Spiro [indene-indoline] compound, preparation method thereof and application of spiro [indene-indoline] compound as bactericide

Abstract

The invention relates to the technical field of medicinal chemistry, and discloses a spiro [indene-indoline] compound, a preparation method thereof and an application of the spiro [indene-indoline] compound as a bactericide, in the air atmosphere, benzaldehyde compounds derived from the third position of indole and amine compounds are mixed, a solvent is added, under the action of a catalyst, stirring reaction is performed at the temperature of 25-60 DEG C, and the spiro [indene-indoline] compound is obtained. The structural formula of the spiro [indene-indoline] compound is shown as a formula I in the specification. The spiro [indene-indoline] compound prepared by the invention has excellent inhibitory activity on four common pathogenic fungi: strawberry botrytis cinerea, valsa mali, cucumber fusarium wilt and peanut root rot, and a new solution is provided for overcoming the drug resistance of fungi, so that the spiro [indene-indoline] compound has important application value in the field of agricultural germ control.

Description

Technical Field

[0001] This invention relates to the field of medicinal chemistry, specifically to a spirocyclic [indole-indoline] compound, its preparation method, and its application as a bactericide. Background Technology

[0002] In global agricultural ecosystems, plant fungal diseases have become a core biological stressor restricting crop yield and quality improvement. These diseases infect crop vascular bundles, cortical tissues, or reproductive organs, leading not only to global food crop losses but also posing food safety risks through toxin secretion, thereby creating a cascading effect on the material cycle and energy flow of agro-ecosystems.

[0003] In agricultural production, fungal diseases affecting different crops exhibit significant host specificity and infection specialization. Apple rot, caused by *Valsa mali*, invades the trunk through wounds, forming ulcerative lesions that lead to necrosis and shedding of the cortex. The vascular transport function of diseased plants is impaired, resulting in a decrease in fruit marketability, making it a devastating disease in major apple-producing areas of northern my country. Fusarium wilt, caused by *Fusarium oxysporum* f.sp. *cucumerinum*, can invade through the roots from the seedling stage to the mature plant stage, spreading systemically via the vascular bundle system. In severely affected fields, mortality rates can reach over 80%, causing significant economic losses. Strawberry gray mold, caused by Botrytis cinerea Pers., has a wide host range. Under low temperature and high humidity conditions, it can infect flowers, fruits, and vegetative organs, leading to diseased fruit. Furthermore, conidia are spread by air currents, forming a reinfection cycle, which has become a major plant protection challenge in greenhouse strawberry production. Peanut root rot caused by Fusarium solani can cause seed rot during seed germination, root cortex rot and plant wilting during the growing season, and fruit rot during pod development, resulting in huge economic losses.

[0004] Meanwhile, agricultural pathogens, with their strong survival and transmission capabilities, often overwinter in the soil, diseased plant debris, or seeds in the form of mycelium and spores, forming opportunistic primary sources of infection. Their spread is widespread, and they can spread through rainwater washing, airflow, or agricultural operations. Once environmental conditions such as temperature and humidity are suitable, they quickly initiate a cycle of contact infection, colonization, and reinfection, causing irreversible damage to crops. This not only significantly reduces the yield and quality of agricultural products, but also leads to a substantial increase in pesticide use and labor costs due to repeated control measures, seriously restricting the sustainable development of the agricultural economy and becoming a key problem that urgently needs to be solved in modern agricultural production.

[0005] Currently, the plant fungal disease control system faces multiple technical challenges. Agricultural control methods, such as field sanitation management (e.g., removal of diseased plant debris), require significant labor and are inefficient in large-scale planting environments. Crop rotation is limited by the intensive use of arable land resources and is difficult to implement in monoculture production areas; for example, the rotation rate in my country's greenhouse vegetable production areas is less than 15%. The colonization efficiency of biocontrol agents (such as Trichoderma and Bacillus) is significantly affected by soil microbial community structure, pH, and organic matter content; efficacy fluctuations can reach 40% in acidic soils or high-organic-matter environments. Metabolites (such as chitinase and antibiotics) have poor stability and are easily inactivated under high temperatures or ultraviolet radiation. Chemical control, due to its rapid and efficient characteristics, has become the most widely used control method in agricultural production. However, the lagging development of highly selective agents has exacerbated the risk of target resistance, and long-term reliance on chemical pesticides has made pathogen resistance increasingly prominent, causing some traditional chemical agents to gradually lose their effectiveness and significantly reducing control efficacy. As drug-resistant pathogens continue to grow, the effectiveness of chemical control continues to decline.

