48 hours, the morphology of a large number of Sf9 cells changed significantly, the cell membrane edges were uneven, the cytoplasm was uneven, the cells showed shrinkage and vacuolation, cell disintegration particles appeared, and the cells did not adhere to the wall; after culturing Sf9 cells with various concentrations of B3 for 72 hours, almost no normal cells were seen in the cell morphology observation pictures.

[0240] This shows that B3 (12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl) matrine butyramide) has significant cell inhibitory activity, can effectively inhibit Sf9 cells, and has a great impact on the cell morphology of Sf9 cells.

[0241] 8. The inhibition rates of different concentrations of B4 on Spodoptera frugiperda Sf9 are shown in Table 8.

[0242] Table 8 Inhibition rate of different concentrations of B4 on Spodoptera frugiperda Sf9

[0243]

[0244] Note: Different letters in the same column of data represent significant differences (n=3, P<0.05).

[0245] The results showed that when Sf9 cells were cultured with various concentrations of B4 (12N-(3-fluorobenzoyl)-18N-(2-fluoroethyl) berberine butyramide), the higher the concentration of B4, the stronger the inhibitory effect (inhibition rate) on Sf9; and as the culture time increased, the inhibitory effect (inhibition rate) on Sf9 gradually increased.

[0246] The results showed that the inhibitory effect of B4 on Sf9 cells was concentration-dependent and time-dependent. After culturing Sf9 cells with 2.mM B4 for 72h, the inhibition rate was 50.48±2.87%; the half inhibitory concentration (IC50) of B4 (12N-(3-fluorobenzoyl)-18N-(2-fluoroethyl)matrine butanamide) was 1.37±1.03. 50 ) is 2.33mM.

[0247] The cell morphology of Sf9 cells after being cultured with different concentrations of B4 is shown in the figure below. Figure 24 As shown, A to D are cell morphology observation pictures of cells cultured with 1.00 mM B4 at 0 h, 24 h, 48 h and 72 h, respectively; E to H are cell morphology observation pictures of cells cultured with 2.00 mM B4 at 0 h, 24 h, 48 h and 72 h, respectively; I to L are cell morphology observation pictures of cells cultured with 3.00 mM B4 at 0 h, 24 h, 48 h and 72 h, respectively; Figure 24 The green circles are normal cells, and the red circles are abnormal cells.

[0248] The results showed that after culturing Sf9 cells with 1.0 mM B4 for 24, 48, and 72 hours, the vast majority of Sf9 cells grew normally. However, when cultured with 2.0 mM or 3.0 mM B4, a small number of Sf9 cells showed incomplete morphology, cell shrinkage, and vacuolation at 48 and 72 hours, indicating that B4 has a certain inhibitory effect on Sf9 cells, but the inhibitory effect is weak.

[0249] Based on the above experimental results, B3 (12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl) berberine butanamide) and A2 (12N-(9-fluorenylmethoxy)-18N-(5-indole) berberine butanamide) have significant inhibitory activity against Spodoptera frugiperda Sf9 cells; after culturing Sf9 cells with 0.05mM B3 for 72h, the inhibition rate of Sf9 cells was 60.70%; after culturing Sf9 cells with 0.10mM A2 for 72h, the inhibition rate of Sf9 cells was 72.47%.

[0250] Example 9 Insecticidal Activity Test of Matrine Diamide Derivatives

[0251] 1. Experimental Methods

[0252] Hatching and cultivation of 1st and 2nd instar worms

[0253] Place two layers of paper towels in a transparent box, use alcohol-sterilized tweezers to evenly disperse the fall armyworm eggs on the paper towels, cover with two paper towels, cover the transparent box with a lid with air holes, and place it in a 28°C incubator for cultivation. During the cultivation process, ensure 8 hours of light per day. After the eggs hatch into insects, chop the agricultural insect artificial feed (purchased from Henan Jiyuan Baiyun Industrial Co., Ltd.) and evenly disperse it in the transparent box. Continue to cultivate, and select healthy, uniformly sized second-instar fall armyworms as the experimental insects of fall armyworm.

