A tanshinone IIA C-1 position esterified derivative, its preparation method and application

Abstract

The present invention belongs to the field of medicinal chemistry, relates to tanshinone C-1 derivatives, and particularly refers to a tanshinone IIA C-1 esterified derivative, its preparation method and application. For the first time, through a thiadiazole heterocyclic intermediate, a series of combined products of tanshinone IIA and tanshinone aromatic acid compounds were designed and synthesized, and a total of 25 tanshinone IIA derivatives were obtained, among which 24 were new compounds. Through thin layer chromatography analysis and solubility property research, compared with tanshinone IIA, the polarity of the C-1 substituted esterified derivatives of tanshinone IIA all increased, and the hydrophilicity of most derivatives increased significantly. This application provides certain reference value for the synthesis of tanshinone IIA derivatives and the application of the combination principle in the structural modification of traditional Chinese medicines. Activity experiments show that the derivatives have the potential of certain prodrugs, providing new possibilities for the development of new cardiovascular drugs and the research of innovative tanshinone drugs.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical chemistry, relates to tanshinone 1-position derivatives, and particularly refers to a tanshinone IIA C-1 position esterification derivative, its preparation method and application. Background Art

[0002] Tanshinone IIA is an important lipophilic active substance in Salvia miltiorrhiza with definite curative effects on cardiovascular diseases. Pharmacological studies have shown that it mainly has the effects of anti-arrhythmia, protecting the myocardium, improving myocardial ischemia and hypoxia, and restoring blood circulation. Although the activity of tanshinone IIA is definite and significant, due to its strong lipophilicity and poor solubility in water, there are not many monomer applications in clinical practice. So far, there is only the sodium tanshinone IIA sulfonate preparation for injection. Although sodium tanshinone IIA sulfonate improves the polarity of tanshinone IIA, increases its water solubility and thus enhances the activity, its short half-life and fast metabolism are its significant disadvantages, and it has irritation. Therefore, how to synthesize better tanshinone IIA derivatives to improve their polarity, increase bioavailability and enhance activity is one of the current research focuses and difficulties in Salvia miltiorrhiza research: because the structure of tanshinone IIA is unique, its strong lipophilicity, the active site is thermally unstable, and it is difficult to introduce other groups on the parent nucleus and other factors lead to the difficulty in synthesizing its derivatives. Patent 201610190543.3 discloses the application of a tanshinone IIA derivative in the preparation of an endothelial cell protection drug. By substituting the 14th position of the main nucleus of tanshinone IIA, it is found that the substitution at this position alleviates the oxidative damage pathway of vascular endothelial cells; the research group also carried out amide substitution at the 15th position and esterification substitution at the 17th position of tanshinone IIA in the early stage to improve the utilization rate of Salvia miltiorrhiza and develop new drugs. Furthermore, it is found that there are significant differences in the pharmacological properties of the drugs prepared with different substituents and different substitution sites. In order to further develop tanshinone IIA derivatives that can improve myocardial cell viability, the research group has carried out long-term exploration. Summary of the Invention

[0003] To solve the above technical problems, the present invention provides a tanshinone IIA C-1 position esterification derivative, its preparation method and application.

[0004] The technical solution of the present invention is realized as follows:

[0005] A tanshinone IIA C-1 position esterification derivative has the structural formula of formula I or formula II as follows:

[0006]

[0007] Formula I or formula II;

[0008]

[0009] Among them, X is arbitrarily selected from any one of halogen and nitrate group-substituted groups;

[0010] R is arbitrarily selected from phenyl, benzyl, phenethyl and phenyl monosubstituted by methoxy, nitro, methyl, halogen group.

[0011] The preparation method of the above-mentioned tanshinone IIA C-1 position esterification derivative comprises the following steps:

[0012] The first step:

[0013] Weigh a certain amount of tanshinone IIA and dissolve it in chlorobenzene, add 1.2 - 2 equivalents of 2,2,6,6-tetramethylpiperidine 1-oxide (TEMPO) and 1.2 - 2 equivalents of haloacetic acid, stir, heat in an oil bath (120 - 130 °C), react for (3 - 6) hours, vacuum concentrate to obtain a solid mixture, separate by silica gel chromatography column to obtain compound 1, namely tanshinone-1-haloacetate.

[0014] The second step:

[0015] At room temperature, dissolve compound 1, 1 - 1.2 equivalents of 2-amino-5-mercapto-1,3,4-thiadiazole, 1.2 - 2 equivalents of potassium iodide, and 1.2 - 2 equivalents of potassium carbonate in organic solvent I, stir and react at 25 - 40 °C for 1 - 2 hours, vacuum concentrate to a solid, dissolve with organic solvent II, extract with water, collect the organic phase, vacuum concentrate, and separate by silica gel chromatography column to obtain compound 2.

[0016] The third step:

[0017] Dissolve compound 2 in organic solvent III, add 1 - 2 equivalents of a series of benzoyl chlorides and 1 - 2 equivalents of triethylamine, stir and react at 25 - 40 °C for 1 - 4 hours, add an appropriate amount of dilute hydrochloric acid (25%) for washing, extract with organic solvent IV, add an appropriate amount of saturated brine for washing, collect the organic phase, wash successively with an appropriate amount of Na 2 CO 3 solution for 1 - 2 times, wash with water for 1 - 2 times, dry with anhydrous magnesium sulfate or anhydrous sodium sulfate, vacuum concentrate to a solid, and separate by silica gel chromatography column to obtain compound 3.

[0018] The above-mentioned haloacetic acid is chloroacetic acid or bromoacetic acid.

[0019] The above-mentioned organic solvent I is any one of acetonitrile, tetrahydrofuran, 1,4-dioxane, pyridine or DMF, and the organic solvent II is ethyl acetate or dichloromethane

[0020] The above-mentioned derivatives of benzoyl chloride are any one of benzoyl chloride, phenylacetyl chloride, phenylpropionyl chloride, 2-methoxybenzoyl chloride, 3-methoxybenzoyl chloride, 4-methoxybenzoyl chloride, 2-nitrobenzoyl chloride, 3-nitrobenzoyl chloride, 4-nitrobenzoyl chloride, 2-methylbenzoyl chloride, 3-methylbenzoyl chloride, 4-methylbenzoyl chloride, 2-fluorobenzoyl chloride, 3-fluorobenzoyl chloride, 4-fluorobenzoyl chloride, 2-chlorobenzoyl chloride, 3-chlorobenzoyl chloride, 4-chlorobenzoyl chloride, 2-bromobenzoyl chloride, 3-bromobenzoyl chloride or 4-bromobenzoyl chloride.

[0021] The above-mentioned organic solvent III is dichloromethane, acetonitrile or chloroform; the organic solvent IV is dichloromethane, chloroform or ethyl acetate.

[0022] Application of the above-mentioned tanshinone IIA C-1 position esterification derivative in preparing a new drug for improving myocardial cell viability.

[0023] The present invention has the following beneficial effects:

[0024] 1. For the first time, a series of combined products of tanshinone IIA and salvia aromatic acid compounds were designed and synthesized through a thiadiazole heterocyclic intermediate, and a total of 25 tanshinone IIA derivatives were obtained, among which 24 were new compounds. Through thin-layer chromatography analysis and solubility property research, compared with tanshinone IIA, the polarities of the esterification derivatives substituted at the C-1 position of tanshinone IIA C are all increased, and the hydrophilicity of most derivatives is significantly increased.

[0025] 2. The present application established an H9c2 myocardial cell H 2 O 2 oxidative stress model to evaluate the cardioprotective activity of tanshinone IIA derivatives. The experimental results show that compared with myocardial cells induced by H 2 O 2 to induce oxidative stress injury (relative myocardial cell viability is 55%), when tanshinone IIA acts on damaged cells at a concentration of 1 μM, the relative myocardial cell viability is about 77.8%; when derivatives T9, T10, and T16 act on damaged cells at a concentration of 1 μM, the relative myocardial cell viabilities are about 80.1%, 84.5%, and 82.2% respectively. Compared with tanshinone IIA, the cardioprotective activities of the three derivatives are all significantly improved, and they have better cardioprotective effects.

[0026] 3. Through the study on the regulation of the contraction and relaxation of isolated rat thoracic aortic vascular rings, the present application evaluated the relaxation effect of tanshinone IIA derivatives on the isolated rat thoracic aortic vascular rings pre-contracted with 1×10 -6 mol / L norepinephrine. The experimental results show that compared with tanshinone IIA, compounds T7, T13, T15, and T19 at a concentration of 2×10 -5When the concentration is [[mol / L]], it has a significant vasodilatory effect on the contraction of the rat thoracic aortic vascular ring induced by norepinephrine bitartrate. Among them, the vasodilatory amplitude of derivative T15 is 42.14 ± 0.025, and that of tanshinone IIA is 22.02 ± 0.0076. The vasodilation of the derivative is significantly improved.