[0006] Therefore, developing active molecules with novel mechanisms of action has become a cutting-edge field in international fungicide research and development, and is also key to ensuring agricultural production safety and achieving green and sustainable development. Summary of the Invention

[0007] The purpose of this invention is to overcome the shortcomings of existing technologies and provide a spirocyclic [indole-indoline] compound, its preparation method, and its application as a fungicide. Under the catalysis of bismuth trifluoromethanesulfonate, benzaldehyde compounds derived at the third position of indole react with amine compounds to successfully construct a spirocyclic [indole-indoline] skeleton. These compounds exhibit excellent inhibitory activity against *Botrytis cinerea*, *Fusarium wilt*, *Fusarium wilt*, and *Fusarium root rot*, providing a new option for antibacterial agents.

[0008] To achieve the above objectives, the technical solution of the present invention is: a spirocyclic [indole-indoline] compound, with the structural formula shown in Formula I:

[0009]

[0010] In formula I, R 1 Selected from methyl and ethyl; R 2 Selected from any one of 4-methyl, 5-methyl, 5-methoxy, 5-fluoro, 6-fluoro, 5-chloro, and 6-chloro; R 3 Ar is selected from any one of hydrogen, methyl, ethyl, and n-propyl; Ar is selected from any one of 3,4-(methylenedioxy)aniline, 3,4-dimethoxyaniline, 3,5-dimethoxyaniline, 3-methoxyaniline, 1-naphthylamine, 2-naphthylamine, and 5-bromo-1-naphthylamine.

[0011] Another technical solution of the present invention is: a method for preparing the spirocyclic [indole-indoline] compound, comprising the following steps: under an air atmosphere, a benzaldehyde compound derived from the third position of indole is mixed with an amine compound, a solvent is added, and the mixture is stirred and reacted at 25-60°C under the action of a catalyst to obtain the spirocyclic [indole-indoline] compound.

[0012] Furthermore, the structural formula of the benzaldehyde compound derived at the third position of indole is shown below:

[0013]

[0014] Among them, R 1 Selected from methyl and ethyl; R 2 It is selected from any one of 4-methyl, 5-methyl, 5-methoxy, 5-fluoro, 6-fluoro, 5-chloro, and 6-chloro.

[0015] Furthermore, the structural formula of the amine compound is shown below:

[0016]

[0017] Among them, R 3 Ar is selected from any one of hydrogen, methyl, ethyl, and n-propyl; Ar is selected from any one of 3,4-(methylenedioxy)aniline, 3,4-dimethoxyaniline, 3,5-dimethoxyaniline, 3-methoxyaniline, 1-naphthylamine, 2-naphthylamine, and 5-bromo-1-naphthylamine.

[0018] Further; the molar ratio of the indole-derived benzaldehyde compound at the third position to the amine compound is 1:(1-1.5), the catalyst is bismuth trifluoromethanesulfonate, and the molar ratio of the catalyst to the indole-derived benzaldehyde compound at the third position is (0.2-0.5):1.

[0019] Furthermore, the solvent is selected from any one of hexafluoroisopropanol, ethanol, methanol, trifluoroethanol, isopropanol, tert-butanol, 1,2-dichloroethane, and tetrahydrofuran.

[0020] Furthermore, the amount of solvent used is 10-20 L of solvent added per mole of benzaldehyde compound derived from the third position of indole.

[0021] Furthermore, the spirocyclic [indole-indoline] compound has any one of the structures shown in formulas I-1 to I-20:

[0022]

[0023] Another technical solution of the present invention is the application of the aforementioned spirocyclic [indole-indoline] compound as a bactericide.

[0024] Furthermore, the spirocyclic [indole-indoline] compounds are used to prepare fungicides that inhibit strawberry gray mold, apple rot fungus, cucumber wilt fungus, or peanut root rot fungus.