[0254] The eggs of Spodoptera frugiperda were replaced with those of Mythimna separata, and the 28°C culture was replaced with a 25°C incubator. The other treatments were the same, and the second-instar Mythimna separata was obtained as the experimental insects of Mythimna separata.

[0255] 2. Insecticidal activity test of matrine diamide derivatives against fall armyworm

[0256] The matrine (Mat-Ac) obtained in Example 1 was prepared according to "Part 14: Leaf Dipping Method of Insecticides in the Agricultural Industry Standard of the People's Republic of China (NY / T1154.14-2008)" to obtain matrine solutions with concentrations of 0.01 mg / mL, 0.05 mg / mL, 0.10 mg / mL, 0.50 mg / mL and 1.00 mg / mL, respectively.

[0257] Fresh bok choy leaves were cut into 3cm×3cm leaves, immersed in a matrine solution until completely moistened, taken out, air-dried, and naturally air-dried to obtain soaked leaves; 5 leaves were immersed in each concentration of matrine solution, and a total of 25 soaked leaves were obtained (5 leaves for each concentration were repeated); 3cm×3cm leaves were immersed in a liquid containing 0.1% (w / w) Tween and 0.01% (w / w) DMSO solution until completely moistened, taken out, air-dried, and naturally air-dried to obtain blank control leaves.

[0258] The fall armyworm experimental group was as follows: the fall armyworm experimental insect bodies (2nd instar fall armyworm) obtained in step 1 were starved for 5 hours, 10 healthy and uniformly sized 2nd instar fall armyworms were selected and placed in a culture box with filter paper, a drug-soaked leaf was placed in the culture box, and the box was placed in a 28°C incubator for culture. The number of dead insects at 72 hours was recorded, and the mortality rate was calculated according to formula II; 5 groups of 2nd instar fall armyworms were repeatedly treated with drug-soaked leaves of various concentrations, and the final mortality rate was expressed as (mean ± standard deviation).

[0259] Formula II: Mortality rate (%) = (number of dead insects / total number of insects) × 100%.

[0260] The fall armyworm blank control group was as follows: the drug-soaked leaves in the fall armyworm experimental group were replaced with blank control leaves, and the rest of the treatments were the same, and the fall armyworm mortality rate (mortality rate) of the fall armyworm blank control group was obtained. (空白) ).

[0261] The adjusted mortality rate of fall armyworm treated with leaves soaked in each concentration in the fall armyworm experimental group was calculated according to formula III;

[0262] Formula III: Corrected mortality rate (%) = {(mortality rate - mortality rate (空白) ) / (1-mortality rate (空白) )}×100%.

[0263] When the mortality rate of the fall armyworm blank control group is greater than 20%, the experimental data of the fall armyworm experimental group is determined to be invalid, and the second-instar fall armyworms are rehatched according to the method shown in step 1 to conduct the insecticidal experiment.

[0264] The matrine acid obtained in Example 1 was replaced by matrine (Mat), A2 prepared in Example 3, A3 prepared in Example 4, B2 prepared in Example 5, B3 prepared in Example 6 and B4 prepared in Example 7, respectively, to obtain the corrected mortality rates of matrine and each matrine bisamide derivative against fall armyworm.

[0265] The toxicity equations of matrine and matrine diamide derivatives to fall armyworm were calculated. The calculation of the toxicity equations is as shown in the prior art (Huang Minsong. Toxicity regression calculation method and introduction to the use of corresponding software [J]. Anhui Agricultural Sciences, 2014, 42(27): 5. DOI: 10.3969 / j.issn.0517-6611.2014.27.030.).

[0266] 3. Insecticidal activity test of matrine diamide derivatives against armyworms

[0267] The experimental insect bodies of Spodoptera frugiperda (2nd instar insects of Spodoptera frugiperda) in step 2 were replaced with experimental insect bodies of Separata separata (2nd instar insects of Separata separata), and the 28°C incubator was replaced with a 25°C incubator. The other treatments were the same, and the corrected mortality and toxicity equations of matrine acid, matrine and each matrine diamide derivative on Separata separata were obtained.

