A chlorogenic acid derivative and its preparation method and application

Abstract

The present invention discloses a chlorogenic acid derivative, its preparation method, and application. The chlorogenic acid derivative has the following molecular structure: it is prepared by reacting chlorogenic acid with 4-aminopyrazolo[3,4-d]pyrimidine and has promising application prospects as a uric acid-lowering drug; #imgabs0#

Description

Technical Field

[0001] The present invention relates to a drug derivative based on a natural product, and in particular to a chlorogenic acid derivative and a preparation method and application thereof. Background Art

[0002] With rapid economic development and a significant improvement in people's living standards, the number of patients suffering from hyperuricemia and gout, commonly known as "diseases of affluence," has been increasing year by year, and the incidence is trending toward younger patients. Gout is a disorder of purine metabolism characterized by decreased uric acid excretion and / or increased uric acid production. It can progress to uremia, renal failure, arteriosclerosis, coronary heart disease, stroke, and other conditions, and in severe cases, can be life-threatening.

[0003] Currently, prevention of hyperuricemia and gout primarily involves limiting high-purine foods and controlling alcohol consumption. Treatment requires both controlling acute arthritis attacks and long-term treatment of hyperuricemia to prevent tophi deposition. Chemical medications are categorized into two types: those for treating gouty arthritis and those for correcting hyperuricemia. The former includes colchicine, nonsteroidal anti-inflammatory drugs, and glucocorticoids, which quickly and effectively relieve pain and reduce inflammation. The latter includes probenecid, benzbromarone (which promotes uric acid excretion), and allopurinol (which inhibits uric acid production). However, these medications have significant adverse reactions, with benzbromarone being associated with severe liver damage and neurological side effects. Allopurinol, the only clinically used xanthine oxidase inhibitor, inhibits uric acid production by inhibiting xanthine oxidase, but it can also cause toxic side effects such as skin allergies, liver damage, and life-threatening hypersensitivity syndrome.

[0004] In response to the serious toxic side effects of existing anti-gout drugs, the development of new alternative drugs based on natural products with good biocompatibility has become a current hot topic and scientific research challenge. Summary of the Invention

[0005] In order to solve the above technical problems, the present invention provides a chlorogenic acid derivative and a preparation method and application thereof.

[0006] The present invention first proposes a chlorogenic acid derivative having the following molecular structure expression:

[0007]

[0008] Chlorogenic acid is a phenolic acid active substance with multiple pharmacological activities, including free radical scavenging, anti-inflammatory, antiviral, and immunomodulatory. It is widely found in plants such as chicory, stevia, and dandelion. This invention uses chlorogenic acid as a raw material for drug structure design. While retaining the main molecular structure of chlorogenic acid, it has developed a chlorogenic acid derivative with good biocompatibility and high uric acid-lowering activity. This is expected to have promising applications in uric acid-lowering drugs and preparations.

[0009] Secondly, the present invention also provides a method for preparing a chlorogenic acid derivative, comprising the following steps:

[0010]

[0011] Chlorogenic acid and 4-aminopyrazolo[3,4-d]pyrimidine are used as raw materials and a condensation reaction is carried out in a solvent to generate a chlorogenic acid derivative.

[0012] As a preferred embodiment of the preparation method of the present invention, the molar ratio of chlorogenic acid to 4-aminopyrazolo[3,4-d]pyrimidine is 1:(0.9-1.2), for example, 1:0.9, 1:0.95, 1:1, 1:1.05, 1:1.1, 1:1.15, etc.;

[0013] Preferably, the solvent is selected from one or more of dichloromethane, chloroform, tetrahydrofuran, acetonitrile, ethanol, and ethyl acetate.

[0014] As a preferred embodiment of the preparation method of the present invention, the condensation reaction is carried out in the presence of a DCC / DMAP catalyst system.

[0015] As a preferred embodiment of the preparation method of the present invention, the DCC / DMAP catalyst system is composed of dicyclohexylcarbodiimide and 4-dimethylaminopyridine in a molar ratio of 1:(0.2-0.5), and the molar ratio of the two is, for example, 1:0.25, 1:0.3, 1:0.35, 1:0.4, 1:0.45, etc.;

[0016] Preferably, the amount of dicyclohexylcarbodiimide used is 1-1.5 times the molar amount of chlorogenic acid, for example, 1.1 times, 1.2 times, 1.3 times, 1.4 times, etc.

[0017] As a preferred embodiment of the preparation method of the present invention, it is characterized in that the reaction conditions of the condensation reaction are stirring the reaction at room temperature for 12-24 hours, for example, 14 hours, 15 hours, 18 hours, 20 hours, 22 hours, etc.

[0018] As a preferred embodiment of the preparation method of the present invention, after the reaction is completed, the insoluble matter is first removed by filtration, and then diluted hydrochloric acid is added to extract the organic phase, which is then purified to obtain the desired product.

