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Optically active bicyclol glucoside and preparation method thereof, and application of optically active bicyclol glucoside in prevention and treatment of liver diseases

An optically active and glucoside technology, applied in the field of medicine, can solve the problems of difficult to obtain optically active bicyclic alcohol glycoside derivatives, unfriendly environment, low yield, etc., and achieve novel preparation method, environmentally friendly method, and high conversion rate Effect

Inactive Publication Date: 2020-06-16
INST OF MATERIA MEDICA AN INST OF THE CHINESE ACAD OF MEDICAL SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] In the analysis of the existing literature, there are currently reports on the glycosylation of bicyclic alcohols by chemical methods, but there are still various deficiencies in the above-mentioned chemical methods, resulting in the difficulty in obtaining optically active bicyclic alcohol glycoside derivatives, low yields and Technical problems such as unfriendly environment

Method used

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  • Optically active bicyclol glucoside and preparation method thereof, and application of optically active bicyclol glucoside in prevention and treatment of liver diseases
  • Optically active bicyclol glucoside and preparation method thereof, and application of optically active bicyclol glucoside in prevention and treatment of liver diseases
  • Optically active bicyclol glucoside and preparation method thereof, and application of optically active bicyclol glucoside in prevention and treatment of liver diseases

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Embodiment 1. Preparation of recombinant glycosyltransferase

[0053] The mutant OleD derived from the glycosyltransferase gene OleD of Streptomyces antibioticus Loki Constructed in the expression vector pET28a, the recombinant plasmid pET28a-OleD Loki Introduced into Escherichia coli BL21, positive transformants were screened by PCR, and sent for sequencing to confirm the correctness of the reading frame.

[0054] The correct positive transformants were inoculated in LB medium containing kanamycin (pET-28a carrier carrying resistance), 37°C, 200rpm shaking culture for 12h; 2), the activated seed solution was again mixed with 100:1 The proportion of inoculation in LB (kanamycin + ) culture medium, select 250mL triangular flask, the volume of each bottle is 50mL (use 500mL triangular flask, the volume of each bottle is 200mL); 3), 37 ℃, 200rpm culture to OD 600 ≈0.6, add IPTG with a final concentration of 0.1mM to each bottle of culture; induce the expression of the t...

Embodiment 2

[0059] Example 2. Enzymatic preparation and structural identification of optically active bicyclol-9-O-β-D-glucosidase

[0060]Cultivate 2.0L of recombinant Escherichia coli, purify by the method described in Example 1 to obtain 16mL of pure enzyme solution, add 2.56mL of bicyclol DMSO solution (total 50.0mg, 0.128mM) and UDPG (156mg, 0.256mM) and mix gently, Reaction at 30°C for 48h. Get 100 μ L reaction solution and carry out HPLC-MS analysis, the result shows, the conversion rate of product I and II is respectively 26.5%, 24.2%, and total conversion rate is 50.7% (HPLC figure sees attached Figure 15 ), MS analysis results showed that the product was bicyclol-9-O-β-D-glucoside. The HPLC analysis conditions of bicyclic alcohol and its glycosidation products are: Agilent 1200 series high-performance liquid phase instrument, Shiseido HPLC reverse phase analysis C 18 Column, flow rate 1.0mL / min, mobile phase is methanol and 0.1% formic acid water system, injection 20μL, gradi...

experiment example 1

[0065] Experimental example 1. Protective effect of optically active bicyclol-9-O-β-D-glucoside on APAP-induced liver cell injury in vitro

[0066] 1. Cell Culture

[0067] HCC HepG 2 Cells were grown in DMEM medium containing 10% fetal bovine serum (containing penicillin 100 U / mL, streptomycin 100 μg / mL) at 37 °C, 5% CO 2 , saturated humidity. Digested and passaged with solution containing 0.25% trypsin and 0.02% EDTA.

[0068] 2. Optically active bicyclol-9-O-β-D-glucoside on HepG 2 cytotoxicity of cells

[0069] The MTT method is used. HepG 2 The cells were seeded in a 96-well cell culture plate, and after 24 hours of culture, 0.5-20 μM bicyclol-9-O-β-D-glucoside I and II were added, and a solvent control group was set at the same time, and 3 parallel wells were set for each concentration. After bicyclol-9-O-β-D-glucoside I and II acted on the cells for 48 hours, discard the culture medium, add 100 μL MTT (0.5 mg / mL) solution to each well, continue to cultivate for 4...

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Abstract

The invention discloses optically active (P)-bicyclol-9-O-beta-D-glucoside and (M)-bicyclol-9-O-beta-D-glucoside applicable to medicines and a preparation method and application thereof. According tothe invention, bicyclol and uridine diphosphate glucose disodium salt are catalyzed by glycosyltransferase to synthesize optically active (P)-bicyclol-9-O-beta-D-glucoside and (M)-bicyclol-9-O-beta-D-glucoside in one step. The bicyclol-9-O-beta-D-glucoside prepared according to the invention can be used for preventing and treating liver diseases.

Description

technical field [0001] The invention belongs to the field of medicine, and in particular relates to an optically active bicyclic alcohol-9-O-β-D-glucoside compound, a preparation method thereof, and an application thereof in preventing and treating liver diseases. Background technique [0002] Institute of Materia Medica, Chinese Academy of Medical Sciences The national first-class new drug with independent intellectual property rights led by scientists such as , is used for the treatment of elevated aminotransferases caused by chronic hepatitis, and the curative effect is remarkable. It was launched in China in 2001 under the trade name Besano, and it was launched in Ukraine and Russia in 2004 and 2016 respectively. The research achievement "Research on National Class I Anti-Hepatitis New Drug Bicyclol" won the second prize of the 2007 National Science and Technology Progress Award. [0003] Chronic viral hepatitis is a common infectious disease that seriously endangers ...

Claims

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Application Information

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IPC IPC(8): C07H15/26C12P19/58C12P19/18A61P1/16A61K31/7048
CPCC07H15/26C12P19/58C12P19/18A61P1/16
Inventor 戴均贵孙华解可波李梅谭圳张凡姜琳陈日道陈大伟
Owner INST OF MATERIA MEDICA AN INST OF THE CHINESE ACAD OF MEDICAL SCI
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