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Nanometer grain preparation method of bionic SiO2 fixing Beta-glucuronide enzyme

A technology of aldolidase and silicon dioxide, which is applied in the direction of immobilization on or in the inorganic carrier, can solve the problems of activity decline and inactivation, and achieve good immobilization ability, mild preparation conditions and high activity maintenance rate Effect

Inactive Publication Date: 2008-05-21
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although this method is simple and easy to implement, the alcohol by-products produced during the process have an adverse effect on the maintenance of the activity of the biological enzyme, resulting in a decrease in activity or even complete inactivation.

Method used

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  • Nanometer grain preparation method of bionic SiO2 fixing Beta-glucuronide enzyme
  • Nanometer grain preparation method of bionic SiO2 fixing Beta-glucuronide enzyme

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] Accurately weigh 1.0mg of β-glucuronidase, dissolve it in 0.05mol / L tris-hydrochloric acid (Tris-HCl) buffer solution, and dilute to 10mL to obtain 0.1mg / mL β-glucuronidase Acidase solution. Weigh 300 mg of protamine sulfate, dissolve it in 0.05 mol / L Tris-HCl buffer solution, and dilute to 20 mL to obtain a 15 mg / mL protamine sulfate solution. Sodium silicate was dissolved in deionized water, and the pH of the solution was adjusted to 7.0 with 2mol / L HCl to prepare a 10mmol / L sodium silicate solution.

[0017] Take 0.5ml each of β-glucuronidase solution and protamine sulfate solution, mix well and add to 7mL freshly prepared sodium silicate solution, let it stand for 5min, set the centrifuge speed to 4500r / min, and centrifuge for 10min . Pour out the supernatant, wash with deionized water, centrifuge for 10 min, and pour out the supernatant. Wash repeatedly 2-3 times. The content of β-glucuronidase in the supernatant was analyzed through the conversion reaction of ...

Embodiment 2

[0021] Accurately weigh 1.0mg of β-glucuronidase, dissolve it in 0.05mol / L tris-hydrochloric acid (Tris-HCl) buffer solution, and dilute to 10mL to obtain 0.1mg / mL β-glucuronidase Acidase solution. Weigh 40 mg of protamine sulfate, dissolve it in 0.05 mol / L Tris-HCl buffer solution, and dilute to 20 mL to obtain a 2 mg / mL protamine sulfate solution. Sodium silicate was dissolved in deionized water, and the pH of the solution was adjusted to 7.0 with 2mol / L HCl to prepare a 30mmol / L sodium silicate solution.

[0022] Take 0.5mL of β-glucuronidase solution and protamine sulfate solution, mix well and add to 7mL freshly prepared sodium silicate solution, let it stand for 10min, set the centrifuge speed to 4500r / min, and centrifuge for 10min . Pour out the supernatant, wash with deionized water, centrifuge for 10 min, and pour out the supernatant. Wash repeatedly 2-3 times. The content of β-glucuronidase in the supernatant was analyzed through the conversion reaction of baical...

Embodiment 3

[0026] Accurately weigh 1.0 mg of β-glucuronidase, dissolve it in deionized water, and dilute to 10 ml to obtain 0.1 mg / ml β-glucuronidase solution. Weigh 300 mg of protamine sulfate, dissolve it in deionized water, and dilute to 20 ml to obtain a 15 mg / ml protamine sulfate solution. Sodium silicate was dissolved in deionized water, and the pH of the solution was adjusted to 7.0 with 2mol / L HCl to prepare a 0.03mmol / L sodium silicate solution.

[0027] Take 0.5mL of β-glucuronidase solution and protamine sulfate solution, mix well and add to 10mL freshly prepared sodium silicate solution, let it stand for 5min, set the centrifuge speed to 4500r / min, and centrifuge for 10min . Pour out the supernatant, wash with deionized water, centrifuge for 10 min, and pour out the supernatant. Wash repeatedly 2-3 times. The content of β-glucuronidase in the supernatant was analyzed through the conversion reaction of baicalin, and it was found that there was no β-glucuronidase in the supe...

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Abstract

The invention discloses a method for preparing nanoparticles of biomimetic silicon dioxide immobilized β-glucuronidase. The method comprises the steps of dissolving protamine sulfate in tris-hydrochloric acid buffer solution or deionized water to prepare a protamine sulfate solution, and dissolving β-glucuronidase in tris-hydroxymethylaminomethane - In hydrochloric acid buffer solution or deionized water, prepare β-glucuronidase solution, mix protamine sulfate solution with β-glucuronidase solution to obtain a mixed solution; dissolve sodium silicate in deionized water or in saline, and adjust the pH value to the sodium silicate solution; add the mixed solution to the sodium silicate solution, stand still, separate, remove the supernatant, wash, and freeze-dry to obtain biomimetic silicon dioxide immobilized β-glucuronic acid Glucosidase nanoparticles. The invention has the advantages of simple preparation process, high activity maintenance rate of the immobilized β-glucuronidase, and good repeated use stability.

Description

technical field [0001] The invention relates to a method for preparing nanoparticles of biomimetic silicon dioxide immobilized β-glucuronidase, which belongs to the enzyme immobilization technology. Background technique [0002] Silica gel is a commonly used carrier for immobilized enzymes. At present, silica is used as a carrier to immobilize enzymes mainly by embedding or adsorption. The adsorption method is simple to operate, but the enzyme is easy to fall off, and the reusability is poor. The high loading rate of the embedding method is conducive to maintaining the structural integrity of the enzyme, thereby improving the activity maintenance rate of the enzyme, and the silica has a mesoporous structure, which is conducive to the transfer of reactants and products. [0003] At present, the sol-gel (sol-gel) method is generally used, using acid or alkali as a catalyst, using the hydrolysis and polycondensation of sodium silicate or organosilane precursors to synthesize ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N11/14
Inventor 吴洪姜忠义李林张羽飞
Owner TIANJIN UNIV
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