[0268] 2. Experimental Results

[0269] 1. Test results of insecticidal activity against fall armyworm

[0270] The corrected mortality results of matrine diamide derivatives against fall armyworm are shown in Table 9.

[0271] Table 9 Corrected mortality results of matrine diamide derivatives against fall armyworm

[0272]

[0273] The results showed that after 72 hours of action of 1.00 mg / mL of matrine (Mat) and matrine acid (Mat-Ac) on the fall armyworm, the corrected mortality rates were both 72.14%, indicating that matrine acid had no improved insecticidal activity compared to matrine; while matrine acid, matrine, A2 prepared in Example 3, A3 prepared in Example 4, B2 prepared in Example 5, B3 prepared in Example 6 and B4 prepared in Example 7 were all able to kill the fall armyworm.

[0274] Among them, the adjusted mortality rate of 1.00 mg / mL A2 on fall armyworm after 72 hours of action was as high as 93.57%, which was more than 20% higher than that of matrine and berberine. The adjusted mortality rate of 0.05 mg / mL A2 on fall armyworm after 72 hours of action was also 82.86%; and the adjusted mortality rates of 0.50 mg / mL B2, B3 and B4 on fall armyworm after 72 hours of action were all higher than 70%, which was similar to the effect of 1.00 mg / mL berberine or matrine acid, indicating that the insecticidal activity of the prepared matrine diamide derivatives was significantly enhanced compared with matrine and berberine, and the same effect could be achieved using a lower dose.

[0275] The results of the toxicity of matrine diamide derivatives against Spodoptera frugiperda are shown in Table 10.

[0276] Table 10 The results of the toxicity of matrine diamide derivatives to fall armyworm

[0277] Test drugs Toxicity Equation R <![CDATA[LC 50 (mg / mL)]]> 95% confidence interval Mat y=0.3203x+0.5702 0.982 0.0165 0.0135-0.0203 Mat-Ac y=0.2849x+0.5254 0.964 0.0128 0.0157-0.0104 A2 y=0.6856x+1.3275 0.960 0.0118 0.0099-0.0141 A3 y=0.3213x-0.0422 0.731 1.3335 - B2 y=0.7218x+0.9536 0.974 0.0479 0.0373-0.0616 B3 y=0.3312x+0.7951 0.977 0.0038 0.0031-0.0038 B4 y=0.5766x+0.8148 0.900 0.0401 0.0311-0.0517

[0278] The results showed that the half-lethal concentration (LC50) of B3 prepared in Example 6 against Spodoptera frugiperda was 50 ) was 0.0038 mg / mL, which was significantly lower than the LC 50 (0.0128) and LC of matrine 50 (0.0165); and A2 prepared in Example 3 also has excellent insecticidal activity against fall armyworm, with a half-lethal concentration (LC50) of 50 ) was 0.0118 mg / mL, which was also less than the half-lethal concentration of matrine and berberine.

[0279] Based on the above results, B3 (12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl) berberine butanamide) prepared in Example 6 and A2 (12N-(9-fluorenylmethoxy)-18N-(5-indole) berberine butanamide) prepared in Example 3 have excellent insecticidal effects on Spodoptera frugiperda and the effect of inhibiting the proliferation of Spodoptera frugiperda Sf9 cells.

[0280] 2. Test results of insecticidal activity against armyworms

[0281] The corrected mortality results of matrine diamide derivatives against armyworms are shown in Table 11.

