Application of laccase in protein gel decoloration

A technology of protein gel and laccase, which is applied in the application field of laccase in the decolorization of protein gel, can solve the problems that the decolorization of dye wastewater is limited to the aqueous environment, a large amount of organic dye waste liquid, and long decolorization time, etc. Achieve good decolorization effect, reduce pollution, and operate green and environmental protection

Inactive Publication Date: 2015-05-20
FUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

To obtain a better decolorization effect, a longer decolorization time (overnight) is required, and the decolorization solution must be replaced several times, which will generate a large amou

Method used

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  • Application of laccase in protein gel decoloration
  • Application of laccase in protein gel decoloration
  • Application of laccase in protein gel decoloration

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Embodiment 1 HYB07 laccase decolorizes different dyes

[0043] (1) HYB07 laccase was used for 12 different dyes (Coomassie Brilliant Blue R250, Methyl Orange, Methyl Red, Basic Fuchsin, Bromothymol Blue, Neutral Red, Crystal Violet, Acid Fuchsin, Congo Red , bromocresol green, methylene blue, rose bengal B) decolorization. Decolorization system 2 mL, containing 50 mM citric acid-disodium hydrogen phosphate buffer (pH 6.0), dye (25-200 mg / L) and enzyme (2 U / mL), decolorized at room temperature for 24 h. From figure 1 It can be seen that after laccase treatment, the decolorization rate of Coomassie Brilliant Blue is the highest, reaching 92.37%.

[0044] (2) Scan the Coomassie Brilliant Blue R250 solution before and after laccase decolorization at full wavelength. From figure 2 It can be seen that the absorption peak of the dye decreases after decolorization.

Embodiment 2

[0045] Example 2 Decolorization of Protein Gel Using Laccase / Mediator System

[0046] (1) Effect of Mediator Types on Protein Gel Decolorization

[0047] Different laccase mediators including ACE (20 μM), SYA (20 μM), HBT (20 μM), ABTS (2 μM) and SYD (20 μM ) on the decolorization of protein gels. From image 3 It can be seen that ABTS has the best effect as a mediator.

[0048] (2) Effect of temperature on protein gel decolorization

[0049] Under the conditions of pH 7.0, enzyme concentration 20 U / mL, and ABTS 2 μM, the effects of different temperatures (20 ℃, 25 ℃, 30 ℃, 35 ℃, 40 ℃, 45 ℃) on the decolorization of protein gels were studied. From Figure 4 It can be seen that 40 °C is the optimum temperature for decolorization.

[0050] (3) Effect of laccase activity on protein gel decolorization

[0051] Under the conditions of pH 7.0 and temperature 40 ℃, the effect of laccase activity (3 U / mL, 6 U / mL, 9 U / mL, 12 U / mL, 15 U / mL, 18 U / mL) on protein gel was studied. ...

Embodiment 3

[0056] Example 3 The detection sensitivity of laccase / mediator decolorization system

[0057] (1) Decolorize for 2 hours under the conditions of pH 7.0, temperature 40 ℃, laccase activity 15 U / mL, and ABTS 2 μM, and detect the detection sensitivity of the laccase / mediator decolorization system. From Figure 8 It can be seen that BSA can still be detected as low as 10 ng.

[0058] (2) Compare the effects of different decolorization methods. From Figure 9 It can be seen that the decolorization effect of the laccase / mediator decolorization system is the best, and there is little difference in the preparation of the decolorization solution with buffer or distilled water. Boiling with water directly for decolorization may easily cause loss of protein signal. Decolorization with water or buffer solution for 2 h at 40°C resulted in a higher background, indicating that laccase not only degrades R-250 in the solution, but also accelerates the decolorization of protein gels. Prote...

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Abstract

The invention belongs to the technical field of biology, more particularly relates to an application of laccase in protein gel decoloration, and specifically relates to an application of laccase in decoloration of protein gel electrophoresis after conventional dyeing. The application aims at decoloring dyed protein gels by using laccase; and the application is environment-friendly, rapid, simple and convenient, does not need organic solvents, and can be used for saving energy and reducing the pollution caused by dye wastewater. According to the application of decoloring the protein gels by using laccase, disclosed by the invention, rapid decoloration can be achieved (a low-background clear protein band can be obtained within 2 hours), simplicity and convenience in operations can be achieved (boiling or decoloring liquid replacement is not needed), and environmental protection can be achieved (organic solvents such as methanol and acetic acid required by conventional decoloring liquid are not used, dye degradation can be achieved while decoloring, and no waste liquid treatment problems and cost can be produced). According to the application, by optimizing factors such as laccase concentration, medium types and concentration and decoloring time, good decoloring effects can be achieved in a relatively short time, and simplicity and convenience in operations and environmental protection can be achieved.

Description

technical field [0001] The invention belongs to the field of biotechnology, and more specifically relates to the application of laccase in protein gel decolorization. Background technique [0002] Laccase (laccase, EC 1.10.3.2) is a copper-containing polyphenol oxidase that widely exists in higher plants, fungi and insects, among which white-rot fungi are considered to be the most efficient producers of laccase. Laccase is a green enzyme. Through oxidation reaction, laccase causes the substrate to lose electrons to form free radicals, and the lost electrons are transferred to molecular oxygen to form water. Laccase has a wide range of substrates and can catalyze the oxidation of various aromatic compounds, but the effect of laccase on different dyes is different, depending on the properties of laccase and the structure of the dye. Small molecule laccase mediators, such as ABTS (2,2-azinobis- (3-ethylbenzothiazoline-6-sulfonic acid), HBT (1-hydroxybenzotriazole), syringaldeh...

Claims

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Application Information

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IPC IPC(8): G01N27/447
Inventor 杨捷杨晓丹林娟叶秀云
Owner FUZHOU UNIV
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