CN107522766B

CN107522766B - Ursolic acid quinolyl hydrazide derivatives with anti-tumor activity and preparation method and application thereof

Info

Publication number: CN107522766B
Application number: CN201710718110.5A
Authority: CN
Inventors: 谷文; 金晓燕; 陶徐兵; 陈浩; 耿艺; 王石发
Original assignee: Nanjing Forestry University
Current assignee: Nanjing Forestry University
Priority date: 2017-08-21
Filing date: 2017-08-21
Publication date: 2020-03-27
Anticipated expiration: 2037-08-21
Also published as: CN107522766A

Abstract

Discloses ursolic acid quinoline hydrazide derivatives with anti-tumor activity, a preparation method and application thereof. The invention relates to ursolic acid quinoline hydrazide heterocyclic derivatives with a structure shown in a general formula I and pharmaceutically acceptable salts thereof:

wherein, I-a: r₁＝H R₂＝CH₃；I‑b：R₁＝OMe R₂＝CH₃；I‑c：R₁＝F R₂＝CH₃；I‑d：R₁＝Cl R₂＝CH₃；I‑e：R₁＝H R₂＝n‑C₄H₉；I‑f：R₁＝OMe R₂＝n‑C₄H₉；I‑g：R₁＝F R₂＝n‑C₄H₉；I‑h：R₁＝Cl R₂＝n‑C₄H₉The ursolic acid quinoline hydrazide heterocyclic derivatives and the pharmaceutically acceptable salts thereof have obvious antitumor activity, and pharmacological experiments show that the ursolic acid quinoline hydrazide heterocyclic derivatives have obvious inhibition effects on human breast cancer cells MDA-MB-231, human cervical cancer cells HeLa and human liver cancer cells SMMC-7721, show low toxicity on human normal liver epithelial cells QSG-7701, and have the potential of developing antitumor drugs.

Description

Ursolic acid quinolyl hydrazide derivatives with anti-tumor activity and preparation method and application thereof

Technical Field

The invention relates to the field of organic synthesis and pharmaceutical chemistry, in particular to ursolic acid quinoline hydrazide heterocyclic derivatives with anti-tumor activity, and a preparation method and application thereof.

Background

Malignant tumors are diseases that seriously threaten human health and social development. According to the latest statistics of the world health organization, 880 million people die from cancer every year around the world, accounting for about 1/6 of the total number of death every year around the world, and the number of Chinese malignant tumors accounts for about 21.8% of the world. The three major approaches of tumor treatment are surgery, radiotherapy (abbreviated as radiotherapy) and chemotherapy (abbreviated as chemotherapy), respectively. Among them, chemotherapy is a common treatment method in which cytotoxic agents such as DNA synthesis inhibitors or cell division inhibitors are used to inhibit tumor cells, but at the same time, these agents also kill normal rapidly proliferating cells, causing symptoms such as infection and bleeding. Therefore, the development of tumor-inhibiting drugs with high selectivity, good safety and high curative effect is an important direction for the research of modern tumor diseases.

Ursolic Acid (UA) is also called ursolic acid and ursolic acid, and is an ursane type pentacyclic triterpene compound. The ursolic acid is widely distributed in plants, is distributed in various plants such as tea trees, fruit trees, medicinal plants, vanilla plants, paulownia and the like, and is statistically separated from 108 plants of 34 families in the nature, so the ursolic acid is a plant active ingredient with rich resources and development potential. Ursolic acid has anticancer, antiinflammatory, antioxidant, antidiabetic, antiviral, hepatoprotective, and immunity enhancing effects. Among them, its anticancer effect is receiving more and more attention. The anticancer action mechanism of ursolic acid includes several aspects, such as cytotoxic action, induced cell necrosis and apoptosis, epidermal growth factor receptor kinase inhibitory action, DNA polymerase and topoisomerase inhibitory action, and anti-tumor angiogenesis action. But the bioavailability of the ursolic acid is low, so that the clinical application of the ursolic acid is limited to a certain extent. Therefore, it is required to improve the anticancer activity and bioavailability of ursolic acid by chemical structure modification. At present, the research on the structure derivatization of ursolic acid at home and abroad mainly focuses on three sites: (1) a carboxyl group at the C-28 position; (2) a ring A is hydroxyl at the C-3 position; (3) a C ring double bond. However, very few reports on ursolic acid heterocyclic derivatives are reported at home and abroad at present.

The quinoline compound refers to quinoline and homologues thereof, is an important nitrogen-containing heterocyclic compound, has good biological activities of antibiosis, tumor resistance and the like, and is one of hot spots for research and development of novel medicaments and pesticides. In the aspect of medical care, quinoline medicines are widely used for treating and preventing diseases such as malaria, ulcer, cancer, HIV virus, schizophrenia and the like. Quinoline is a ring nucleus with strong pharmacological activity, and after quinoline rings are introduced into molecules of some compounds, the quinoline nucleus can show the activities of antibiosis, antiphlogosis, analgesia, tumor resistance, virus resistance and the like. The hydrazide compound is a compound containing nitrogen heteroatom, and many compounds containing hydrazide groups have biological activities of antivirus, anti-inflammation, anticancer and the like, and are a very important pharmacophore in drug development. Therefore, quinoline rings with different substituents are introduced to the ursolic acid molecule A ring, the carboxyl group is structurally modified, hydrazide groups are introduced, and a biological activity test is performed, so that the novel ursolic acid quinoline hydrazide derivative with better biological activity has important chemical and biological significance.

Disclosure of Invention

In order to overcome the defects in the prior art, the invention aims to provide ursolic acid quinoline hydrazide heterocyclic derivatives with high anti-tumor activity and anti-tumor activity, and a preparation method and application thereof.

In order to achieve the technical purpose, the technical scheme adopted by the invention is as follows: the invention relates to ursolic acid quinoline hydrazide derivatives with anti-tumor activity and pharmaceutically acceptable salts thereof:

wherein, I-a: r₁＝H R₂＝CH₃；I-b：R₁＝OMe R₂＝CH₃；I-c：R₁＝F R₂＝CH₃；I-d：R₁＝ClR₂＝CH₃；

I-e：R₁＝H R₂＝n-C₄H₉；I-f：R₁＝OMe R₂＝n-C₄H₉；I-g：R₁＝F R₂＝n-C₄H₉；I-h：R₁＝Cl R₂＝n-C₄H₉

The preparation method of the ursolic acid quinoline hydrazide derivatives with anti-tumor activity comprises the following steps:

(1) the ursolic acid is subjected to Jones reagent oxidation reaction to obtain 3-oxidized ursolic acid, which has a structure shown in a general formula II:

(2) reducing o-nitrobenzaldehyde with different substituents under the action of Fe/HCl to obtain corresponding o-aminobenzaldehyde, carrying out condensation reaction on 3-oxidized ursolic acid and o-aminobenzaldehyde with different substituents under alkaline condition and nitrogen atmosphere to obtain corresponding ursolic acid quinoline containing different substituents, wherein the structure is shown in a general formula III:

wherein, III-a: r₁＝H；III-b：R₁＝OMe；III-c：R₁＝F；III-d：R₁＝Cl；

(3) Obtaining corresponding ursolic acid quinoline acyl chloride compounds under the action of thionyl chloride, wherein the ursolic acid quinoline has a structure shown in a general formula IV:

wherein, IV-a: r₁＝H；IV-b：R₁＝OM_e；IV-c：R₁＝F；IV-d：R₁＝Cl；

(4) The acyl chloride compound of ursolic acid quinoline with different substituents reacts with acethydrazide and hydrazine valerate under alkaline conditions to obtain corresponding hydrazide derivatives of ursolic acid quinoline, which have the structure shown in the general formula I:

Further, in the step (1), synthesis of 3-oxoursolic acid II

Adding ursolic acid and acetone into a 500mL round-bottom flask, stirring and dissolving, stirring and reacting in ice water for 15min, slowly dropwise adding a Jones reagent, heating to room temperature, stirring and reacting for 5h, adding isopropanol, stirring and reacting for 30min, filtering out a precipitate after the reaction is finished, collecting a filtrate, and recrystallizing a light yellow green viscous solid obtained by concentrating the filtrate under reduced pressure with methanol to obtain a white needle crystal to obtain 3-oxoursolic acid II; the molar volume ratio of the ursolic acid to the acetone is 0.0184 mol/L; the volume ratio of the acetone to the Jones reagent to the isopropanol is 250: 1.9: 90;

in the step (2), synthesis of ursolic acid quinoline heterocyclic derivative III-a

Stirring 2-nitrobenzaldehyde in ethanol, acetic acid and distilled water, slowly heating to 50 ℃, adding reduced iron powder and 600 mu L concentrated hydrochloric acid after complete dissolution, continuously heating to 85 ℃, carrying out reflux reaction for 1h, stopping the reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining filtrate, extracting for 3 times by using dichloromethane, collecting an organic phase, washing for 3 times, washing a saturated sodium bicarbonate solution and the concentrated hydrochloric acid once respectively, then removing water by using anhydrous sodium sulfate, and carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, namely a 2-aminobenzaldehyde crude product, which is directly used for synthesizing unsubstituted ursolic acid quinoline; the molar ratio of the 2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the ethanol to the acetic acid to the distilled water is 2:2: 1; the molar volume of the 2-nitrobenzaldehyde and the total volume of the ethanol, the acetic acid and the distilled water is 0.15 mol/L; the volume ratio of the concentrated hydrochloric acid to the total volume of the ethanol, the acetic acid and the distilled water is 0.6: 20;

dropwise adding 2.4mL of saturated potassium hydroxide ethanol solution into 3-oxidized ursolic acid II and 2-aminobenzaldehyde ethanol solution in a 50mL three-neck round-bottom flask under the protection of nitrogen, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction solution into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, carrying out separation and purification by using a silica gel column, wherein the solvent is a petroleum ether acetone system, the volume ratio of petroleum ether to acetone is 150:1, and preparing a purified compound ursolic acid quinoline heterocyclic derivative III-a;

the molar ratio of the 3-oxoursolic acid to the ethanol solution of 2-aminobenzaldehyde is 2: 3; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.35 g/mL;

in the step (3), the synthesis of ursolic acid quinoline acylchloride compound IV-a