[0282] Table 11 Corrected mortality results of matrine diamide derivatives against armyworms

[0283]

[0284] The results showed that after 72 hours of treatment with matrine (Mat) and matrine acid (Mat-Ac) at 1.00 mg / mL, the adjusted mortality rates were 32.63% and 55.79%, respectively. The insecticidal activity of matrine against armyworms was higher than that of matrine. In addition, A2 prepared in Example 3, A3 prepared in Example 4, B2 prepared in Example 5, B3 prepared in Example 6 and B4 prepared in Example 7 were all able to kill fall armyworms. The prepared B3 (12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl) matrine butanamide) and the prepared A2 (12N-(9-fluorenylmethoxyacyl)-18N-(5-indole) matrine butanamide) in Example 3 both have excellent insecticidal activity against sticky insects. The corrected mortality rate of B3 at a concentration of 0.50 mg / mL against sticky insects after 72 hours of action reached 76.84%, and the corrected mortality rate of A2 at a concentration of 0.50 mg / mL against sticky insects after 72 hours of action reached 74.74%.

[0285] The results of the toxicity of matrine diamide derivatives against Mythimna separata are shown in Table 12.

[0286] Table 12 The results of the toxicity of matrine diamide derivatives to Mythimna separata

[0287] Test drugs Toxicity Equation R <![CDATA[LC 50 (mg / mL)]]> 95% confidence interval Mat y=0.4689x-0.4608 0.967 ＞1 - Mat-Ac y=0.4004x+0.1099 0.994 0.5309 0.4323-0.6520 A2 y=0.3607x+0.8048 0.975 0.0060 0.0049-0.0073 A3 y=0.2375x-0.4379 0.906 ＞1 - B2 y=0.3511x+0.0021 0.832 1.0000 - B3 y=0.4323x+0.9529 0.972 0.0062 0.0051-0.0075 B4 y=0.3190x-1.1912 0.835 ＞1 -

[0288] The results showed that the half-lethal concentration (LC50) of B3 prepared in Example 6 was 50 ) was 0.0062 mg / mL, which was significantly lower than the LC 50 (0.5309) and LC of matrine 50 (>1); The half lethal concentration (LC50) of A2 (12N-(9-fluorenylmethoxy)-18N-(5-indole) matrine butyramide) prepared in Example 3 against Mythimna separata 50 ) was 0.0060 mg / mL, which was also significantly lower than the half-lethal concentration of matrine and berberine to armyworms.

[0289] Based on the above results, B3 (12N-(3-fluorobenzoyl)-18N-(4-fluorophenyl) berberine butanamide) prepared in Example 6 and A2 (12N-(9-fluorenylmethoxy)-18N-(5-indole) berberine butanamide) prepared in Example 3 have excellent insecticidal effects on armyworms.

[0290] Finally, it should be noted that the above embodiments are intended only to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Those skilled in the art will readily appreciate that other variations or modifications may be made based on the above descriptions and concepts. It is not necessary and impossible to provide an exhaustive list of all possible implementations. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention shall be included within the scope of protection of the claims of the present invention.

Claims

1. A class of matrine bisamide derivatives, characterized in that: The chemical structural formulas of the matrine bisamide derivatives are shown in Formula VIII to Formula XI; 2. The matrine bisamide derivative according to claim 1, characterized in that The chemical structural formula of the matrine bisamide derivative is shown in Formula VIII or Formula XI.

3. Use of the matrine bisamide derivative according to any one of claims 1 to 2 in the preparation of products for killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells.

4. The use according to claim 3, characterized in that The agricultural pests are Lepidoptera agricultural pests.

5. The use according to claim 4, characterized in that The lepidopteran agricultural pests are fall armyworm and / or armyworm.

6. The use according to claim 3, characterized in that The agricultural pest cells are Sf9 cells.

7. An insecticide and / or pesticide for killing agricultural pests and / or inhibiting the proliferation of agricultural pest cells, characterized in that: The active ingredient is a matrine bisamide derivative represented by a chemical formula such as Formula VIII or XI; When the matrine bisamide derivative having the chemical structure shown in Formula VIII is used as the active ingredient, the concentration of the active ingredient in the product is 0.02 mM; When the matrine bisamide derivative having the chemical structure shown in Formula XI is used as the active ingredient, the concentration of the active ingredient in the product is 0.01 mM;

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