CN110759961A - Ursolic acid indolyquinone amide derivatives and preparation method and application thereof - Google Patents

Abstract

The invention discloses ursolic acid indoloquinone amide derivatives, and a preparation method and application thereof. The invention relates to ursolic acid indoloquinone amide derivatives I-a to I-I with a structure shown in a general formula (I) and pharmaceutically acceptable salts thereof:

Description

Ursolic acid indolyquinone amide derivatives and preparation method and application thereof

Technical Field

The invention relates to the technical field of organic synthesis and pharmaceutical chemistry, in particular to ursolic acid indoloquinone amide derivatives 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 main means of tumor therapy include surgery, radiotherapy (abbreviated as radiotherapy), chemotherapy (abbreviated as chemotherapy), cellular immunotherapy, and the like. Chemotherapy is a common treatment method in which cytotoxic drugs 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, faster proliferating cells, causing symptoms such as bone marrow suppression, infection, bleeding, and the like. 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 oldenlandia diffusa, glossy privet fruit, dark plum, selfheal and the like, and is statistically separated from 108 plants in 34 families in the nature, so the ursolic acid is a plant active ingredient with abundant resources and development potential. Ursolic acid has antiviral, antibacterial, antifungal, hepatoprotective, antiinflammatory, and antitumor effects. Among them, its antitumor effect has been 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. The research on the structural modification of ursolic acid mainly focuses on the modification of A-ring, C-ring and C28 carboxyl. At present, very few reports about ursolic acid heterocyclic derivatives are reported at home and abroad.

Indoloquinone belongs to indole alkaloids, is an endogenous natural active compound existing in animals, plants and marine organisms, is also a main active component in natural indigo naturalis, and has pharmacological activities of resisting bacteria and atherosclerosis, reducing cholesterol, inhibiting vascular endothelial cell growth factor, resisting cancer, early warning Parkinson disease, regulating balance of acetylcholine and dopamine in brain, and the like. It is also an important medical intermediate, and can be used for synthesizing indirubin with anticancer effect, and tryptanthrin and its derivatives which are natural products with antibacterial, anti-inflammatory and anticancer activities. In addition, several natural products or synthetic compounds having indoloquinone structure also show significant antitumor effects. For example, mitomycin C is a spectral antitumor antibiotic isolated and extracted from Streptomyces caecum culture solution, has therapeutic effect on various cancers, and is clinically suitable for digestive tract cancers such as gastric cancer, intestinal cancer, liver cancer, pancreatic cancer and the like, and is also effective on lung cancer, breast cancer, cervical cancer, chorionic epithelial cancer and the like. Therefore, the indoloquinone is a very potential pharmacophore in the research and development of antitumor drugs. If the ring A of the ursolic acid molecule is introduced with indoloquinone groups, the carboxyl groups are further derivatized, and different amide groups are introduced; the synthesized series derivatives are tested for anti-tumor activity, for example, the novel ursolic acid indoloquinone derivatives with better anti-tumor activity are obtained, which has important chemical and biological significance for researching and developing novel anti-tumor drugs.

At present, an ursolic acid indoloquinone amide derivative with high anti-tumor activity, a preparation method and an application thereof are lacked.

Disclosure of Invention

In order to overcome the defects in the prior art, the invention aims to provide ursolic acid indoloquinone amide derivatives with high 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 indoloquinone amide derivatives I-a to I-I with a structure shown in a general formula (I) and pharmaceutically acceptable salts thereof:

wherein, I-a: r is H, n is 1; i-b: r is CH₃，n＝1；I-c：R＝C₂H₅，n＝1；

I-d：R＝H，n＝2；I-e：R＝CH₃，n＝2；I-f：R＝C₂H₅，n＝2；

I-g：R＝H，n＝3；I-h：R＝H₃，n＝3；I-i：R＝C₂H₅，n＝3。

The preparation method of the ursolic acid indoloquinone amide derivatives I-a to I-I with the structure shown in the general formula (I) 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) 3-oxoursolic acid and 3, 5-dimethoxy phenylhydrazine are subjected to Fishier indole synthesis reaction to obtain a 3, 5-dimethoxy indole derivative, which has a structure shown in a general formula III:

(3) the indole derivative, phosphorus oxychloride and N, N-dimethylformamide are subjected to Vilsmeier-Haack formylation reaction to obtain a 3, 5-dimethoxy-2-aldehyde indole derivative which has a structure shown in a general formula IV:

(4) the compound IV is subjected to oxidation reaction with hydrogen peroxide to obtain ursolic acid indoloquinone derivative with a structure shown in a general formula V:

(5) carrying out amidation reaction on the compound V and different fatty amines to obtain the ursolic acid indoloquinone amide derivative with the structure shown in the general formula VI:

wherein, I-a: r is equal to the content of H, n is equal to the content of 1; i-b: r ≡ CH₃，n≡1；I-c：R≡C₂H₅，n≡1；

I-d：R＝H，n＝2；I-e：R＝CH₃，n＝2；I-f：R＝C₂H₅，n＝2；

I-g：R≡H，n≡3；I-h：R≡CH₃，n≡3；I-i：R≡C₂H₅，n≡3

Further, in step (2), 0.33g (3mmol) of 3, 5-dimethoxyaniline was weighed out and dissolved in 3mL of 20% hydrochloric acid in a single-neck round-bottom flask. Then 0.28g (4mmol) of sodium nitrite is weighed and dissolved in 0.7mL of water, and slowly dropped into the reaction flask under the ice bath condition, and the reaction is stirred for 1h under the ice bath condition. Weighing 1.35g (6mmol) of stannous chloride, dissolving in 1.8mL of concentrated hydrochloric acid, slowly dripping into the flask in which the reaction in the previous step is finished under the ice bath condition, stirring for 2 hours at room temperature, and performing suction filtration after the reaction is finished, wherein the solid is 3, 5-dimethoxy phenylhydrazine; the molar ratio of the 3, 5-dimethoxyaniline to the sodium nitrite to the stannous chloride is 3: 4: 6.