Dissolving the compound III-a with 5mL of benzene, slowly dropwise adding thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4 hours, stopping the reaction, and evaporating the benzene and the thionyl chloride in the reaction liquid to obtain a light yellow solid compound IV-a for subsequent reaction; the molar ratio of the compound III-a to the thionyl chloride and benzene is 0.15:3: 56;

in the step (4), synthesis of ursolic acid quinoline hydrazide heterocyclic derivative I-a

Dissolving the obtained ursolic acid quinoline hydrazide heterocyclic derivative IV-a in 5mL of diethyl ether, cooling to 0 ℃, dissolving acethydrazide and triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring and reacting for 6h, pouring reaction liquid into an ice water mixture after the reaction is finished, extracting for 3 times by using dichloromethane after ice is melted, collecting an organic phase, washing for 3 times, washing saturated sodium bicarbonate solution and concentrated brine for one time respectively, removing water by using anhydrous sodium sulfate, performing reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying by using a silica gel column, selecting a petroleum ether acetone system as a solvent, wherein the volume ratio of petroleum ether to acetone is 30:1, and obtaining a purified compound ursolic acid quinoline hydrazide heterocyclic derivative I-a; the molar ratio of the IV-a, the acethydrazide, the triethylamine, the diethyl ether and the dichloromethane is 0.15:0.225:0.3:48: 76;

further, in the step (1), synthesis of 3-oxoursolic acid II

Adding ursolic acid and acetone into a 500mL round-bottom flask, stirring and dissolving, stirring and reacting in ice water for 15min, slowly dropwise adding 1.872mL Jones reagent, heating to room temperature, stirring and reacting for 5h, adding isopropanol, stirring and reacting for 30min, filtering out precipitates after the reaction is finished, collecting filtrate, and recrystallizing pale yellow green viscous solid obtained by concentrating the filtrate under reduced pressure with methanol to obtain white needle crystals to obtain 3-oxidized ursolic acid II; the molar volume ratio of the ursolic acid to the acetone is 0.0184 mol/L; the volume ratio of acetone to jones reagent to isopropanol is 250: 1.9: 90, respectively;

the mass ratio of the 3-oxoursolic acid to the ethanol solution of 2-aminobenzaldehyde is 2: 3; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.35 g/mL;

in the step (4), synthesis of ursolic acid quinoline hydrazide heterocyclic derivatives I-e

Dissolving the obtained compound IV-a in 5mL of diethyl ether, cooling to 0 ℃, dissolving hydrazine valerate and triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring reaction liquid into an ice water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine for one time respectively, removing water with anhydrous sodium sulfate, decompressing, concentrating and removing an organic solvent to obtain a yellow solid, separating and purifying the yellow solid with a silica gel column, wherein the solvent adopts a petroleum ether acetone system, the volume ratio of petroleum ether to acetone is 80:1, and obtaining a purified compound, namely the ursolic acid quinoline hydrazide heterocyclic derivative I-e; the molar ratio of the IV-a, the hydrazine valerate, the triethylamine, the diethyl ether and the dichloromethane is 0.15:0.225:0.3:48: 76.

Further, in the step (1), synthesis of 3-oxoursolic acid II

in the step (2), synthesis of ursolic acid quinoline heterocyclic derivative III-b

Adding 5-hydroxy-2-nitrobenzaldehyde, methyl iodide, potassium carbonate and N, N-dimethylformamide into a 50mL round-bottom flask, stirring at room temperature for reacting for 8h, pouring a reaction solution into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid which is 100% of 5-methoxy-2-nitrobenzaldehyde; the molar ratio of the 5-hydroxy-2-nitrobenzaldehyde to the methyl iodide to the potassium carbonate is 4:6: 6; the volume ratio of the methyl iodide to the N, N-dimethylformamide is 0.4: 10;

stirring 5-methoxy-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water, slowly heating to 50 ℃, adding reduced iron powder and 600 mu L concentrated hydrochloric acid after complete dissolution, continuously heating to 85 ℃, carrying out reflux reaction for 1h, after the reaction is stopped, carrying out suction filtration on the mixed solution, washing filter residues with water, combining the filtrate, extracting with dichloromethane for 3 times, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated hydrochloric acid once respectively, then removing water with anhydrous sodium sulfate, and carrying out reduced pressure concentration to remove the organic solvent to obtain a yellow solid which is a 5-methoxy-2-aminobenzaldehyde crude product which is directly used for synthesizing methoxyursolic acid quinoline; the mass-volume ratio of the 5-methoxy-2-nitrobenzaldehyde to the total volume of the ethanol, the acetic acid and the distilled water is 0.02 g/mL; the volume ratio of the ethanol to the acetic acid to the distilled water is 2:2: 1; the molar ratio of the 5-methoxy-2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the total volume of the ethanol, the acetic acid and the distilled water to the volume of the concentrated hydrochloric acid is 175: 6;

dropwise adding 0.445g (1mmol) of 3-oxidized ursolic acid II and 0.225g (1.5mmol) of 5-methoxy-2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 100:1, so as to obtain a purified compound ursolic acid quinoline heterocyclic derivative III-b; the molar ratio of the 3-oxoursolic acid to the ethanol solution of the 5-methoxy-2-aminobenzaldehyde is 2: 3; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.22 g/mL;

in the step (3), the synthesis of ursolic acid quinoline acylchloride compound IV-b

Dissolving 0.085g (0.15mmol) of the compound III-b with 5mL of benzene, then slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain a light yellow solid compound IV-b for subsequent reaction; the molar ratio of the compound III-b to the thionyl chloride and the benzene is 0.15:3: 56;

in the step (4), synthesis of ursolic acid quinoline hydrazide heterocyclic derivative I-b

Dissolving the obtained compound IV-b in 5mL of diethyl ether, cooling to 0 ℃, dissolving 17.1mg (0.225mmol) of acethydrazide and 45 mu L (0.3mmol) of triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein a petroleum ether-acetone system is adopted as the solvent, and the volume ratio of petroleum ether to acetone is 30:1, so as to obtain a purified compound I-b of quinoline hydrazide heterocyclic derivatives of ursolic acid; the molar ratio of IV-b, acethydrazide, triethylamine, diethyl ether and dichloromethane is 0.15:0.225:0.3:48: 76;

further, in the step (1), synthesis of 3-oxoursolic acid II

Adding 0.668g (4mmol) of 5-hydroxy-2-nitrobenzaldehyde, 400 mu L (6mmol) of methyl iodide, 0.832g (6mmol) of potassium carbonate and 10mL of N, N-dimethylformamide into a 50mL round bottom flask, stirring at room temperature for reacting for 8h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once, removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid which is 5-methoxy-2-nitrobenzaldehyde (0.52g, 100%); the molar ratio of the 5-hydroxy-2-nitrobenzaldehyde to the methyl iodide to the potassium carbonate is 4:6: 6; the volume ratio of the methyl iodide to the N, N-dimethylformamide is 0.4: 10;

stirring 0.36g (2mmol) of 5-methoxy-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2:2:1, 17.5mL), slowly heating to 50 ℃, completely dissolving, adding 1.12g (20mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining filtrates, extracting 3 times with dichloromethane, collecting an organic phase, washing 3 times with water, washing saturated sodium bicarbonate solution and concentrated hydrochloric acid once respectively, removing water with anhydrous sodium sulfate, concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid which is a 5-methoxy-2-aminobenzaldehyde crude product (0.24g), and directly using the yellow solid for synthesizing methoxyursolic acid quinoline; the mass-volume ratio of the 5-methoxy-2-nitrobenzaldehyde to the total volume of the ethanol, the acetic acid and the distilled water is 0.02 g/mL; the volume ratio of the ethanol to the acetic acid to the distilled water is 2:2: 1; the molar ratio of the 5-methoxy-2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the total volume of the ethanol, the acetic acid and the distilled water to the volume of the concentrated hydrochloric acid is 175: 6;

in the step (4), synthesis of ursolic acid quinoline hydrazide heterocyclic derivatives I-f

Dissolving the obtained compound IV-b in 5mL of diethyl ether, cooling to 0 ℃, dissolving 25.0mg (0.225mmol) of hydrazine valerate and 45 mu L (0.3mmol) of triethylamine in 2mL of dichloromethane, dropwise adding the solution into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring the reaction solution into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein the solvent is a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 50:1, so as to obtain a purified compound I-f of quinoline heterocyclic derivatives of ursolic acid; the molar ratio of the IV-b, the hydrazine valerate to the triethylamine, the diethyl ether and the dichloromethane is 0.15:0.225:0.3:48: 76.