[0027] 4. This application provides certain reference value for the synthesis of tanshinone IIA derivatives and the application of the principle of combination in the structural modification of traditional Chinese medicine. The activity experiment shows that the derivative has the potential of a certain prodrug, providing new possibilities for the development of new cardiovascular drugs and the research of innovative drugs from Salvia miltiorrhiza. Brief Description of the Drawings

[0028] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, the following drawings are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can be obtained based on these drawings.

[0029] Figure 1 It is the synthesis route of the C-1 esterified derivative of tanshinone IIA of this application.

[0030] Figure 2 It is the diagram of the vasodilatory change of sodium nitroprusside on the contraction of the vascular ring induced by norepinephrine. Detailed Embodiments

[0031] The following will combine the embodiments of the present invention to clearly and completely describe the technical solutions of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, rather than all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts belong to the scope of protection of the present invention.

[0032] The present invention uses tanshinone IIA isolated from the traditional Chinese medicine Salvia miltiorrhiza as the starting material to synthesize the tanshinone derivatives shown by the general formula. Specifically, tanshinone IIA is the starting material for preparing this kind of compound, and tanshinone IIA can be isolated from the ethanol extract of the traditional Chinese medicine Salvia miltiorrhiza.

[0033] The structural formula of tanshinone IIA is:

[0034]

[0035] The preparation of tanshinone IIA (compound DIIA) uses the following method:

[0036] The salvia miltiorrhiza extract was dissolved in 75% ethanol, extracted with petroleum ether: ethyl acetate = 3:1, and then separated by column chromatography repeatedly to obtain tanshinone IIA. (DIIA, petroleum ether: ethyl acetate = 8:1, Rf = 0.4;)

[0037] The tanshinone compounds represented by the general formula (I) can be obtained from tanshinone IIA through 1 - 3 steps of reaction according to the reaction process in Example 1 (see the reaction flow chart, Figure 1 ).

[0038] Example 1

[0039] Preparation method of compound T1:

[0040]

[0041] To a solution of compound DIIA (295 mg) in chlorobenzene (10 mL), TEMPO (315 mg) and chloroacetic acid (190 mg) were added successively. After addition, the gas was evacuated (N 2 ), and then the reaction was carried out at room temperature for 3 hours. After the reaction was completed, it was concentrated to a solid, and column chromatography (petroleum ether: ethyl acetate = 6:1) was carried out to obtain compound T1 (274 mg, 71%).

[0042] Compound T1: Red needle - shaped crystals. 1 H NMR (400 MHz, Chloroform - d) δ7.75 (dd, 2H), 7.26 (d, J = 1.4 Hz, 1H), 6.63–6.39 (m, 1H), 4.22–3.79 (dd, J = 14.4 Hz, 2H), 2.26 (d, J = 1.4 Hz, 3H), 2.26–2.21 (m, 1H), 2.07–1.86 (m, 2H), 1.65–1.53 (m, 1H), 1.41 (s, 3H), 1.29 (s, 3H). 13 C NMR (101 MHz, Chloroform - d) δ182.87, 174.82, 166.28, 160.95, 150.70, 141.73, 136.79, 134.31, 128.53, 126.90, 123.32, 121.42, 120.30, 69.28, 40.97, 34.77, 32.07, 31.51, 30.97, 24.51, 8.77. HRMS:C 21 H 20 ClO 5 for[M + H] + , calculated 387.09993, found 388.25336. m.p. 197~198℃.

[0043] Example 2

[0044] Preparation method of compound T2:

[0045]

[0046] To a solution of compound DIIA (295 mg) in bromobenzene (10 mL), TEMPO (315 mg) and chloroacetic acid (190 mg) were added successively. After addition, the gas was evacuated (N 2 ), and then the reaction was carried out at room temperature for 3 hours. After the reaction was completed, it was concentrated to a solid, and column chromatography (petroleum ether: ethyl acetate = 6:1) was carried out to obtain compound T2 (274 mg, 71%).

[0047] Compound T2: Red needle-like crystals. 1 H NMR (600 MHz, Chloroform-d) δ 7.80–7.67 (m, 2H), 6.51 (t, J = 3.5 Hz, 1H), 3.85–3.76 (dd, J = 12.0 Hz, 2H), 2.26 (d, J = 1.3 Hz, 3H), 2.01–1.91 (dd, J = 8.0 Hz, 2H), 1.58 (t, J = 7.8 Hz, 2H), 1.41 (s, 3H), 1.29 (s, 3H). 13 C NMR (151 MHz, Chloroform-d) δ 182.81, 174.82, 166.09, 160.97, 150.74, 141.71, 136.83, 134.29, 128.52, 126.90, 123.29, 121.42, 120.30, 69.10, 34.77, 32.07, 31.52, 30.98, 26.20, 24.33, 8.76. HRMS: C 21 H 20 BrO 5 for [M+H] + , calculated 431.04941, found 431.05142. m.p. 186~187 °C.

[0048] Example 3

[0049] Preparation of compound T3:

[0050]

[0051] To a solution of compound T1 (90 mg) in THF (5 mL), 2-amino-5-mercapto-1,3,4-thiadiazole (36 mg), potassium carbonate (50 mg), and potassium iodide (60 mg) were added successively. After addition, the gas was evacuated (N2 ), and then react for 1 hour at room temperature. After the reaction is completed, dilute with water (10 mL), extract three times with DCM (3 x 8 mL), then wash successively with saturated sodium chloride solution, dry over anhydrous sodium sulfate, concentrate and perform column chromatography (petroleum ether: ethyl acetate = 2:3) to obtain compound T3 (102 mg, 91%).

[0052] The structural analysis data of compound T3 are as follows:

[0053] Compound T3: Red solid. 1 H NMR (400 MHz, Chloroform-d) δ 7.75 (dd, 2H), δ 7.25 (d, J = 1.5 Hz, 1H), 6.55 (t, J = 3.4 Hz, 1H), 5.46 (s, 2H), 3.92–3.80 (dd, J = 15.6 Hz, 2H), 2.26 (d, J = 1.2 Hz, 3H), 2.04 (s, 1H), 1.95–1.87 (m, 2H), 1.60–1.51 (m, 1H), 1.40 (s, 3H), 1.27 (s, 3H). 13 C NMR (151 MHz, Chloroform-d) δ 182.70, 174.85, 167.22, 161.28, 150.89, 147.14, 141.86, 136.87, 134.50, 128.46, 126.65, 124.52, 124.03, 123.43, 121.37, 120.21, 119.11, 69.03, 36.23, 34.91, 31.94, 31.48, 30.99, 24.48, 8.86. HRMS: C 23 H 22 N 3 O 5 S 2 for [M+H] + , calculated 484.10009, found 484.09869. m.p. 178~179 °C.

[0054] Example 4

[0055] Preparation of compound T4:

[0056]

[0057] To a solution of compound T2 (42 mg) in acetonitrile (8 mL), subsequently add silver nitrate (35 mg). After addition, evacuate and replace the gas (N 2), and then react at 80 °C for 8 hours. After the reaction is completed, filter, dilute with water (10 mL), extract three times with DCM (3×8 mL), then wash with saturated sodium chloride solution in sequence, dry with anhydrous sodium sulfate, concentrate and recrystallize with ethanol to obtain compound T4 (18 mg, 44%).

[0058] The structural analysis data of compound T4 are as follows:

[0059] Compound T4: Red solid. 1 H NMR (600 MHz, Chloroform-d) δ 7.80–7.68 (dd, J = 5.2, 5.6 Hz, 2H), 6.54 (t, J = 3.6 Hz, 1H), 4.87 (dd, J = 11.6, 11.2 Hz, 2H), 2.26 (s, 3H), 2.05–1.97 (m, 1H), 1.85 (td, J = 13.7, 2.9 Hz, 1H), 1.59 (dt, J = 14.1, 3.9 Hz, 1H), 1.44 (t, J = 7.3 Hz, 1H), 1.40 (s, 3H), 1.29 (s, 3H). 13 C NMR (151 MHz, Chloroform-d) δ 182.96, 174.78, 164.76, 160.89, 150.69, 141.78, 136.30, 134.39, 128.58, 126.84, 123.46, 121.44, 120.34, 69.38, 67.31, 34.71, 31.98, 31.54, 30.94, 24.65, 8.74. HRMS: C 21 H 20 NO 8 for [M+H] + , calculated 414.11889, found 414.12048. m.p. 165~166 °C.