weighing 0.441g (1mmol) of 3-oxoursolic acid, dissolving in 10mL of absolute ethanol, adding the 3, 5-dimethoxyphenylhydrazine (3mmol) reacted in the previous step into the system, adding 0.5mL of concentrated hydrochloric acid, and refluxing at 85 ℃ for 3 h. After the reaction was completed, the reaction was poured into a beaker containing 20mL of ice water, and when the ice was melted, extraction was carried out, and the solvent was distilled off. Separating and purifying by silica gel column chromatography, gradient eluting with petroleum ether/acetone (20:1-5:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound III; the molar ratio of the 3-oxoursolic acid to the 3, 5-dimethoxyphenylhydrazine is 1: 3.

further, in step (3), 0.24g (1.6mmol) of POCl was added₃Dropwise adding into 0.2mL (2mmol) of DMF cooled with ice water under stirring to react for 0.5h to form a coordination complex, slowly adding 0.59g (1mmol) of compound III into the complex reaction solution, stirring at 30 deg.C to react for 3h to generate an intermediate, adding the intermediate reaction solution into the stirred ice water to hydrolyze for 2h, melting ice, extractingAnd the solvent was distilled off. Separating and purifying by silica gel column chromatography, gradient eluting with petroleum ether/acetone (20:1-10:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound IV; the POCl₃DMF, compound III in a molar ratio of 1.6: 2: 1.

further, in step (4), 0.2g (0.31mmol) of Compound IV was dissolved in 50mL of methanol, and 3mL of 30% H was added₂O₂And 3 drops of concentrated hydrochloric acid, stirring the mixed solution in an ice bath for reacting for 3 hours, pouring the mixed solution into 50mL of ice water, melting the ice, extracting, and distilling to remove the solvent. Separating and purifying by silica gel column chromatography, gradient eluting with petroleum ether/acetone (15:1-8:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound V; the pure compound IV: 30% H₂O₂: the molar ratio of concentrated hydrochloric acid was 1:96.8: 15.6.

Further, in step (5), 30mg (0.05mmol) of compound V was dissolved in 2mL of dichloromethane, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) were added to the reaction system, and after stirring at room temperature for reaction for 30min, 4.8mg (0.08mmol) of ethylenediamine was added. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, dissolving the filtrate in acetonitrile for overnight standing at 4 ℃ after vacuum concentration, further filtering out precipitate, removing the solvent by vacuum concentration, performing gradient elution by using dichloromethane/methanol (30: 1-10:1), combining product components, performing vacuum concentration, removing the solvent, and preparing a pure compound I-a; the HOBt: DCC: pure compound V: the molar ratio of ethylenediamine was 7:7:5: 8.

Further, in the step (5), the different aliphatic amines are ethylenediamine, N-dimethylethylenediamine, N-diethylethylenediamine, 1, 3-propylenediamine, 3-dimethylaminopropylamine, 3-diethylaminopropylamine, 1, 4-butanediamine, 4-dimethylaminobutylamine, 4-diethylaminobutylamine;

in the step (5), when the ethylenediamine is used, the pure compound I-a is prepared; when the N, N-dimethyl ethylenediamine is used, a pure compound I-b is prepared; when the N, N-diethyl ethylenediamine is used, pure compounds I-c are prepared; when the 1, 3-propane diamine is used, obtaining pure compounds I-d; when the 3-dimethylaminopropylamine is present, obtaining the pure compound I-e; when the 3-diethylaminopropylamine is used, pure compounds I-f are prepared; when the 1, 4-butanediamine is used, preparing pure compound I-g; when the 4-dimethylamino butylamine is used, pure compounds I-h are prepared; when the 4-diethylaminobutylamine is used, the pure compound I-I is obtained.

The invention relates to an application of ursolic acid indoloquinone amide derivative with a structure shown in formula I and pharmaceutically acceptable salt thereof in preparing a medicament for treating tumors.

Furthermore, the tumors are human liver cancer cells HepG2 and SMMC-7721.

Has the advantages that: the ursolic acid indoloquinone derivative has antitumor activity, and pharmacological experiments show that the ursolic acid indoloquinone derivative has obvious inhibition effect on human liver cancer cells HepG2 and SMMC-7721, wherein the compound I-h has the best in-vitro antitumor activity.

Compared with the prior art, the invention has the following advantages:

the invention obtains 3, 5-dimethoxy indole derivatives by oxidizing C-3 site hydroxyl on an ursolic acid A ring into carbonyl and then reacting with 3, 5-dimethoxy phenylhydrazine, then generates indoloquinone groups by Vilsmeier-Haack formylation reaction and oxidation reaction, and then carries out amidation reaction on C-28 site carboxyl of the ursolic acid indoloquinone derivatives to obtain the corresponding ursolic acid indoloquinone amide derivatives. The derivatives have novel structures and are not reported at home and abroad. Has potential value in 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 relates to ursolic acid indoloquinone amide derivatives I-a to I-I with a structure shown in a general formula (I) and pharmaceutically acceptable salts thereof:

wherein, I-a: r is H, n is 1; i-b: r ═CH₃，n＝1；I-c：R＝C₂H₅，n＝1；

I-d：R＝H，n＝2；I-e：R＝CH₃，n＝2；I-f：R＝C₂H₅，n＝2；

I-g：R＝H，n＝3；I-h：R＝CH₃，n＝3；I-i：R＝C₂H₅，n＝3。

The preparation method of the ursolic acid indoloquinone amide derivatives I-a to I-I with the structure shown in the general formula (I) 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) 3-oxoursolic acid and 3, 5-dimethoxy phenylhydrazine are subjected to Fishier indole synthesis reaction to obtain a 3, 5-dimethoxy indole derivative, which has a structure shown in a general formula III:

(3) the indole derivative, phosphorus oxychloride and N, N-dimethylformamide are subjected to Vilsmeier-Haack formylation reaction to obtain a 3, 5-dimethoxy-2-aldehyde indole derivative which has a structure shown in a general formula IV:

(4) the compound IV is subjected to oxidation reaction with hydrogen peroxide to obtain ursolic acid indoloquinone derivative with a structure shown in a general formula V:

(5) carrying out amidation reaction on the compound V and different fatty amines to obtain the ursolic acid indoloquinone amide derivative with the structure shown in the general formula VI:

wherein, I-a: r is H, n is 1; i-b: r is CH₃，n＝1；I-c：R＝C₂H₅，n＝1；

I-d：R＝H，n＝2；I-e：R＝CH₃，n＝2；I-f：R＝C₂H₅，n＝2；

I-g：R＝H，n＝3；I-h：R＝CH₃，n＝3；I-i：R＝C₂H₅，n＝3。

In the step (1), 2g (4.6mmol) of ursolic acid and 250mL of acetone are added into a 500mL round-bottom flask, stirred and dissolved, then stirred and reacted in ice water for 15min, 1.87mL of Jones reagent is slowly dripped, the temperature is raised to room temperature, stirred and reacted for 5h, then 90mL of isopropanol is added, stirred and reacted for 30min, after the reaction is finished, the precipitate is filtered out, the filtrate is collected, the filtrate is decompressed and concentrated, and the light yellow green viscous solid obtained by the filtrate is recrystallized by methanol to obtain white needle-shaped crystals, namely 3-oxoursolic acid (II).

