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Preparation method for wine brewing yeast cell culture secretion and use for the same in resisting DNA virus and promoting hepatocyte growth

A technology of Saccharomyces cerevisiae cells and Saccharomyces cerevisiae, applied in antiviral agents, biochemical equipment and methods, and microbial-based methods, can solve problems that hinder the research of anti-HBV drugs, and achieve the goal of promoting and repairing mechanical damage of liver cells Effect

Inactive Publication Date: 2008-02-13
302 MILITARY HOSPITAL OF CHINA +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the past, due to the limitation of HBV by the narrow host range and the lack of in vitro culture system, the research of anti-HBV drugs has been hindered.

Method used

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  • Preparation method for wine brewing yeast cell culture secretion and use for the same in resisting DNA virus and promoting hepatocyte growth

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1. Preparation of Yeast Cell Culture Secretion

[0022] 1. Materials

[0023] PD medium: 15 grams of hydrolyzed milk protein (purchased from Beijing Shuangxuan Microbial Medium Products Factory)

[0024] Glucose 10 grams (Beijing Boda Biotechnology Co., Ltd.)

[0025] Sodium chloride 9 grams (Beijing Boda Biotechnology Co., Ltd.)

[0026] Dissolve in 1000 ml of sterilized water, and sterilize with 8 pounds of high pressure for 40 minutes. pH 7.5.

[0027] Saccharomyces cerevisiae CICC1001 (purchased from China Industrial Microorganism Culture Collection Management Center)

[0028] 2. Method

[0029] Pick a single colony of Saccharomyces cerevisiae, inoculate in 5ml of PD culture medium, and shake overnight at 30°C; transfer 1 ml to 500ml of PD culture medium, shake at 28°C for 48 hours; use a 0.2 micron filter membrane Sterilize by filtration.

Embodiment 2

[0030] Embodiment 2. in vitro antiviral test

[0031] 1. Materials

[0032] Yeast cell culture secretion: prepared according to the method in Example 1, and concentrated 10 times.

[0033] Human liver cancer 2215 cells (2.2.15 cells): donated by Professor Fu Xixian from the First Hospital of Beijing Medical University from the Department of Biochemistry, Mount Sinai University School of Medicine, USA.

[0034] Culture medium: DMEM, containing 380 μg / ml G418 (GIBCO, USA) and 100 IU / ml each of penicillin and streptomycin,

[0035] 10% mycoplasma-free calf serum (purchased from Hangzhou Sijiqing Institute of Bioengineering Materials) 03% glutamine, pH value about 7.0.

[0036] 2. Method

[0037] 1. Determination of the secretion of HBsAg and HBeAg by 2215 cells:

[0038] Divide 2215 cells into 1×10 5 Each / ml inoculated culture bottle (10ml / bottle), the medium was changed every 3 days and the supernatant was collected, stored at -20°C, collected 8 times in total, and then the...

Embodiment 3

[0055] Example 3. Saccharomyces cerevisiae cell culture secretion fluid promotes the growth of liver cells and inhibits the effect of liver fibrosis cells

[0056] 1. Materials

[0057] Adult male Wistar rats, weighing 200±20g, provided by the Experimental Animal Center of our hospital,

[0058] Type IV collagenase, product of SIGMA company,

[0059] Culture medium: DMEM, containing 380 μg / ml G418 (GIBCO, USA) and 100 IU / ml each of penicillin and streptomycin,

[0060] 10% mycoplasma-free calf serum (purchased from Hangzhou Sijiqing Institute of Bioengineering Materials) 03% glutamine, pH value about 7.0.

[0061] 2. Method

[0062] 1. Separation of hepatocytes: Separation and cultivation of hepatocytes The separation method of rat hepatocytes adopts the in situ two-step perfusion method. Wistar rats were taken, anesthetized with 0.5% pentobarbital sodium, fixed, and opened after aseptic treatment. Portal vein cannulation. Perfuse the Ca-free 2+ , Mg 2+ Ionic Hanks solut...

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Abstract

The invention discloses a method for preparing Saccharomyces cerevisiae cell culture secretion and the function of the secretion in DNA virus resistance and acceleration of hepatocyte growth, pertaining to the field of bio-pharmaceutics. The following method is adopted to prepare Saccharomyces cerevisiae cell culture secretion: monoclonal colony of Saccharomyces cerevisiae is selected and inoculated into 5ml PD culture medium, pendulated under 30 DEG C for a night; 1 ml is inoculated into 500ml PD culture medium, and pendulated under 28-30 DEG C for 40-60h; 0.2-micron filter membrane is used for filtration sterilization to achieve the product. The invention has the advantages that Saccharomyces cerevisiae cell culture secretion after cultivation by culture medium and purification has eminent function in DNA virus resistance and acceleration of hepatocyte growth, and is a novel medicine to cure hepatitis B and liver-injury diseases.

Description

technical field [0001] The invention relates to a preparation method of brewer's yeast cell culture secretion liquid and its application of anti-DNA virus and promoting liver cell growth, belonging to the field of biomedicine. Background technique [0002] Hepatitis B virus (HBV) belongs to DNA virus, and the acute and chronic hepatitis, hepatic necrosis and hepatocellular carcinoma caused by it are extremely harmful to human health, and the number of people who die from liver cancer reaches more than 1 million every year. In the past, the research of anti-HBV drugs was hindered because of the narrow host range of HBV and the lack of in vitro culture system. In recent years, with the successive establishment of HBV DNA transfected cell lines, necessary conditions have been provided for the screening of anti-HBV drugs. Contents of the invention [0003] The first technical problem to be solved by the present invention is to provide a Saccharomyces cerevisiae cell culture s...

Claims

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Application Information

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IPC IPC(8): C12N1/18A61K36/06A61P31/12A61P1/16C12R1/865
Inventor 貌盼勇刘洪
Owner 302 MILITARY HOSPITAL OF CHINA
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