In the step (1), synthesis of 3-Oxaprotinic acid II

in the step (2), synthesis of ursolic acid quinoline heterocyclic derivative III-c

Stirring 0.51g (3mmol) of 5-fluoro-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2:2:1, 20mL), slowly heating to 50 ℃, completely dissolving, adding 1.68g (30mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining the filtrate, extracting 3 times with dichloromethane, collecting an organic phase, washing 3 times with water, washing the saturated sodium bicarbonate solution and the concentrated hydrochloric acid once respectively, removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove the organic solvent to obtain a yellow solid which is a 5-fluoro-2-aminobenzaldehyde crude product (0.43g) which is directly used for synthesizing ursolic acid quinoline fluoride; the molar ratio of the 5-fluoro-2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the ethanol to the acetic acid to the distilled water is 2:2: 1; the molar volume of the 5-fluoro-2-nitrobenzaldehyde with the total volume of ethanol, acetic acid and distilled water is 0.15 mol/L; the volume ratio of the concentrated hydrochloric acid to the total volume of the ethanol, the acetic acid and the distilled water is 0.6: 20;

dropwise adding 0.67g (1.5mmol) of 3-oxoursolic acid II and 0.35g (2.5mmol) of 5-fluoro-2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen into 2mL of saturated potassium hydroxide ethanol solution, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein a petroleum ether-acetone system is adopted as a solvent, and the volume ratio of petroleum ether to acetone is 150:1, so as to obtain a purified compound ursolic acid quinoline heterocyclic derivative III-c; the molar ratio of the 3-oxoursolic acid to the 5-fluoro-2-aminobenzaldehyde in the ethanol solution is 3: 5; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.34 g/mL;

in the step (3), the synthesis of ursolic acid quinoline acylchloride compound IV-c

Dissolving 0.084g (0.15mmol) of the compound III-c with 5mL of benzene, then slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain a light yellow solid compound IV-c for subsequent reaction; the molar ratio of the compound III-c to the thionyl chloride and benzene is 0.15:3: 56;

in the step (4), synthesis of ursolic acid quinoline hydrazide heterocyclic derivative I-c

Dissolving the obtained compound IV-c in 5mL of diethyl ether, cooling to 0 ℃, dissolving 17.1mg (0.225mmol) of acethydrazide and 45 mu L (0.3mmol) of triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein the solvent is a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 30:1, thus obtaining a purified compound I-c of quinoline heterocyclic derivatives of ursolic acid; the molar ratio of IV-c, acethydrazide, triethylamine, diethyl ether and dichloromethane is 0.15:0.225:0.3:48: 76;

further, in the step (1), synthesis of 3-oxoursolic acid II

dropwise adding 0.67g (1.5mmol) of 3-oxoursolic acid II and 0.35g (2.5mmol) of 5-fluoro-2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen into 2mL of saturated potassium hydroxide ethanol solution, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, and carrying out separation and purification by using a silica gel column, wherein the solvent adopts a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 150:1, thus obtaining a purified compound III-c; the molar ratio of the 3-oxoursolic acid to the 5-fluoro-2-aminobenzaldehyde in the ethanol solution is 3: 5; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.34 g/mL;

in the step (4), synthesis of ursolic acid quinoline hydrazide heterocyclic derivative I-g

Dissolving the obtained compound IV-c in 5mL of diethyl ether, cooling to 0 ℃, dissolving 25.0mg (0.225mmol) of hydrazine valerate and 45 mu L (0.3mmol) of triethylamine in 2mL of dichloromethane, dropwise adding the solution into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring the reaction solution into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein the solvent is a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 80:1, so as to obtain a purified compound I-g of quinoline heterocyclic derivatives of ursolic acid; the molar ratio of the IV-c, the hydrazine valerate, the triethylamine, the diethyl ether and the dichloromethane is 0.15:0.225:0.3:48: 76.

In the step (1), synthesis of 3-Oxaprotinic acid II

in the step (2), synthesis of ursolic acid quinoline heterocyclic derivatives III-d

Stirring 0.55g (3mmol) of 5-chloro-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2:2:1, 20mL), slowly heating to 50 ℃, completely dissolving, adding 1.68g (30mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining filtrate, extracting 3 times with dichloromethane, collecting organic phase, washing 3 times with water, washing saturated sodium bicarbonate solution and concentrated hydrochloric acid once respectively, removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove organic solvent to obtain a yellow solid which is 5-chloro-2-aminobenzaldehyde crude product (0.45g) which is directly used for synthesizing ursolic acid quinoline chloride; the molar ratio of the 5-chloro-2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the ethanol to the acetic acid to the distilled water is 2:2: 1; the molar volume of the 5-chloro-2-nitrobenzaldehyde with the total volume of ethanol, acetic acid and distilled water is 0.15 mol/L; the volume ratio of the concentrated hydrochloric acid to the total volume of the ethanol, the acetic acid and the distilled water is 0.6: 20;

dropwise adding 0.67g (1.5mmol) of 3-oxoursolic acid II and 0.38g (2.5mmol) of 5-chloro-2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, and carrying out separation and purification by using a silica gel column, wherein the solvent adopts a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 150:1, thus obtaining purified compounds III-d; the molar ratio of the 3-oxoursolic acid to the ethanol solution of the 5-chloro-2-aminobenzaldehyde is 3: 5; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.34 g/mL;

in the step (3), synthesis of ursolic acid quinoline acylchloride compound IV-d

Dissolving 0.086g (0.15mmol) of the compound III-d with 5mL of benzene, then slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain a light yellow solid compound IV-d for subsequent reaction; the molar ratio of the compound III-d to the thionyl chloride and benzene is 0.15:3: 56;

in the step (4), synthesis of ursolic acid quinoline hydrazide heterocyclic derivatives I-d

Dissolving the obtained compound IV-d in 5mL of diethyl ether, cooling to 0 ℃, dissolving 17.1mg (0.225mmol) of acethydrazide and 45 mu L (0.3mmol) of triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein a petroleum ether-acetone system is adopted as the solvent, and the volume ratio of petroleum ether to acetone is 30:1, so as to obtain a purified compound I-d of quinoline hydrazide heterocyclic derivatives of ursolic acid; the molar ratio of IV-d, acethydrazide, triethylamine, diethyl ether and dichloromethane is 0.15:0.225:0.3:48: 76;

further, in the step (1), synthesis of 3-oxoursolic acid II

dropwise adding 0.67g (1.5mmol) of 3-oxoursolic acid II and 0.38g (2.5mmol) of 5-chloro-2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, and carrying out separation and purification by using a silica gel column, wherein the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 150:1, thus obtaining a purified compound ursolic acid quinoline heterocyclic derivative III-d; the molar ratio of the 3-oxoursolic acid to the ethanol solution of the 5-chloro-2-aminobenzaldehyde is 3: 5; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.34 g/mL;

in the step (3), synthesis of ursolic acid quinoline acylchloride compound IV-d

Dissolving 0.086g (0.15mmol) of the compound III-d with 5mL of benzene, then slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain a light yellow solid compound IV-d for subsequent reaction; the molar ratio of the compound III-d to thionyl chloride and benzene is 0.15:3: 56;

in the step (4), synthesis of ursolic acid quinoline hydrazide heterocyclic derivatives I-h

Dissolving the obtained compound IV-d in 5mL of diethyl ether, cooling to 0 ℃, dissolving 25.0mg (0.225mmol) of hydrazine valerate and 45 mu L (0.3mmol) of triethylamine in 2mL of dichloromethane, dropwise adding the solution into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring the reaction solution into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid with a silica gel column, wherein the solvent is a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 80:1, so as to obtain a purified compound I-h of quinoline hydrazide heterocyclic derivatives of ursolic acid; the molar ratio of the IV-d, the hydrazine valerate, the triethylamine, the diethyl ether and the dichloromethane is 0.15:0.225:0.3:48: 76.

The invention relates to an application of ursolic acid quinoline hydrazide heterocyclic derivatives with a structure shown in formula I and pharmaceutically acceptable salts thereof in preparing medicaments for treating tumors.

Furthermore, the tumor is any one of human breast cancer cell MDA-MB-231, human cervical cancer cell HeLa or human liver cancer cell SMMC-7721.

Has the advantages that: the ursolic acid quinoline derivative has anti-tumor activity, and pharmacological experiments show that the ursolic acid quinoline derivative has obvious inhibition effect on human breast cancer cells MDA-MB-231, human cervical cancer cells HeLa and human liver cancer cells SMMC-7721, and shows low toxicity on human normal liver epithelial cells QSG-7701.

The invention is characterized in that after C-3 hydroxyl on an ursolic acid A ring is oxidized into carbonyl and condensed with anthranilate aldehyde with different substituents, quinoline heterocycles with different substituents are introduced to obtain corresponding ursolic acid quinoline, and C-28 carboxyl of the ursolic acid parent of the ursolic acid quinoline with different substituents reacts with acethydrazide or hydrazine valerate to obtain the corresponding ursolic acid quinoline heterocycle derivative. The derivatives have novel structures and are not reported at home and abroad. Has the value of developing antitumor drugs.

Detailed Description

The invention is further illustrated by the following examples. It should be understood that these examples are illustrative and exemplary of the present invention, and are not intended to limit the scope of the present invention in any way.

The invention provides an ursolic acid quinoline hydrazide heterocyclic derivative with a structure shown in a general formula I and pharmaceutically acceptable salts thereof:

The invention relates to ursolic acid quinoline hydrazide derivatives with anti-tumor activity, a preparation method and application thereof, which comprise the following steps:

(1) ursolic acid is subjected to Jones reagent oxidation reaction to 3-oxidized ursolic acid, and has a structure shown in a general formula II:

wherein, III-a: r₁＝H；III-b：R₁＝OMe；III-c：R₁＝F；III-d：R₁＝Cl

wherein, IV-a: r₁＝H；IV-b：R₁＝OM_e；IV-c：R₁＝F；IV-d：R₁＝Cl

The tumor is any one of human breast cancer cell MDA-MB-231, human cervical cancer cell HeLa or human liver cancer cell SMMC-7721, and the normal cell is human normal liver epithelial cell QSG-7701.

Example 1

Synthesis of 3-Oxoastolic acid II

Adding 2g (4.6mmol) of ursolic acid and 250mL of acetone into a 500mL round-bottom flask, stirring for dissolving, stirring for reaction in ice water for 15min, slowly dropwise adding 1.872mL of Jones reagent, heating to room temperature, stirring for reaction for 5h, adding 90mL of isopropanol, stirring for reaction for 30min, filtering out a precipitate after the reaction is finished, collecting a filtrate, and recrystallizing a light yellow green viscous solid obtained by concentrating the filtrate under reduced pressure by using methanol to obtain a white needle-like crystal, namely 3-oxoursolic acid II (1.2g, 65.6%).

