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Magnetic immobilized cross-linking cellulase aggregates (CLEAs), preparation method and application thereof

A technology of cellulase and aggregates, applied in the direction of immobilization on or in the inorganic carrier, on or in the organic carrier, etc., can solve the problems that the enzyme immobilization technology has not been reported, and improve the operating performance Poor, Vitality-enhancing, Ease of Handling Effects

Inactive Publication Date: 2012-03-28
SOUTH CHINA UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, fully considering the advantages and disadvantages of the two technologies, the enzyme immobilization technology combining the two technologies has not been reported.

Method used

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  • Magnetic immobilized cross-linking cellulase aggregates (CLEAs), preparation method and application thereof
  • Magnetic immobilized cross-linking cellulase aggregates (CLEAs), preparation method and application thereof
  • Magnetic immobilized cross-linking cellulase aggregates (CLEAs), preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Preparation of Magnetic Chitosan Microspheres

[0052] Take by weighing 1.2g chitosan and dissolve in 40mL volume concentration and be 3% acetic acid solution, be mixed with the chitosan acidic colloid solution that mass volume ratio is 3%, add 0.3g Fe 3 o 4 Nanoparticles, 30KHz ultrasonic dispersion for 30min to form a uniformly dispersed magnetic chitosan colloidal solution. Under slow stirring, the above-mentioned magnetic chitosan solution was added to the oil phase system (composed of 160mL liquid paraffin, 4.2g Span 80 and 2.1g Tween 80), stirred at a speed of 1000r / min for 30min, then dropped into the volume Concentration of 3% glutaraldehyde solution, the final volume concentration of glutaraldehyde is 0.2%, continue to stir at 1000r / min for 1h at 40°C, and then adjust the pH value to 11 with 3mol / L NaOH. Then continue to react at 80° C. for 2 h, and finally collect magnetic chitosan composite microspheres under magnet adsorption, wash with acetone, petroleum ...

Embodiment 2

[0054] Preparation of Magnetic Chitosan Microspheres

[0055] Take by weighing 0.8g chitosan and dissolve in 40mL volume concentration and be 2% acetic acid solution, be mixed with the chitosan acidic colloid solution that mass volume ratio is 3%, add 0.8g Fe 3 o 4 Nanoparticles, 30KHz ultrasonic dispersion for 30min to form a uniformly dispersed magnetic chitosan colloidal solution. Under slow stirring, the above-mentioned magnetic chitosan solution was added to the oil phase system (composed of 160mL liquid paraffin, 5g Span 80 and 2.5g Tween 80), stirred at a speed of 1000r / min for 30min, and the volume concentration 3% glutaraldehyde solution, the final volume concentration of glutaraldehyde is 0.2%, and the reaction is continued at 40° C. with stirring at 1000 r / min for 1 h, and then the pH value is adjusted to 11 with 3 mol / L NaOH. Then continue to react at 60° C. for 2 h, and finally collect magnetic chitosan composite microspheres under magnet adsorption, wash with a...

Embodiment 3

[0057] (1) Precipitation of cellulase aggregates

[0058] 0.45g of magnetic chitosan composite microspheres prepared in Example 1 were added to 10mL of phosphate buffer (pH7.5, 0.05mol / L) and soaked for 12h to fully swell the microspheres, and then added to 5mL with a concentration of 3mg / L after centrifugation. In mL cellulase solution (prepared with pH 7.5, 0.05mol / L phosphate buffer), add 5 mL of ammonium sulfate solution with a saturation of 60% at 25°C for 3 hours, and then add 5 mL of ammonium sulfate solution at 10°C for 2 hours. The free cellulase that is not adsorbed by the magnetic chitosan microspheres will form cellulase aggregates and precipitate on the surface of the magnetic chitosan composite microspheres.

[0059] (2) Preparation of immobilized cross-linked cellulase aggregates

[0060] Add a volume concentration of 3% glutaraldehyde solution to the precipitated enzyme solution, the final concentration of glutaraldehyde in the solution is 0.1%, and carry out ...

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Abstract

The invention discloses magnetic immobilized cross-linking cellulase aggregates (CLEAs), a preparation method and application thereof. The preparation method comprises the following steps of: first, fully swelling magnetic chitosan composite microspheres by using phosphate buffer, adding a cellulase solution, and adsorbing for 1 to 5 hours at the temperature of 10 to 25 DEG C; then, precipitating 0.5 to 2 hours at the temperature of 4 to 25 DEG C by adding a precipitating agent; and finally, adding a glutaric dialdehyde solution into a precipitated system, and performing a cross-linking reaction for 3 to 15 h at the temperature of 25 to 40 DEG C, thus obtaining the magnetic immobilized CLEAs. The preparation method provided by the invention is low in cost and is easy to operate. In the preparation method, a magnetic immobilized enzyme technology and a cross-linking enzyme aggregate technology are combined organically, not only the activity of the magnetic immobilized enzyme is improved, but also the problem of poor operating performance of the CLEAs is solved. The obtained magnetic immobilized CLEAs which have high enzyme activity and high operating performance have stable characteristics, can be widely applied by serving as biological enzyme catalysts, and are particularly suitable for a large-scale enzyme reactor.

Description

technical field [0001] The invention relates to the field of biochemical industry, in particular to a magnetically immobilized cross-linked cellulase aggregate and its preparation method and application. Background technique [0002] Cellulase is a general term for a class of multicomponent enzymes that can degrade cellulose into glucose. They work synergistically to decompose cellulose to produce oligosaccharides and cellobiose, which are finally hydrolyzed into glucose. They are widely used in food, Textile, feed, oil exploration, extraction of traditional Chinese medicine and other fields. The most popular application of cellulase is to decompose fibrous substances to produce biofuels, so as to replace the large amount of corn crops currently consumed in the production of biofuels, relieve the pressure on food consumption, and act as a sustainable substitute for petroleum. However, the fermentation activity of cellulase is low, and the hydrolysis efficiency is 1-2 orders...

Claims

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Application Information

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IPC IPC(8): C12N11/14C12N11/10C12N11/04
Inventor 李冰李琳谢晓灵李晓玺陈玲苏健裕
Owner SOUTH CHINA UNIV OF TECH
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