Acylhydrazone compound, preparation method thereof and application of acylhydrazone compound in anthracnose and soft rot of camellia oleifera

Abstract

The invention belongs to the field of chemical medicine synthesis, and particularly relates to an acylhydrazone compound, a preparation method thereof and application of the acylhydrazone compound in camellia oleifera anthracnose and soft rot. The structure of the acylhydrazone compound is shown as a formula I, and the acylhydrazone compound is directly prepared from citronellal and formylhydrazine through a one-step reaction. The invention provides the acylhydrazone compound, the IC50 of the acylhydrazone compound on liver cancer can reach 7.0 mu M, and a new solution is provided for research and development of drugs for treating liver cancer. The EC50 (mg / L) of the strain can reach 1.65 when the strain is used in an oil-tea camellia anthracnose strain inhibition experiment, the EC50 (mg / L) of the strain can reach 1.38 when the strain is used in an oil-tea camellia soft rot strain inhibition experiment, and a new solution is provided for controlling oil-tea camellia anthracnose and soft rot. Meanwhile, the invention provides a synthesis method of citronellal and formylhydrazine through one-step reaction, and the synthesis method has the advantages of high yield, high raw material conversion rate, high selectivity and the like. And the formula I of the # imgabs0 # is shown in the specification.

Description

Technical Field

[0001] The present invention belongs to the field of chemical drug synthesis, and particularly relates to a hydrazone compound, a preparation method thereof, and applications in Camellia oleifera anthracnose and soft rot disease. Background Art

[0002] Liver cancer is the most common malignant tumor disease. According to statistics, in 2020, there were 906,000 newly diagnosed liver cancer patients globally (accounting for 4.7% of all newly diagnosed cancers), ranking 6th in all cancer incidences and 3rd in all cancer deaths. In China, the incidence and mortality rates of liver cancer both account for more than 50% of the global total and about 70% of the Asian total. In recent years, although the early diagnosis technology and treatment level of liver cancer have been continuously improved, the 5-year survival rate of patients is still not ideal. Therefore, studying the in vitro inhibitory effect of compounds on liver cancer HepG2 cells can lay a research foundation for the development of related drugs.

[0003] Camellia oleifera anthracnose and Camellia oleifera soft rot disease are the main factors affecting the growth and development of Camellia oleifera. Camellia oleifera anthracnose often causes serious fruit drop and flower bud drop of Camellia oleifera, and even the whole plant withers. In each province (region), the yield of Camellia oleifera seeds is often reduced by 10 - 30% due to this disease, and can reach 40 - 50% in severely affected areas. Camellia oleifera soft rot disease is another important disease in Camellia oleifera production areas, which harms the leaves and fruits of Camellia oleifera, often causing a large amount of flower and fruit drop, resulting in significant economic losses. Therefore, studying the inhibitory effect of compounds on Camellia oleifera anthracnose strains and soft rot strains can lay a research foundation for the development of related drugs. Summary of the Invention

[0004] The purpose of the present invention is to solve the deficiencies of the prior art and provide a hydrazone compound, the IC 50 (μM) of which in the HepG2 cell inhibition experiment can reach 7.0, providing a new solution for the drug treatment of liver cancer; and in the agricultural field, the EC50 (mg / L) of which in the Camellia oleifera anthracnose strain inhibition experiment can reach 1.65, and the EC50 (mg / L) in the Camellia oleifera soft rot strain inhibition experiment can reach 1.38, providing a new solution for controlling Camellia oleifera anthracnose and soft rot disease. The specific technical solution is as follows: A hydrazone compound, the structure of which is shown in Formula I: Formula I.

[0005] Acylhydrazone compounds contain the acylhydrazone group (-CONHN=CH-), and exhibit excellent and diverse biological activities, such as insecticidal, herbicidal, plant growth regulation, bactericidal, anti-inflammatory, analgesic, anti-tuberculosis, etc. Therefore, acylhydrazone compounds have a wide range of applications in the fields of pesticides, medicine, biology, agriculture, etc. However, there is no report on the preparation of citronellal acylhydrazone compounds and their inhibition of HepG2 tumor cells, and there is also no report on their use in inhibiting Camellia oleifera anthracnose and soft rot.