Example 2

Synthesis of ursolic acid quinoline heterocyclic derivative (III-a)

Stirring 0.45g (3mmol) of 2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2:2:1, 20mL), slowly heating to 50 ℃, completely dissolving, adding 1.68g (30mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining the filtrate, extracting 3 times with dichloromethane, collecting an organic phase, washing 3 times with water, washing the saturated sodium bicarbonate solution and the concentrated hydrochloric acid once respectively, then removing water with anhydrous sodium sulfate, and carrying out reduced pressure concentration to remove the organic solvent to obtain a yellow solid, namely a 2-aminobenzaldehyde crude product (0.41g), which is directly used for synthesizing the unsubstituted ursolic acid quinoline.

Adding 0.78g (1.8mmol) of 3-oxoursolic acid II and 0.41g (2.7mmol) of 2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen dropwise, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, and separating and purifying the yellow solid by using a silica gel column, wherein the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 150:1 to obtain a purified compound III-a (0.71g, 68%).

M.p.243-245℃；¹H NMR(500MHz,CDCl₃):δ0.90(s,3H),0.96(s,3H),0.95(d,J＝5.80Hz,3H),1.15(s,3H),1.27(d,J＝7.75Hz,3H),1.28-1.38(m,3H),1.40(s,3H),1.43(s,3H),1.49-1.76(m,12H),1.92-2.28(m,5H),2.58(d,J＝15.30Hz,1H),2.96(d,J＝15.70Hz,1H),5.35(s,1H),7.40(t,J＝7.30Hz,1H),7.58(t,J＝7.15Hz,1H),7.67(d,J＝9.60Hz,1H),7.68(s,1H),7.99(d,J＝8.35Hz,1H)；IR(KBr,cm^-1):2924,2854,1695,1607,1454,1378,1087,1041,1015,973；ESI-MS:m/z[M+H]⁺:540.39；Anal.Calcd.for C₃₇H₄₉NO₂:C82.33,H 9.15,N 2.59；found:C 82.29,H 9.17,N 2.61.

Example 3

Synthesis of ursolic acid quinoline heterocyclic derivative (III-b)

A50 mL round bottom flask was charged with 0.668g (4mmol) of 5-hydroxy-2-nitrobenzaldehyde, 400. mu.L (6mmol) of methyl iodide, 0.832g (6mmol) of potassium carbonate and 10mL of N, N-dimethylformamide, stirred at room temperature for 8h, after the reaction was completed, the reaction mixture was poured into an ice-water mixture, after the ice was melted, the organic phase was collected by extraction with dichloromethane 3 times, washed with water 3 times, once with a saturated sodium bicarbonate solution and once with concentrated brine, then removed with anhydrous sodium sulfate, and concentrated under reduced pressure to remove the organic solvent to give 5-methoxy-2-nitrobenzaldehyde as a yellow solid (0.52g, 100%).

Stirring 0.36g (2mmol) of 5-methoxy-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2:2:1, 17.5mL), slowly heating to 50 ℃, completely dissolving, adding 1.12g (20mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining filtrates, extracting with dichloromethane for 3 times, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated hydrochloric acid once respectively, then removing water with anhydrous sodium sulfate, concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid, namely a 5-methoxy-2-aminobenzaldehyde crude product (0.24g), which is directly used for synthesizing methoxyursolic acid quinoline.

Adding 0.445g (1mmol) of 3-oxidized ursolic acid II and 0.225g (1.5mmol) of 5-methoxy-2-aminobenzaldehyde in a 50mL three-neck round-bottom flask in ethanol solution dropwise under the protection of nitrogen, refluxing at 85 ℃ for 24h, pouring the reaction solution into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid, separating and purifying by using a silica gel column, wherein the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 100:1, and the pure compound III-b (0.43g, 70%) is prepared.

M.p.255-258℃；¹H NMR(300MHz,CDCl₃):δ0.88(s,3H),0.92(s,3H),0.95(d,J＝6.75Hz,3H),1.15(s,3H),1.27(d,J＝6.55Hz,3H),1.28-1.38(m,3H),1.40(s,3H),1.43(s,3H),1.49-1.76(m,12H),1.92-2.28(m,5H),2.56(d,J＝15.03Hz,1H),2.93(d,J＝15.18Hz,1H),3.90(s,3H,OCH₃),5.36(s,1H),6.96(s,1H),7.25(d,J＝7.65Hz,1H),7.60(s,1H),7.91(d,J＝8.80Hz,1H)；IR(KBr,cm^-1):2926,2857,1728,1693,1457,1385,1314,1277,1078,1030,972；ESI-MS:m/z[M+H]⁺:570.40；Anal.Calcd.for C₃₈H₅₁NO₃:C 80.10,H 9.02,N2.46；found:C 80.13,H 9.04,N 2.59.

Example 4

Synthesis of ursolic acid quinoline heterocyclic derivative (III-c)

Stirring 0.51g (3mmol) of 5-fluoro-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2:2:1, 20mL), slowly heating to 50 ℃, completely dissolving, adding 1.68g (30mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining the filtrate, extracting 3 times with dichloromethane, collecting an organic phase, washing 3 times with water, washing the saturated sodium bicarbonate solution and the concentrated hydrochloric acid once respectively, then removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove the organic solvent to obtain a yellow solid which is a 5-fluoro-2-aminobenzaldehyde crude product (0.43g) which is directly used for synthesizing ursolic acid quinoline fluoride.

Adding 0.67g (1.5mmol) of 3-oxoursolic acid II and 0.35g (2.5mmol) of 5-fluoro-2-aminobenzaldehyde in a 50mL three-neck round-bottom flask under the protection of nitrogen into 2mL of saturated potassium hydroxide ethanol solution dropwise, refluxing at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane after ice is melted, collecting an organic phase for 3 times, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid, separating and purifying with a silica gel column, wherein the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 150:1 to obtain a purified compound III-c (0.51g, 62%).

M.p.235-237℃；¹H NMR(300MHz,CDCl₃):δ0.88(s,3H),0.91(s,3H),0.93(d,J＝6.20Hz,3H),1.14(s,3H),1.26(d,J＝6.40Hz,3H),1.28-1.38(m,3H),1.36(s,3H),1.40(s,3H),1.49-1.76(m,11H),1.92-2.28(m,6H),2.57(d,J＝15.51Hz,1H),2.94(d,J＝15.66Hz,1H),5.33(s,1H),7.27(d,J＝8.50Hz,1H),7.37(t,J＝8.50Hz,1H),7.62(s,1H),7.97(dd,J＝8.91,5.61Hz,1H)；IR(KBr,cm^-1):2959,2925,2855,1695,1492,1457,1378,1213,1147,1077,968,829；ESI-MS:m/z[M+H]⁺:558.38；Anal.Calcd.for C₃₇H₄₈FNO₂:C 79.67,H 8.67,N2.51；found:C 79.68,H 8.65,N 2.53.

Example 5

Synthesis of ursolic acid quinoline heterocyclic derivative (III-d)

Stirring 0.55g (3mmol) of 5-chloro-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2:2:1, 20mL), slowly heating to 50 ℃, completely dissolving, adding 1.68g (30mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining the filtrate, extracting 3 times with dichloromethane, collecting an organic phase, washing 3 times with water, washing the saturated sodium bicarbonate solution and the concentrated hydrochloric acid once respectively, removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove the organic solvent to obtain a yellow solid, namely a 5-chloro-2-aminobenzaldehyde crude product (0.45g), which is directly used for synthesizing the chloroquinoline ursolate.

Adding 0.67g (1.5mmol) of 3-oxoursolic acid II and 0.38g (2.5mmol) of 5-chloro-2-aminobenzaldehyde in an ethanol solution of 0.67g (1.5mmol) of 3-oxoursolic acid II in a 50mL three-neck round-bottom flask dropwise under the protection of nitrogen, refluxing at 85 ℃ for 24h, pouring the reaction solution into an ice-water mixture after the reaction is finished, extracting with dichloromethane after ice is melted, collecting an organic phase for 3 times, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid, and separating and purifying the yellow solid by using a silica gel column, wherein the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 150:1 to obtain a purified compound III-d (0.58g, 64%).

M.p.247-249℃；¹H NMR(300MHz,CDCl₃):δ0.89(s,3H),0.91(s,3H),0.96(d,J＝6.40Hz,3H),0.98(s,3H),1.26(d,J＝6.40Hz,3H),1.28-1.38(m,3H),1.36(s,3H),1.40(s,3H),1.49-1.76(m,12H),1.92-2.28(m,5H),2.57(d,J＝14.94Hz,1H),2.95(d,J＝15.75Hz,1H),5.35(s,1H),7.51(d,J＝8.88Hz,1H),7.59(s,1H),7.65(s,1H),7.92(d,J＝8.94Hz,1H)；IR(KBr,cm^-1):2924,2855,1695,1480,1404,1379,1262,1183,1070,967,920,828；ESI-MS:m/z[M+H]⁺:574.35；Anal.Calcd.for C₃₇H₄₈ClNO₂:C 77.39,H 8.43,N 2.44；found:C77.40,H 8.45,N 2.47.

Example 6

Synthesis of ursolic acid quinoline acylchloride compound (IV-a)

Dissolving 0.081g (0.15mmol) of the compound III-a with 5mL of benzene, slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain the compound IV-a which is light yellow solid for subsequent reaction

Example 7

Synthesis of ursolic acid quinoline acylchloride compound (IV-b)

Dissolving 0.085g (0.15mmol) of the compound III-b in 5mL of benzene, slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain the compound IV-b as a light yellow solid for subsequent reaction.

Example 8

Synthesis of ursolic acid quinoline acylchloride compound (IV-c)

Dissolving 0.084g (0.15mmol) of the compound III-c in 5mL of benzene, slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain the compound IV-c as a light yellow solid for subsequent reaction.

