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Preparation for amylovorin of bacillus and application thereof

A technology of exopolysaccharide and Bacillus subtilis, which is applied in the field of biopharmaceuticals to achieve the effects of high yield, easy quality control and simple ingredients

Inactive Publication Date: 2007-02-07
NANJING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

According to the literature survey, there are few reports of anti-tumor bacterial exopolysaccharides

Method used

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  • Preparation for amylovorin of bacillus and application thereof
  • Preparation for amylovorin of bacillus and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0018] Embodiment one: the fermentation condition of Bacillus subtilis and the preparation method of fermented liquid

[0019] Medium composition: peptone 10g, beef extract 5g, sodium chloride 10g, add water to 1 liter, pH7.0.

[0020] Preparation of seed liquid: Take a little strain stored in a -80°C refrigerator in 4ml of liquid medium, and cultivate overnight at 37°C on a shaker at 100 rpm.

[0021] Preparation of fermentation broth: 4ml of seed liquor was inoculated into a 1L Erlenmeyer flask containing 400ml of culture medium, cultured on a shaking table at 37°C and 100rpm for 48h.

Embodiment 2

[0022] Example 2: Preparation of exopolysaccharide

[0023] The Bacillus subtilis fermentation broth was sterilized by centrifugation at 4000rpm for 30min, then concentrated under reduced pressure to an appropriate volume, then added 3 times the volume of 95% ethanol to precipitate, the precipitate was washed twice with acetone and ethyl acetate, dissolved in water, and repeated by Sevag method Remove the protein, finally use 3 times the volume of 95% ethanol to precipitate, and dry at 60°C to obtain a dry matter of 2.1 g / L. Infrared spectrum at 3600-3200cm -1 、3000-2800cm -1 and 1400-1000cm -1 The presence of characteristic absorption peaks of polysaccharides indicates that the chemical properties of the substance are polysaccharides; the anthrone sulfate method and the sulfuric acid phenol method are positive, further verifying that the substance is a polysaccharide compound; there are no ultraviolet absorption peaks at 260nm and 280nm, indicating that the substance Nucle...

Embodiment 3

[0024] Example 3: Effect of exopolysaccharides on the phagocytic activity of mouse peritoneal macrophages in vitro

[0025] One day in advance, inject 1ml of 6% starch broth into the peritoneal cavity of mice to induce the production of macrophages. 6 cells / ml; then add 200 μl per well into a 96-well micro-reaction plate, adhere at 37°C for 2 hours; then wash off non-adherent cells with normal saline, add 200 μl of 0.1% neutral red normal saline solution to each well, and different dilutions 20 μl of polysaccharide solution with a concentration of 100 μg / ml, so that the final concentration of polysaccharides is 0, 25, 50, 100 μg / ml, and phagocytosis at 37 ° C for 45 minutes; after that, discard the neutral red and wash away the extracellularly adhered neutral red with physiological saline, add isopropyl hydrochloride Cells were lysed with alcohol and neutral red was dissolved, and the absorbance value at 570nm was measured to reflect the phagocytic activity of macrophages. Th...

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Abstract

the invention discloses a preparing method of extracellular polysaccharide of Bacillus subtilis and application in the tumour facet, which is characterized by the following: inhibiting tumour from growing; accelerating the growth of macrophage, leucocyte and lymph cell; fitting for health or immune adjusting drug.

Description

1. Technical field [0001] The invention belongs to the technical field of biopharmaceuticals, and in particular relates to a preparation method and application of a bacillus exopolysaccharide isolated from the marine intertidal zone. 2. Background technology [0002] Malignant tumors are frequently-occurring and common diseases that threaten human health. According to data released by the World Health Organization in 2003, there were 10 million malignant tumor patients worldwide in 2000, and 6.2 million deaths due to malignant tumors, of which about 1.8 million were in China. 10,000-2 million, accounting for 1 / 5 of the world's total, and 1.4-1.5 million deaths, accounting for 1 / 4 of the world's total. The prevention and treatment of tumor diseases has become one of the biggest challenges facing human health in the 21st century. [0003] Most of the drugs currently used for tumor treatment are cytotoxic drugs. Although they have high activity and can directly kill tumor cells...

Claims

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Application Information

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IPC IPC(8): C12P19/04C08B37/00A61K31/715A61K35/74A61P35/00A61P37/02A61K35/742
Inventor 刘常宏薛雅蓉汤城杨欢陈欣幸颖沈悦斐郑冠群
Owner NANJING UNIV
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