Preparation method and application of enzyme-integrated rare earth coordination polymer

A technology of coordination polymers and rare earths, which is applied in the preparation and application of fluorescent probes, can solve the problems of nano-probes in their infancy, improve detection sensitivity and selectivity, improve catalytic activity and stability, and eliminate interference Effect

Pending Publication Date: 2021-11-09
JIANGXI NORMAL UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Even so, most of the current research focuses on using coordination polymers as carriers to enhance the catalytic p

Method used

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  • Preparation method and application of enzyme-integrated rare earth coordination polymer
  • Preparation method and application of enzyme-integrated rare earth coordination polymer
  • Preparation method and application of enzyme-integrated rare earth coordination polymer

Examples

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Example Embodiment

[0027] A method of preparing an enzyme integrated rare earth coordination polymer, combined with a rare earth coordination polymer having fluorescent properties, specifically comprising the steps:

[0028] (1) Put metal salt nitrate (TB (NO 3 ) 3 Aqueous solution was pre-agit of 20 μg / mL to 25 μg / ml of superoxide dismalization enzyme (SOD).

[0029] (2) The aqueous solution of ligand triphosphate (ATP) was added. After 3 h, it was centrifuged at a rotational speed of 13000 rpm for 10 min, and the solid product was separated, wherein the metal salt nitrate (TB (NO) 3 ) 3 ) The molar ratio of adenosine adenosine (ATP) with ligand is 8: 3;

[0030] (3) The solid product separated in step (2) first wash 3 times with water, and then the resulting final solid product was separated after 12 h, and then the obtained enzyme was incorporated into a rare earth. Component polymer.

Example Embodiment

[0032] Example 1

[0033] Follow the steps below to prepare SOD @ ATP / TB-CPBA:

[0034] (1) 5.5 mg of nitric acid was dissolved in 4 ml of water to give a nitrate water solution;

[0035] (2) Solve 0.1 mg SOD in 0.1 ml of water, resulting in an aqueous solution of SOD;

[0036] (3) 6.1 mg of ATP is dissolved in 4 ml of water to give an ATP aqueous solution;

[0037] (4) After mixing 4 ml of nitrate aqueous solution was mixed with 0.1 ml of SOD aqueous solution, stirred for 30 min; then added an aqueous solution of ATP, followed by stirring for 3 h hetero;

[0038] (5) 13.3 mg of CPBA was dissolved in 0.4 ml of water to give a CPBA aqueous solution;

[0039] (6) Take the above step (4), 3.6 ml of SOD @ ATP / TB (0.1 mg / mL) aqueous solution was mixed with 0.4 ml of CPBA aqueous solution, stirred after 12 h (13000 rpm, 10min), and Wash 3 times with water to get SOD @ ATP / TB-CPBA.

Example Embodiment

[0040] Example 2

[0041] In combination with the water-soluble tetrazol salt reagent (WST-1), the superoxide anion is detected, and the absorbance corresponding to the maximum absorption peak is recorded by the UV spectrometer to detect the superoxide anion catalytic activity of the SOD. Its method steps are:

[0042] (1) As a cushioning solution, a total volume of 200 μL, a reaction temperature of 37 ° C.

[0043] (2) Super oxygenation from KO dissolved in dimethyl sulfoxide (DMSO) solution 2 get;

[0044] (3) Use 96-well plates, respectively, the reaction holes are numbered A, B, C, D; where

[0045]The formulation of the solution in the reaction holes A is: a HEPBS buffer solution containing 10 μm WST-1 reagent, a water bath at 37 ° C for 20 min;

[0046] The formulation method of the solution in the reaction hole B is: a HEPBS buffer solution comprising 10 μm WST-1 reagent and a 50 μm superoxide anion solution, and a 37 ° C water bath reaction for 20 min;

[0047] The formula...

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Abstract

The invention relates to the field of preparation and application of fluorescent probes, in particular to a preparation method and application of an enzyme-integrated rare earth coordination polymer. According to the invention, an enzyme is combined with a rare earth coordination polymer with fluorescence characteristics to construct the enzyme-integrated rare earth coordination polymer fluorescent probe; and the enzyme-integrated rare earth coordination polymer fluorescent probe is applied to time-resolved fluorescence detection of superoxide anions and overcomes the problems that existing fluorescent probes are insoluble in water, poor in light stability and serious in background interference in actual detection. The rare earth coordination polymer can protect an enzyme structure from being damaged by environmental factors, and the catalytic activity and stability of the enzyme are greatly improved. In a superoxide anion detection process, the enzyme can specifically recognize superoxide anions and quench fluorescence of the rare earth coordination polymer through an intramolecular charge transfer process. The interference of background fluorescence can be eliminated by the long fluorescence lifetime of the rare earth fluorescence, so the time-resolved fluorescence detection is realized. The method has the advantages of simplicity and convenience in operation, low cost, high efficiency and the like.

Description

technical field [0001] The invention relates to the field of preparation and application of fluorescent probes, in particular to a method for combining rare earth coordination polymers with enzymes to form complex probes. Background technique [0002] Superoxide anion (O 2 ·- ) is a type of reactive oxygen species (ROS), which play an important role in cell growth and metabolism. However, O 2 ·- The overexpression of can cause oxidative stress in the organism, leading to serious diseases such as rheumatoid arthritis, cardiovascular disease and cancer. Therefore, for O 2 ·- The highly sensitive and selective detection of methionine is of great significance for evaluating its role in physiological and pathological processes. Currently, a variety of O 2 ·- Detection techniques such as electron spin resonance, electrochemistry, chemiluminescence, and fluorescence. Among them, fluorescence analysis is often considered as an ideal analysis method because of its advantage...

Claims

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Application Information

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IPC IPC(8): C08G83/00G01N21/64
CPCC08G83/008G01N21/643
Inventor 谭宏亮翁宇豪朱巧玉
Owner JIANGXI NORMAL UNIVERSITY
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