Example 9

Synthesis of ursolic acid quinoline acylchloride compound (IV-d)

Dissolving 0.086g (0.15mmol) of the compound III-d in 5mL of benzene, slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain the compound IV-d as a light yellow solid for subsequent reaction.

Example 10

Synthesis of ursolic acid quinoline hydrazide heterocyclic derivative (I-a)

The compound IV-a obtained in example 6 is dissolved in 5mL of diethyl ether, cooled to 0 ℃, 17.1mg (0.225mmol) of acethydrazide and 45 μ L (0.3mmol) of triethylamine are dissolved in 2mL of dichloromethane and added dropwise into a reaction vessel, the temperature is slowly raised to room temperature and stirred for reaction for 6 hours, after the reaction is finished, the reaction liquid is poured into an ice-water mixture, after ice is melted, dichloromethane is used for extraction for 3 times to collect an organic phase, water washing is carried out for 3 times, saturated sodium bicarbonate solution and concentrated brine are respectively washed once and then water is removed by anhydrous sodium sulfate, organic solvent is removed by decompression and concentration to obtain yellow solid, the yellow solid is separated and purified by a silica gel column, the solvent adopts a petroleum ether acetone system, the volume ratio of petroleum ether to acetone is 30:1, and the purified compound I-a (0.076g, 84.1%) is obtained.

M.p.267-269℃；¹H NMR(500MHz,CDCl₃):δ0.83(s,3H),0.87(s,3H),0.93(d,J＝6.40Hz,3H),0.98(d,J＝5.05Hz,3H),1.18(s,3H),1.42(s,3H),1.44(s,3H),1.50-1.94(m,15H),2.03(s,3H),2.06-2.37(m,4H),2.59(d,J＝15.45Hz,1H),2.96(d,J＝15.35Hz,1H),5.60(s,1H),7.41(t,J＝7.55Hz,3H),7.58(t,J＝7.65Hz,1H),7.69(d,J＝8.50Hz,1H),7.70(s,1H),8.00(d,J＝8.50Hz,1H),8.77(d,J＝7.10Hz,1H,NH),8.95(d,J＝7.15Hz,1H,NH)；IR(KBr,cm^-1):3251,2953,2924,2854,1618,1492,1458,1378,1081,968,759；ESI-MS:m/z[M+H]⁺:596.43；Anal.Calcd.for C₃₉H₅₃N₃O₂:C 78.61,H 8.97,N 7.05,found:C 78.60,H 8.98,N 7.08.

Example 11

Synthesis of ursolic acid quinoline hydrazide heterocyclic derivative (I-b)

The compound IV-b obtained in example 7 is dissolved in 5mL of diethyl ether, cooled to 0 ℃, 17.1mg (0.225mmol) of acethydrazide and 45 μ L (0.3mmol) of triethylamine are dissolved in 2mL of dichloromethane and added dropwise into a reaction vessel, the temperature is slowly raised to room temperature and stirred for reaction for 6 hours, after the reaction is finished, the reaction liquid is poured into an ice-water mixture, after ice is melted, dichloromethane is used for extraction for 3 times to collect an organic phase, water washing is carried out for 3 times, saturated sodium bicarbonate solution and concentrated brine are respectively washed once and then water is removed by anhydrous sodium sulfate, organic solvent is removed by decompression and concentration to obtain yellow solid, the yellow solid is separated and purified by a silica gel column, the solvent adopts a petroleum ether acetone system, the volume ratio of petroleum ether to acetone is 30:1, and the purified compound I-b (0.043g, 57.6%) is obtained.

M.p.290-293℃；¹H NMR(500MHz,CDCl₃):δ0.83(s,3H),0.87(s,3H),0.92(d,J＝7.25Hz,3H),0.97(d,J＝7.50Hz,3H),1.18(s,3H),1.39(s,3H),1.42(s,3H),1.47-1.93(m,15H),2.03(s,3H),2.05-2.18(m,4H),2.57(d,J＝15.10Hz,1H),2.92(d,J＝15.30Hz,1H),3.91(s,3H,OCH₃),5.59(s,1H),6.96(d,J＝2.55Hz,1H),7.25(d,J＝7.65Hz,1H),7.60(s,1H),7.91(d,J＝8.80Hz,1H),8.78(m,1H,NH),8.94(m,1H,NH)；IR(KBr,cm^-1):3249,2953,2925,2855,1621,1492,1457,1378,1222,1080,1030,828,668；ESI-MS:m/z[M+H]⁺:626.44；Anal.Calcd.for C₄₀H₅₅N₃O₃:C 76.76,H 8.86,N 6.71,found:C 76.73,H 8.88,N 6.75.

Example 12

Synthesis of ursolic acid quinoline hydrazide heterocyclic derivative (I-c)

The compound IV-c obtained in example 8 is dissolved in 5mL of diethyl ether, cooled to 0 ℃, 17.1mg (0.225mmol) of acethydrazide and 45 μ L (0.3mmol) of triethylamine are dissolved in 2mL of dichloromethane and added dropwise into a reaction vessel, the temperature is slowly raised to room temperature and stirred for reaction for 6 hours, after the reaction is finished, the reaction liquid is poured into an ice-water mixture, after ice is melted, dichloromethane is used for extraction for 3 times to collect an organic phase, water washing is carried out for 3 times, saturated sodium bicarbonate solution and concentrated brine are respectively washed once and then water is removed by anhydrous sodium sulfate, organic solvent is removed by decompression and concentration to obtain yellow solid, the yellow solid is separated and purified by a silica gel column, the solvent adopts a petroleum ether acetone system, the volume ratio of petroleum ether to acetone is 30:1, and purified compound I-c (0.071g, 88.4%) is obtained.

M.p.274-278℃；¹H NMR(500MHz,CDCl₃):δ0.83(s,3H),0.89(s,3H),0.93(d,J＝6.40Hz,3H),0.97(d,J＝6.25Hz,3H),1.18(s,3H),1.40(s,3H),1.43(s,3H),1.45-1.93(m,15H),2.04(s,3H),2.06-2.19(m,4H),2.57(d,J＝15.45Hz,1H),2.95(d,J＝15.50Hz,1H),5.69(s,1H),7.29(dd,J＝9.05,2.65Hz,1H),7.35(dt,J＝8.95,2.90Hz,1H),7.64(s,1H),7.98(dd,J＝9.15,5.40Hz,1H),8.89(d,J＝7.00Hz,1H,NH),8.96(d,J＝7.00Hz,1H,NH)；IR(KBr,cm^-1):3230,2952,2925,2855,1616,1492,1456,1378,1212,1148,1077,969,829,802；ESI-MS:m/z[M+H]⁺:614.42；Anal.Calcd.for C₃₉H₅₂FN₃O₂:C 76.31,H 8.54,N 6.85,found:C 76.30,H 8.55,N 6.87.

Example 13

Synthesis of ursolic acid quinoline hydrazide heterocyclic derivative (I-d)

The compound IV-d obtained in example 9 is dissolved in 5mL of diethyl ether, cooled to 0 ℃, 17.1mg (0.225mmol) of acethydrazide and 45 μ L (0.3mmol) of triethylamine are dissolved in 2mL of dichloromethane and added dropwise into a reaction vessel, the temperature is slowly raised to room temperature and stirred for reaction for 6 hours, after the reaction is finished, the reaction liquid is poured into an ice-water mixture, after ice is melted, dichloromethane is used for extraction for 3 times to collect an organic phase, water washing is carried out for 3 times, saturated sodium bicarbonate solution and concentrated brine are respectively washed once and then water is removed by anhydrous sodium sulfate, organic solvent is removed by decompression and concentration to obtain yellow solid, the yellow solid is separated and purified by a silica gel column, the solvent adopts a petroleum ether acetone system, the volume ratio of petroleum ether to acetone is 30:1, and purified compound I-d (0.078g, 74.3%) is obtained.

M.p.270-273℃；¹H NMR(300MHz,CDCl₃):δ0.83(s,3H),0.88(s,3H),0.93(d,J＝6.33Hz,3H),0.98(d,J＝6.78Hz,3H),1.18(s,3H),1.40(s,3H),1.42(s,3H),1.54-1.95(m,15H),2.03(s,3H),2.08-2.23(m,4H),2.58(d,J＝15.84Hz,1H),2.95(d,J＝15.69Hz,1H),5.60(s,1H),7.51(d,J＝8.91Hz,1H),7.61(s,1H),7.67(s,1H),7.79(d,J＝7.50Hz,2H),7.92(d,J＝9.00Hz,1H),8.65(bis,1H,NH),8.95(bis,1H,NH)；IR(KBr,cm^-1):3234,2949,2924,2855,1616,1479,1456,1378,1184,1070,968,919,828,753；ESI-MS:m/z[M+H]⁺:630.38；Anal.Calcd.for C₃₉H₅₂ClN₃O₂:C 74.32,H 8.32,N 6.67,found:C 74.37,H 8.30,N6.65.

Example 14

Synthesis of ursolic acid quinoline hydrazide heterocyclic derivative (I-e)

The compound IV-a obtained in example 6 is dissolved in 5mL of diethyl ether, cooled to 0 ℃, 25.0mg (0.225mmol) of hydrazine valerate and 45 μ L (0.3mmol) of triethylamine are dissolved in 2mL of dichloromethane and added dropwise into a reaction vessel, the mixture is slowly heated to room temperature and stirred for reaction for 6 hours, after the reaction is finished, the reaction liquid is poured into an ice-water mixture, after ice is melted, dichloromethane is used for extraction for 3 times to collect an organic phase, water washing is carried out for 3 times, saturated sodium bicarbonate solution and concentrated brine are respectively washed once and then water is removed by anhydrous sodium sulfate, organic solvent is removed by decompression and concentration to obtain yellow solid, the yellow solid is separated and purified by a silica gel column, the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 80:1, and purified compound I-e (0.056g, 70.2%) is obtained.

