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Method for simulating peroxidase by manganese dioxide nanosheet for detection of reductive biological molecules

A manganese dioxide, biomolecular technology, applied in the fields of nano biomaterials and analytical chemistry, can solve the problems of volatile inactivation, poor stability of natural enzymes, etc.

Active Publication Date: 2016-11-09
XI AN JIAOTONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Therefore, the study of simulated enzymes can solve the problems of poor stability, variability and inactivation of natural enzymes.

Method used

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  • Method for simulating peroxidase by manganese dioxide nanosheet for detection of reductive biological molecules
  • Method for simulating peroxidase by manganese dioxide nanosheet for detection of reductive biological molecules
  • Method for simulating peroxidase by manganese dioxide nanosheet for detection of reductive biological molecules

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Experimental program
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Effect test

Embodiment 1

[0038] Based on the method of the present invention to detect reducing biomolecule-glutathione GSH

[0039] (1) Add 100μL of GSH solution of known concentration to 100μL of manganese dioxide nanosheet solution and 0.5mL of NaAc-HAc (0.2M) solution for 10 minutes at room temperature;

[0040] (2) Add the mixed solution in (1) to 200μL TMB solution (1.0mM) and react for 5min at room temperature;

[0041] (3) Detect the absorbance value of the solution in (2) at 650nm with a dual-beam UV-visible spectrophotometer, and mark it as A. Use ultrapure water instead of GSH solution as a blank experiment, and its absorbance value at 650nm is recorded as A 0 .

[0042] (4) With absorbance net signal △A=A 0 -A is used as a quantitative indicator to draw a GSH standard curve to obtain a linear equation (refer to figure 2 ) And detect the linear range (1μM-15μM) and detection limit (0.3μM) of GSH.

[0043] (5) Repeat the above steps (1), (2), (3) of the GSH solution to be tested to obtain the A valu...

Embodiment 2

[0046] Based on the method of the present invention to detect reducing biomolecules-ascorbic acid AA

[0047] (1) Add 100μL of AA solution of known concentration to 100μL of manganese dioxide nanosheet solution and 0.5mL of NaAc-HAc (0.2M) solution for 10 minutes at room temperature;

[0048] (2) Add the mixed solution in (1) to 200μL TMB solution (1.0mM) and react for 5min at room temperature;

[0049] (3) Detect the absorbance value of the solution in (2) at 650nm with a dual-beam UV-visible spectrophotometer, and mark it as A. Use ultrapure water instead of AA solution as a blank experiment, and its absorbance value at 650nm is recorded as A 0 .

[0050] (4) With absorbance net signal △A=A 0 -A is used as a quantitative indicator to draw the AA standard curve to obtain a linear equation (refer to image 3 ) And detection AA linear range (3-100μM) and detection limit (0.8μM).

[0051] (5) Repeat the above steps (1), (2), (3) of the AA solution to be tested to obtain the A value of A...

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Abstract

The invention relates to a method for simulating peroxidase by manganese dioxide nanosheet for detection of reductive biological molecules. The peroxidase simulated by manganese dioxide nanosheet can perform catalytic oxidation on substrates of 3,3',5,5'-tetramethyl benzidine TMB, 2,2-azino-di(3-ethyl-benzothiazoles-6-sulfonic acid) diammonium salt ABTS and o-phenylenediamine OPD, and changes the color from colorless to blue, green and orange respectively, at the same time, the manganese dioxide nanosheet can sensitively and selectively perform an oxidation reduction reaction with reductive biological molecules such as glutathione and ascorbic acid, oxidation product concentration of the substrates such as TMB, ABTS and OPD is changed, and then, the reductive biological molecules such as glutathione and ascorbic acid are subjected to quantitative determination through a colorimetric analysis method. The method has the characteristics of simple operation, high sensitivity, good reappearance and high selectivity; a detection linear scope of glutathione is 1-15 [mu]M, the detection limit is 0.3 [mu]M; the detection linear scope of ascorbic acid is 3-100 [mu]M, the detection limit is 0.8 [mu]M; and the method can be used for detecting various phenolic compounds.

Description

Technical field [0001] The invention belongs to the technical field of nano-biological materials and analytical chemistry, and particularly relates to a method for detecting reducing biomolecules by manganese dioxide nano-sheet simulating oxidase. Background technique [0002] Enzymes are macromolecular substances with biocatalytic function, which have strong substrate specificity and high catalytic efficiency under mild conditions. However, the extraction and purification process of natural enzymes is time-consuming and complicated, high cost, and poor stability. At the same time, the catalytic activity of natural enzymes is easily denatured and inactivated by environmental influences, which severely restricts its further application. Mimic enzymes are substances with similar functions to natural enzymes, and can still maintain high stability under harsher conditions. Therefore, the research on mimic enzymes can solve the problems of poor stability and volatile inactivation of ...

Claims

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Application Information

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IPC IPC(8): G01N31/10G01N21/78G01N21/33
CPCG01N21/33G01N21/78G01N31/10
Inventor 孟令杰刘晶党东锋曹瑞军
Owner XI AN JIAOTONG UNIV
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