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A method for detecting glutathione and/or hydrogen peroxide based on a colorimetric biosensor

A colorimetric sensor, glutathione technology, applied in the field of analysis and detection, can solve the problems of high cost, low sensitivity, cumbersome steps, etc., to improve stability and accuracy, increase practicability, excellent stability and repeatability Effects of Using Properties

Active Publication Date: 2022-05-03
NANJING MEDICAL UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The purpose of the present invention is to overcome the disadvantages of low sensitivity, time-consuming detection, high cost and cumbersome steps of the existing detection method, and provide a method for preparing h-Fe with high peroxidase activity based on a one-pot method. 3 o 4 A method for detecting glutathione and / or hydrogen peroxide with @ppy's sensitive colorimetric sensor, which can detect GSH and / or H in a simple, rapid and highly sensitive manner 2 o 2

Method used

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  • A method for detecting glutathione and/or hydrogen peroxide based on a colorimetric biosensor
  • A method for detecting glutathione and/or hydrogen peroxide based on a colorimetric biosensor
  • A method for detecting glutathione and/or hydrogen peroxide based on a colorimetric biosensor

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Embodiment 1

[0050] A one-pot method for the preparation of h-Fe with high peroxidase activity 3 o 4 The method of @ppy's sensitive colorimetric sensor to detect hydrogen peroxide is as follows:

[0051] Step (a), construct magnetic composite material h-Fe 3 o 4 @ppy: first disperse 0.04g CTAB into 40mL H by continuous stirring 2 In O, a uniform and transparent surfactant solution is formed; the 60mgFe 3 o 4 NPs, 90 μL of pyrrole monomer and 40 μL of heme solution (1 mM, prepared by dissolving heme in water), were dropped into the surfactant solution respectively; after stirring for 2 hours, 2 mL of 0.2M FeCl 3 The solution was slowly dropped into the above mixture, and reacted at room temperature for 24 hours to obtain a black product, washed with ethanol and deionized water in sequence, dried in vacuum, and stored in a refrigerator at 4°C;

[0052] Step (b), magnetic composite material h-Fe 3 o 4 Catalysis of @ppy: mix 400μL of hydrogen peroxide with different concentrations wit...

Embodiment 2

[0055] A one-pot method for the preparation of h-Fe with high peroxidase activity 3 o 4 The method of @ppy's sensitive colorimetric sensor to detect hydrogen peroxide is as follows:

[0056] Step (a), construct magnetic composite material h-Fe 3 o 4 @ppy (with embodiment 1 step (a));

[0057] Step (b), magnetic composite material h-Fe 3 o 4 The catalysis of @ppy (with embodiment 1 step (b));

[0058] Step (c), construction of hydrogen peroxide colorimetric biosensor: Magnetic separation of magnetic composite material h-Fe 3 o 4 @ppy, to get the supernatant, add 20 μL of 3 M sulfuric acid, as in figure 2 A. figure 2 B, with a UV spectrophotometer, measure the maximum UV absorbance value of the reaction solution at 450nm, and establish a standard curve: A=0.0037C H2O2 +0.1812, R 2 = 0.994.

[0059] Step (d), sample detection: process the sample with unknown concentration according to step (b) and step (c), detect the maximum ultraviolet absorption value of the samp...

Embodiment 3

[0061] A one-pot method for the preparation of h-Fe with high peroxidase activity 3 o 4 The method of @ppy's sensitive colorimetric sensor to detect glutathione, the steps are as follows:

[0062] Step (a), construct magnetic composite material h-Fe 3 o 4 @ppy (same as embodiment 1 step (a)).

[0063] Step (b), construction of GSH colorimetric sensor: 400 μ L H 2 o 2 , TMB, h-Fe 3 o 4 @ppy, 400μL glutathione solution of different concentrations (prepared by dissolving glutathione in water) and sodium citrate buffer (5mM, pH 3.0) were mixed, the final reaction solution volume was 1mL, and the H in the system 2 o 2 The final concentration is 0.2mM, the final concentration of TMB is 0.2mM, h-Fe 3 o 4 The final concentration of @ppy is 60μg / mL, react at 45°C for 40 minutes; magnetically separate the magnetic composite material h-Fe 3 o 4 @ppy, add 20 μL of 3 M sulfuric acid to the supernatant and observe the color change; as Figure 4 A. Figure 4 B. Colorimetrically...

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Abstract

The invention discloses a method for detecting glutathione and / or hydrogen peroxide based on a colorimetric biosensor, comprising: (a), constructing a magnetic composite material h-Fe 3 o 4 @ppy; (b), h‑Fe 3 o 4 @ppy, TMB and different concentrations of H 2 o 2 Incubate the reaction, measure the UV absorption curve, and establish a hydrogen peroxide standard curve; (c), record the maximum UV absorption value of the sample to be tested according to step (b), and substitute it into the standard curve to obtain H 2 o 2 concentration. The present invention also includes (d), h-Fe 3 o 4 @ppy、H 2 o 2 , TMB and different concentrations of glutathione to incubate the reaction to separate h‑Fe 3 o 4 @ppy, add sulfuric acid to establish a glutathione standard curve; (e), measure the maximum UV absorption value of the sample to be tested according to step (d), and substitute it into the standard curve to obtain the GSH concentration. The invention can detect glutathione and hydrogen peroxide simply, quickly and sensitively.

Description

technical field [0001] The invention belongs to the field of analysis and detection, and relates to a colorimetric biosensor for detecting glutathione (GSH) and / or hydrogen peroxide (H 2 o 2 ) method, more specifically, relates to a one-pot method based on the preparation of high peroxidase activity h-Fe 3 o 4 A method for the detection of glutathione and / or hydrogen peroxide with sensitive colorimetric sensors by @ppy. Background technique [0002] Enzymes are widely used to catalyze reactions in cascades. Natural enzymes have limited their further applications due to their high preparation costs, strict storage requirements, and limited reaction conditions. Compared with natural enzymes, mimetic enzymes have the advantages of low cost, high stability, adjustable catalytic activity, large-scale preparation and easy surface modification. In particular, simulated nanozymes are widely used in the fields of medicine, biological detection, food and industry. In recent deca...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/78G01N21/33
CPCG01N21/78G01N21/33
Inventor 周学敏杨威翁晨园李晓芸费建文许蔚严孝强朱婉莹
Owner NANJING MEDICAL UNIV
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