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Method for preparing immobilized laccase in electrospun fiber membrane

A technology for immobilizing laccase and electrospinning fibers, which is applied in the directions of immobilization on/in organic carriers, fiber processing, textiles and papermaking, etc. problems, to achieve the effects of good stability and reusability, protection of enzyme activity, and high enzyme loading capacity

Inactive Publication Date: 2010-03-17
BEIJING NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In this case, although the fiber membrane has a large specific surface area, the enzyme can only be immobilized on the surface of the fiber membrane in a single layer, the enzyme load is relatively low, and the enzyme falls off seriously during the reaction, resulting in poor reusability of the enzyme; if Directly increase the enzyme loading by chemical cross-linking method, the enzyme activity will be greatly affected
However, there is no research on the immobilization of laccase by this method.

Method used

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  • Method for preparing immobilized laccase in electrospun fiber membrane
  • Method for preparing immobilized laccase in electrospun fiber membrane

Examples

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preparation example Construction

[0028] The preparation steps of the enzyme-loaded electrospun fiber membrane include:

[0029] 1) Dissolve polylactic acid-caprolactone copolymer (P(LA / CL), 7 / 3) particles with a molecular weight of 100,000 in dichloromethane, add block copolymer surfactant F108, mix evenly and stir Obtain copolymer mixed gel in 2 hours, wherein the consumption of block copolymer F108 is 10% of polylactic acid-caprolactone copolymer (P(LA / CL), 7 / 3) weight;

[0030] 2) Weigh 10 mg of Versicolor versicolor laccase and dissolve it in 1 ml of phosphate buffer solution with a pH value of 3.5, and shake until completely dissolved. In order to observe the distribution of laccase in the electrospun fiber using laser confocal scanning microscope, the concentration of 10 mg / ml of laccase solution;

[0031]3) Add the laccase solution obtained in step 2) into the copolymer mixed gel, and mix on a vortex mixer for 20 minutes until a homogeneous emulsion is formed, wherein the volume ratio of the enzyme ...

Embodiment 1

[0046] 0.6 g of poly(lactic acid-caprolactone) copolymer (P(LA / CL), 7 / 3) was dissolved in 12 g of dichloromethane, then 0.06 g of block copolymer surfactant F108 was added, mixed evenly and stirred for 2 hours A copolymer mixed gel was obtained. Weigh 10 mg of Versicolor versicolor laccase and dissolve it in 1 ml of phosphate buffered saline solution with a pH value of 3.5, and shake until completely dissolved. Add 0.1 ml of the above laccase solution or 10 mg / ml laccase solution labeled with fluorescein isothiocyanate (FITC) into the copolymer mixed gel, and mix on a vortex mixer for 20 minutes until a homogeneous gel is formed. quality lotion. The emulsion is then introduced into a high-voltage electrospinning device to be spun. Adjust the high-voltage power supply voltage to 10 kV and the receiving distance to 14 cm to obtain a stable and continuous jet stream, and collect the fiber product on the receiving plate covered with aluminum foil. During the whole process of sp...

Embodiment 2

[0049] 0.6 g of poly(lactic acid-caprolactone) copolymer (P(LA / CL), 7 / 3) was dissolved in 12 g of dichloromethane, then 0.06 g of block copolymer surfactant F108 was added, mixed evenly and stirred for 2 hours A copolymer mixed gel was obtained. Weigh 10 mg of Versicolor versicolor laccase and dissolve it in 1 ml of phosphate buffered saline solution with a pH value of 3.5, and shake until completely dissolved. Take 0.15 ml of the above-mentioned laccase solution or a 10 mg / ml laccase solution labeled with fluorescein isothiocyanate (FITC) and add it to the copolymer mixed gel, and mix on a vortex mixer for 20 minutes until a homogeneous gel is formed. quality lotion. The emulsion is then introduced into a high-voltage electrospinning device to be spun. Adjust the voltage of the high-voltage power supply to 11 kV, and the receiving distance is 14 cm to obtain a stable and continuous jet stream, and collect the fiber product on the receiving plate covered with aluminum foil. ...

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Abstract

The invention belongs to the enzyme immobilization field and in particular relates to a method for preparing immobilized laccase in electrospun fiber membrane by utilizing emulsion electrospinning technology. The method is characterized by in-situ embedding and immobilizing the laccase in the electrospun fiber by the emulsion electrospinning technology. The carrier material of the immobilized laccase is polylactic acid-caprolactone copolymer (P(LA / CL), 7 / 3, namely the ratio of polylactic acid to polycaprolactone during synthesis) with good biocompatibility. The method specifically comprises two steps of preparing the enzyme-carrying electrospun fiber membrane and immobilizing the enzyme-carrying electrospun fiber membrane by crosslinking. The first step is as follows: first preparing polymer solution and laccase solution into homogeneous emulsion, then carrying out electrospinning on the emulsion and obtaining the enzyme-carrying electrospun fiber membrane a few hours later. The secondstep is as follows: exposing the membrane in glutaradehyde saturated vapor to be crosslinked, thus obtaining the immobilized laccase in the electrospun fiber membrane. The invention provides the new,simple and efficient method preparing the immobilized laccase, and the immobilized laccase obtained by the method has high catalytic activity and good stability and features low preparation cost.

Description

technical field [0001] The invention belongs to the field of enzyme immobilization, and specifically relates to a method for preparing polylactic acid-caprolactone copolymer (P(LA / CL), 7 / 3) electrospun fiber membrane to immobilize laccase by using emulsion electrospinning technology. Background technique [0002] Laccase (Laccase, EC 1.10.3.2) is a copper-containing polyphenol oxidase discovered in 1883 by Japanese scholars from the secretion of Lac sumac. With the deepening of research, it has been confirmed that laccase exists in many organisms, including higher plants, microorganisms (mainly fungi, partly bacteria) and insects. Laccases generally contain four copper ions, which can be divided into three types according to their spectral and magnetic characteristics: Type I Cu 2+ and type II Cu 2+ One each, single-electron acceptor, paramagnetic; Type III Cu 4+ Two, which are two-electron acceptors, are diamagnetic. Type I Cu 2+ Blue in color with a characteristic abs...

Claims

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Application Information

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IPC IPC(8): C12N11/08C12N11/04D01D5/00
Inventor 牛军峰代云容刘佳殷立峰沈珍瑶
Owner BEIJING NORMAL UNIVERSITY
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