[0006] The present invention also provides a preparation method of the above-mentioned acylhydrazone compounds. The main method is that under the condition of using methanol as a solvent, the raw materials citronellal and formylhydrazine are directly prepared into the target compound through a one-step reaction. This synthesis reaction has the advantages of high yield, high conversion rate of raw materials, high selectivity, etc. The specific process of the reaction between citronellal and formylhydrazine is as follows: React formylhydrazine with citronellal at 40 °C - 60 °C. After the reaction is complete, cool it to room temperature, then add ethyl acetate, remove the solvent, load the sample by dry method, and obtain the acylhydrazone compound through column chromatography analysis. Among them, the eluent used for column chromatography separation is composed of petroleum ether and ethyl acetate with a volume ratio of 5:1. Under the same conditions, when using ethyl acetate, ethanol, toluene, etc. as solvents, the reaction yield, raw material conversion rate, content of the target compound, etc. are all lower than those under the reaction conditions when using methanol as a solvent.

[0007] Preferably, the molar ratio of formylhydrazine to citronellal is 1:(1 - 1.5). More preferably, it is 1:1.2. When the molar ratio of the materials is lower than 1:1 or higher than 1:1.5, there are residues of acylhydrazine raw materials or citronellal raw materials, and there are many side reactions, the content of the target compound in the product is low, and the reaction yield is greatly affected.

[0008] When the above-mentioned acylhydrazone compounds are used in the preparation of drugs for treating liver cancer or in the preparation of drugs for treating Camellia oleifera anthracnose and soft rot, they can be in the form of a pharmaceutical composition, which in addition to including the above-mentioned acylhydrazone compounds, also includes other pharmaceutically acceptable adjuvants. The adjuvants can be solvents or carriers; for example, water and oils (peanut oil, soybean oil, mineral oil, etc.).

[0009] The beneficial effects of the present invention are as follows: The present invention provides an acylhydrazone compound. According to the data results of the examples of the present invention, its IC 50 for liver cancer can reach 7.0 μM, providing a new solution for the drug research and development of liver cancer. Its EC50 (mg / L) in the inhibition experiment of Camellia oleifera anthracnose strains can reach 1.65, and its EC50 (mg / L) in the inhibition experiment of Camellia oleifera soft rot strains can reach 1.38, providing a new solution for controlling Camellia oleifera anthracnose and soft rot. At the same time, the present invention proposes a synthesis method of one-step reaction of citronellal and formylhydrazine, which has the advantages of high yield, high conversion rate of raw materials, high selectivity, etc. Brief Description of the Drawings

[0010] Figure 1 Shown is the 13 13C NMR spectrum of the compound of Formula I; Figure 2 Shown is the 1 1H NMR spectrum of the compound of Formula I. Detailed Description of the Invention

[0011] The following will clearly and completely describe the concept of the present invention and the technical effects produced in combination with the embodiments, so as to fully understand the purpose, solution and effects of the present invention.

[0012] Example 1: A hydrazone compound, the structure of which is shown in Formula I: Formula I.

[0013] Its preparation method includes the following steps: Add formohydrazide (1 eqiv (equivalent)), citronellal (1.2 eqiv) and 5 mL of methanol into a 10 mL round-bottom flask, and react with full stirring at 50 °C using an HJ-2A magnetic stirring reactor, and monitor the reaction by TLC until it is complete. After the reaction is complete, the system is cooled to room temperature, ethyl acetate is added, the solvent is evaporated to dryness using a RE-301 rotary evaporator, passed through a 20 mm×50 mm chromatography column, and then loaded onto the column by dry method, and separated by column chromatography (eluent, petroleum ether:ethyl acetate = 5:1) to obtain the target compound N'-(3,7-dimethyloct-6-en-1-ylidene)formohydrazide.

[0014] The compound was detected using a Bruker-400Hz nuclear magnetic resonance spectrometer and a Thermo-QE high-resolution liquid chromatography-mass spectrometry instrument, and its carbon spectrum and hydrogen spectrum are respectively as Figure 1-2 shown, and the specific results are as follows: Brown oily liquid, yield 32%; 1 1HNMR (400 MHz, CDCl3) δ 10.72 (d, J J = 8.8 Hz, 1H), 8.62 (d, J J = 8.8 Hz, 1H), 7.31(t, J J = 6.0 Hz, 1H), 5.08 (t, J= 7.2 Hz, 1H), 2.30 - 2.24 (m, 1H), 2.13 - 1.96 (m, 3H), 1.77 - 1.72 (m, 1H), 1.68 (s, 3H), 1.60 (s, 3H), 1.42 - 1.18 (m, 2H), 0.94 (d, J = 3.2 Hz, 3H). 13 C NMR (100 MHz, CDCl3) δ 165.7, 150.2, 131.5, 124.2, 39.3, 36.7, 30.8, 25.6, 25.4, 19.5, 17.6.