[0060] Example 5

[0061] Preparation of compound T5:

[0062]

[0063] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), add triethylamine (0.2 mmol) and benzoyl chloride (0.1 mmol) in sequence. After addition, evacuate and replace with gas (N 2), and then react at room temperature for 2 hours. After the reaction is completed, dilute with water (10 mL), extract three times with DCM (3×8 mL), then wash successively with saturated sodium chloride solution, dry over anhydrous sodium sulfate, concentrate and separate by column chromatography (petroleum ether:ethyl acetate = 2:1), and recrystallize with ethanol to obtain compound T5 (38 mg, 65%).

[0064] Compound T5: Red solid. 1 H NMR(400MHz,Chloroform-d)δ12.17(s,1H),8.15(d,J=7.2Hz,2H),7.70–7.46(m,5H),7.04(d,J=1.5Hz,1H),6.55–6.39(m,1H),4.19–3.93(dd,J=15.6Hz,2H),2.20(d,J=14.3Hz,1H),2.10(s,3H),1.98–1.74(m,2H),1.49(d,J=13.6Hz,1H),1.28(s,3H),1.20(s,3H). 13 C NMR(101MHz,Chloroform-d)δ182.33,174.47,166.81,165.09,161.30,160.72,158.66,150.44,141.62,137.76,136.58,134.04,133.39,130.63,128.81,128.62,128.43,128.32,126.53,123.15,121.27,120.16,68.91,35.38,34.62,31.95,31.34,30.94,24.45,8.58.HRMS:C 30 H 26 N 3 O 6 S 2 for[M+H] + ,calculated588.12630,found 588.12512.m.p.204~206℃.

[0065] Example 6

[0066] Preparation of compound T6:

[0067]

[0068] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), add triethylamine (0.2 mmol) and phenylacetyl chloride (0.1 mmol) successively. After addition, evacuate and refill with N 2), and then react at room temperature for 2 hours. After the reaction is completed, dilute with water (10 mL), extract three times with DCM (3×8 mL), then wash with saturated sodium chloride solution successively, dry over anhydrous sodium sulfate, concentrate and separate by column chromatography (petroleum ether:ethyl acetate = 2:1), and recrystallize with ethanol to obtain compound T6 (38 mg, 65%).

[0069] Compound T6: Red solid. 1 H NMR (400 MHz, Chloroform-d) δ 12.73 (s, 1H), 7.48 (dt, J = 24.0, 7.9 Hz, 4H), 7.34 (t, J = 7.3 Hz, 2H), 7.28 (d, J = 7.1 Hz, 1H), 7.13 (s, 1H), 6.40 (t, J = 3.5 Hz, 1H), 4.15 (d, 1H, J = 16.0 Hz), 3.93 (d, J = 15.8 Hz, 1H), 3.87 (s, 2H), 2.27–2.18 (m, 1H), 2.16 (s, 3H), 1.93 (t, J = 14.2 Hz, 1H), 1.81 (t, J = 13.3 Hz, 1H), 1.48 (d, J = 13.1 Hz, 1H), 1.31 (s, 3H), 1.20 (s, 3H). 13 C NMR (101 MHz, Chloroform-d) δ 182.43, 174.62, 169.31, 166.77, 160.81, 160.65, 158.75, 150.45, 141.72, 136.62, 134.15, 133.49, 129.58, 128.80, 127.6, 128.30, 127.50, 126.48, 124.45, 123.16, 121.24, 120.16, 68.99, 42.54, 35.69, 34.61, 31.97, 31.38, 30.90, 24.49, 8.76. HRMS: C 31 H 28 N 3 O 6 S 2 for [M+H] + , calculated 602.14195, found 602.14081. m.p. 195~197℃.

[0070] Example 7

[0071] Preparation of compound T7:

[0072]

[0073] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and phenylpropionyl chloride (0.1 mmol) were successively added. After addition, the mixture was evacuated and refilled with N 2 1

[0074] Compound T7: Red solid. 1 H NMR (600 MHz, Chloroform-d) δ 7.61 - 7.52 (m, 2H), 7.22 - 7.16 (m, 5H), 7.06 (s, 1H), 6.34 (s, 1H), 4.02 (d, J = 16.2 Hz, 1H), 3.85 (d, J = 16.8 Hz, 1H), 2.96 (t, J = 8.4 Hz, 2H), 2.84 (t, J = 8.4 Hz, 2H), 2.14 (d, J = 13.2 Hz, 1H), 2.06 (s, 3H), 1.90 (m 1H), 1.85 (m, 1H), 1.78 (m, 1H), 1.45 (m, 1H), 1.30 (s, 3H), 1.16 (s, 3H).

[0075] 13 C NMR (151 MHz, Chloroform-d) δ 182.32, 176.65, 174.43, 170.67, 166.74, 160.76, 158.58, 150.50, 141.84, 140.17, 136.65, 133.34, 134.21, 133.15, 128.47, 128.32, 126.32, 123.11, 121.22, 120.19, 118.54, 117.62, 68.94, 37.40, 35.56, 34.67, 31.99, 31.45, 30.97, 29.70, 24.49, 8.74. HRMS: C 32 H 30 N 3 O 6 S 2 for [M+H] + , calculated 616.15760, found 616.15649. m.p. 182~185 °C.

[0076] Example 8

[0077] Preparation of compound T8:

[0078]

[0079] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 2-methoxybenzoyl chloride (0.1 mmol) were successively added. After addition, the mixture was evacuated and refilled with N 2 ), and then reacted at room temperature for 2 hours. After the reaction was completed, it was diluted with water (10 mL), extracted with DCM three times (3×8 mL), then washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After concentration, it was separated by column chromatography (petroleum ether:ethyl acetate = 2:1) and recrystallized from ethanol to obtain compound T8 (38 mg, 65%).

[0080] Compound T8: Red solid. 1 H NMR (400 MHz, Chloroform-d) δ 11.14 (s, 1H), 8.23 (dd, J = 7.9, 1.9 Hz, 1H), 7.70 (s, 2H), 7.63–7.56 (m, 1H), 7.21–7.13 (m, 2H), 7.09 (d, J = 8.4 Hz, 1H), 6.48 (d, J = 3.9 Hz, 1H), 4.11 (s, 3H), 4.07 (d, J = 11.2 Hz, 1H), 3.98 (d, J = 15.6 Hz, 1H), 2.25 (d, J = 1.3 Hz, 3H), 1.97 (m, 1H), 1.94–1.84 (m, 2H), 1.59–1.52 (m, 1H), 1.39 (s, 3H), 1.26 (s, 3H).

[0081] 13 C NMR (151 MHz, Chloroform-d) δ 185.65, 179.76, 169.54, 166.78, 164.56, 162.32, 158.76, 155.32, 150.66, 147.78, 145.67, 142.34, 141.54, 135.09, 134.89, 133.98, 132.90, 125.33, 123.12, 122.67, 121.85, 117.89, 68.95, 56.44, 35.84, 34.73, 32.01, 31.50, 29.70, 24.48, 8.77. HRMS: C 31 H 28 N 3 O 7 S 2 for [M+H] +, calculated 618.13687, found 618.13574. m.p. 126~129℃.

[0082] Example 9

[0083] Preparation of Compound T9:

[0084]

[0085] To a solution of Compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 3-methoxybenzoyl chloride (0.1 mmol) were successively added. After addition, the mixture was evacuated and refilled with N 2 ), and then reacted at room temperature for 2 hours. After the reaction was completed, it was diluted with water (10 mL), extracted three times with DCM (3×8 mL), then washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (petroleum ether: ethyl acetate = 2:1), and recrystallized from ethanol to obtain Compound T9 (38 mg, 65%).

[0086] Compound T9: Red solid. 1 H NMR (400 MHz, Chloroform-d) δ 11.91 (s, 1H), 7.75 (d, J = 7.7 Hz, 1H), 7.65–7.55 (m, 3H), 7.43 (t, J = 7.8 Hz, 1H), 7.10 (d, J = 8.1 Hz, 2H), 6.48 (s, 1H), 4.17–3.96 (dd, J = 15.6 Hz, 2H), 3.85 (s, 3H), 2.19 (d, J = 16.3 Hz, 1H), 2.12 (s, 3H), 1.92 (m, 1H), 1.81 (m, 1H), 1.49 (m, 1H), 1.30 (s, 3H), 1.21 (s, 3H).

[0087] 13 C NMR (151 MHz, Chloroform-d) δ 182.32, 177.63, 167.83, 166.83, 165.44, 159.83, 155.4, 153.43, 150.94, 147.67, 145.56, 141.54, 136.78, 135.09, 132.90, 130.89, 129.87, 128.98, 126.78, 125.45, 123.12, 121.85, 68.95, 56.44, 35.84, 34.73, 32.01, 31.50, 29.70, 24.48, 8.77. HRMS: C 31 H 28 N 3 O7 S 2 for [M+H] + , calculated 618.13687, found 618.13574. m.p. 126~129℃. m.p. 190~193℃.