[0025] The beneficial effects of this invention are:

[0026] (1) The spirocyclic [indole-indoline] compounds prepared in this invention have not been reported. Under the condition of Lewis acid as catalyst, electron-rich amines are more likely to nucleophilically attack indole aldehyde derivatives, resulting in aldehyde-amine condensation to generate imine intermediates, followed by [4+2] cycloaddition reaction, thereby realizing the construction of spirocyclic [indole-indoline] skeleton molecules.

[0027] (2) The reaction environment of this invention is relatively mild, without the need for harsh high temperature, high pressure or extreme acid and alkalinity conditions, which reduces the requirements for reaction equipment, reduces equipment costs and safety risks, and can effectively avoid side reactions that may occur under violent reaction conditions, ensuring the stability of the reaction system. It can also achieve efficient conversion to the target product in a short time, which greatly improves production efficiency and provides an important method for synthesizing spirocyclic [indole-indoline] compounds with specific structures and functions.

[0028] (3) The spirocyclic [indole-indoline] compounds prepared in this invention have excellent inhibitory activity against four common pathogenic fungi: strawberry gray mold, apple rot fungus, cucumber wilt fungus and peanut root rot fungus, providing a new solution to overcome fungal resistance, and thus have important application value in the field of agricultural pathogen control. Attached Figure Description

[0029] Figure 1 An image showing the inhibition of the fungus that causes cucumber wilt;

[0030] Figure 2 An illustration of the inhibition of bacteria that cause apple rot;

[0031] Figure 3 An image showing the inhibition of the root rot pathogen in peanuts;

[0032] Figure 4 An image showing the inhibition of gray mold pathogens in strawberries;

[0033] Figure 5 The 1H NMR spectrum of compound I-1, the product of Example 1;

[0034] Figure 6 The image shows the carbon NMR spectrum of compound I-1, the product of Example 1. Detailed Implementation

[0035] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Unless otherwise defined, all technical and scientific terms used in this specification have the same meaning as commonly understood by those skilled in the art to which this invention pertains.

[0036] Unless otherwise specified, the experimental methods used in the following examples are conventional methods; unless otherwise specified, the reagents, materials, instruments, etc. used in the following examples are all commercially available.

[0037] Example 1:

[0038] Preparation of a spirocyclic [indole-indoline] compound

[0039] The chemical reaction formula is shown below:

[0040]

[0041] The steps are as follows:

[0042] 0.1 mmol of 2-((1H-indol-3-yl)methyl)benzaldehyde, 0.12 mmol of benzo[d][1,3]dioxacyclopentadien-5-amine, and 0.02 mmol of bismuth trifluoromethanesulfonate were placed in a reaction flask, and 1 mL of isopropanol was added. The system temperature was controlled at 25 °C, and the reaction was stirred. The reaction was monitored by spotting the sample onto a thin-layer chromatography plate until the reactants were completely reacted. After the reaction was completed, the solid product was collected by filtration and washed with isopropanol to obtain 29.5 mg of the target product I-1 (16-methyl-9b,10,15b,16-tetrahydro-5H-[1,3]dioxacyclopenten[4,5-g]indo[1,2-b]indol[3,2-c]quinoline), a white solid, yield: 80%, melting point (mp): 225-226 °C. 1H NMR (500MHz, CDCl3) δ7.27 (d, J=2.0Hz, 1H), 7.26–7.17 (m, 3H), 7.15 (td, J=7. 5,1.5Hz,1H),6.74(dd,J=7.5,1.5Hz,1H),6.65(td,J=7.5,1.0Hz,1H),6.61(s ,1H),6.59(d,J=8.0Hz,1H),6.16(s,1H),5.83(dd,J=9.0,1.5Hz,2H),4.44(s, 1H),3.88(s,1H),3.37(d,J=16.5Hz,1H),3.14(d,J=16.0Hz,1H),2.76(s,3H). 13 C NMR (126MHz, CDCl3) δ151.73,148.25,143.57,142.77,139.64,139.39,135.84,128.47,128.28,127.29,124.66,124.15 ,122.09,118.77,111.21,110.64,108.39,100.69,97.80,73.22,65.22,54.77,42.50,33.96.HRMS(ESI-TOF):m / z[M+H] + calcd for C 24 H 21 N2O2:369.1598, found:369.1596.