In step (2), 0.33g (3mmol) of 3, 5-dimethoxyaniline was weighed out and dissolved in 3mL of 20% hydrochloric acid in a single-neck round-bottom flask. Then 0.28g (4mmol) of sodium nitrite is weighed and dissolved in 0.7mL of water, and slowly dropped into the reaction flask under the ice bath condition, and the reaction is stirred for 1h under the ice bath condition. Weighing 1.35g (6mmol) of stannous chloride, dissolving in 1.8mL of concentrated hydrochloric acid, slowly dripping into the flask in which the reaction in the previous step is finished under the ice bath condition, stirring for 2 hours at room temperature, and performing suction filtration after the reaction is finished, wherein the solid is 3, 5-dimethoxy phenylhydrazine; the molar ratio of the 3, 5-dimethoxyaniline to the sodium nitrite to the stannous chloride is 3: 4: 6.

weighing 0.441g (1mmol) of 3-oxoursolic acid, dissolving in 10mL of absolute ethanol, adding the 3, 5-dimethoxyphenylhydrazine (3mmol) reacted in the previous step into the system, adding 0.5mL of concentrated hydrochloric acid, and refluxing at 85 ℃ for 3 h. After the reaction was completed, the reaction was poured into a beaker containing 20mL of ice water, and when the ice was melted, extraction was carried out, and the solvent was distilled off. Separating and purifying by silica gel column chromatography, gradient eluting with petroleum ether/acetone (20:1-5:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound III; the molar ratio of the 3-oxoursolic acid to the 3, 5-dimethoxyphenylhydrazine is 1: 3.

in step (3), 0.24g (1.6mmol) of POCl was added₃Dropwise adding the mixture into 0.2mL (2mmol) of DMF which is stirred and cooled by ice water to react for 0.5h to form a coordination complex, slowly adding 0.59g (1mmol) of the compound III into the complex reaction solution, stirring at 30 ℃ for reacting for 3h to generate an intermediate, adding the intermediate reaction solution into the stirred ice water to hydrolyze for 2h, melting the ice, extracting, and distilling to remove the solvent. Separating and purifying by silica gel column chromatography, gradient eluting with petroleum ether/acetone (20:1-10:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound IV; the POCl₃DMF, compound III in a molar ratio of 1.6: 2: 1.

in step (4), 0.2g (0.31mmol) of Compound IV was dissolved in 50mL of methanol, and 3mL of 30% H was added₂O₂And 3 drops of concentrated hydrochloric acid, stirring the mixed solution in an ice bath for reacting for 3 hours, pouring the mixed solution into 50mL of ice water, melting the ice, extracting, and distilling to remove the solvent. Separating and purifying by silica gel column chromatography, gradient eluting with petroleum ether/acetone (15:1-8:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound V; the pure compound IV: 30% H₂O₂: the molar ratio of concentrated hydrochloric acid was 1:96.8: 15.6.

In step (5), 30mg (0.05mmol) of Compound V was dissolved in 2mL of dichloromethane, and HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) were added to the reaction system, and the reaction was stirred at room temperature for 30min, and then 4.8mg (0.08mmol) of ethylenediamine was added thereto. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, dissolving the filtrate in acetonitrile for overnight standing at 4 ℃ after vacuum concentration, further filtering out precipitate, removing the solvent by vacuum concentration, performing gradient elution by using dichloromethane/methanol (30: 1-10:1), combining product components, performing vacuum concentration, removing the solvent, and preparing a pure compound I-a; the HOBt: DCC: pure compound V: the molar ratio of ethylenediamine was 7:7:5: 8.

In the step (5), the different aliphatic amines are ethylenediamine, N-dimethylethylenediamine, N-diethylethylenediamine, 1, 3-propylenediamine, 3-dimethylaminopropylamine, 3-diethylaminopropylamine, 1, 4-butylenediamine, 4-dimethylaminobutylamine, 4-diethylaminobutylamine;

in the step (5), when the ethylenediamine is used, the pure compound I-a is prepared; when the N, N-dimethyl ethylenediamine is used, a pure compound I-b is prepared; when the N, N-diethyl ethylenediamine is used, pure compounds I-c are prepared; when the 1, 3-propane diamine is used, obtaining pure compounds I-d; when the 3-dimethylaminopropylamine is present, obtaining the pure compound I-e; when the 3-diethylaminopropylamine is used, pure compounds I-f are prepared; when the 1, 4-butanediamine is used, preparing pure compound I-g; when the 4-dimethylamino butylamine is used, pure compounds I-h are prepared; when the 4-diethylaminobutylamine is used, the pure compound I-I is obtained.

The invention relates to an application of ursolic acid indoloquinone amide derivative with a structure shown in formula I and pharmaceutically acceptable salt thereof in preparing a medicament for treating tumors.

The tumor is human liver cancer cell HepG2 and SMMC-7721.

Example 1

EXAMPLE 13 Synthesis of Oxyaursolic acid (II) of the present invention

Adding 2g (4.6mmol) of ursolic acid and 250mL of acetone into a 500mL round-bottom flask, stirring for dissolving, then stirring for reaction in ice water for 15min, slowly dropwise adding 1.87mL of Jones reagent, heating to room temperature, stirring for reaction for 5h, then 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-shaped crystal, namely 3-oxoursolic acid (II) (1.2g, 65.6%).

