New application of resveratrol oligomer in reducing uric acid

Abstract

The invention provides a new application of a resveratrol oligomer in reducing uric acid. The research finds that the resveratrol oligomer separated from the Chinese herbaceous peony seed extract has XOD inhibitory activity, and can be used for treating hyperuricemia and gout or gout complications caused by hyperuricemia.

Description

Technical Field

[0001] The invention belongs to the field of medicines or health products, and specifically relates to a new use of resveratrol oligomers in reducing uric acid. Background Art

[0002] Uric acid is the final metabolite of purine compounds in humans, and purine metabolism disorders lead to hyperuricemia. Under normal purine diet conditions, fasting blood uric acid levels in men and women on two different days are higher than 416 μmol / L, which is called hyperuricemia. At present, the treatment of hyperuricemia, gout and gout complications is mainly through the control of uric acid in the blood. Its mechanism of action mainly includes the following two aspects: (1) effectively inhibiting the formation of uric acid by inhibiting the activity of xanthine oxidase (XO), and representative drugs include allopurinol and febuxostat; (2) promoting the excretion of uric acid, and representative drugs include probenecid and benzbromarone. However, the above drugs usually have large toxic side effects, and the tolerance of the above drugs is generally low. In summary, these problems limit the clinical application of these drugs to a certain extent. Therefore, it is of great significance to study new drugs for the treatment of gout.

[0003] Oligo-resveratrol compounds have a wide range of sources, and their main reported activities are similar to those of resveratrol, including antioxidant, anti-aging, liver protection, antibacterial, antiviral, anti-allergic, etc. There are no related reports on the use of oligo-resveratrol compounds in the field of uric acid lowering. Summary of the invention

[0004] The technical problem solved by the present invention is to provide a new use of resveratrol oligomers in reducing uric acid.

[0005] To solve the above technical problems, the present invention is achieved through the following technical solutions: The present invention provides the use of resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof in the preparation of medicines with uric acid lowering effects.

[0006] Preferably, in the use of the above-mentioned resveratrol oligomer and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof in the preparation of a drug having a uric acid-lowering effect, the resveratrol oligomer is selected from suffruticosol A or suffruticosol B.

[0007] Further preferred is the use of the resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof in the preparation of a drug having a uric acid-lowering effect. The resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof are prepared by conventional processes and adding conventional excipients to form clinically acceptable tablets, capsules, powders, mixtures, pills, granules, syrups, patches, suppositories, aerosols, ointments or injections.

[0008] Another aspect of the present invention provides the use of resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof in the preparation of medicines for treating gout.

[0009] Preferably, in the use of the above-mentioned resveratrol oligomer and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof in the preparation of a medicament for treating gout, the resveratrol oligomer is selected from suffruticosol A or suffruticosol B.

[0010] Further preferred is the use of the above-mentioned resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof in the preparation of medicaments for treating gout. The resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof are prepared by conventional processes and adding conventional excipients to form clinically acceptable tablets, capsules, powders, mixtures, pills, granules, syrups, patches, suppositories, aerosols, ointments or injections. Implementation

[0011] The present invention is further described in detail below through specific implementation modes.

[0012] Xanthine, xanthine oxidase, allopurinol, analytical grade anhydrous ethanol, chloroform, methanol, ethyl acetate, distilled water, dimethyl sulfoxide, potassium dihydrogen phosphate, and potassium hydrogen phosphate are all commercially available products. Peony seeds were purchased from Bozhou medicinal materials market.

[0013] Example 1 Preparation of resveratrol oligomers suffruticosol A and suffruticosol B 800 g of dried peony seeds were crushed and then heated and refluxed with 70% ethanol aqueous solution, with a solid-liquid ratio of 1:8 (weight ratio), an extraction time of 30 minutes, and extracted three times. The extracts were combined, filtered, and concentrated under reduced pressure to obtain an extract. Part of the extract was concentrated under reduced pressure to dryness and marked as extract E01. The remaining extract was extracted with three times the volume of ethyl acetate, extracted three times, and the ethyl acetate layers were combined and concentrated under reduced pressure to obtain ethyl acetate extract EA02. The obtained ethyl acetate extract was separated using a silica gel column, and chloroform methanol (5:1, V:V) was used as an eluent for elution. The eluent of the 2nd to 4th column volume was collected and concentrated to dryness under reduced pressure to obtain EAFr24. EAFr24 was further separated and purified using an Agilent SD-1 preparative liquid chromatography, using a reversed-phase C8 chromatographic column and isocratic elution with a 50% methanol aqueous solution. According to the liquid phase detection spectrum collection, compounds suffruticosol A and suffruticosol B were obtained in turn. Structural confirmation data of the compound 1 H NMR and 13 C NMR refers to the following literature: [1] Sarker SD, Whiting P, Dinan L, et al. Identification and Ecdysteroid AntagonistActivity of Three Resveratrol Trimers (Suffruticosols A, B and C) from Paeonia suffruticosa[J]. Tetrahedron, 1999, 55(2):513–524.

[0014] Example 2 In vitro inhibition experiment of benzoylpeonidin compounds on xanthine oxidase 1. Experimental methods 1.1 Solution preparation Preparation of phosphate buffered saline (PB) Prepare K2HPO4 (weigh 5.23g K2HPO4 and dissolve it in 400mL water) and KH2PO4 solution (weigh 1.02g KH2PO4 and dissolve it in 100mL water) in advance, then accurately measure 324mL K2HPO4 and 76mL KH2PO4, mix them evenly, and you will get 50mM, pH7.5 phosphate buffer PB as the test solution for the blank control group.