[0088] Example 10

[0089] Preparation of Compound T10:

[0090]

[0091] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 4 - methoxybenzoyl chloride (0.1 mmol) were added successively. After addition, the mixture was evacuated and refilled with N 2 ), and then reacted at room temperature for 2 hours. After the reaction was completed, it was diluted with water (10 mL), extracted with DCM three times (3×8 mL), then washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (petroleum ether: ethyl acetate = 2:1), and recrystallized from ethanol to obtain compound T10 (38 mg, 65%).

[0092] Compound T10: Red solid. 1 H NMR (600 MHz, Chloroform - d) δ8.19 (m, 1H), 7.72 (d, J = 7.2 Hz, 1H), 7.71–7.66 (dd, 2H), 7.66–7.53 (m, J = 5.6 Hz, 1H), 7.19–7.14 (m, 1H), 6.44 (d, J = 3.2 Hz, 1H), 4.41–4.30 (m, 2H), 4.14 (dd, J = 16.0, 11.6 Hz, 1H), 2.19 (s, 3H), 2.15 (s, 3H), 1.93 (d, J = 14.7 Hz, 1H), 1.87 (d, J = 13.4 Hz, 1H), 1.58–1.53 (m, 1H), 1.38 (s, 3H), 1.24 (s, 3H).

[0093] 1313C NMR(151MHz,Chloroform-d)δ182.38,174.60,166.68,164.84,160.83,159.59,158.77,158,05,157.60,154.78,150.62,141.81,136.67,134.29,130.75,128.33,126.51,123.20,121.28,120.18,114.20,113.71,68.97,55.56,42.17,35.54,34.70,31.99,29.70,24.48,8.71.HRMS:C 31 H 28 N 3 O 7 S 2 for[M+H] + ,calculated 618.13687,found618.13715.m.p.124~127℃.

[0094] Example 11

[0095] Preparation of Compound T11:

[0096]

[0097] To a solution of compound T3(0.1 mmol) in DCM(3 mL), triethylamine(0.2 mmol) and 2-nitrobenzoyl chloride(0.1 mmol) were added successively. After addition, the mixture was evacuated and filled with N 2 2), and then reacted at room temperature for 2 hours. After the reaction was completed, the mixture was diluted with water(10 mL), extracted with DCM three times(3×8 mL), then washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After concentration, it was separated by column chromatography(petroleum ether:ethyl acetate = 2:1) and recrystallized from ethanol to obtain compound T11(38 mg, 65%).

[0098] Compound T11: Red solid. 1 1H NMR(400MHz,Chloroform-d)δ8.13(m,1H),7.78–7.67(m,4H),7.60(s,1H),7.19(s,1H),6.43(s,1H),4.34(dd,J=12.4Hz,2H),2.15(d,J=5.4Hz,3H),1.94–1.83(m,2H),1.56(m,2H),1.40(s,3H),1.25(s,3H).

[0099] 1313C NMR (151 MHz, Chloroform-d) δ 183.43, 179.87, 167.66, 166.70, 157.88, 159.90, 157.88, 144.67, 143.22, 142.78, 141.84, 137.33, 136.27, 135.87, 133.23, 131.31, 126.33, 125.56, 124.54, 123.25, 121.22, 119.98, 69.04, 42.07, 34.72, 31.97, 31.50, 30.96, 23.73, 8.73. HRMS: C 30 H 25 N 4 O 8 S 2 for [M+H] + , calculated 633.11138, found 633.11078. m.p. 148~150 °C.

[0100] Example 12

[0101] Preparation of Compound T12:

[0102]

[0103] To a solution of Compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 3-nitrobenzoyl chloride (0.1 mmol) were added successively. After addition, the mixture was evacuated and refilled with N 2 2, and then reacted at room temperature for 2 hours. After the reaction was completed, it was diluted with water (10 mL), extracted with DCM three times (3×8 mL), then washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After concentration, it was separated by column chromatography (petroleum ether: ethyl acetate = 2:1) and recrystallized from ethanol to obtain Compound T12 (38 mg, 65%).

[0104] Compound T12: Red solid. 11H NMR (400 MHz, Chloroform-d) δ 8.91 (s, 1H), 8.67 (d, J = 7.7 Hz, 1H), 8.43 (d, J = 5.8 Hz, 1H), 7.84 (t, J = 7.9 Hz, 1H), 7.70–7.57 (m, 2H), 7.09 (s, 1H), 6.41 (s, 1H), 4.10–3.89 (dd, J = 16.4 Hz, 2H), 2.06 (s, 3H), 1.99–1.80 (m, 2H), 1.53 (d, J = 14.7 Hz, 1H), 1.44 (t, J = 7.2 Hz, 1H), 1.36 (s, 3H), 1.23 (s, 3H).

[0105] 13 13C NMR (151 MHz, Chloroform-d) δ 181.78, 176.98, 173.09, 169.98, 166.95, 160.98, 159.34, 156.78, 155.56, 153.43, 145.99, 137.11, 135.27, 133.45, 132.09, 130.99, 129.58, 127.30, 125.45, 126.46, 124.56, 117.89, 61.96, 42.07, 33.56, 32.77, 31.236, 29.70, 28.80, 8.27. HRMS: C 30 H 25 N 4 O 8 S 2 for [M+H] + , calculated 633.11138, found 633.10992. m.p. 145~146 °C.

[0106] Example 13

[0107] Preparation of Compound T13:

[0108]

[0109] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 4-nitrobenzoyl chloride (0.1 mmol) were added successively. After addition, the mixture was evacuated and filled with N 2 2, and then reacted at room temperature for 2 hours. After completion of the reaction, the mixture was diluted with water (10 mL), extracted with DCM three times (3×8 mL), washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (petroleum ether: ethyl acetate = 2:1), and recrystallized from ethanol to obtain compound T13 (38 mg, 65%).

[0110] Compound T13: Red solid. 1 H NMR(400MHz,Chloroform-d)δ8.44–8.28(m,4H),7.69–7.56(dd,J=8.4,8.0Hz,2H),7.08(s,1H),6.43(s,1H),4.15–3.95(dd,J=16.0Hz,2H),2.21(d,J=13.5Hz,1H),2.01(s,3H),1.97–1.80(m,2H),1.52(s,1H),1.34(s,3H),1.22(s,3H).

[0111] 13 C NMR(151MHz,Chloroform-d)δ182.71,174.86,169.89,167.17,161.19,159.89,154.64,153.45,152.64,150.84,141.78,138.98,137.46,136.89,134.36,128.44,126.71,124.48,123.97,123.33,121.35,120.21,68.95,36.32,34.53,31.51,30.97,30.21,24.17,8.78.HRMS:C 30 H 24 N 4 O 8 S 2 for[M+H] + ,calculated 633.11138,found633.10986.m.p.193~194℃.

[0112] Example 14

[0113] Preparation of Compound T14:

[0114]

[0115] To a solution of compound T3(0.1mmol) in DCM(3mL), triethylamine(0.2mmol) and 2-methylbenzoyl chloride(0.1mmol) were added successively. After addition, the mixture was evacuated and refilled with N 2 ), and then reacted at room temperature for 2 hours. After the reaction was completed, the reaction mixture was diluted with water(10mL), extracted with DCM three times(3ⅹ8mL), then washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography(petroleum ether:ethyl acetate = 2:1), and recrystallized from ethanol to obtain compound T14(38mg, 65%).

[0116] Compound T14: Red solid. 1 H NMR(600MHz,Chloroform-d)δ7.69(d,J=11.1Hz,3H),7.39(t,J=6.8Hz,1H),7.32(t,J=7.5Hz,1H),7.25(s,1H),7.15(d,J=1.5Hz,1H),6.50(t,J=3.5Hz,1H),4.03–3.94(dd,J=10.8,10.2Hz,2H),2.50(s,3H),2.21(t,J=5.5Hz,1H),2.16(d,J=1.3Hz,3H),1.95(tt,J=14.4,3.2Hz,1H),1.85(td,J=13.5,2.6Hz,1H),1.53(d,J=13.7Hz,1H),1.35(s,3H),1.24(d,J=4.2Hz,3H).

[0117] 13 C NMR(151MHz,Chloroform-d)δ183.39,175.52,170.09,169.76,161.76,159.33,155.58,150.41,143.09,137.73,136.35,133.19,130.02,129.66,128.95,127.77,126.36,123.53,120.47,119.68,118.30,117.45,69.31,35.57,34.77,32.02,30.99,29.75,24.51,20.81,8.80.HRMS:C 31 H 28 N 3 O 6 S 2 for[M+H] + ,calculated 602.14195,found602.14008.m.p.182~183℃.