Example 2

Synthesis of ursolic acid 3, 5-dimethoxyindole derivative (III)

3, 5-Dimethoxyaniline 0.33g (3mmol) was weighed out and dissolved in 3mL of 20% hydrochloric acid in a single-neck round-bottom flask. Then 0.28g (4mmol) of sodium nitrite is weighed and dissolved in 0.7mL of water, and slowly dropped into the reaction flask under the ice bath condition, and the reaction is stirred for 1h under the ice bath condition. Weighing stannous chloride 1.35g (6mmol) and dissolving in concentrated hydrochloric acid 1.8mL, slowly dripping into the flask in which the reaction in the previous step is finished under the ice bath condition, stirring for 2 hours at room temperature, and performing suction filtration after the reaction is finished, wherein the solid is 3, 5-dimethoxy phenylhydrazine.

Weighing 0.441g (1mmol) of 3-oxoursolic acid, dissolving in 10mL of absolute ethanol, adding the 3, 5-dimethoxyphenylhydrazine (3mmol) reacted in the previous step into the system, adding 0.5mL of concentrated hydrochloric acid, and refluxing at 85 ℃ for 3 h. After the reaction is finished, pouring the reactant into a beaker filled with 20mL of ice water, and extracting with dichloromethane for 3 times with 20mL each time after the ice is melted; the combined organic phases were washed 3 times with 20mL portions of water, 1 time with 20mL portions of saturated sodium bicarbonate solution and finally with 20mL portions of saturated sodium chloride solution. Dried over anhydrous sodium sulfate, and the solvent was distilled off. Separating and purifying by silica gel column chromatography, eluting with petroleum ether/acetone gradient (20:1-5:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound III 0.39g with yield of 66.3%.

M.p.227～230℃；¹H NMR(300MHz,CDCl₃):δ0.88(s,3H),0.90(d,J＝6.5Hz, 3H),0.93(d,J＝10.9Hz,3H),0.97(s,3H),1.13(s,3H),1.15(s,3H),1.27(s,3H), 1.32～1.90(m,16H),1.98(m,1H),2.10～2.17(m,2H),2.19(d,J＝14.9Hz,1H),2.25 (d,J＝10.3Hz,1H),3.07(d,J＝15.5Hz,1H),3.81(s,3H,OCH₃),3.86(s,3H, OCH₃),5.36(t,J＝3.3Hz,1H),6.15(s,1H),6.42(s,1H),7.55(s,1H)；IR(KBr, cm^-1):3420,2924,2866,1696,1627,1575,1457,1260,1202,1150,806；MS(ESI): m/z[M+H]⁺:604.8；Anal.Calcd.for C₃₈H₅₃NO₄:C77.64；H 9.09；N 2.38；found:C 77.69；H 9.06；N 2.36.

Example 3

Synthesis of ursolic acid 3, 5-dimethoxy-2-aldehyde indole derivative (IV)

0.24g (1.6mmol) of POCl₃Dropwise adding the mixture into 0.2mL (2mmol) of DMF which is stirred and cooled by ice water to react for 0.5h to form a coordination complex, slowly adding 0.59g (1mmol) of the compound III into the complex reaction solution, stirring at 30 ℃ to react for 3h to generate an intermediate, adding the intermediate reaction solution into the stirred ice water to hydrolyze for 2h, melting the ice, and extracting with dichloromethane for 3 times, wherein each time is 20 mL; the combined organic phases were washed 3 times with 20m of water each timeL, washed 1 more times with 20mL of saturated sodium bicarbonate solution and finally once with 20mL of saturated sodium chloride solution. Dried over anhydrous sodium sulfate, and the solvent was distilled off. Separating and purifying by silica gel column chromatography, eluting with petroleum ether/acetone gradient (20:1-10:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound IV,0.46g, with yield of 74.7%.

M.p.260～263℃；¹H NMR(300MHz,CDCl₃):δ0.88(s,3H),0.90(d,J＝6.5 Hz,3H),0.93(d,J＝10.9Hz,3H),0.97(s,3H),1.13(s,3H),1.15(s,3H),1.27(s, 3H),1.32～1.90(m,16H),1.98(m,1H),2.10～2.17(m,2H),2.19(d,J＝14.9Hz,1H), 2.25(d,J＝10.3Hz,1H),3.02(d,J＝15.6Hz,1H),3.95(s,3H,OCH₃),3.97(s,3H, OCH₃),5.36(t,J＝3.3Hz,1H),6.08(s,1H),9.92(s,1H),10.33(s,1H)；IR(KBr, cm^-1):3428,3338,3296,2926,2848,1694,1637,1579,1575,1458,1250,1215,993, 791；MS(ESI):m/z[M+H]⁺:616.8；Anal.Calcd.forC₃₉H₅₃NO₅:C 76.06；H 8.67；N 2.27；found:C 75.98；H 8.73；N 2.29.

Example 4

Synthesis of ursolic acid indoloquinone derivative (V)

0.2g (0.31mmol) of Compound IV was dissolved in 50mL of methanol, and 3mL of 30% H was added₂O₂And 3 drops of concentrated hydrochloric acid, stirring the mixed solution in an ice bath for reacting for 3 hours, pouring the mixed solution into 50mL of ice water, and extracting with dichloromethane for 3 times with 30mL of dichloromethane each time after the ice is melted. The combined organic phases are washed 3 times with 30mL of water, 1 time with 30mL of saturated sodium bicarbonate solution and finally with 30mL of saturated sodium chloride solution. Dried over anhydrous sodium sulfate, and the solvent was distilled off. Separating and purifying by silica gel column chromatography, gradient eluting with petroleum ether/acetone (15:1-8:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound V0.11 g with yield of 60.3%.

M.p.296～299℃；¹H NMR(300MHz,CDCl₃):δ0.88(s,3H),0.90(d,J＝6.5 Hz,3H),0.93(d,J＝10.9Hz,3H),0.97(s,3H),1.13(s,3H),1.15(s,3H),1.27(s, 3H),1.32～1.90(m,15H),1.98(m,1H),2.10～2.17(m,2H),2.19(d,J＝14.9Hz,1H), 2.25(d,J＝10.3Hz,1H),3.02(d,J＝15.6Hz,1H),3.81(s,3H,OCH₃),5.33(t,J＝ 3.3Hz,1H),6.65(s,1H),10.41(s,1H)；IR(KBr,cm^-1):3254,3201,2925,2869, 1695,1632,1596,1491,1452,1380,1232,1114,1028,839,784；MS(ESI):m/z [M-H]^-:586.4；Anal.Calcd.for C₃₇H₄₉NO₅:C75.61；H 8.40；N 2.38；found:C 75.65； H 8.42；N 2.36.

