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Naturally symbiotic mixed culture and method for producing coumaric acid esterase by degrading straws by using naturally symbiotic mixed culture

A mixed culture, coumaric acid ester technology, applied in the directions of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve problems such as the inability to achieve high enzymatic activity of coumaric acid esterase, and prevent bacteria from being affected. Pollution, ease of promotion, and the effect of improving efficiency

Active Publication Date: 2021-03-26
GANSU ACAD OF SCI INST OF BIOLOGY
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Problems solved by technology

The inventor first studied the anaerobic fermentation of the mixed culture of grazing yak rumen anaerobic fungi and methanogens with wheat straw, corn straw and rice straw as substrates (Wei Yaqin. Yak rumen anaerobic fungi mixed with methanogens) Research on the diversity of cultures and their fiber degradation characteristics [D]. 2016.), by detecting gas production, polysaccharide hydrolase activity, various esterase activities, dry matter degradation rate, phenolic acid release, methane and acetic acid production Evaluate the effectiveness of the co-culture of anaerobic fungi and methanogens in degrading straw, among which the co-cultures of anaerobic fungi and methanogens that efficiently degrade the above three straws are N.frontalis Yak16+M.ruminantium, Piromyces Yak18+M.ruminantium and O.joyonii Yak7+M.ruminantium respectively degrades the three major straws, and the coumaric esterase activity produced is lower than 5.0mU, which cannot achieve the effect of producing high enzyme activity coumaric esterase

Method used

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  • Naturally symbiotic mixed culture and method for producing coumaric acid esterase by degrading straws by using naturally symbiotic mixed culture

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Experimental program
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Effect test

Embodiment 1

[0037] Embodiment 1, strain screening and identification

[0038] 1. Collection of yak rumen juice

[0039] The research site is located in the grazing pasture of Xinghai County, Hainan Tibetan Autonomous Region, Qinghai Province. The annual average temperature is -1-1.3°C, the highest temperature is 26°C, the lowest temperature is -30°C, and the altitude is 3100-4300m. It belongs to the cold plateau climate type with drastic temperature changes. With strong sunshine and abundant rainfall, alpine meadow is the main grassland type. Wild pastures mainly include fescue, Kobresia, Kobresia Tibetan, and Kobresia tenifolia, which are the main dominant species in this area.

[0040] 20 healthy male yaks (age 4-5 years old, body weight 250±50kg) were used as experimental animals in the whole grazing yaks that ate natural forage and did not supplementary feeding. Insert one end of the stainless steel gastric tube into the rumen through the opener, and the other end is connected to an...

Embodiment 2

[0074] Embodiment two, the preparation of the mixed culture YakQH5 bacterial agent of anaerobic fungus (Neocallimastix frontalis) and methanogen (Methanobrevibacter gottschalkii)

[0075] Pipette 1 mL of YakQH5, a mixed culture of anaerobic fungi (Neocallimastix frontalis) and methanogens (Methanobrevibacter gottschalkii), into 9 mL of anaerobic medium with air-dried and crushed wheat straw as a substrate in a Heinz anaerobic tube, and add 0.1 mL of compound Antibiotics (1600IU / mL penicillin and 2000IU / mL streptomycin sulfate), anaerobic culture at 39°C for 72 hours, the growth peak will be reached, and the fermentation broth is a highly active bacterial agent at this time.

Embodiment 3

[0076] Embodiment 3, mixed culture YakQH5 anaerobic fermentation degrades straw to produce coumaric acid esterase

[0077] 45mL of liquid basic medium was placed in a 100mL anaerobic fermentation bottle, and 0.5g of dried and pulverized wheat straw, corn straw, rice straw, oat straw and sorghum straw were used as substrates. Deoxygenated, autoclaved. Draw 5mL of the mixed culture YakQH5 obtained in Example 1 and inoculate them into the above-mentioned anaerobic medium with stalks respectively, and add 0.5mL compound antibiotics (1600IU / mL penicillin and 2000IU / mL streptomycin sulfate) at the same time , 39°C anaerobic culture for 5 days. A total of 3 parallel experiments were set up, and the activity of coumaric acid esterase in the anaerobic bottle was measured every 24 hours.

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Abstract

The invention relates to the field of renewable energy sources of biotechnology, in particular to a naturally symbiotic mixed culture YakQH5 composed of anaerobic fungi (Neocallimastix frontalis) andmethane bacteria (Methanobrevibacter gottschalkii). The mixed culture is preserved in china general microbiological culture collection center on 09 March 2020, the preservation number is CGMCC No.19299, and the mixed culture YakQH5 can be used for fermenting straws to produce coumaric acid esterase. The invention specifically discloses a fermentation method by using the mixed culture. According tothe mixed culture and the method, the activity of coumaric acid esterase generated by degrading sorghum straws can reach 20.5 mU, and composite antibiotics are added in a fermentation process, so that a mixed culture system can be prevented from being polluted by bacteria, the anaerobic fermentation efficiency is improved, and the important industrial application value is achieved.

Description

technical field [0001] The invention relates to the field of biotechnology renewable energy, in particular to a naturally symbiotic natural mixed culture composed of anaerobic fungi and methane bacteria and a method for producing coumaric acid esterase by degrading straw. Background technique [0002] Crop straw is the main source of agricultural waste non-point source pollution, and the annual output of crop straw in my country is high. Conventional treatment methods such as open burning and burial will pollute the environment and cause a lot of energy waste. However, a large number of straw resources in our country are in the state of high consumption, high pollution and low utilization rate. Lignocellulose is the main component of straw, and lignocellulose is mainly composed of cellulose, hemicellulose and lignin. The covalent bonds between lignin and hemicellulose embed cellulose molecules in it, and the ether bonds and carbon-carbon bonds in lignin form macromolecular ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N1/14C12N9/18C12R1/01C12R1/645
CPCC12N1/14C12N1/20C12N9/18
Inventor 魏亚琴李伟王治业
Owner GANSU ACAD OF SCI INST OF BIOLOGY
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