[0019] As a preferred embodiment of the preparation method of the present invention, the concentration of the dilute hydrochloric acid is 0.1-2 mol / L, for example, 0.5 mol / L, 1 mol / L, 1.5 mol / L, 2 mol / L, etc.

[0020] The present invention does not impose any limitation on the specific purification method of the target product, which can be any one or more of distillation, water washing, recrystallization, column chromatography, or a conventional combination of these with other known separation and purification methods.

[0021] Again, the present invention provides a use of the chlorogenic acid derivative as described above or the chlorogenic acid derivative prepared by the method described above in the medical field, especially in uric acid-lowering drugs or preparations.

[0022] The chlorogenic acid derivative provided by the present invention has high uric acid-lowering activity, a simple preparation method, and good biocompatibility, and is expected to become a new therapeutic drug for hyperuricemia and gout. DETAILED DESCRIPTION

[0023] The present invention will be further described below through specific examples. The examples of the present invention are only for illustration of the present invention and do not limit the scope of the present invention.

[0024] Unless otherwise specified, the raw materials and reagents in the present invention can be purchased from commercial sources.

[0025] [Example 1]

[0026]

[0027] Chlorogenic acid (0.35 g, 1 mmol), 4-aminopyrazolo[3,4-d]pyrimidine (0.14 g, 1 mmol), dicyclohexylcarbodiimide (0.21 g, 1 mmol), and 4-dimethylaminopyridine (0.02 g, 0.2 mmol) were dissolved in dichloromethane (10 mL) and stirred at room temperature for 12 h. After completion of the reaction, the insoluble matter was removed by filtration, and then the organic phase was extracted with 1 mol / L dilute hydrochloric acid. The organic phase was dried over anhydrous sodium sulfate, rotary evaporated, and separated and purified by silica gel column chromatography (dichloromethane:methanol = 100:1) to obtain the target product, a chlorogenic acid derivative (yield 74%).

[0028] 1H NMR (500MHz, Chloroform-d): δ13.23(s,1H), δ9.64(s,1H),8.90(s,1H),8.58(s,1H),8.41(s,1H),7.57- 7.50(m,1H),7.01(d,J=1.9Hz,1H),7.00-6.95(m,1H),6.95(d,J=9.0Hz,1H),6.80(d,J=8.3Hz,1H),6.21 (d,J=16.3Hz,1H),5.42(s,1H),5.38-5.30(m,1H),4.18(ddtd,J=7.5,6.1,5.0,3.7Hz,1H),4.11(d,J=6. 2Hz,1H),3.79(ddd,J=8.1,5.9,4.9Hz,1H),3.28(d,J=5.9Hz,1H),2.66-2.34(m,2H),2.29-1.97(m,2H).

[0029] [Example 2]

[0030] Chlorogenic acid (0.35 g, 1 mmol), 4-aminopyrazolo[3,4-d]pyrimidine (0.12 g, 0.9 mmol), dicyclohexylcarbodiimide (0.25 g, 1.2 mmol), and 4-dimethylaminopyridine (0.06 g, 0.5 mmol) were dissolved in ethyl acetate (10 mL) and stirred at room temperature for 24 h. After completion of the reaction, the insoluble matter was removed by filtration, and then the organic phase was extracted with 0.5 mol / L dilute hydrochloric acid. The organic phase was dried over anhydrous sodium sulfate, rotary evaporated, and separated and purified by silica gel column chromatography (dichloromethane:methanol = 100:1) to obtain the target product, a chlorogenic acid derivative (yield 72%).

[0031] [Example 3]

[0032] Chlorogenic acid (0.35 g, 1 mmol), 4-aminopyrazolo[3,4-d]pyrimidine (0.16 g, 1.2 mmol), dicyclohexylcarbodiimide (0.31 g, 1.5 mmol), and 4-dimethylaminopyridine (0.04 g, 0.33 mmol) were dissolved in ethanol (10 mL) and stirred at room temperature for 18 h. After completion of the reaction, the insoluble matter was removed by filtration, and then the organic phase was extracted with 2 mol / L dilute hydrochloric acid. The organic phase was dried over anhydrous sodium sulfate, rotary evaporated, and separated and purified by silica gel column chromatography (dichloromethane:methanol = 100:1) to obtain the target product, a chlorogenic acid derivative (yield 75%).

[0033] <Cytotoxicity test>

[0034] The cytotoxicity test of PC12 cells was performed using the CCK-8 method, wherein the experimental group was cultured with the chlorogenic acid derivative prepared in Example 1; the control group was cultured with chlorogenic acid. The results showed that at a drug concentration of 0.5-20 μmol / L, there was no significant change in the OD values ​​at 450 nm between the experimental and control groups at the same concentration.