M.p.278-281℃；¹H NMR(500MHz,CDCl₃):δ0.81(s,3H),0.87(s,3H),0.94(d,J＝6.40Hz,3H),0.97(d,J＝7.40Hz,3H),1.18(s,3H),1.42(s,3H),1.44(s,3H),1.49-1.78(m,18H),1.92-2.20(m,8H),2.23(t,J＝7.45Hz,2H),2.59(d,J＝15.50Hz,1H),2.96(d,J＝15.30Hz,1H),5.62(s,1H),7.41(t,J＝7.45Hz,1H),7.58(t,J＝7.65Hz,1H),7.68(d,J＝8.55Hz,1H),7.70(s,1H),7.99(d,J＝8.45Hz,1H),8.71(d,J＝7.35Hz,1H,NH),9.06(d,J＝7.40Hz,1H,NH)；IR(KBr,cm^-1):3223,2954,2924,2855,1615,1492,1457,1378,1190,1075,969,753；ESI-MS:m/z[M+H]⁺:638.48；Anal.Calcd.for C₄₂H₅₉N₃O₂:C 79.08,H 9.32,N6.59,found:C 79.12,H 9.33,N 6.57.

Example 15

Synthesis of ursolic acid quinoline hydrazide heterocyclic derivative (I-f)

The compound IV-b obtained in example 7 is dissolved in 5mL of diethyl ether, cooled to 0 ℃, 25.0mg (0.225mmol) of hydrazine valerate and 45 μ L (0.3mmol) of triethylamine are dissolved in 2mL of dichloromethane and added dropwise into a reaction vessel, the mixture is slowly heated to room temperature and stirred for reaction for 6 hours, after the reaction is finished, the reaction liquid is poured into an ice-water mixture, after ice is melted, the organic phase is extracted by dichloromethane for 3 times, the organic phase is washed by water for 3 times, a saturated sodium bicarbonate solution and concentrated brine are respectively washed by once, then water is removed by anhydrous sodium sulfate, the organic solvent is removed by decompression and concentration to obtain a yellow solid, the yellow solid is separated and purified by a silica gel column, the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 50:1, and the purified compound I-f (0.051g, 46.4%) is obtained.

M.p.282-285℃；¹H NMR(500MHz,CDCl₃):δ0.81(s,3H),0.89(s,3H),0.94(d,J＝6.45Hz,3H),0.98(d,J＝6.40Hz,3H),1.18(s,3H),1.39(s,3H),1.42(s,3H),1.47-1.76(m,18H),1.91-2.10(m,4H),2.18-2.25(m,4H),2.56(d,J＝15.70Hz,1H),2.92(d,J＝16.20Hz,1H),3.90(s,3H,OCH₃),5.61(s,1H),6.96(s,1H),7.24(d,J＝8.95Hz,1H),7.59(s,1H),7.88(d,J＝9.20Hz,1H),8.71(d,J＝7.60Hz,1H,NH),9.07(d,J＝7.30Hz,1H,NH)；IR(KBr,cm^-1):3226,2954,2924,2854,1618,1492,1459,1379,1222,1080,1032,964,829,805；ESI-MS:m/z[M+H]⁺:668.48；Anal.Calcd.for C₄₃H₆₁N₃O:C 81.21,H 9.67,N 6.61,found:C81.24,H 9.65,N 6.62.

Example 16

Synthesis of ursolic acid quinoline hydrazide heterocyclic derivative (I-g)

The compound IV-c obtained in example 8 is dissolved in 5mL of diethyl ether, cooled to 0 ℃, 25.0mg (0.225mmol) of hydrazine valerate and 45 μ L (0.3mmol) of triethylamine are dissolved in 2mL of dichloromethane and added dropwise into a reaction vessel, the mixture is slowly heated to room temperature and stirred for reaction for 6 hours, after the reaction is finished, the reaction solution is poured into an ice-water mixture, after ice is melted, the organic phase is extracted 3 times by dichloromethane, the organic phase is collected after washing for 3 times, saturated sodium bicarbonate solution and concentrated brine are washed once respectively, then water is removed by anhydrous sodium sulfate, organic solvent is removed by decompression and concentration to obtain yellow solid, the yellow solid is separated and purified by a silica gel column, the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 80:1, and purified compound I-g (0.083g, 7.2%) is obtained.

M.p.289-291℃；¹H NMR(500MHz,CDCl₃):δ0.81(s,3H),0.88(s,3H),0.93(d,J＝6.40Hz,3H),0.97(d,J＝6.55Hz,3H),1.18(s,3H),1.40(s,3H),1.42(s,3H),1.45-1.78(m,18H),1.92-2.19(m,8H),2.23(t,J＝7.45Hz,2H),2.57(d,J＝15.50Hz,1H),2.95(d,J＝15.55Hz,1H),5.61(s,1H),7.29(dd,J＝9.00,2.35Hz,1H),7.35(dt,J＝8.55,2.35Hz,1H),7.64(s,1H),7.98(dd,J＝9.05,5.45Hz,1H),8.73(d,J＝7.25Hz,1H,NH),9.06(d,J＝7.35Hz,1H,NH)；IR(KBr,cm^-1):3238,2954,2925,2870,1614,1493,1456,1378,1286,1213,1148,1077,969,829,801；ESI-MS:m/z[M+H]⁺:656.47；Anal.Calcd.for C₄₂H₅₈FN₃O₂:C76.91,H 8.91,N 6.41,found:C 76.86,H 8.92,N 6.45.

Example 17

Synthesis of ursolic acid quinoline hydrazide heterocyclic derivative (I-h)

The compound IV-d obtained in example 9 is dissolved in 5mL of diethyl ether, cooled to 0 ℃, 25.0mg (0.225mmol) of hydrazine valerate and 45 μ L (0.3mmol) of triethylamine are dissolved in 2mL of dichloromethane and added dropwise into a reaction vessel, the temperature is slowly raised to room temperature and stirred for reaction for 6 hours, after the reaction is finished, the reaction liquid is poured into an ice-water mixture, after ice is melted, dichloromethane is used for extraction for 3 times to collect an organic phase, water washing is carried out for 3 times, saturated sodium bicarbonate solution and concentrated brine are respectively washed once and then water is removed by anhydrous sodium sulfate, organic solvent is removed by decompression and concentration to obtain yellow solid, the yellow solid is separated and purified by a silica gel column, the solvent adopts a petroleum ether acetone system, and the volume ratio of petroleum ether to acetone is 80:1, and purified compound I-h (0.084g, 70.4%) is obtained.

M.p.303-307℃；¹H NMR(500MHz,CDCl₃):δ0.82(s,3H),0.88(s,3H),0.91(d,J＝7.35Hz,3H),0.94(d,J＝6.25Hz,3H),1.22(s,3H),1.38(s,3H),1.41(s,3H),1.49-1.79(m,19H),1.93(m,1H),2.08(m,2H),2.19(m,2H),2.24(m,2H),2.58(d,J＝15.20Hz,1H),2.96(d,J＝15.30Hz,1H),5.62(s,1H),7.52(d,J＝9.00,1H),7.62(s,1H),7.67(s,1H),7.93(d,J＝7.90Hz,1H),8.76(d,J＝6.60Hz,1H,NH),9.07(d,J＝6.90Hz,1H,NH)；IR(KBr,cm^-1):3230,2953,2925,2869,1614,1479,1457,1379,1183,1069,965,918,828；ESI-MS:m/z[M+H]⁺:672.43；Anal.Calcd.for C₄₂H₅₈ClN₃O₂:C 75.02,H 8.69,N 6.25,found:C 75.00,H 8.73,N6.24.

Test 1

Screening for antitumor Activity in vitro

The cell lines are selected as follows: human breast cancer cell MDA-MB-231, human cervical cancer cell HeLa, human liver cancer cell SMMC-7721 and human normal liver epithelial cell QSG-7701.

The experimental method comprises the following steps:

taking the cells with good logarithmic growth phase, digesting with trypsin to obtain 5 × 10⁴cells/mL suspension. Transferring the cell suspension into 96-well culture plate at 100 μ L per well, standing at 37 deg.C and 5% CO₂Culturing for 24h under the condition.

Preparing a mother solution of a tested derivative with DMSO (dimethyl sulfoxide) at a certain concentration, and diluting the derivative mother solution into diluents with different action concentrations by using a DMEM (DMEM) culture medium or a 1640 culture medium. Old medium was removed and 100. mu.L of each well of DMEM medium or 1640 medium containing serum and drug at different concentrations was added. A blank control group and a positive control etoposide (VP-16) control group are additionally arranged. After 72h of drug action, 10. mu.L of MTT solution (5mg/mL) was added to each well and incubation was continued for 4 h.

And (3) absorbing supernatant in each hole, adding 100 mu L of DMSO into each hole, oscillating for 5min to fully dissolve crystals, measuring the light absorption value (OD value) of each hole at 540nm by using an enzyme-labeling instrument, and calculating the proliferation inhibition rate of the cells: the inhibition ratio (%) × (1-mean OD value in drug administration group/mean OD value in blank control group) × 100%. Data processing and calculation of median Inhibitory Concentration (IC) for cancer cell proliferation were performed using SPSS16.0 software₅₀) The results are shown in Table 1. Table 1 shows the results of the in vitro proliferation inhibition effect of ursolic acid quinoline hydrazide heterocyclic derivatives on MDA-MB-231, HeLa and SMMC-7721 cells.

