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Application of theaflavin-3,3'-digallate in preparation of anti-inflammatory medicines

A technology of double gallate, anti-inflammatory drug, applied in the field of biomedicine

Inactive Publication Date: 2018-02-23
GUANGZHOU JINAN BIOMEDICINE RES & DEV CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The application of theaflavin-3,3'-digallate in the preparation of anti-inflammatory drugs has not been reported so far

Method used

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  • Application of theaflavin-3,3'-digallate in preparation of anti-inflammatory medicines
  • Application of theaflavin-3,3'-digallate in preparation of anti-inflammatory medicines
  • Application of theaflavin-3,3'-digallate in preparation of anti-inflammatory medicines

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 MTT method detects the toxicity of TF-3 to cells

[0024] The specific steps of using the MTT method to detect the toxicity of TF-3 to cells are:

[0025] (1) RAW 264.7, U937, LO-2 cell plating: Evenly spread 100 μL of RAW264.7, U937, LO-2 cells in the logarithmic growth phase in a 96-well plate, and then put it in 37°C, 5% CO 2 Cultivate in a constant temperature incubator until the cell density reaches 80-90%, discard the medium, wash the cells once with PBS buffer, and discard the PBS solution.

[0026] (2) Drug gradient dilution: the drug mother solution was diluted with serum-free DMEM medium into drug working solutions of different concentrations, and the concentrations were 200, 100, 50, 25, 12.5, 6.25, and 3.125 μM in sequence.

[0027] (3) Dosing treatment: 100 μL TF-3 dilution was added to each well, and three replicate wells were set for each drug concentration, and a blank control group, a normal cell control group and a solvent control group were...

Embodiment 2

[0032] Embodiment 2 establishes the application model of medicine described in the present invention

[0033] (1) RAW 264.7 cell inflammatory response model

[0034] (1) Dilute the cell suspension to 2×10 5 Cells / mL density, add 1 mL per well into a sterile 12-well plate, shake gently to ensure that the cells are evenly attached to the wall.

[0035] (2) Use serum-free DMEM medium to dilute the 1 mg / mL LPS stock solution to a working solution with a final concentration of 100 ng / mL.

[0036] (3) After culturing overnight, observe the state and confluence of the cells. When the state is good and the density reaches 80-90%, discard the medium, set a normal cell control group without LPS, add serum-free medium, and add to the remaining wells containing Serum-free DMEM medium with 100ng / ml LPS and different concentrations of drugs was placed in a cell culture incubator to continue culturing.

[0037] (4) After 6 hours, total cellular RNA was extracted, reverse-transcribed into ...

Embodiment 3

[0043] Example 3 Detection of inflammatory gene expression level

[0044] (1) Extraction of total RNA from cells

[0045] (1) Pre-preparation: Put on the lab coat, mask and gloves; pre-cool the high-speed refrigerated centrifuge for 10 minutes to lower the temperature to 4°C; prepare 1000 μL, 200 μL, and 10 μL pipette tips without RNase; get ready Chloroform, isopropanol, and reconstituted 75% ethanol.

[0046] (2) Cell lysis: Wash the drug-treated cells 3 times with PBS solution, discard the PBS solution, immediately add 1mL Trizol reagent, place at room temperature for 5min, and repeatedly blow vigorously with a 1mL pipette to fully lyse the cells. Transfer to a 1.5mL EP tube without RNase.

[0047] (3) Phase separation: add 200 μL of chloroform, oscillate on a vortex shaker or vigorously invert to mix, and stand at room temperature for 5 minutes. Centrifuge at 12000g for 15min at 4°C.

[0048] (4) RNA precipitation: Carefully pipette 300-500 μL supernatant (do not suck ...

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Abstract

The invention belongs to the technical field of biomedicine, and in particular relates to the application of theaflavin-3,3'-digallate in the preparation of anti-inflammatory drugs. The present invention uses two inflammatory cell models to demonstrate for the first time the anti-inflammatory activity of the small molecule monomer compound TF-3; the present invention establishes a mouse acute lung injury inflammation model for the first time to prove that TF-3 inhibits the prevention and treatment of lung injury inflammation in vivo Applications. The experimental results prove that: TF‑3 can effectively inhibit the expression of inflammatory factors TNF‑α, IL‑1β and IL‑6 genes, and can inhibit the production of inflammatory cytokines in the serum of mice with acute lung injury, and can prevent the infiltration of inflammatory cells, Treat lung damage. The medium and small molecule monomer compound theaflavin-3,3'-digallate body of the present invention has low toxicity and remarkable anti-inflammatory effect, and overcomes the shortcomings of existing anti-inflammatory drugs such as high toxicity and easy drug resistance. question.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to the application of theaflavin-3,3'-digallate in the preparation of anti-inflammatory drugs. Background technique [0002] Inflammation is a defense response produced by the body under various foreign pathogenic microorganisms or stress stress conditions. Excessive inflammatory response is the basis of many diseases and is closely related to the occurrence and development of many diseases. Millions of inflammatory patients around the world suffer from pain every year. Therefore, the development of low-toxicity and effective anti-inflammatory drugs is an increasingly urgent need for inflammatory patients. [0003] Theaflavin-3,3'-digallate (theaflavin-3,3'-digallate, TF3) is a kind of substance with benzotropolone structure, which is formed by benzocyclization of catechin; Its molecular formula is C 43 h 32 o 20 , the molecular weight is 868.70, CAS NO.33377-72...

Claims

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Application Information

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IPC IPC(8): A61K31/353A61P29/00A61P11/00
CPCA61K31/353
Inventor 王一飞袁晓吴燕婷
Owner GUANGZHOU JINAN BIOMEDICINE RES & DEV CENT
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