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Method for rapidly determining xylanase in fermentation liquor

A xylanase and rapid assay technology, applied in the biological field, can solve the problems of slow assay speed and achieve the effect of reducing human error

Inactive Publication Date: 2014-07-02
NORTHWEST A & F UNIV
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  • Abstract
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  • Claims
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Problems solved by technology

Most of the research reports on xylanase at home and abroad adopt this method, but its disadvantage is that the determination speed is slow, and the spectrophotometer can only analyze one sample at a time.

Method used

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  • Method for rapidly determining xylanase in fermentation liquor
  • Method for rapidly determining xylanase in fermentation liquor

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Embodiment

[0016] A rapid assay method for xylanase in fermentation broth, prepare xylose standard solution, make and measure xylanase activity standard curve, the result is as follows figure 1 shown. The curve relationship is: Y=95.281X+3.7833(R 2 =0.9952), where Y is the xylan content (μg), and X is the absorbance.

[0017] Specifically, four roughages of wheat straw, corn straw, rice straw and Leymus chinensis were selected as substrates. Weigh 80 mg of substrate and 9.0 mL of culture medium into a Hungate culture tube, and sterilize with damp heat at 121°C for 20 min. The fungi were inoculated under anaerobic conditions, and the inoculated medium was continuously cultured at 39°C for 5 days, with 4 replicates for each substrate. At the same time, there are substrate blanks and controls for each substrate during incubation. After culturing for 5 days and centrifuging at 1000×g for 10 min at 4°C, the xylanase activity in the fermentation broth in the tube was measured. The results...

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Abstract

The invention relates to a method for rapidly determining xylanase in fermentation liquor. The method comprises the following steps: centrifuging 1000*g of fermentation liquor for 10 minutes, taking the supernatant, determining the activity of xylanase, and preheating properly diluted supernatant, 10g / L of xylanase solution and 50mM of sodium phosphate buffer solution with the pH value of 7.0 at the temperature of 39 DEG C for 15-30 minutes, adding 75micronL of sodium phosphate buffer solution and 50 micronL of xylanase solution into 75micronL of diluted supernatant, reacting at the temperature of 39 DEG C for 15-30 minutes, adding 300micronL of DNS solution and ending the reaction; heating in a boiling water bath for 5 minutes, and cooling to room temperature; putting 200-300 micronL of solution on a dried and clean 96-pore ELISA plate, detecting a light absorption value at 530-550nm by using a full-automatic microplate reader, and calculating the activity of xylanase according to a standard curve of xylose. According to the technology, a method for measuring the xylanase in the fermentation liquor is revised, the activity of the xylanase can be rapidly, greatly and accurately measured, and personal errors are reduced. Meanwhile, fewer samples can be detected.

Description

technical field [0001] The invention relates to a rapid assay method for xylanase in fermentation broth, belonging to the field of biotechnology. Background technique [0002] Xylan is a poly-five-carbon sugar, an important component of plant hemicellulose, which accounts for one-third of the total plant carbohydrates, and is the second most abundant regenerative plant after cellulose in nature. material resources. Although xylan is a type of hemicellulose that widely exists in plants, since xylan cannot be digested and absorbed in the digestive tract of animals, it also increases the viscosity of chyme and hinders the absorption of nutrients, especially fat and protein. Digestion and absorption, and reduce the conversion rate of feed, has strong anti-nutritional properties, thus limiting the application of many feeds. Xylanase can degrade xylan into xylooligosaccharides and xylose successively, thereby eliminating the anti-nutritional effect of xylan, improving the utiliz...

Claims

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Application Information

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IPC IPC(8): G01N21/31
Inventor 曹阳春姚军虎王腊梅魏筱诗李宗军孙菲菲刘凯曹志昂王卫涛李飞
Owner NORTHWEST A & F UNIV
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