[0118] Example 15

[0119] Preparation of compound T15:

[0120]

[0121] To a solution of compound T3(0.1mmol) in DCM(3mL), triethylamine(0.2mmol) and 3-methylbenzoyl chloride(0.1mmol) were added successively. After addition, the mixture was evacuated and refilled with N 2), and then react at room temperature for 2 hours. After the reaction is completed, dilute with water (10 mL), extract three times with DCM (3×8 mL), then wash with saturated sodium chloride solution successively, dry with anhydrous sodium sulfate, concentrate and separate by column chromatography (petroleum ether: ethyl acetate = 2:1), and recrystallize with ethanol to obtain compound T15 (38 mg, 65%).

[0122] Compound T15: Red solid. 1 H NMR(600MHz,Chloroform-d)δ7.96–7.92(m,1H),7.89(s,1H),7.63–7.54(m,J=5.6Hz 2H),7.46–7.36(m,2H),7.06(s,1H),6.47(t,J=3.4Hz,1H),4.00(dd,J=15.7Hz,2H),2.43(s,3H),2.23–2.17(m,1H),2.07(s,3H),1.92(tt,J=14.3,3.1Hz,1H),1.87–1.78(m,1H),1.49(d,J=13.8Hz,1H),1.29(s,3H),1.20(s,3H).

[0123] 13 C NMR(151MHz,Chloroform-d)δ182.39,174.52,166.78,165.25,161.51,160.75,158.82,150.41,141.59,138.81,136.51,134.20,134.02,130.61,128.91,128.74,128.33,126.55,125.67,123.18,121.30,120.15,68.90,35.72,34.63,33.45,31.96,30.91,24.46,21.27,8.59.HRMS:C 31 H 28 N 3 O 6 S 2 for[M+H] + ,calculated 602.14195,found602.14172.m.p.145~148℃.

[0124] Example 16

[0125] Preparation of compound T16:

[0126]

[0127] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 4-methylbenzoyl chloride (0.1 mmol) were successively added. After addition, the mixture was evacuated and refilled with N 2 ), and then reacted at room temperature for 2 hours. After the reaction was completed, the reaction mixture was diluted with water (10 mL) and extracted with DCM three times (3×8 mL). Then it was washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After concentration, it was separated by column chromatography (petroleum ether:ethyl acetate = 2:1) and recrystallized from ethanol to obtain compound T16 (38 mg, 65%).

[0128] Compound T16: Red solid. 1 H NMR (600 MHz, Chloroform-d) δ 8.01 (d, J = 7.9 Hz, 2H), 7.63–7.57 (dd, J = 5.6, 5.2 Hz, 2H), 7.30 (d, J = 7.8 Hz, 2H), 7.08 (s, 1H), 6.48 (t, J = 3.5 Hz, 1H), 4.10 (d, J = 15.8 Hz, 1H), 4.01 (d, J = 15.8 Hz, 1H), 2.39 (s, 3H), 2.10 (s, 3H), 2.04 (s, 1H), 1.94–1.89 (m, 1H), 1.82 (t, J = 14.9 Hz, 1H), 1.52–1.46 (m, 1H), 1.29 (s, 3H), 1.20 (s, 3H). 13 C NMR (151 MHz, Chloroform-d) δ 182.41, 174.53, 166.79, 164.83, 161.31, 160.77, 158.71, 150.48, 144.31, 141.60, 136.62, 134.05, 132.43, 130.32, 129.53, 128.53, 128.34, 127.87, 126.58, 123.15, 121.31, 120.18, 68.93, 35.49, 34.63, 31.95, 31.33, 30.94, 24.44, 21.65, 8.62. HRMS: C 31 H 28 N 3 O 6 S 2 for [M+H] + , calculated 602.14195, found 602.14093. m.p. 157~159 °C.

[0129] Example 17

[0130] Preparation of compound T17:

[0131]

[0132] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 2-fluorobenzoyl chloride (0.1 mmol) were successively added. After addition, the mixture was evacuated and refilled with N 2 ), and then reacted at room temperature for 2 hours. After the reaction was completed, it was diluted with water (10 mL), extracted with DCM three times (3×8 mL), then washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (petroleum ether:ethyl acetate = 2:1), and recrystallized from ethanol to obtain compound T17 (38 mg, 65%).

[0133] Compound T17: Red solid. 1 H NMR (400 MHz, Chloroform-d) δ 10.35 (s, 1H), 8.19–8.08 (m, 1H), 7.69 (s, 2H), 7.66–7.59 (m, 1H), 7.37 (t, J = 7.6 Hz, 1H), 7.27 (s, 1H), 7.18 (s, 1H), 6.48 (s, 1H), 4.17–3.89 (dd, J = 16.0, 15.6 Hz, 2H), 2.26 (s, 1H), 2.23 (s, 3H), 2.01–1.82 (m, 2H), 1.60–1.51 (m, 1H), 1.39 (s, 3H), 1.25 (s, 3H).

[0134] 13 C NMR (151 MHz, Chloroform-d) 13 C NMR (151 MHz, Chloroform-d) δ 182.64, 174.74, 166.85, 161.78, 160.96, 159.62, 150.71, 141.67, 136.82, 135.48, 134.31, 132.71, 128.42, 126.74, 125.28, 124.03, 124.00, 123.22, 121.40, 120.27, 118.64, 116.80, 69.03, 35.66, 34.75, 31.97, 31.52, 31.00, 24.48, 8.79. HRMS: C 30 H 25 FN 3 O 6 S 2 for [M+H] + , calculated 606.11688, found 606.11566. m.p. 135~138 °C.

[0135] Example 18

[0136] Preparation of Compound T18:

[0137]

[0138] To a solution of Compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 3-fluorobenzoyl chloride (0.1 mmol) were successively added. After addition, the mixture was evacuated and refilled with N 2 ), and then reacted at room temperature for 2 hours. After the reaction was completed, it was diluted with water (10 mL), extracted with DCM three times (3×8 mL), then washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (petroleum ether:ethyl acetate = 2:1), and recrystallized from ethanol to obtain Compound T18 (38 mg, 65%).

[0139] Compound T18: Red solid. 1 H NMR (600 MHz, Chloroform-d) δ 7.97 (d, J = 7.6 Hz, 1H), 7.85 (d, J = 9.3 Hz, 1H), 7.66–7.57 (dd, J = 5.2 Hz 2H), 7.52 (s, 2H), 7.10 (s, 1H), 6.46 (s, 1H), 4.11 (d, J = 15.9 Hz, 1H), 4.01 (d, J = 15.9 Hz, 1H), 2.19 (s, 1H), 2.11 (s, 3H), 1.92 (d, J = 14.0 Hz, 1H), 1.85 (d, J = 13.5 Hz, 1H), 1.51 (d, J = 13.0 Hz, 1H), 1.32 (s, 3H), 1.22 (s, 3H).

[0140] 13 C NMR (151 MHz, Chloroform-d) δ 182.28, 174.46, 166.86, 164.08, 163.55, 161.91, 160.79, 150.52, 141.78, 136.62, 134.17, 132.77, 130.73, 130.68, 128.32, 126.51, 124.64, 123.21, 121.30, 120.62, 120.19, 115.89, 68.98, 35.39, 34.68, 31.98, 31.43, 31.00, 24.50, 8.72. HRMS: C 30 H 25 FN 3 O 6 S 2 for [M+H]+ , calculated 606.11688, found 606.11749. m.p. 125 - 128 °C.

[0141] Example 19

[0142] Preparation of Compound T19:

[0143]

[0144] To a solution of Compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 4-fluorobenzoyl chloride (0.1 mmol) were added successively. After addition, the air was evacuated (N 2 ), and then the reaction was carried out at room temperature for 2 hours. After the reaction was completed, it was diluted with water (10 mL), extracted with DCM three times (3 x 8 mL), then washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (petroleum ether: ethyl acetate = 2:1), and recrystallized from ethanol to obtain Compound T19 (38 mg, 65%).

[0145] Compound T19: Red solid. 1 H NMR (400 MHz, Chloroform-d) δ 12.48 (s, 1H), 8.29 - 8.15 (m, 2H), 7.66 - 7.49 (dd, J = 8.0 Hz, 2H), 7.20 (t, J = 8.4 Hz, 2H), 7.02 (s, 1H), 6.47 (s, 1H), 4.28 - 3.87 (dd, J = 13.2, 16.0 Hz, 2H), 2.20 (d, J = 13.6 Hz, 1H), 2.02 (s, 2H), 1.96 - 1.79 (m, 2H), 1.56 - 1.46 (m, 1H), 1.30 (s, 3H), 1.21 (s, 3H). 13 C NMR (151 MHz, Chloroform-d) δ 182.29, 174.45, 166.86, 166.76, 165.17, 164.11, 161.80, 160.80, 150.55, 141.69, 136.56, 134.18, 131.59, 131.53, 128.33, 126.77, 126.51, 123.22, 121.29, 120.15, 116.13, 115.99, 69.09, 35.50, 34.69, 31.96, 31.36, 31.00, 24.50, 8.63. HRMS: C 30 H 25 F N 3 O 6 S 2for [M+H] + , calculated 606.11688, found 606.11505. m.p. 196~198℃.