Example 5

Synthesis of ursolic acid indoloquinone amide derivative (I-a)

30mg (0.05mmol) of Compound V was dissolved in 2mL of dichloromethane, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) were added to the reaction system, and the reaction mixture was stirred at room temperature for 30min, and then 4.8mg (0.08mmol) of ethylenediamine was added thereto. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, concentrating the filtrate under reduced pressure, dissolving the filtrate with acetonitrile, standing the filtrate at 4 ℃ overnight, further filtering out precipitate, concentrating under reduced pressure to remove the solvent, performing gradient elution with dichloromethane/methanol (30: 1-10:1), combining product components, concentrating under reduced pressure to remove the solvent to obtain a pure compound I-a,23.9mg, wherein the yield is 76%.

M.p.307～309℃；¹H NMR(500MHz,CDCl₃):δ0.82(s,3H),0.89(d,J＝7.2 Hz,3H),0.95(d,J＝7.3Hz,3H),0.98(s,3H),1.14(s,3H),1.22(s,3H),1.27(s,3H), 1.29～2.30(m,21H),2.32(brs,2H),2.97(t,J＝6.9Hz,2H),3.04(m,1H),3.69(m, 1H),3.82(s,3H,OCH₃),5.38(t,J＝3.3Hz,1H),5.67(s,1H),6.93(s,1H),9.38(s, 1H)；IR(KBr，cm^-1):3405,2964,2921,2852,1638,1589,1482,1440,1381,1232, 1102,1039,837,788；MS(ESI):m/z[M+H]⁺:630.4；Anal.Calcd.forC₃₉H₅₅N₃O₄:C 74.37；H 8.80；N 6.67；found:C 74.31；H 8.83；N 6.71.

Example 6

Synthesis of Ursolic acid indoloquinone amide derivative (I-b)

30mg (0.05mmol) of Compound V was dissolved in 2mL of methylene chloride, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) and added to the reaction system, and the reaction mixture was stirred at room temperature for 30min, and then 7.1mg (0.08mmol) of N, N-dimethylethylenediamine was added thereto. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, dissolving the filtrate in acetonitrile for overnight standing at 4 ℃ after vacuum concentration, further filtering out precipitate, removing the solvent by vacuum concentration, performing gradient elution by using dichloromethane/methanol (30: 1-10:1), combining product components, performing vacuum concentration, removing the solvent, and preparing a pure compound I-b, wherein the yield is 82%, and 27.0 mg.

M.p.:332～335℃；¹H NMR(500MHz,CDCl₃):δ0.83(s,3H),0.90(d,J＝7.1 Hz,3H),0.95(d,J＝7.2Hz,3H),0.98(s,3H),1.15(s,3H),1.20(s,3H),1.28(s,3H), 1.36～2.33(m,21H),2.42(s,6H),2.64(t,J＝12.5Hz,2H),3.06(d,J＝16.9Hz,1H), 3.27(m,1H),3.79(s,3H,OCH₃),5.37(t,J＝4.2Hz,1H),5.67(s,1H),6.93(s,1H), 9.32(s,1H)；IR(KBr，cm^-1): 3401,2973,1927,1865,1639,1587,1483,1443,1378,1236,1099,1020,800,732；MS(ESI):m/z[M+H]⁺:658.4；Anal.Calcd.forC₄₁H₅₉N₃O₄:C 74.85；H 9.04；N 6.39； found:C74.78；H 9.01；N 6.32.

Example 7

Synthesis of ursolic acid indoloquinone amide derivative (I-c)

30mg (0.05mmol) of Compound V was dissolved in 2mL of methylene chloride, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) and added to the reaction system, and the reaction mixture was stirred at room temperature for 30min, and then 9.3mg (0.08mmol) of N, N-diethylethylenediamine was added thereto. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, concentrating the filtrate under reduced pressure, dissolving the filtrate with acetonitrile, standing the filtrate at 4 ℃ overnight, further filtering out precipitate, concentrating under reduced pressure to remove the solvent, performing gradient elution with dichloromethane/methanol (30: 1-10:1), combining the product components, concentrating under reduced pressure to remove the solvent to obtain a pure compound I-b, wherein the yield is 73 percent.

M.p.:322～324℃；¹H NMR(500MHz,CDCl₃):δ0.84(s,3H),0.86(d,J＝7.0 Hz,3H),0.89(d,J＝3.1Hz,3H),0.95(s,3H),1.07(t,J＝6.4,6H),1.11(s,3H), 1.21(s,3H),1.29(s,3H),1.25～2.23(m,21H),2.60～2.65(m,6H),2.79(d,J＝11.3 Hz,1H),3.08(m,1H),3.81(s,3H,OCH₃),5.37(t,J＝3.3Hz,1H),5.64(s,1H), 6.94(s,1H),9.38(s,1H)； IR(KBr,cm^-1):3411,2965,2911,1636,1641,1591,1479,1452,1374,1236,1103,1023,802,736；MS(ESI):m/z[M+H]⁺:686.5；Anal.Calcd.for C₄₃H₆₃N₃O₄:C 75.29；H 9.26；N 6.13；found:C 75.23；H 9.21；N 6.18.

Example 8

Synthesis of Ursolic acid indoloquinone amide derivative (I-d)

30mg (0.05mmol) of Compound V was dissolved in 2mL of methylene chloride, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) and added to the reaction system, and the reaction mixture was stirred at room temperature for 30min, and 5.9mg (0.08mmol) of 1, 3-propanediamine was added thereto. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, concentrating the filtrate under reduced pressure, dissolving the filtrate with acetonitrile, standing the filtrate at 4 ℃ overnight, further filtering out precipitate, concentrating under reduced pressure to remove the solvent, performing gradient elution with dichloromethane/methanol (30: 1-10:1), combining product components, concentrating under reduced pressure to remove the solvent to obtain a pure compound I-d, wherein the yield is 76%, and the product is 24.5 mg.