[0015] Example 2: An acylhydrazone compound, the structure of which is shown in Formula II: Formula II.

[0016] Its preparation method includes the following steps: Add 4-fluorobenzohydrazide (1 eqiv), citronellal (1.2 eqiv) and 5 mL of methanol into a 10 mL round-bottom flask, and stir well at 50 °C using an HJ-2A magnetic stirring reactor for reaction. Monitor the reaction by TLC until it is complete. After the reaction is complete, cool the system to room temperature, add ethyl acetate, rotary evaporate the solvent using an RE-301 rotary evaporator, pass through a 20 mm × 50 mm chromatography column, and then perform dry loading. Column chromatography separation (eluent, petroleum ether:ethyl acetate = 5:1) to obtain the target compound.

[0017] The compound was detected using a Bruker-400Hz nuclear magnetic resonance spectrometer and a Thermo-QE high-resolution liquid chromatography-mass spectrometry instrument. The characterization results of the compound: Brown oily liquid, yield 84%; 1 H NMR (400 MHz, CDCl3) δ 11.30 (s, 1H), 7.96 - 7.83 (m, 3H), 6.99 (t, J = 3.6 Hz, 2H), 5.00 (s, 1H), 2.27 - 2.20 (m, 1H), 2.06 - 2.01 (m, 1H), 1.92 - 1.89 (m, 2H), 1.67 - 1.55 (m, 4H), 1.55 (s, 3H), 1.33 - 1.26 (m, 1H), 1.13 - 1.16 (m, 1H), 0.84 (d, J = 3.2 Hz, 3H). 1313C NMR (100 MHz, CDCl3) δ 164.9 (d, 1 J CF J = 251.0 Hz), 163.8, 153.5, 131.4, 130.3 (d, 3 J CF J = 8.7 Hz), 129.1, 124.2, 115.3 (d, 2 J CF J = 21.6 Hz), 39.7, 36.8, 31.1, 25.6, 25.4, 19.4, 17.5。

[0018] Example 3: An acylhydrazone compound, the structure of which is shown in Formula III: Formula III.

[0019] Its preparation method includes the following steps: Add benzohydrazide (1 eqiv), citronellal (1.2 eqiv) and 5 mL of methanol into a 10 mL round-bottom flask, and stir thoroughly at 50 °C using an HJ-2A magnetic stirring reactor for reaction. Monitor the reaction by TLC until it is complete. After the reaction is complete, cool the system to room temperature, add ethyl acetate, spin-dry the solvent using a RE-301 rotary evaporator, pass through a 20 mm × 50 mm chromatography column, and then perform dry loading. Column chromatography separation (eluent, petroleum ether:ethyl acetate = 5:1) to obtain the target compound N'-(3,7-dimethyloct-6-en-1-ylidene)benzohydrazide.

[0020] The compound was detected using a Bruker-400Hz nuclear magnetic resonance spectrometer and a Thermo-QE high-resolution liquid chromatography-mass spectrometry instrument. The characterization results of the compound: Brown oily liquid, yield 89%; 1 1H NMR (400 MHz, CDCl3) δ10.9 (s, 1H), 7.86-7.76 (m, 3H), 7.41 (t, J J = 6.8 Hz, 1H), 7.30 (t, J J = 7.6 Hz,2H), 5.02 (t, J= 6.8 Hz, 1H), 2.28 - 2.22 (m, 1H), 2.11 - 2.04 (m, 1H), 1.97 - 1.87(m, 2H), 1.67 - 1.65 (m, 4H), 1.56 (s, 3H), 1.34 - 1.27 (m, 1H), 1.19 - 1.10 (m,1H), 0.85 (d, J = 3.2 Hz, 3H). 13 C NMR (100 MHz, CDCl3) δ 164.2, 152.4, 133.3,131.8, 131.5, 128.5, 127.4, 124.3, 39.5, 36.9, 31.2, 25.7, 25.4, 19.5, 17.6.