[0146] Example 20

[0147] Preparation of Compound T20:

[0148]

[0149] To a solution of Compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 2-chlorobenzoyl chloride (0.1 mmol) were successively added. After addition, the air was evacuated (N 2 ), and then the reaction was carried out at room temperature for 2 hours. After the reaction was completed, water (10 mL) was added for dilution, and the mixture was extracted with DCM three times (3×8 mL). Then it was washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After concentration, it was separated by column chromatography (petroleum ether:ethyl acetate = 2:1) and recrystallized from ethanol to obtain Compound T20 (38 mg, 65%).

[0150] Compound T20: Red solid. 1 H NMR (600 MHz, Chloroform-d) δ 7.79 (d, J = 7.5 Hz, 1H), 7.70 (s, 2H), 7.54–7.40 (m, 3H), 7.16 (s, 1H), 6.48 (t, J = 3.5 Hz, 1H), 4.08–4.00 (d, J = 12.8 Hz, 1H), 3.99–3.89 (d, J = 18.4 Hz, 1H), 2.23 (d, J = 17.9 Hz, 1H), 2.18–2.12 (m, 3H), 2.01–1.91 (m, 1H), 1.85 (t, J = 13.5 Hz, 1H), 1.54 (d, J = 17.3 Hz, 1H), 1.38 (s, 3H), 1.25 (s, 3H). 13 C NMR (151 MHz, Chloroform-d) δ 182.57, 174.68, 166.72, 164.05, 160.88, 150.64, 141.84, 141.70, 136.75, 136.67, 134.33, 134.28, 132.87, 132.10, 131.60, 130.83, 130.74, 128.40, 128.34, 126.68, 123.26, 120.24, 69.00, 42.18, 35.44, 34.70, 32.00, 30.96, 24.48, 8.70. HRMS: C 30 H 25 ClN3 O 6 S 2 for [M+H] + , calculated 622.08733, found 622.08612. m.p. 137~139℃.

[0151] Example 21

[0152] Preparation of Compound T21:

[0153]

[0154] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 3-chlorobenzoyl chloride (0.1 mmol) were added successively. After addition, the mixture was evacuated and refilled with N 2 ), and then reacted at room temperature for 2 hours. After the reaction was completed, it was diluted with water (10 mL), extracted with DCM three times (3×8 mL), then washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After concentration, it was separated by column chromatography (petroleum ether: ethyl acetate = 2:1) and recrystallized from ethanol to obtain compound T21 (38 mg, 65%).

[0155] Compound T21: Red solid. 1 H NMR (600 MHz, Chloroform-d) δ8.11–8.05 (m, 2H), 7.63 (d, J = 8.3 Hz, 1H), 7.58 (d, J = 8.2 Hz, 1H), 7.54 (d, J = 7.0 Hz, 1H), 7.49 (t, J = 7.8 Hz, 1H), 7.09 (s, 1H), 6.45 (d, J = 3.6 Hz, 1H), 4.12–3.95 (dd, J = 10.4, 10.8 Hz, 2H), 2.22 (dd, J = 18.5, 5.8 Hz, 1H), 2.09 (s, 3H), 1.96–1.80 (m, 2H), 1.51 (d, J = 10.0 Hz, 1H), 1.33 (s, 3H), 1.22 (s, 3H).1H), 1.33 (s, 3H), 1.22 (s, 3H). 1313C NMR (151 MHz, Chloroform-d) δ 182.31, 174.49, 166.72, 163.91, 161.45, 160.77, 159.20, 150.52, 141.70, 136.58, 135.01, 134.12, 133.35, 132.50, 130.23, 128.75, 128.30, 126.71, 126.54, 123.16, 121.29, 120.17, 68.93, 58.49, 35.61, 34.66, 31.96, 31.42, 24.45, 8.62. HRMS: C 30 H 25 ClN 3 O 6 S 2 for [M+H] + , calculated 622.08733, found 622.08936. m.p. 138~140 °C.

[0156] Example 22

[0157] Preparation of Compound T22:

[0158]

[0159] To a solution of Compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 4-chlorobenzoyl chloride (0.1 mmol) were added successively. After addition, the mixture was evacuated and filled with N 2 ), and then reacted at room temperature for 2 hours. After the reaction was completed, the reaction mixture was diluted with water (10 mL) and extracted with DCM three times (3×8 mL). Then it was washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After concentration, it was separated by column chromatography (petroleum ether: ethyl acetate = 2:1) and recrystallized from ethanol to obtain Compound T22 (38 mg, 65%).

[0160] Compound T22: Red solid. 11H NMR (600 MHz, Chloroform-d) δ 8.16–8.01 (m, 2H), 7.93–7.82 (m, 2H), 7.74–7.68 (dd, J = 5.6, 5.2 Hz, 2H), 7.19 (s, 1H), 6.44 (d, J = 3.4 Hz, 1H), 4.34 (d, J = 15.0 Hz, 1H), 4.33 (d, J = 15.0 Hz, 1H), 2.24–2.23 (m, 1H), 2.17 (s, 3H), 1.95 (dt, J = 14.0, 2.9 Hz, 2H), 1.58–1.55 (m, 1H), 1.41 (s, 3H), 1.25 (s, 3H). 13 13C NMR (151 MHz, Chloroform-d) δ 182.22, 174.41, 170.36, 166.81, 164.28, 161.66, 160.77, 159.04, 150.55, 141.68, 140.01, 139.90, 136.59, 134.16, 131.51, 130.29, 129.12, 128.30, 126.50, 123.19, 121.29, 120.15, 69.06, 35.47, 34.69, 31.96, 31.01, 29.75, 24.50, 8.64. HRMS: C 30 H 25 ClN 3 O 6 S 2 for [M+H] + , calculated 622.08733, found 622.08594. m.p. 136~137 °C.

[0161] Example 23

[0162] Preparation of Compound T23:

[0163]

[0164] To a solution of Compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 2-bromobenzoyl chloride (0.1 mmol) were added successively. After addition, the mixture was evacuated and refilled with N 2 2, and then reacted at room temperature for 2 hours. After completion of the reaction, the reaction mixture was diluted with water (10 mL), extracted with DCM three times (3 × 8 mL), washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (petroleum ether: ethyl acetate = 2:1), and recrystallized from ethanol to obtain Compound T23 (38 mg, 65%).

[0165] Compound T23: Red solid. 1 H NMR (600 MHz, Chloroform-d) δ 7.98–7.78 (m, 3H), 7.63–7.52 (dd, J=6.2 Hz, 2H), 7.34 (m, 1H), 7.16 (s, 1H), 6.45 (t, J=3.4 Hz, 1H), 4.32 (d, J=12.6 Hz, 1H), 4.33 (s, J=12.0 Hz, 1H), 2.25–2.23 (m, 1H), 2.15 (s, 3H), 1.98 (m, 2H), 1.58–1.54 (m, 1H), 1.37 (s, 3H), 1.22 (s, 3H). 13 C NMR (151 MHz, Chloroform-d) δ 184.47, 175.34, 166.75, 164.73, 162.86, 158.83, 149.58, 141.09, 139.22, 138.69, 137.08, 136.76, 135.34, 133.67, 131.09, 130.34, 129.05, 126.33, 122.01, 121.19, 119.28, 117.76, 62.07, 39.67, 35.66, 33.09, 32.98, 29.78, 29.09, 24.55, 8.47. HRMS: C 30 H 25 ClN 3 O 6 S 2 for [M+H] + , calculated 666.03534, found 666.03552. m.p. 136~137 °C.

[0166] Example 24

[0167] Preparation of Compound T24:

[0168]

[0169] To a solution of compound T3 (0.1 mmol) in DCM (3 mL), triethylamine (0.2 mmol) and 3-bromobenzoyl chloride (0.1 mmol) were added successively. After addition, the air was evacuated (N 2 ), and then the reaction was carried out at room temperature for 2 hours. After the reaction was completed, it was diluted with water (10 mL), extracted with DCM three times (3×8 mL), then washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography (petroleum ether: ethyl acetate = 2:1), and recrystallized from ethanol to obtain compound T24 (38 mg, 65%).

[0170] Compound T24: Red solid. 1 H NMR(600MHz,Chloroform-d)δ8.32(s,1H),7.78–7.70(m,3H),7.46(t,J=7.7Hz,1H),7.37(t,J=7.8Hz,1H),7.25(s,1H),6.43(d,J=3.2Hz,1H),4.20–3.91(dd,J=12.0,16.2Hz,2H),2.25(d,J=7.4Hz,3H),2.13(s,1H),1.76–1.61(m,2H),1.58–1.52(m,1H),1.33(s,3H),1.25(s,3H).