M.p.307～309℃；¹H NMR(500MHz,CDCl₃):δ0.81(s,3H),0.87(d,J＝8.2 Hz,3H),0.94(d,J＝7.9Hz,3H),0.98(s,3H),1.14(s,3H),1.21(s,3H),1.25(s,3H), 1.29～1.98(m,23H),2.12(s,2H),2.16(t,J＝6.9Hz,2H),2.83(m,1H),3.04(m,1H), 3.87(s,3H,OCH₃),5.38(t,J＝3.3Hz,1H),5.65(s,1H),6.93(s,1H),9.36(s,1H)； IR(KBr，cm^-1)：3407,2963,2923,2851,1635,1587,1487,1444,1382,1228,1103, 1033,845,802；MS(ESI):m/z[M+H]⁺:644.4；Anal.Calcd.forC₄₀H₅₇N₃O₄:C 74.61；H 8.92；N 6.53；found:C 74.67；H 8.89；N6.57.

Example 9

Synthesis of ursolic acid indoloquinone amide derivative (I-e)

30mg (0.05mmol) of Compound V was dissolved in 2mL of dichloromethane, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) were added to the reaction system, and the reaction mixture was stirred at room temperature for 30min, and 8.2mg (0.08mmol) of 3-dimethylaminopropylamine was added thereto. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, concentrating the filtrate under reduced pressure, dissolving the filtrate with acetonitrile, standing the filtrate at 4 ℃ overnight, further filtering out precipitate, concentrating under reduced pressure to remove the solvent, performing gradient elution with dichloromethane/methanol (30: 1-10:1), combining product components, concentrating under reduced pressure to remove the solvent to obtain a pure compound I-e, wherein the yield is 86 percent, and the content of the product is 29.0 mg.

M.p.285～288℃；¹H NMR(300MHz,CDCl₃):δ0.84(s,3H),0.91(d,J＝6.3 Hz,3H),0.95(d,J＝6.3Hz,3H),0.97(s,3H),1.14(s,3H),1.21(s,3H),1.29(s,3H), 1.34～2.30(m,23H),2.32(s,6H),2.43(t,J＝6.9Hz,2H),2.75(d,J＝14.9Hz,1H), 3.09(m,1H),3.81(s,3H,OCH₃),5.37(t,J＝3.3Hz,1H),5.67(s,1H),6.94(s,1H), 9.38(s,1H)；IR(KBr,cm^-1):3305,2948,2925,2869,1695,1633,1526,1459,1381, 1285,1204,1050,796；MS(ESI):m/z[M+H]⁺:673.0；Anal.Calcd.for C₄₂H₆₁N₃O₄: C 75.07；H 9.15；N 6.25；found:C 75.03；H 9.18；N 6.27.

Example 10

Synthesis of ursolic acid indoloquinone amide derivative (I-f)

30mg (0.05mmol) of Compound V was dissolved in 2mL of dichloromethane, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) were added to the reaction system, and the reaction was stirred at room temperature for 30min, and then 10.4mg (0.08mmol) of 3-diethylaminopropylamine was added. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, concentrating the filtrate under reduced pressure, dissolving the filtrate with acetonitrile, standing the filtrate at 4 ℃ overnight, further filtering out precipitate, concentrating under reduced pressure to remove the solvent, performing gradient elution with dichloromethane/methanol (30: 1-10:1), combining product components, concentrating under reduced pressure to remove the solvent to obtain a pure compound I-f, wherein the yield is 81%, and the pure compound I-f is 28.4 mg.

M.p.335～337℃；¹H NMR(500MHz,CDCl₃):δ0.78(s,3H),0.86(d,J＝5.3Hz, 3H),0.94(d,J＝5.3Hz,3H),0.98(s,3H),1.06(t,J＝6.3Hz,6H),1.14(s,3H),1.19 (s,3H),1.24(s,3H),1.19～2.30(m,23H),2.42(t,J＝6.7Hz,2H),2.53(q,J＝6.5Hz, 4H),2.75(d,J＝14.3Hz,1H),3.08(m,1H),3.81(s,3H,OCH₃),5.36(t,J＝3.7Hz, 1H),5.65(s,1H),6.95(s,1H),9.67(s,1H)； IR(KBr,cm^-1):3407,2917,2920,2861,1639,1587,1490,1436,1378,1220,1095,1031,798, 736；MS(ESI):m/z[M+H]⁺:700.5；Anal.Calcd.for C₄₄H₆₅N₃O₄:C75.50；H 9.36；N 6.00；found:C 75.58；H 9.30；N 6.07.

Example 11

Synthesis of Ursolic acid indoloquinone amide derivative (I-g)

30mg (0.05mmol) of Compound V was dissolved in 2mL of methylene chloride, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) and added to the reaction system, and the reaction mixture was stirred at room temperature for 30min, and then 7.1mg (0.08mmol) of 1, 4-butanediamine was added thereto. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, concentrating the filtrate under reduced pressure, dissolving the filtrate with acetonitrile, standing the filtrate at 4 ℃ overnight, further filtering out precipitate, concentrating under reduced pressure to remove the solvent, performing gradient elution with dichloromethane/methanol (30: 1-10:1), combining product components, concentrating under reduced pressure to remove the solvent to obtain a pure compound I-g,23.7mg, wherein the yield is 72%.

M.p.343～345℃；¹H NMR(500MHz,CDCl₃):δ0.84(s,3H),0.89(d,J＝7.0 Hz,3H),0.95(d,J＝7.0Hz,3H),0.97(s,3H),1.14(s,3H),1.21(s,3H),1.29(s,3H), 1.76～2.50(m,25H),2.72(m,2H),2.93(m,2H),3.07(s,2H),3.89(s,3H,OCH₃), 5.38(t,J＝3.3Hz,1H),5.64(s,1H),6.93(s,1H),9.38(s,1H)；IR(KBr,cm^-1):3407, 2981,2927,2865,1631,1951,1487,1436,1370,1232,1031,802,748；MS(ESI):m/z [M+H]⁺:658.4；Anal.Calcd.forC₄₁H₅₉N₃O₄:C 74.85；H 9.04；N 6.39；found:C 74.92；H 9.08；N 6.32.

Example 12

Synthesis of ursolic acid indoloquinone amide derivative (I-h)

30mg (0.05mmol) of Compound V was dissolved in 2mL of dichloromethane, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) were added to the reaction system, and the reaction mixture was stirred at room temperature for 30min, and 9.3mg (0.08mmol) of 4-dimethylaminobutylamine was added thereto. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, dissolving the filtrate in acetonitrile for overnight standing at 4 ℃ after vacuum concentration, further filtering out precipitate, removing the solvent by vacuum concentration, performing gradient elution by using dichloromethane/methanol (30: 1-10:1), combining product components, performing vacuum concentration, removing the solvent, and preparing a pure compound I-h,27.8mg, wherein the yield is 81%.