[0043] Example 2-20

[0044] Following the preparation method of Example 1: 0.1 mmol of an indole-derived benzaldehyde compound at the 3-position, 0.12 mmol of an amine compound, and 0.02 mmol of bismuth trifluoromethanesulfonate were placed in a reaction flask, and 1 mL of isopropanol was added. The system temperature was controlled at 25°C, and the reaction was stirred. The reaction was monitored by spotting the sample onto a thin-layer chromatography plate until the reactants reacted completely. After the reaction was complete, the solid product was collected by filtration and washed with isopropanol to obtain the target product. See Table 1. The structural characterization data of the product are shown in Table 2.

[0045] Table 1. Indole-derived benzaldehyde compounds and amine compounds used in Examples 2-20, and the target products.

[0046]

[0047]

[0048]

[0049]

[0050]

[0051] Table 2. Structural characterization data of products from Examples 2-20

[0052]

[0053]

[0054]

[0055]

[0056]

[0057]

[0058]

[0059]

[0060]

[0061]

[0062] II. Antibacterial Activity Test of Spirocyclic [Indole-Indoline] Compounds

[0063] Four product compounds I-1, I-7, I-14 and I-16 prepared in Examples 1, 7, 14 and 16 were selected as test samples for antibacterial activity testing.

[0064] The testing procedure was as follows: Four pathogenic fungi were selected: *Fusarium oxysporum* f. sp. *cucumerinum*, *Valsa mali*, *Fusarium solani*, and *Botrytiscinerea* Pers. The antifungal activity of the compounds was tested using the mycelial growth rate method (Ann. Appl. Biol., 2008, 152, 369). All the tested fungi were provided by the Shandong Provincial Engineering Technology Research Center for Biological Pesticides.

[0065] First, weigh 39g of potato dextrose agar into a wide-mouth flask, add 1000mL of distilled water to prepare the culture medium, and then sterilize it in an autoclave at 121℃ for 30 minutes. Weigh 1mg of each test sample and dissolve it in 10mL of acetone to prepare a 100mg / L antibacterial stock solution. Take 5mL of the stock solution and dilute it to 10mL with acetone to obtain a 50mg / L antibacterial agent. Prepare a series of antibacterial agents at 25mg / L and 12.5mg / L using this gradient dilution method. Take 5mL of each concentration of antibacterial agent and add it to 50mL of sterilized culture medium, mix thoroughly, and then evenly distribute the mixture into 5 autoclaved petri dishes. After the culture medium cools and solidifies, inoculate the above four pathogens with an inoculation loop, seal the petri dishes with sealing film, and transfer them to a 25℃ incubator for incubation and observation. When the colony coverage area in the control group culture medium without added antibacterial agent reached 80%, the colony diameter of each experimental group was measured using the cross-cross method.

[0066] The antibacterial activity formula is as follows:

[0067] Inhibition rate = (Coronavirus diameter in control group - Coronavirus diameter in experimental group) / Coronavirus diameter in control group × 100%;

[0068] The experimental results are shown in Table 3:

[0069] Table 3 Antibacterial rates of each compound

[0070]

[0071] As shown in Table 3, compounds I-1, I-7, I-14 and I-16 all exhibited good inhibitory activity against Fusarium wilt of cucumber, Fusarium rot of apple, Fusarium root rot of peanut and gray mold of strawberry.

[0072] For *Fusarium wilt*, the pathogen of cucumber, compounds I-1 and I-14 showed no growth at any of the three tested concentrations. Compounds I-7 and I-16 also achieved inhibition rates of over 50% at lower concentrations, indicating that these compounds have good inhibitory effects against *Fusarium wilt*. Figure 1 The diagram shows the hyphal growth of the four compounds.

[0073] For apple rot pathogens, such as Figure 2 As shown, compounds I-1, I-14, and I-16 exhibited 100% inhibition rates at the three tested concentrations, demonstrating excellent antibacterial activity. Compound I-7 showed good antibacterial activity at high concentrations, but its antibacterial activity decreased with decreasing concentration.

[0074] For peanut root rot pathogens, such as Figure 3As shown, at the lowest inhibitory concentration of 12.5 mg / L, compound I-14 showed no mycelial growth and an inhibition rate of 100%. Compounds I-1, I-7, and I-16 showed good antibacterial effects at high concentrations but poor antibacterial effects at low concentrations.