[0035] <In vivo animal experiments>

[0036] ① Animal preparation and grouping: Adult male SD rats were divided into 6 groups, 10 in each group, namely model group, low-dose experimental group, medium-dose experimental group, high-dose experimental group, positive control group, and raw material control group;

[0037] ② Drug preparation: First, prepare a 0.8 wt% CMC-Na solution, dissolve the chlorogenic acid derivative prepared in Example 1 in the CMC-Na solution, and prepare a low-dose experimental group solution, a medium-dose experimental group solution, and a high-dose experimental group solution with drug concentrations of 0.5 mg / mL, 1 mg / mL, and 2 mg / mL, respectively; then dissolve benzbromarone in the CMC-Na solution to prepare a positive control group solution with a drug concentration of 1 mg / mL; equal amounts of chlorogenic acid and 4-aminopyrazolo[3,4-d]pyrimidine were dissolved in the CMC-Na solution to prepare a raw material control group solution with a total drug concentration of 1 mg / mL.

[0038] ③Dosing experiment: Rats were intragastrically administered with a dose of 10 mL / kg per day according to their body weight for 7 consecutive days; the specific feeding method was as follows: the model group was given 0.8 wt% CMC-Na solution; the low-dose experimental group, medium-dose experimental group, and high-dose experimental group were given experimental group drug solutions with concentrations of 0.5 mg / mL, 1 mg / mL, and 2 mg / mL, respectively; the positive control group was given a positive control group drug solution with a concentration of 1 mg / mL; and the raw material control group was given a raw material control group drug solution with a total concentration of 1 mg / mL.

[0039] ④ Sample Processing and Sampling: On day 8, 30 minutes after the last administration, rats in each group received an intraperitoneal injection of 50 mg / ml hypoxanthine solution (solvent: 0.8 wt% CMC-Na solution) to establish a hyperuricemia model. The dose was 1000 mg / kg. Thirty minutes after administration, the rats were immediately euthanized and their eyes enucleated for blood collection. After serum exuded, the rats were centrifuged at 15,000 rpm for 4 minutes. The upper serum layer was aspirated and placed into a capped sample tube. The volume was then adjusted to the desired volume with normal saline. Serum uric acid levels were measured on a Beckman LX20 fully automated biochemical analyzer. Data are expressed as mean ± standard deviation. The test results are shown in Table 1.

[0040] Table 1. Blood uric acid content test results

[0041]

[0042] From the above test results, it can be seen that the chlorogenic acid derivatives provided by the present invention can exert the effect of lowering uric acid in a short period of time at a dosage of 5-20 mg / kg, and have almost zero biological toxicity; and the uric acid lowering level at a lower dosage is significantly better than that of the control group using chlorogenic acid and 4-aminopyrazolo[3,4-d]pyrimidine, which produces unexpected effect changes compared with the known technology, and has obvious technical advantages.

[0043] The above is only a preferred embodiment of the present invention. It should be pointed out that ordinary technicians in this field can make several improvements and supplements without departing from the method of the present invention. These improvements and supplements should also be considered within the scope of protection of the present invention.

Claims

1. A chlorogenic acid derivative, characterized in that It has the following molecular structure expression: 。 2. A method for preparing the chlorogenic acid derivative according to claim 1, characterized in that: The following steps are involved: ； Chlorogenic acid and 4-aminopyrazolo[3,4-d]pyrimidine are used as raw materials and a condensation reaction is carried out in a solvent to generate a chlorogenic acid derivative.

3. The method for preparing the chlorogenic acid derivative according to claim 2, characterized in that: The molar ratio of chlorogenic acid to 4-aminopyrazolo[3,4-d]pyrimidine is 1:(0.9-1.2).

4. The method for preparing the chlorogenic acid derivative according to claim 3, characterized in that: The solvent is selected from one or more of dichloromethane, chloroform, tetrahydrofuran, acetonitrile, ethanol, and ethyl acetate.

5. The method for preparing a chlorogenic acid derivative according to any one of claims 2 to 4, characterized in that: The condensation reaction is carried out in the presence of a DCC / DMAP catalyst system.

6. The method for preparing the chlorogenic acid derivative according to claim 5, characterized in that: The DCC / DMAP catalyst system consists of dicyclohexylcarbodiimide and 4-dimethylaminopyridine in a molar ratio of 1:(0.2-0.5).

7. The method for preparing the chlorogenic acid derivative according to claim 6, characterized in that: The amount of dicyclohexylcarbodiimide used is 1-1.5 times the molar amount of chlorogenic acid.

8. The method for preparing a chlorogenic acid derivative according to any one of claims 2 to 4, characterized in that: The reaction conditions of the condensation reaction are stirring and reacting at room temperature for 12-24 hours.

9. The method for preparing a chlorogenic acid derivative according to any one of claims 2 to 4, characterized in that: After the reaction is completed, the insoluble matter is first removed by filtration, and then diluted hydrochloric acid is added to extract the organic phase, which is purified to obtain the target product.

10. The method for preparing the chlorogenic acid derivative according to claim 9, characterized in that: The concentration of the dilute hydrochloric acid is 0.1-2 mol / L.

11. Use of the chlorogenic acid derivative according to claim 1 or the chlorogenic acid derivative prepared by the method according to any one of claims 2 to 10 in the preparation of a uric acid-lowering drug or preparation.

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