M.p.343～345℃；¹H NMR(500MHz,CDCl₃):δ0.81(s,3H),0.86(d,J＝6.3 Hz,3H),0.91(d,J＝15.9Hz,3H),0.96(s,3H),1.13(s,3H),1.20(s,3H),1.25(s, 3H),1.40～2.49(m,25H),2.63(s,6H),2.86(t,J＝6.9Hz,2H),3.06(d,J＝16.4Hz, 1H),3.33(m,1H),3.80(s,3H,OCH₃),5.35(t,J＝3.3Hz,1H),5.63(s,1H),6.92(s, 1H),9.93(s,1H)；IR(KBr，cm^-1):3400,3973,2927,2861,1645,1587,1547,1440, 1374,1236,1111,1033,806,740；MS(ESI):m/z[M+H]⁺:686.5；Anal.Calcd.for C₄₃H₆₃N₃O₄:C 75.29；H 9.26；N 6.13；found:C75.35；H 9.30；N 6.08.

Example 13

Synthesis of ursolic acid indoloquinone amide derivative (I-I)

30mg (0.05mmol) of Compound V was dissolved in 2mL of dichloromethane, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) were added to the reaction system, and the reaction mixture was stirred at room temperature for 30min, and 11.5mg (0.08mmol) of 4-diethylaminobutylamine was added thereto. Stir at room temperature overnight. Filtering out precipitate after the reaction is finished, dissolving the filtrate in acetonitrile for overnight standing at 4 ℃ after vacuum concentration, further filtering out precipitate, removing the solvent by vacuum concentration, performing gradient elution by using dichloromethane/methanol (30: 1-10:1), combining product components, performing vacuum concentration, removing the solvent, and preparing a pure compound I-f,27.8mg with the yield of 78%.

M.p.349～351℃；¹H NMR(500MHz,CDCl₃):δ0.83(s,3H),0.88(d,J＝6.3 Hz,3H),0.94(d,J＝6.3Hz,3H),0.98(s,3H),1.06(t,J＝6.3Hz,6H),1.14(s,3H), 1.19(s,3H),1.24(s,3H),1.19～2.30(m,23H),2.42(t,J＝6.7Hz,2H),2.53(q,J＝ 6.5Hz,4H),2.75(m,2H),3.08(m,1H),3.66(m,1H),3.81(s,3H,OCH₃),5.36(t,J ＝3.7Hz,1H),5.65(s,1H),6.95(s,1H),9.67(s,1H)；IR(KBr，cm^-1):3408,2970, 2911,2865,1641,1573,1480,1448,1374,1220,1107,1027,802,724；MS(ESI):m/z [M+H]⁺:714.5；Anal.Calcd.for C₄₅H₆₇N₃O₄:C 75.69；H 9.46；N 5.88；found:C 75.60；H 9.51；N 5.85.

Example 14

Screening for antitumor Activity in vitro

The cell lines are selected as follows:

human liver cancer cells HepG2 and SMMC-7721; human normal liver cell line 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 of HepG2, SMMC-7721 and QSG-7701 cells by ursolic acid indoloquinone amide derivatives. The results of the antitumor activities of the compounds I-a to I-I are shown in Table 1:

TABLE 1 results of antitumor Activity of Compounds I-a to I-I

^aNT not tested

As shown in Table 1, the synthesized ursolic acid indoloquinone amide derivatives show different degrees of inhibition effects on two tumor cells (HepG2 and SMMC-7721), the compounds I-e and I-f have stronger cytotoxic activity on two liver cancer cells, and the compounds I-b, I-c, I-d, I-h and I-I also show moderate cytotoxic activityWhereas the antitumor activity of compounds I-a and I-g is relatively weak. IC of compounds I-e on HepG2 and SMMC-7721 cells, among others₅₀The values are respectively 6.38 +/-0.28 and 5.71 +/-0.37 mu M, the activity of the product is equivalent to that of positive control etoposide, and the IC of I-e on normal liver cells QSG-7701 is equivalent to that of I-e₅₀The value was 38.05 ± 1.02 μ M, indicating that the cytotoxicity of the compound on normal liver cells was significantly lower than on tumor cells. Preliminary structure-activity relationship analysis shows that the amide side chain structure of the series of compounds has certain influence on the antitumor activity. The three-carbon side chain derivatives (I-d, I-e and I-f) are more active than the two-carbon side chain and four-carbon side chain derivatives with respect to the alkyl chain length; for the amine group on the side chain, the activity of the derivative containing the dimethylamino group and the diethylamino group is better, while the activity of the derivative containing the amino group is weaker, for example, the activity of the compounds I-e and I-f is obviously stronger than that of the compounds I-d. The results show that the compounds have obvious inhibition effect on liver cancer cells and have the potential of developing anticancer 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 (10)

1. Ursolic acid indolyquinone amide derivatives I-a to I-I with structures shown in general formula (I) and pharmaceutically acceptable salts thereof:

2. the preparation method of ursolic acid indoloquinone amide derivatives I-a to I-I with the structure shown in the general formula (I) in claim 1, which is characterized by comprising 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) 3-oxoursolic acid and 3, 5-dimethoxy phenylhydrazine are subjected to Fishier indole synthesis reaction to obtain a 3, 5-dimethoxy indole derivative, which has a structure shown in a general formula III:

(3) the indole derivative, phosphorus oxychloride and N, N-dimethylformamide are subjected to Vilsmeier-Haack formylation reaction to obtain a 3, 5-dimethoxy-2-aldehyde indole derivative which has a structure shown in a general formula IV:

(4) the compound IV is subjected to oxidation reaction with hydrogen peroxide to obtain ursolic acid indoloquinone derivative with a structure shown in a general formula V:

(5) carrying out amidation reaction on the compound V and different fatty amines to obtain the ursolic acid indoloquinone amide derivative with the structure shown in the general formula VI:

3. the method for preparing ursolic acid indoloquinone amide derivatives I-a to I-I with the structure shown in the general formula (I) according to claim 2, wherein: in the step (1), 2g (4.6mmol) of ursolic acid and 250mL of acetone are added into a 500mL round-bottom flask, stirred and dissolved, then stirred and reacted in ice water for 15min, 1.87mL of Jones reagent is slowly dripped, the temperature is raised to room temperature, stirred and reacted for 5h, then 90mL of isopropanol is added, stirred and reacted for 30min, after the reaction is finished, the precipitate is filtered out, the filtrate is collected, the filtrate is decompressed and concentrated, and the light yellow green viscous solid obtained by the filtrate is recrystallized by methanol to obtain white needle-shaped crystals, namely 3-oxoursolic acid (II).