[0171] 13 C NMR(151MHz,Chloroform-d)δ188.91,178.82,175.41,172.33,169.87,159.64,155.43,150.05,141.73,136.41,134.16,133.26,131.91,131.53,130.51,130.04,128.91,128.71,127.16,123.12,122.75,122.55,68.95,46.83,34.99,31.97,30.98,28.78,24.44,8.68.HRMS:C 30 H 25 BrN 3 O 6 S 2 for[M+H] + ,calculated 666.03681,found666.03552.m.p.122~124℃.Example 25

[0172] Preparation of Compound T25:

[0173]

[0174] To a solution of compound T3(0.1 mmol) in DCM(3 mL), triethylamine(0.2 mmol) and 4-bromobenzoyl chloride(0.1 mmol) were added successively. After addition, the mixture was evacuated and refilled with N 2 ), and then reacted at room temperature for 2 hours. After completion of the reaction, the reaction mixture was diluted with water(10 mL), extracted with DCM three times(3×8 mL), washed successively with saturated sodium chloride solution, dried over anhydrous sodium sulfate, concentrated, and separated by column chromatography(petroleum ether: ethyl acetate = 2:1), and recrystallized from ethanol to give compound T25(38 mg, 65%).

[0175] Compound T25: Red solid.

[0176] 1 H NMR(600MHz, Chloroform-d) δ8.03(d, J=8.5Hz, 2H), 7.64(d, J=8.7Hz, 2H), 7.60(dd, J=20.2, 3.5Hz, 2H), 7.05(s, 1H), 6.47(d, J=3.4Hz, 1H), 4.09(d, J=15.9Hz, 1H), 4.01(d, J=16.0Hz, 1H), 2.23(s, 1H), 2.05(d, J=3.3Hz, 3H), 1.93(d, J=14.3Hz, 1H), 1.87(d, J=13.2Hz, 1H), 1.51(d, J=14.2Hz, 1H), 1.31(s, 3H), 1.21(s, 3H).

[0177] 13 C NMR(151MHz, Chloroform-d) δ182.30, 174.47, 166.72, 165.01, 164.38, 161.77, 160.81, 158.82, 150.59, 141.70, 136.62, 134.20, 132.19, 131.64, 130.38, 129.37, 128.78, 128.35, 126.55, 123.23, 121.33, 120.18, 69.14, 35.49, 34.71, 31.97, 31.39, 29.75, 24.50, 8.67. HRMS: C 30 H 25 BrN 3 O 6 S 2 for [M+H] + , calculated 666.03681, found 666.03522. m.p. 136~137℃.

[0178] Application Example 1 Evaluation of the Cardioprotective Activity of Tanshinone IIA Derivatives

[0179] I. Experimental Purpose

[0180] Using the CCK-8 reagent method to detect the toxicity of tanshinone IIA and its derivatives to H9c2 cardiomyocytes, establish an H2O2 oxidative stress injury model of H9c2 cardiomyocytes, detect the effect of tanshinone IIA derivatives on cardiomyocyte activity, and preliminarily evaluate their cardioprotective activity.

[0181] II. Experimental Methods

[0182] 1. Toxicity Detection of Cell Culture and Its Derivatives on H9c2 Cardiomyocytes

[0183] Culture H9c2 cardiomyocytes in DMEM medium containing fetal bovine serum (10%) in a CO 2 incubator (5% CO 2 , 95% relative humidity) at 37°C. Use CCK-8 reagent to detect the toxicity of tanshinone IIA and its derivatives on H9c2 cardiomyocytes at 1 μM, 5 μM, and 10 μM. The results are shown in Table 1.

[0184] Table 1 Toxicity of Tanshinone IIA on Normal Cardiomyocytes ( n = 3)

[0185]

[0186]

[0187] Continued Table

[0188]

[0189]

[0190] Continued Table

[0191]

[0192]

[0193] Note: Compared with the normal group, **P < 0.05, **P < 0.01

[0194] It can be seen from the data in Table 1 that the viability of most tanshinone IIA derivatives decreased significantly (P < 0.01) at concentrations of 5 μM and 10 μM, while at a concentration of 1 μM, all tanshinone IIA derivatives had no significant effect on cardiomyocyte viability (P > 0.05), proving that tanshinone IIA derivatives have no cytotoxicity at this concentration. Therefore, the active experimental concentration was selected as 1 μM.

[0195] 2. Establishment of H 2 O 2 Oxidative Stress Model of H9c2 Cardiomyocytes

[0196] Culture H9c2 cells in a culture dish. When the cell state is observed to be good under a microscope and the cell number reaches more than 80%, use a hemocytometer for cell counting, adjust the cell concentration to 5 × 10 3Inoculate each well with cells and seed them in a 96-well plate. After culturing for 24 h, discard the original culture medium and add 200 μL of H 2 O 2 culture medium with a concentration of 300 μM, and then culture in an incubator at 37 °C for 1 h. Observe the cell morphology. If cell shrinkage, decreased refractive index, and cell vacuolization occur, it indicates that the H9c2 cardiomyocyte H 2 O 2 oxidative stress model is successfully established.

[0197] 3. Cell viability assay

[0198] After culturing the cells in a 96-well plate for 24 h, establish the H 2 O 2 oxidative stress model of cardiomyocytes. After 1 h, add the sample solution of tanshinone IIA and its derivatives, and then culture for another 24 h. Subsequently, discard the original culture medium, add 200 μL of CCK-8 solution to each well, shake well, incubate in an incubator at 37 °C for 4 h, detect the absorbance of each well at 450 nm, and calculate the relative cardiomyocyte viability according to the ratio of the absorbance of the administered cardiomyocytes to that of the normal cardiomyocytes.

[0199] 4. Experimental grouping

[0200] (1) Blank group: H9c2 cardiomyocytes cultured normally.

[0201] (2) Model group: Treat the normally cultured H9c2 cardiomyocytes with 200 μL of 300 μM H 2 O 2 solution.

[0202] (3) Tanshinone IIA group: After modeling for 1 h, add 1 μM of complete culture medium containing tanshinone IIA when administering the drug.

[0203] (4) Sample group: After modeling for 1 h, add 1 μM of complete culture medium containing each tanshinone IIA derivative when administering the drug.

[0204] III. Experimental results

[0205] Use Graphpad Prism 8 software for data analysis. One-way ANOVA is used for multiple groups, and the t-test is used for comparison between two groups. P < 0.05 indicates that the experimental data have statistical differences.

[0206] Table 1 Toxicity of tanshinone IIA to normal cardiomyocytes ( n = 3)

[0207]

[0208] Note: Compared with the normal group, **P < 0.01; compared with the model group,# P < 0.05, ## P < 0.01

[0209] As can be seen from the data shown in Table 2, compared with normal cells, the relative viability of cardiomyocytes in the model group decreased significantly, about 55.3% of the blank group (P < 0.01), indicating that the 2 O 2 oxidative stress model was successfully established. Compared with the model group, when the concentration of tanshinone IIA was 1 μM, the viability of damaged cells was improved, and the relative viability of cardiomyocytes was about 77.8% of the blank group (P < 0.05), indicating that tanshinone IIA itself has a certain protective effect on cardiomyocytes. Correspondingly, except for compounds T3, T5, T8, and T4, the relative viability of the remaining tanshinone IIA at a concentration of 1 μM was above 55% (P < 0.05), indicating that these compounds can all improve the viability of damaged cardiomyocytes. Among them, the relative viability of compound T9 at a concentration of 1 μM was about 80.1% (P < 0.01), the relative viability of compound T10 at a concentration of 1 μM was about 84.5% (P < 0.01), and the relative viability of T16 at a concentration of 1 μM was about 82.2% (P < 0.01). Compared with tanshinone IIA, the cardioprotective activities of the three compounds were significantly improved.

[0210] Application Example 2 Study on the Regulation of Vasodilation and Vasoconstriction of Tanshinone IIA Derivatives on Isolated Thoracic Aortic Rings of Rats

[0211] I. Experimental Purpose

[0212] Using the isolated vascular perfusion technique, the change in vascular ring tension was detected by a bioenergy signal acquisition system, and derivatives with significant vasodilatory activity on blood vessels were screened out using isolated thoracic aortic rings of rats pre - constricted with norepinephrine.