[0021] Example 4: The HepG2 cell suspension was inoculated into a 96 - well plate at 100 μL per well (1×10 4 / well) and incubated overnight. The test compound was used, and solutions containing different concentrations (5, 10, 20 μmol / L) were added respectively. The control group was added with an equal volume of 0.1% DMSO. They were incubated for 24, 48, and 72 h respectively. After the incubation, 20 μL of MTT solution (5 mg / mL) was added and incubated in the incubator for 4 h. After the experiment, the supernatant was aspirated, 150 μL of DMSO was added, and the precipitate was dissolved by shaking at 37 °C for 10 min. The absorbance OD value at 570 nm was measured using a microplate reader.

[0022] The specific results are shown in Table 1.

[0023] Table 1 Compound IC50 (μM) Example 1 7.0 Example 2 23.0 Example 3 25.4 As can be seen from Table 1, the compound with the structure of formula I of the present invention has good inhibitory activity against HepG2, while not all compounds with similar structures have this inhibitory activity.

[0024] Example 5 After autoclaving the PDA solid culture, pour an equal amount into a petri dish. For the basic test compound, add solutions with different concentrations (1, 3, 5, 7, 10 mg / L) to the PDA medium. Add an equal amount of sterile water to the control group. Invert a camellia anthracnose fungal cake with consistent growth in the center of the culture medium plate containing compounds with different concentrations, and culture it upside down in an incubator at a constant temperature of 25°C. After 7 days, use the "cross-cross method" to measure the hyphal growth diameter in each plate, calculate the average hyphal growth diameter and relative inhibition rate at each concentration gradient, and conduct statistical analysis in DPS to obtain the virulence regression equation and EC50 value. The method for camellia soft rot disease is the same as the above experimental process.

[0025] The specific results are shown in Table 2 as follows.

[0026] Table 2 Compound EC50 (mg / L) Anthrax EC50 (mg / L) Soft Rot Example 1 1.65 1.38 Example 2 5.74 6.67 Example 3 4.95 6.91 As can be seen from Table 2, the compound with the structure of Formula I in the present invention has good inhibitory activity against camellia anthracnose and soft rot diseases, while not all compounds with similar structures possess this inhibitory activity.

[0027] As mentioned above, these are only the preferred embodiments of the present invention. The present invention is not limited to the above-mentioned implementation manners. As long as the same means are used to achieve the technical effects of the present invention, they should fall within the protection scope of the present invention. Within the protection scope of the present invention, various different modifications and changes can be made to its technical solutions and / or implementation manners.

Claims

1. An acylhydrazone compound, characterized in that, Its structure is shown in Formula I: Formula I.

2. A method for preparing the acylhydrazone compound according to claim 1, characterized in that, It includes the following steps: It is prepared by reacting citronellal with formylhydrazine.

3. The preparation method according to claim 2, characterized in that, The molar ratio of the formylhydrazine to citronellal is 1:(1 - 1.5).

4. The preparation method according to claim 3, characterized in that, The molar ratio of the formylhydrazine to citronellal is 1:1.

2.

5. The preparation method according to claim 2, characterized in that, The specific process of the reaction between citronellal and formylhydrazine is as follows: React formylhydrazine and citronellal under the condition of 40 °C - 60 °C. After the reaction is complete, cool it to room temperature, then add ethyl acetate, remove the solvent, load the sample by dry method, and analyze by column chromatography to obtain the acylhydrazone compound.

6. The preparation method according to claim 5, characterized in that, The eluent used for column chromatography separation is composed of petroleum ether and ethyl acetate with a volume ratio of 5:

1.

7. The preparation method according to claim 2, characterized in that The reaction is carried out under the condition that methanol is used as the solvent.

8. Use of the acylhydrazone compound according to claim 1 in the preparation of a medicament for treating liver cancer.

9. Use of the acylhydrazone compound according to claim 1 in the preparation of a medicament for treating camellia anthracnose and / or soft rot.

10. A pharmaceutical composition, characterized in that, It includes the acylhydrazone compound according to claim 1 and also includes other pharmaceutically acceptable adjuvants.

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