[0075] For strawberry gray mold, such as Figure 4 As shown, the four compounds tested had good antibacterial effects, and no pathogens grew at any of the three tested concentrations, indicating that the compounds had good antibacterial effects against strawberry gray mold.

[0076] From an applied research perspective, the functional value of the spirocyclic [indole-indoline] compounds of this invention is reflected in two aspects: First, as raw material properties for direct synthesis, the molecular skeleton of these compounds, due to its specific electron cloud distribution and functional group activity, can be directly incorporated into the synthetic pathway of antibacterial drugs. Target molecules can be constructed through classic organic reactions such as acylation and alkylation, shortening the drug development cycle. For example, in the preparation of agricultural fungicides, its spirocyclic structure can specifically bind to target enzymes within pathogenic bacteria, achieving inhibitory effects on fungal growth. Second, as a core parent structure for structural derivation, based on the spirocyclic [indole-indoline] skeleton, chemical modifications can be made by introducing halogen atoms, heterocyclic groups, or polar functional groups (such as hydroxyl and carboxyl groups).

[0077] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the present invention in any way. Other variations and modifications may be made without departing from the technical solutions described in the claims.

Claims

1. A spirocyclic [indole-indoline] compound, characterized in that, The structural formula is shown in Formula I: In formula I, R 1 Selected from methyl and ethyl; R 2 Selected from any one of 4-methyl, 5-methyl, 5-methoxy, 5-fluoro, 6-fluoro, 5-chloro, and 6-chloro; R 3 Ar is selected from any one of hydrogen, methyl, ethyl, and n-propyl; Ar is selected from any one of 3,4-(methylenedioxy)aniline, 3,4-dimethoxyaniline, 3,5-dimethoxyaniline, 3-methoxyaniline, 1-naphthylamine, 2-naphthylamine, and 5-bromo-1-naphthylamine.

2. The spirocyclic [indole-indoline] compound according to claim 1, characterized in that, The spirocyclic [indole-indoline] compounds have any one of the structures shown in formulas I-1 to I-20:

3. A method for preparing the spirocyclic [indole-indoline] compound according to claim 1, characterized in that, The steps are as follows: In an air atmosphere, benzaldehyde compounds derived from the third position of indole are mixed with amine compounds, a solvent is added, and the mixture is stirred and reacted at 25-60°C under the action of a catalyst to obtain spirocyclic [indole-indoline] compounds.

4. The preparation method according to claim 3, characterized in that, The structural formula of the benzaldehyde compound derived from the indole third position is shown below: Among them, R 1 Selected from methyl and ethyl; R 2 It is selected from any one of 4-methyl, 5-methyl, 5-methoxy, 5-fluoro, 6-fluoro, 5-chloro, and 6-chloro.

5. The preparation method according to claim 3, characterized in that, The structural formula of the amine compound is shown below: Among them, R 3 Ar is selected from any one of hydrogen, methyl, ethyl, and n-propyl; Ar is selected from any one of 3,4-(methylenedioxy)aniline, 3,4-dimethoxyaniline, 3,5-dimethoxyaniline, 3-methoxyaniline, 1-naphthylamine, 2-naphthylamine, and 5-bromo-1-naphthylamine.

6. The preparation method according to claim 3, characterized in that: The molar ratio of the indole-derived benzaldehyde compound at the third position to the amine compound is 1:(1-1.5), the catalyst is bismuth trifluoromethanesulfonate, and the molar ratio of the catalyst to the indole-derived benzaldehyde compound at the third position is (0.2-0.5):

1.

7. The preparation method according to claim 3, characterized in that: The solvent is selected from any one of hexafluoroisopropanol, ethanol, methanol, trifluoroethanol, isopropanol, tert-butanol, 1,2-dichloroethane, and tetrahydrofuran.

8. The preparation method according to claim 3, characterized in that: The amount of solvent used is 10-20 L of solvent per mole of benzaldehyde compound derived from the third position of indole.

9. The application of the spirocyclic [indole-indoline] compound of claim 1 as a bactericide.

10. The application according to claim 9, characterized in that: The spirocyclic [indole-indoline] compounds are used to prepare fungicides that inhibit strawberry gray mold, apple rot fungus, cucumber wilt fungus, or peanut root rot fungus.

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