[0015] Preparation of allopurinol solution Weigh 5 mg of allopurinol and dissolve it in 1 mL of DMSO to make a stock solution. After it is completely dissolved, dilute it with PB to 1.5 μg / mL and 10 μg / mL, and store it in aliquots of 1 mL / tube in a -20°C refrigerator as the test solution for the positive control group.

[0016] Take 1 tube for each experiment without dilution and use directly after thawing.

[0017] Among them, the allopurinol solution with a concentration of 1.5 μg / mL was used as the allopurinol low-dose group, and the allopurinol solution with a concentration of 10 μg / mL was used as the allopurinol high-dose group.

[0018] Preparation of xanthine oxidase solution According to the enzyme amount and volume indicated on the xanthine oxidase product package, dilute it with PB to 100U / L. After preparation, dispense it into 8mL / tube and store it in a -20℃ environment.

[0019] Take one tube each time and dilute it to 25 U / L with PB for later use.

[0020] Preparation of substrate xanthine solution Accurately weigh 4.56 mg of xanthine, place in a 50 mL volumetric flask, add 150 µL of 1 M sodium hydroxide, 30 mL of PB, and sonicate until the xanthine is completely dissolved and becomes clear and transparent, then dilute to the mark with PB to obtain a 600 µM xanthine solution.

[0021] Preparation of test solution 20 mg of the extracts E01, EA02, EAFr24, and compounds suffruticosol A and suffruticosol B (HPLC purity ≥ 98%) prepared in Example 1 were weighed respectively and added into a 1.5 mL centrifuge tube, and then 1 mL of DMSO was added and ultrasonically dissolved into a mother solution, and 50 μL of the mother solution and 950 μL of PB were drawn and mixed to obtain a solution of the test drug.

[0022] 1.2 Reaction and termination of enzyme and substrate In the experimental group, 400 μL of xanthine oxidase (XOD) solution and 200 μL of test solution were added to the 1.5 mL centrifuge tube in sequence; in the blank control group, 400 μL of xanthine oxidase (XOD) solution and 200 μL of test solution of the blank control group were added to the 1.5 mL centrifuge tube in sequence; in the positive control group, 400 μL of xanthine oxidase (XOD) solution and 200 μL of test solution of the positive control group, low-dose group and high-dose group were added to the 1.5 mL centrifuge tube in sequence; then, each group performed the following experimental operations: incubated in a 37°C water bath for 10 min, 400 μL of xanthine (XAN) solution was added to start the reaction, after reacting in a 37°C water bath for 10 min, 80 μL of 1 M hydrochloric acid was added to terminate the reaction, and then 70-75 μL of 1 M NaOH solution was added to adjust the pH to neutral.

[0023] 1.3 Centrifugation and machine testing After the treated test solution of each group was centrifuged at 12000 rpm for 10 min at room temperature, 600 μL of the supernatant was aspirated into a liquid phase bottle, and the uric acid value of each group of test solution was detected by HPLC.

[0024] 2. Experimental data detection and processing The xanthine oxidase inhibition rate was calculated according to the following formula: inhibition rate = [(uric acid value of the blank control group - uric acid value of the test solution) / uric acid value of the blank control group] * 100.

[0025] 3. Experimental results The specific experimental results of the inhibition rate of each benzoylpeonidin compound on xanthine oxidase are shown in Table 1.

[0026] Table 1 Specific experimental results of the inhibition rate of extracts E01, EA02, EAFr24, compounds suffruticosol A and suffruticosol B on xanthine oxidase Group Inhibition rate (%) Positive control low dose group 40.1% Positive control high dose group 97.2% Experimental group E01 40.7% Experimental group EA02 56.8% Experimental group EAFr24 88.3% suffruticosol A 98.3% suffruticosol B 97.8% The results in Table 1 show that the compounds suffruticosol A and suffruticosol B of the present invention both have good xanthine oxidase inhibitory effects, and the XOD inhibition rate is higher than that of the extract group EAFr24, indicating that the compounds suffruticosol A and suffruticosol B are uric acid-lowering active substances in the extract, have significant xanthine oxidase inhibitory effects, and can be used as potential uric acid-lowering drugs.

Claims

1. Use of resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof in the preparation of uric acid-lowering drugs.

2. Use of the resveratrol oligomer and a pharmaceutically acceptable salt, ester, prodrug or solvate thereof according to claim 1 in the preparation of a drug having a uric acid-lowering effect, characterized in that: The resveratrol oligomer is selected from suffruticosol A or suffruticosol B.

3. Use of the resveratrol oligomer and its pharmaceutically acceptable salt, ester, prodrug or solvate according to claim 1 or 2 in the preparation of a drug having a uric acid-lowering effect, characterized in that: Resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof are prepared into clinically acceptable tablets, capsules, powders, mixtures, pills, granules, syrups, patches, suppositories, aerosols, ointments or injections by conventional processes and adding conventional excipients.

4. Use of resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof in the preparation of medicines for treating gout.

5. Use of the resveratrol oligomer and its pharmaceutically acceptable salt, ester, prodrug or solvate according to claim 4 in the preparation of a drug for treating gout, characterized in that: The resveratrol oligomer is selected from suffruticosol A or suffruticosol B.

6. Use of the resveratrol oligomer and its pharmaceutically acceptable salt, ester, prodrug or solvate according to claim 4 or 5 in the preparation of a drug for treating gout, characterized in that: Resveratrol oligomers and pharmaceutically acceptable salts, esters, prodrugs or solvates thereof are prepared into clinically acceptable tablets, capsules, powders, mixtures, pills, granules, syrups, patches, suppositories, aerosols, ointments or injections by conventional processes and adding conventional excipients.