TABLE 1

^aNT: not tested

As shown in Table 1, the synthesized ursolic acid quinoline hydrazide heterocyclic derivatives have stronger inhibition effect on the three tumor cells, wherein the compounds I-a, I-b, I-c and I-d have stronger inhibition effect on MDA-MB-231 cells and are stronger than the positive control etoposide, wherein the inhibition activity of the compounds I-d on the MDA-MB-231 cells is strongest, and the IC is stronger than that of the compounds I-d on the MDA-MB-231 cells₅₀0.12 +/-0.01 mu mol/L; the compounds I-a, I-b, I-c and I-d show better inhibition effect on HeLa cells than etoposide, wherein the inhibition effect of the compounds I-d on the HeLa cells is optimal, and IC₅₀0.08 +/-0.01 mu mol/L; the compounds I-a, I-b, I-c and I-d have stronger proliferation inhibition effect on SMMC-7721 cells, wherein the proliferation inhibition effect of the compounds I-d on the SMMC-7721 cells is optimal, and IC is₅₀0.34 +/-0.03 mu mol/L; in the synthesized ursolic acid quinoline hydrazide derivatives, the compounds I-a, I-b, I-c and I-d all show low toxicity to normal cell QSG-7701 cells, and IC₅₀Are all made of>40 mu mol/L, wherein the IC of the compound I-d with the strongest inhibition effect on the proliferation of three tumor cells on QSG-7701 cells₅₀43.76 +/-1.73 mu mol/L, and has obviously lower cytotoxicity to normal cells than tumor cells. The results show that the compounds have obvious inhibition effect on breast cancer, cervical cancer and liver cancer cells and have development resistancePotential for cancer drugs.

The foregoing shows and describes the general principles, essential features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, which are described in the foregoing description only for the purpose of illustrating the principles of the present invention, but that various changes and modifications may be made therein without departing from the spirit and scope of the invention as defined by the appended claims, specification, and equivalents thereof.

Claims

1. Ursolic acid quinoline hydrazide derivatives with anti-tumor activity and pharmaceutically acceptable salts thereof:

wherein, I-a: r₁＝H R₂＝CH₃；I-b：R₁＝OMe R₂＝CH₃；I-c：R₁＝F R₂＝CH₃；I-d：R₁＝Cl R₂＝CH₃；

I-e：R₁＝H R₂＝n-C₄H₉；I-f：R₁＝OMe R₂＝n-C₄H₉；I-g：R₁＝F R₂＝n-C₄H₉；I-h：R₁＝CI R₂＝n-C₄H₉

2. The preparation method of ursolic acid quinoline hydrazide derivatives with anti-tumor activity as claimed in claim 1, which is characterized by comprising the following steps:

wherein, III-a: r₁＝H；IH-b：R₁＝OMe；III-c：R₁＝F；III-d：R₁＝Cl；

wherein, IV-a: r₁＝H；IV-b：R₁＝OMe；IV-c：R₁＝F；IV-d：R₁＝Cl；

3. The preparation method of ursolic acid quinoline hydrazide derivatives with anti-tumor activity as claimed in claim 2, wherein:

in the step (1), synthesis of 3-Oxaprotinic acid II

Stirring 2-nitrobenzaldehyde in ethanol, acetic acid and distilled water, slowly heating to 50 ℃, adding reduced iron powder and 600 mu L concentrated hydrochloric acid after complete dissolution, continuously heating to 85 ℃, carrying out reflux reaction for 1h, stopping the reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining filtrate, extracting for 3 times by using dichloromethane, collecting an organic phase, washing for 3 times, washing a saturated sodium bicarbonate solution and the concentrated hydrochloric acid once respectively, then removing water by using anhydrous sodium sulfate, and carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, namely a 2-aminobenzaldehyde crude product, which is directly used for synthesizing unsubstituted ursolic acid quinoline; the molar ratio of the 2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the ethanol to the acetic acid to the distilled water is 2: 1; the molar volume of the 2-nitrobenzaldehyde and the total volume of the ethanol, the acetic acid and the distilled water is 0.15 mol/L; the volume ratio of the concentrated hydrochloric acid to the total volume of the ethanol, the acetic acid and the distilled water is 0.6: 20;

the molar ratio of the 3-oxoursolic acid to the ethanol solution of the 2-aminobenzaldehyde is 2: 3; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.35 g/mL;

Dissolving the compound III-a with 5mL of benzene, slowly dropwise adding thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4 hours, stopping the reaction, and evaporating the benzene and the thionyl chloride in the reaction liquid to obtain a light yellow solid compound IV-a for subsequent reaction; the mol ratio of the compound III-a to the thionyl chloride and the benzene is 0.15:3: 56;

Dissolving the obtained ursolic acid quinoline hydrazide heterocyclic derivative IV-a in 5mL of diethyl ether, cooling to 0 ℃, dissolving acethydrazide and triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring and reacting for 6h, pouring reaction liquid into an ice water mixture after the reaction is finished, extracting 3 times by using dichloromethane after ice is melted, collecting an organic phase, washing 3 times by using water, washing a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water by using anhydrous sodium sulfate, performing reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying by using a silica gel column, selecting a petroleum ether acetone system as a solvent, wherein the volume ratio of petroleum ether to acetone is 30:1, and preparing a purified compound ursolic acid quinoline hydrazide heterocyclic derivative I-a; the molar ratio of the IV-a, the acethydrazide, the triethylamine, the diethyl ether and the dichloromethane is 0.15:0.225:0.3:48: 76.

4. The preparation method of ursolic acid quinoline hydrazide derivatives with anti-tumor activity as claimed in claim 2, wherein:

in the step (1), synthesis of 3-Oxaprotinic acid II

Adding ursolic acid and acetone into a 500mL round-bottom flask, stirring and dissolving, stirring and reacting in ice water for 15min, slowly dropwise adding 1.872mL Jones reagent, heating to room temperature, stirring and reacting for 5h, adding isopropanol, stirring and reacting for 30min, filtering out precipitates after the reaction is finished, collecting filtrate, and recrystallizing pale yellow green viscous solid obtained by concentrating the filtrate under reduced pressure with methanol to obtain white needle crystals to obtain 3-oxidized ursolic acid II; the molar volume ratio of the ursolic acid to the acetone is 0.0184 mol/L; the volume ratio of the acetone to the Jones reagent to the isopropanol is 250: 1.9: 90;

dropwise adding 2.4mL of saturated potassium hydroxide ethanol solution into 3-oxidized ursolic acid II and 2-aminobenzaldehyde ethanol solution in a 50mL three-neck round-bottom flask under the protection of nitrogen, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction solution into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, carrying out separation and purification by using a silica gel column, wherein the solvent is a petroleum ether acetone system, the volume ratio of petroleum ether to acetone is 150:1, and preparing a purified compound ursolic acid quinoline heterocyclic derivative III-a; the molar ratio of the 3-oxoursolic acid to the ethanol solution of the 2-aminobenzaldehyde is 2: 3; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.35 g/mL;

Dissolving the obtained compound IV-a in 5mL of diethyl ether, cooling to 0 ℃, dissolving hydrazine valerate and triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring reaction liquid into an ice water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine for one time respectively, removing water with anhydrous sodium sulfate, decompressing, concentrating and removing an organic solvent to obtain a yellow solid, separating and purifying the yellow solid with a silica gel column, wherein the solvent is a petroleum ether acetone system, the volume ratio of petroleum ether to acetone is 80:1, and obtaining a purified compound, namely the ursolic acid quinoline hydrazide heterocyclic derivative I-e; the molar ratio of the IV-a, the hydrazine valerate, the triethylamine, the diethyl ether and the dichloromethane is 0.15:0.225:0.3:48: 76.

5. The preparation method of ursolic acid quinoline hydrazide derivatives with anti-tumor activity as claimed in claim 2, wherein:

in the step (1), synthesis of 3-Oxaprotinic acid II

Adding 5-hydroxy-2-nitrobenzaldehyde, methyl iodide, potassium carbonate and N, N-dimethylformamide into a 50mL round-bottom flask, stirring at room temperature for reacting for 8h, pouring a reaction solution into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid which is 100% of 5-methoxy-2-nitrobenzaldehyde; the molar ratio of the 5-hydroxy-2-nitrobenzaldehyde, the methyl iodide and the potassium carbonate is 4: 6; the volume ratio of the methyl iodide to the N, N-dimethylformamide is 0.4: 10;

stirring 5-methoxy-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water, slowly heating to 50 ℃, adding reduced iron powder and 600 mu L concentrated hydrochloric acid after complete dissolution, continuously heating to 85 ℃, carrying out reflux reaction for 1h, after the reaction is stopped, carrying out suction filtration on the mixed solution, washing filter residues with water, combining the filtrate, extracting with dichloromethane for 3 times, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated hydrochloric acid once respectively, then removing water with anhydrous sodium sulfate, and carrying out reduced pressure concentration to remove the organic solvent to obtain a yellow solid which is a 5-methoxy-2-aminobenzaldehyde crude product which is directly used for synthesizing methoxyursolic acid quinoline; the mass-volume ratio of the 5-methoxy-2-nitrobenzaldehyde to the total volume of the ethanol, the acetic acid and the distilled water is 0.02 g/mL; the volume ratio of the ethanol to the acetic acid to the distilled water is 2: 1; the molar ratio of the 5-methoxy-2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the total volume of the ethanol, the acetic acid and the distilled water to the volume of the concentrated hydrochloric acid is 175: 6;

dropwise adding 0.445g (1mmol) of 3-oxidized ursolic acid II and 0.225g (1.5mmol) of 5-methoxy-2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein the solvent is a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 100:1, thus obtaining a purified compound of ursolic acid quinoline heterocyclic derivatives III-b; the molar ratio of the 3-oxoursolic acid to the ethanol solution of the 5-methoxy-2-aminobenzaldehyde is 2: 3; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.22 g/mL;

Dissolving 0.085g (0.15mmol) of the compound III-b with 5mL of benzene, then slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain a light yellow solid compound IV-b for subsequent reaction; the mol ratio of the compound III-b to the thionyl chloride and the benzene is 0.15:3: 56;

Dissolving the obtained compound IV-b in 5mL of diethyl ether, cooling to 0 ℃, dissolving 17.1mg (0.225mmol) of acethydrazide and 45 mu L (0.3mmol) of triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein a petroleum ether-acetone system is adopted as the solvent, and the volume ratio of petroleum ether to acetone is 30:1, so as to obtain a purified compound I-b of quinoline hydrazide heterocyclic derivatives of ursolic acid; the molar ratio of IV-b, acethydrazide, triethylamine, diethyl ether and dichloromethane is 0.15:0.225:0.3:48: 76.