4. The method for preparing ursolic acid indoloquinone amide derivatives I-a to I-I with the structure shown in the general formula (I) according to claim 2, wherein: in the step (2), 0.33g (3mmol) of 3, 5-dimethoxyaniline is dissolved in 3mL of 20% hydrochloric acid in a single-neck round-bottom flask; weighing 0.28g (4mmol) of sodium nitrite, dissolving in 0.7mL of water, slowly dropping the sodium nitrite into a reaction flask under an ice bath condition, stirring for reaction for 1h under the ice bath condition, weighing 1.35g (6mmol) of stannous chloride, dissolving in 1.8mL of concentrated hydrochloric acid, slowly dropping into the flask after the last reaction under the ice bath condition, stirring for 2h at room temperature, and performing suction filtration after the reaction is completed to obtain a solid 3, 5-dimethoxy phenylhydrazine; the molar ratio of the 3, 5-dimethoxyaniline to the sodium nitrite to the stannous chloride is 3: 4: 6.

weighing 0.441g (1mmol) of 3-oxoursolic acid, dissolving in 10mL of absolute ethanol, adding the 3, 5-dimethoxyphenylhydrazine (3mmol) reacted in the previous step into the system, adding 0.5mL of concentrated hydrochloric acid, and refluxing at 85 ℃ for 3 h; after the reaction is finished, pouring the reactant into a beaker filled with 20mL of ice water, melting the ice, extracting, distilling to remove the solvent, separating and purifying by silica gel column chromatography, performing gradient elution by using petroleum ether/acetone (20:1-5:1), combining product components, concentrating under reduced pressure, and removing the solvent to obtain a purified compound III; the molar ratio of the 3-oxoursolic acid to the 3, 5-dimethoxyphenylhydrazine is 1: 3.

5. the method for preparing ursolic acid indoloquinone amide derivatives I-a to I-I with the structure shown in the general formula (I) according to claim 2, wherein: in step (3), 0.24g (1.6mmol) of POCl was added₃Dropwise adding into 0.2mL (2mmol) of DMF which is stirred and cooled by ice water for reaction for 0.5h to form a coordination complex, slowly adding 0.59g (1mmol) of compound III into the complex reaction solution, and stirring at 30 ℃ for reaction for 3h to generate an intermediateAfter the reaction, adding the intermediate reaction solution into stirred ice water for hydrolysis for 2 hours, melting the ice, extracting, and distilling to remove the solvent; separating and purifying by silica gel column chromatography, gradient eluting with petroleum ether/acetone (20:1-10:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound IV; the POCl₃DMF, compound III in a molar ratio of 1.6: 2: 1.

6. the method for preparing ursolic acid indoloquinone amide derivatives I-a to I-I with the structure shown in the general formula (I) according to claim 2, wherein: in step (4), 0.2g (0.31mmol) of Compound IV was dissolved in 50mL of methanol, and 3mL of 30% H was added₂O₂And 3 drops of concentrated hydrochloric acid, stirring the mixed solution in an ice bath for reaction for 3 hours, pouring the mixed solution into 50mL of ice water, melting the ice, extracting, and distilling to remove the solvent; separating and purifying by silica gel column chromatography, gradient eluting with petroleum ether/acetone (15:1-8:1), mixing the product components, concentrating under reduced pressure, and removing solvent to obtain pure compound V; the pure compound IV: 30% H₂O₂: the molar ratio of concentrated hydrochloric acid was 1:96.8: 15.6.

7. The method for preparing ursolic acid indoloquinone amide derivatives I-a to I-I with the structure shown in the general formula (I) according to claim 2, wherein: in step (5), 30mg (0.05mmol) of compound V was dissolved in 2mL of dichloromethane, HOBt (10mg, 0.07mmol) and DCC (13mg, 0.07mmol) were added to the reaction system, and after stirring at room temperature for reaction for 30min, 4.8mg (0.08mmol) of ethylenediamine was added; stirring at room temperature overnight, filtering out precipitate after reaction, dissolving the filtrate in acetonitrile at 4 ℃ after vacuum concentration, standing overnight, further filtering out precipitate, removing solvent by vacuum concentration, performing gradient elution with dichloromethane/methanol (30: 1-10:1), combining product components, performing vacuum concentration, removing solvent, and obtaining a purified compound I-a; the HOBt: DCC: pure compound V: the molar ratio of ethylenediamine was 7:7:5: 8.

8. The method for preparing ursolic acid indoloquinone amide derivatives I-a to I-I with the structure shown in the general formula (I) according to claim 6, wherein: in the step (5), the different aliphatic amines are ethylenediamine, N-dimethylethylenediamine, N-diethylethylenediamine, 1, 3-propylenediamine, 3-dimethylaminopropylamine, 3-diethylaminopropylamine, 1, 4-butylenediamine, 4-dimethylaminobutylamine, 4-diethylaminobutylamine;

in the step (5), when the ethylenediamine is used, the pure compound I-a is prepared; when the N, N-dimethyl ethylenediamine is used, a pure compound I-b is prepared; when the N, N-diethyl ethylenediamine is used, pure compounds I-c are prepared; when the 1, 3-propane diamine is used, obtaining pure compounds I-d; when the 3-dimethylaminopropylamine is present, obtaining the pure compound I-e; when the 3-diethylaminopropylamine is used, pure compounds I-f are prepared; when the 1, 4-butanediamine is used, preparing pure compound I-g; when the 4-dimethylamino butylamine is used, pure compounds I-h are prepared; when the 4-diethylaminobutylamine is used, the pure compound I-I is obtained.

9. The use of the ursolic acid indoloquinone amide derivative with the structure shown in formula I and its pharmaceutically acceptable salt in claim 1 in the preparation of medicaments for treating tumors.

10. Use according to claim 9, characterized in that: the tumor is human liver cancer cell HepG2 and SMMC-7721.