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Method for producing heat-resistant superoxide dismutase (SOD) by using thermobifida fusca

A thermosporidium, superoxide technology, applied in the field of bioengineering, can solve the problems of poor thermal stability of natural SOD, cumbersome extraction process, easy to be deactivated by heat, etc. heat labile effect

Inactive Publication Date: 2015-06-10
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

According to current reports, SOD can be produced by microbial fermentation, but mainly intracellular enzymes (Diao Zhimin, Yao Yinxia, ​​Zhang Wenjing. Research on yeast SOD fermentation conditions [J]. Journal of Qinghai Normal University (Natural Science Edition), 2002,4 :48-51; Yang Mingyan, Guo Ailian, Shen Jian, et al. Mutation breeding and fermentation conditions of high-yield SOD yeast [J]. Food Science, 2005,26(10):147-150), the extraction process is relatively cumbersome
[0003] Natural SOD has poor thermal stability and is easily deactivated by heat during processing and use

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0014] Inoculate the slant strain into a 250ml conical flask containing 50ml of culture medium, the culture medium consists of: glucose 20g / l, beef extract 10g / l, peptone 10g / l, NH 4 NO 3 10g / l, corn steep liquor 12g / l, natural pH. 45℃, fermented for 72 hours. After the fermentation, centrifuge 50 ml of the fermentation broth at 10,000 rpm to remove the bacterial cells. The supernatant crude enzyme solution was used to determine the pH and temperature parameters of the enzyme. Take 0.5ml of the supernatant crude enzyme solution and put it in 3.5ml of citric acid-phosphate buffer with pH 4, 5, 6, 7, and 8, respectively. At 45 °C, the SOD activity was determined to be 2200, 2300, 2500, 2300, 2200U / mL. Therefore, the optimum reaction pH of this SOD is 6. Take 0.5ml of the supernatant crude enzyme solution and mix it with 3.5ml of pH 6 citric acid-phosphate buffer. The enzyme activities were measured at 25, 35, 45, 55, and 65°C to be 2100, 2300, 2500, and 2000, respectively. ...

Embodiment 2

[0016] Inoculate the slant strain into a 250ml conical flask containing 50ml of seed medium, the seed medium consists of: glucose 20g / l, beef extract 10g / l, peptone 10g / l, NH 4 NO 3 10g / l, corn steep liquor powder 12g / l, natural pH, sterilized at 121 for 20min, shaker rotation speed 150rpm, cultured at 45°C for 48 hours. There are 10 identical flasks in this process, and a total of 500ml of seed solution is obtained.

[0017] Inoculate 500ml of seed liquid into a 50L fermentation tank, and the fermentation medium consists of: glucose 20g / l, beef extract 10g / l, peptone 3g / l, NH 4 NO 3 10g / l, corn steep liquor powder 6g / l, natural pH, fermentation temperature of 45°C, liquid filling volume of 70%, and ventilation volume of 1:1.5v / v / m. The fermentation time is 3 days. Under this condition, the final SOD activity in the fermentation broth was 2450 U / ml.

Embodiment 3

[0019] The cultured 1L seed solution was inoculated into a 10L fermenter. The composition of seeds and fermentation medium are: fructose 30g / l, beef extract 10g / l, corn steep liquor powder 20g / l, K 2 HPO 4 4g / l, MgSO 4 7H 2 0.5g / l, pH natural. The fermentation temperature was 20°C, the liquid filling volume was 70%, the ventilation rate was 1:0.3 v / v / m, and the fermentation time was 5 days. Under these conditions, the final SOD activity in the fermentation broth was 2500 U / ml.

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Abstract

The invention relates to a method for producing superoxide dismutase (SOD) by using thermobifida fusca and belongs to the technical field of bioengineering. According to the method, SOD is produced from thermobifida fusca under the condition of liquid-state fermentation and is an ectoenzyme. The yield of heat-resistant SOD can reach 2500U / ml to the maximum, so that the method has a good industrial application prospect.

Description

technical field [0001] The production of superoxide dismutase (SOD) by using Thermosporum tanus belongs to the technical field of bioengineering. The invention relates to the use of strain Thermobifida fusca with the preservation number of ATCC 66938 to obtain high temperature resistant SOD through liquid fermentation. Background technique [0002] Superoxide dismutase (superoxide dismutase, SOD) has a good defense against oxygen toxicity and can enhance the body's ability to resist radiation damage. In recent years, it has become a novel anti-inflammatory drug for the treatment of inflammation, tumor and autoimmune diseases; an additive for sun protection, anti-aging and anti-inflammatory in the cosmetic and food industries, and a biochemical reagent for probes of certain diseases. SOD widely exists in animals, plants, all aerobic microorganisms and a few anaerobic microorganisms. At present, SOD is mainly extracted from animal blood (such as pig blood or bovine blood) (Y...

Claims

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Application Information

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IPC IPC(8): C12N9/02C12R1/01
CPCC12N9/0089C12Y115/01001
Inventor 丁彦蕊牟兆琳王雪芹饶榕
Owner JIANGNAN UNIV
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