[0213] II. Experimental Method

[0214] 1. Preparation of Medicinal Solutions

[0215] The tanshinone IIA derivative samples were accurately weighed and dissolved in DMSO to prepare the concentrations required for the experiment. At the same time, norepinephrine bitartrate (NE) and sodium nitroprusside were also prepared into the concentrations required for the experiment. K - H solution (Krebs - Henseleit): 118 mmol / L sodium chloride, 4.7 mmol / L potassium chloride, 2.5 mmol / L calcium chloride, 1.2 mmol / L sodium dihydrogen phosphate, 1.2 mmol / L magnesium sulfate, 25 mmol / L sodium bicarbonate, 10 mmol / L glucose, pH 7.4, stored at 4 °C for later use.

[0216] 2. Preparation of Isolated Thoracic Aortic Rings of Rats

[0217] After the SD rats were decapitated and sacrificed, the chest cavity was quickly opened, and the thoracic aorta was removed and immediately placed in pre-oxygenated K-H solution at 4°C. The blood vessels were washed and the connective tissue and adipose tissue around the blood vessels were dissected, and then divided into blood vessel rings with a length of 3-5 mm. Subsequently, one end of the blood vessel ring was suspended in a 7 mL K-H solution bath maintained at (37.0 ± 0.5)°C and continuously bubbled with 95% O 2 and 5% CO 2 The other end was connected to a muscle tension transducer and transmitted into a computer through a multi-channel physiological signal acquisition and analysis system to record the contraction and relaxation states of the blood vessel rings. The blood vessel rings were equilibrated for 15 min in a tension-free state, then the basal tension was adjusted to 0.75 g and equilibrated for 20 min, and then the basal tension was adjusted to 1.5 g and equilibrated for 60 min. During this period, the K-H solution was changed every 15 minutes. SPF-grade male SD rats, weighing 200-300 g, were purchased from Liaoning Changsheng Biotechnology Co., Ltd., and the animal license number was: NO. SCXK (Liao) 2020-0001. During the experiment, the animals had free access to food and water.

[0218] 3. Detection of the integrity of the endothelial function of blood vessel rings

[0219] After the tension was stabilized, NE was added to the bath to make its concentration reach 1×10 -6 mol / L to stimulate the contraction of the blood vessel rings; after reaching the maximum contraction tension and stabilizing, sodium nitroprusside was added again, Figure 2 As shown in the figure of the relaxation change of the blood vessel ring contraction induced by norepinephrine, to make its concentration reach 1×10 -5 mol / L, observe the change of the value, and calculate the relaxation amplitude of the blood vessel ring. The ratio of the difference between the maximum contraction value and the relaxation value after adding sodium nitroprusside to the difference between the maximum contraction value and the basal tension value > 70% indicates that the blood vessel ring has good activity and can proceed to the next experiment; if the relaxation amplitude < 70%, it indicates that the degree of endothelial injury of the blood vessel ring is relatively large, and the blood vessel ring should be discarded.

[0220] 4. Study on the effect of tanshinone IIA derivatives on the thoracic aorta blood vessel rings of rats

[0221] After screening the blood vessel rings in the previous step and selecting the blood vessel rings with relatively intact endothelium, the K-H solution in the bath was changed, rinsed and stimulated. After changing several times, the bath was finally made up to 7 mL. After the blood vessel rings were stabilized again, NE was added again to make its concentration reach 1×10 -6 mol / L to stimulate blood vessel contraction. After reaching the maximum contraction tension value and stabilizing, tanshinone IIA derivative solution was added respectively to make the concentration of the tanshinone IIA derivative solution reach 2×10 -5mol / L. After the tension value stabilizes, record the data and calculate the relaxation rate of the derivative. Denote the tension after norepinephrine-induced contraction equilibrium as T NE , and denote the tension after adding the derivative as T x .

[0222] The relaxation rate of the rat thoracic aorta = (T NE - T x ) / T NE

[0223] 5. Experimental grouping

[0224] (1) Tanshinone IIA derivative group: Study on the effect of tanshinone IIA derivative at 2×10 -5 mol / L on the contraction of the thoracic aortic vascular ring pre-stimulated by 1×10 -6 mol / L NE

[0225] (2) Blank control group: Add an equal amount of DMSO instead of tanshinone IIA derivative

[0226] (3) Tanshinone IIA group: Study on the effect of tanshinone IIA at 2×10 -5 mol / L on the contraction of the thoracic aortic vascular ring pre-stimulated by 1×10 -6 mol / L NE

[0227] III. Experimental results

[0228] Use SPSS 21.0 software to process and analyze the data. The experimental data are all expressed as the percentage (%) of the relative value of the change in the contraction amplitude of the vascular ring, mean ± standard deviation is the test result. One-way ANOVA is used for comparison among multiple groups, and t-test is used for analysis of the differences between groups

[0229] The relaxation effects of tanshinone IIA derivative and tanshinone IIA at a concentration of 2×10 -5 mol / L on norepinephrine-induced contraction of the rat thoracic aortic vascular ring are shown in Table 4.1

[0230] Table 4.1 Relaxation effects of tanshinone IIA and its derivative at a concentration of 2×10 -5 mol / L on NE-induced contraction of the vascular ring

[0231]

[0232]

[0233] As can be seen from the results listed in Table 4.1, compared with tanshinone IIA, the relaxation effects of compounds T7, T13, T15, and T19 on the contraction of rat thoracic aortic vascular rings induced by NE were enhanced at a concentration of 2×10 -5 mol / L.

[0234] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention shall be included within the protection scope of the present invention.

Claims

1. A tanshinone IIA C-1 esterified derivative having any one of the following structural formulas: 、 、 、 、 、 、 。 2. A method for preparing the tanshinone IIA C-1 esterified derivative according to claim 1, characterized in that, the steps are as follows: (1) Dissolve tanshinone IIA in chlorobenzene, then add 2,2,6,6-tetramethylpiperidine 1-oxyl and haloacetic acid, and react in an oil bath under stirring until complete, then concentrate under vacuum to obtain a solid mixture, and separate by silica gel column chromatography to obtain Compound 1; (2) Dissolve Compound 1, 2-amino-5-mercapto-1,3,4-thiadiazole, potassium iodide and potassium carbonate in organic solvent I, stir until the reaction is complete, concentrate to a solid under vacuum, dilute with water, then extract, wash and dry and concentrate with organic solvent II, and then perform column chromatography to obtain Compound 2; (3) Dissolve Compound 2 in organic solvent III, then add a derivative of benzoyl chloride and triethylamine, stir the reaction, wash with dilute hydrochloric acid, extract with organic solvent IV, then wash with saturated brine, collect the organic phase, wash, dry and concentrate, and then separate by silica gel column chromatography to obtain Compound 3; The derivative of benzoyl chloride is any one of propionyl chloride, 3-methoxybenzoyl chloride, 4-methoxybenzoyl chloride, 4-nitrobenzoyl chloride, 3-methylbenzoyl chloride, 4-methylbenzoyl chloride or 4-fluorobenzoyl chloride.

3. The method for preparing the tanshinone IIA C-1 esterified derivative according to claim 2, characterized in that: In the step (1), the molar ratio of tanshinone IIA, 2,2,6,6-tetramethylpiperidine 1-oxyl and haloacetic acid is 1:1.2 - 2:1.2 - 2; the temperature of the oil bath reaction is 120 - 130 °C and the time is 3 - 6 h.

4. The method for preparing the tanshinone IIA C-1 esterified derivative according to claim 3, characterized in that: The haloacetic acid is chloroacetic acid or bromoacetic acid.

5. The method for preparing the tanshinone IIA C-1 esterified derivative according to claim 2, characterized in that: In the step (2), the molar ratio of Compound 1, 2-amino-5-mercapto-1,3,4-thiadiazole, potassium iodide and potassium carbonate is 1:1.2 - 2:1.2 - 2:1.2 - 2, and the temperature of the stirring reaction is 25 - 40 °C and the time is 1 - 2 h.

6. The method for preparing the tanshinone IIA C-1 esterified derivative according to claim 5, characterized in that: The organic solvent I is any one of acetonitrile, tetrahydrofuran, 1,4-dioxane, pyridine or DMF, and the organic solvent II is ethyl acetate or dichloromethane.

7. The method for preparing the tanshinone IIA C-1 esterified derivative according to claim 2, characterized in that: In the step (3), the molar ratio of Compound 2, the derivative of benzoyl chloride and triethylamine is 1:1 - 2:1 - 2; the temperature of the stirring reaction is 25 - 40 °C and the time is 1 - 2 h.

8. The method for preparing the tanshinone IIA C-1 esterified derivative according to claim 7, characterized in that: The organic solvent III is dichloromethane, acetonitrile or chloroform; the organic solvent IV is dichloromethane, chloroform or ethyl acetate.

9. Use of the tanshinone IIA C-1 position esterification derivative according to claim 1 in the preparation of a myocardial protection drug, characterized in that: The tanshinone IIA C-1 position esterified derivative is , and one or more of them.

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