6. The ursolic acid quinoline hydrazide derivatives with anti-tumor activity and the preparation method thereof as claimed in claim 2, wherein:

in the step (1), synthesis of 3-Oxaprotinic acid II

Adding 0.668g (4mmol) of 5-hydroxy-2-nitrobenzaldehyde, 400 mu L (6mmol) of methyl iodide, 0.832g (6mmol) of potassium carbonate and 10mL of N, N-dimethylformamide into a 50mL round bottom flask, stirring at room temperature for reacting for 8h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once, removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid which is 5-methoxy-2-nitrobenzaldehyde (0.52g, 100%); the molar ratio of the 5-hydroxy-2-nitrobenzaldehyde, the methyl iodide and the potassium carbonate is 4: 6; the volume ratio of the methyl iodide to the N, N-dimethylformamide is 0.4: 10;

stirring 0.36g (2mmol) of 5-methoxy-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2: 1, 17.5mL), slowly heating to 50 ℃, completely dissolving, adding 1.12g (20mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining filtrates, extracting for 3 times with dichloromethane, collecting an organic phase, washing for 3 times, washing with saturated sodium bicarbonate solution and concentrated hydrochloric acid once respectively, removing water with anhydrous sodium sulfate, concentrating under reduced pressure to remove an organic solvent to obtain a yellow solid which is a 5-methoxy-2-aminobenzaldehyde crude product (0.24g), and directly using the yellow solid for synthesizing methoxyursolic acid quinoline; the mass-volume ratio of the 5-methoxy-2-nitrobenzaldehyde to the total volume of the ethanol, the acetic acid and the distilled water is 0.02 g/mL; the volume ratio of the ethanol to the acetic acid to the distilled water is 2: 1; the molar ratio of the 5-methoxy-2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the total volume of the ethanol, the acetic acid and the distilled water to the volume of the concentrated hydrochloric acid is 175: 6;

7. The ursolic acid quinoline hydrazide derivatives with anti-tumor activity and the preparation method thereof as claimed in claim 2, wherein:

in the step (1), synthesis of 3-Oxaprotinic acid II

Stirring 0.51g (3mmol) of 5-fluoro-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2: 1, 20mL), slowly heating to 50 ℃, completely dissolving, adding 1.68g (30mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining the filtrate, extracting 3 times with dichloromethane, collecting an organic phase, washing 3 times with water, washing the saturated sodium bicarbonate solution and the concentrated hydrochloric acid once respectively, removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove the organic solvent to obtain a yellow solid which is a 5-fluoro-2-aminobenzaldehyde crude product (0.43g) which is directly used for synthesizing ursolic acid quinoline fluoride; the molar ratio of the 5-fluoro-2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the ethanol to the acetic acid to the distilled water is 2: 1; the molar volume of the 5-fluoro-2-nitrobenzaldehyde with the total volume of ethanol, acetic acid and distilled water is 0.15 mol/L; the volume ratio of the concentrated hydrochloric acid to the total volume of the ethanol, the acetic acid and the distilled water is 0.6: 20;

dropwise adding 0.67g (1.5mmol) of 3-oxoursolic acid II and 0.35g (2.5mmol) of 5-fluoro-2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen into 2mL of saturated potassium hydroxide ethanol solution, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein a petroleum ether-acetone system is adopted as a solvent, and the volume ratio of petroleum ether to acetone is 150:1, so as to obtain a purified compound ursolic acid quinoline heterocyclic derivative III-c; the molar ratio of the 3-oxoursolic acid to the ethanol solution of the 5-fluoro-2-aminobenzaldehyde is 3: 5; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.34 g/mL;

Dissolving 0.084g (0.15mmol) of the compound III-c with 5mL of benzene, then slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain a light yellow solid compound IV-c for subsequent reaction; the molar ratio of the compound III-c to the thionyl chloride and the benzene is 0.15:3: 56;

Dissolving the obtained compound IV-c in 5mL of diethyl ether, cooling to 0 ℃, dissolving 17.1mg (0.225mmol) of acethydrazide and 45 mu L (0.3mmol) of triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein the solvent is a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 30:1, thus obtaining a purified compound I-c of quinoline heterocyclic derivatives of ursolic acid; the molar ratio of IV-c, acethydrazide, triethylamine, diethyl ether and dichloromethane is 0.15:0.225:0.3:48: 76.

8. The ursolic acid quinoline hydrazide derivatives with anti-tumor activity and the preparation method thereof as claimed in claim 2, wherein:

in the step (1), synthesis of 3-Oxaprotinic acid II

dropwise adding 0.67g (1.5mmol) of 3-oxoursolic acid II and 0.35g (2.5mmol) of 5-fluoro-2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen into 2mL of saturated potassium hydroxide ethanol solution, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, and carrying out separation and purification by using a silica gel column, wherein the solvent adopts a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 150:1, thus obtaining a purified compound III-c; the molar ratio of the 3-oxoursolic acid to the ethanol solution of the 5-fluoro-2-aminobenzaldehyde is 3: 5; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.34 g/mL;

9. The ursolic acid quinoline hydrazide derivatives with anti-tumor activity and the preparation method thereof as claimed in claim 2, wherein:

in the step (1), synthesis of 3-Oxaprotinic acid II

Stirring 0.55g (3mmol) of 5-chloro-2-nitrobenzaldehyde in ethanol, acetic acid and distilled water (2: 1, 20mL), slowly heating to 50 ℃, completely dissolving, adding 1.68g (30mmol) of reduced iron powder and 600 mu L of concentrated hydrochloric acid, continuously heating to 85 ℃, refluxing and reacting for 1h, stopping reaction, carrying out suction filtration on the mixed solution, washing filter residue with water, combining filtrate, extracting 3 times with dichloromethane, collecting organic phase, washing 3 times with water, washing saturated sodium bicarbonate solution and concentrated hydrochloric acid once respectively, removing water with anhydrous sodium sulfate, and concentrating under reduced pressure to remove organic solvent to obtain a yellow solid which is 5-chloro-2-aminobenzaldehyde crude product (0.45g) which is directly used for synthesizing ursolic acid quinoline chloride; the molar ratio of the 5-chloro-2-nitrobenzaldehyde to the reduced iron powder is 1: 10; the volume ratio of the ethanol to the acetic acid to the distilled water is 2: 1; the molar volume of the 5-chloro-2-nitrobenzaldehyde with the total volume of ethanol, acetic acid and distilled water is 0.15 mol/L; the volume ratio of the concentrated hydrochloric acid to the total volume of the ethanol, the acetic acid and the distilled water is 0.6: 20;

dropwise adding 0.67g (1.5mmol) of 3-oxoursolic acid II and 0.38g (2.5mmol) of 5-chloro-2-aminobenzaldehyde ethanol solution into a 50mL three-neck round-bottom flask under the protection of nitrogen, carrying out reflux reaction at 85 ℃ for 24h, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with saturated sodium bicarbonate solution and concentrated brine once respectively, then removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, and carrying out separation and purification by using a silica gel column, wherein the solvent adopts a petroleum ether-acetone system, and the volume ratio of petroleum ether to acetone is 150:1, thus obtaining a purified compound III-d; the molar ratio of the 3-oxoursolic acid to the ethanol solution of the 5-chloro-2-aminobenzaldehyde is 3: 5; the mass-volume ratio of the 3-oxoursolic acid to the saturated potassium hydroxide ethanol solution is 0.34 g/mL;

in the step (3), synthesis of ursolic acid quinoline acylchloride compound IV-d

Dissolving 0.086g (0.15mmol) of the compound III-d with 5mL of benzene, then slowly dropwise adding 200 mu L (3mmol) of thionyl chloride, slowly heating to 80 ℃, refluxing for reaction for about 4h, stopping the reaction, and distilling off the benzene and the thionyl chloride in the reaction liquid to obtain a light yellow solid compound IV-d for subsequent reaction; the mol ratio of the compound III-d to the thionyl chloride and the benzene is 0.15:3: 56;

Dissolving the obtained compound IV-d in 5mL of diethyl ether, cooling to 0 ℃, dissolving 17.1mg (0.225mmol) of acethydrazide and 45 mu L (0.3mmol) of triethylamine in 2mL of dichloromethane, dropwise adding the mixture into a reaction container, slowly heating to room temperature, stirring for reaction for 6 hours, pouring the reaction liquid into an ice-water mixture after the reaction is finished, extracting with dichloromethane for 3 times after ice is melted, collecting an organic phase, washing with water for 3 times, washing with a saturated sodium bicarbonate solution and concentrated brine once respectively, removing water with anhydrous sodium sulfate, carrying out reduced pressure concentration to remove an organic solvent to obtain a yellow solid, separating and purifying the yellow solid by using a silica gel column, wherein a petroleum ether-acetone system is adopted as the solvent, and the volume ratio of petroleum ether to acetone is 30:1, so as to obtain a purified compound I-d of quinoline hydrazide heterocyclic derivatives of ursolic acid; the molar ratio of IV-d, acethydrazide, triethylamine, diethyl ether and dichloromethane is 0.15:0.225:0.3:48: 76.

10. The ursolic acid quinoline hydrazide derivatives with anti-tumor activity and the preparation method thereof as claimed in claim 2, wherein:

in the step (1), synthesis of 3-Oxaprotinic acid II

in the step (3), synthesis of ursolic acid quinoline acylchloride compound IV-d

11. The use of ursolic acid quinoline hydrazide heterocyclic derivatives with the structure shown in formula I in claim 1 and pharmaceutically acceptable salts thereof in the preparation of drugs for treating tumors.

12. Use according to claim 10, characterized in that: the tumor is any one of human breast cancer, cervical cancer or liver cancer.