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Fluorescent probe and application thereof in dynamic detection of mercaptan

A fluorescent probe and dynamic detection technology, applied in the field of fluorescent probes, can solve the problems of inability to accurately provide thiol concentration changes, inability to be dynamic, and lack of application prospects

Active Publication Date: 2014-02-12
ZHANGJIAGANG IND TECH RES INST CO LTD DALIAN INST OF CHEM PHYSICS CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this fluorescent probe can only respond to changes in the concentration of thiols statically, and cannot respond dynamically and continuously to changes in the concentration of thiols in cells.
Since they can only statically respond to changes in intracellular thiols, the above probes cannot accurately provide information on changes in thiol concentrations caused by reactive oxygen species in biological systems, and have no application prospects

Method used

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  • Fluorescent probe and application thereof in dynamic detection of mercaptan
  • Fluorescent probe and application thereof in dynamic detection of mercaptan
  • Fluorescent probe and application thereof in dynamic detection of mercaptan

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Embodiment 1 (synthesis of probe):

[0059] like image 3 As shown, the structure of the probe compound used in the examples is represented by the code A, and the fluorescein precursor used in the synthesis of the probe compound is represented by the code F.

[0060] Synthesis of A: 0.6g of F (available for purchase) and 2.5g of 2-piperazinoyl-1-phenylselenide were dissolved in 25ml of anhydrous dimethyl sulfoxide, and stirred at room temperature for 85 hours. The solvent was evaporated under vacuum, and the obtained solid was purified by column chromatography to obtain the target compound A.

[0061] 1 H NMR (400MHz, d 6 -DMSO)δ(ppm):11.97(br,1H),10.13(s,2H),9.77(s,1H),7.98-7.97(d,1H),7.80-7.77(m,2H),7.43-7.42 (m,3H),7.20-7.18(m,1H),6.68-6.67(d,2H),6.59-6.58(m,3H),4.05-4.00(b r,4H),3.78(br,2H),3.43 (br,2H). 13 C NMR (100MHz, d 6 -DMSO)δ(ppm):181.50,181.37,171.88,168.49,167.92,159.43,159.26,151.80,147.81,142.67,142.54,136.29,131.89,131.73,131.63,130.64,129.25,128...

Embodiment 2

[0063] Embodiment 2 (A is to the selectivity of thiol):

[0064] The pH was controlled with PBS buffer solution. Add 2.0μM A to a 10ml colorimetric tube, then add 20mMPBS, dilute the volume to 10ml with ultrapure water, shake the solution, add the above working solution into a fluorescent dish to measure the fluorescence spectrum. Fluorescence intensity changes with pH as Figure 4 shown. Figure 4 It shows that there is no obvious change in the fluorescence intensity of A near the physiological pH, that is, A can be used to detect thiols in a system with a pH of 7.0-10.0.

[0065] In order to simulate physiological conditions as much as possible, the following experiments were carried out under the condition of PH=7.4 (PBS buffer solution, the concentration is 20mM).

[0066] Add 2.0μM probe to a 10ml colorimetric tube, then add 20mM PBS pH7.4, add ultrapure water to 10ml, shake well, and then add various analytes (see the attached figure for the amount of each analyte, bl...

Embodiment 3

[0067] Embodiment 3 (A is to the quantitative detection of mercaptan):

[0068] Add 2.0μM A to a 10ml colorimetric tube, then add 20mM PBS pH7.4, add ultrapure water to 10ml, shake well, and then add different concentrations of thiols. Shake the solution evenly, pour the working solution into a fluorescent dish to measure the fluorescence spectrum, take the maximum value of each fluorescence spectrum, and input it into the software OriginPro8.0 to obtain a linear working curve.

[0069] Thiol concentration after constant volume: 0, 0.2, 0.4, 0.8, 1.0μM.

[0070] Figure 6 (a) shows the change of fluorescence intensity of the system with the change of thiol concentration, indicating that the fluorescence of the system is obviously enhanced with the increase of thiol concentration; Figure 6 (b) represents the linear fitting curve of the fluorescence intensity changing with the thiol concentration, and the linear regression constant of the linear fitting curve is 0.99488, indi...

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Abstract

The invention provides a fluorescent probe and an application thereof in dynamic detection of mercaptan, and relates to a fluorescent probe of which the fluorescence is strengthened in the presence of mercaptan and back to the original state in the presence of oxidizing substances. The fluorescent probe can be used for selectively and dynamically detecting mercaptan in cells. The fluorescent probe is characterized in that fluorochrome with certain overlapping in absorption and emission is adopted as a fluorescent matrix, such as fluorescein and rhodamine; and a dual-selenium bond structure is introduced to the fluorescent matrix as an active center in the reaction with mercaptan, thus realizing the selective detection of mercaptan; a reaction product of a dual-selenium compound and mercaptan reacts with active oxygen such as hydrogen peroxide, hypochlorous acid or nitrosyl peroxide and the like to regenerate the dual-selenium compound, and the reversibility of probe molecule is realized due to the fluorescence quenching property; and simultaneously, the fluorescent property of the probe molecule can be changed through the variation of the distance between fluorophores before and after the reaction of the dual-selenium compound and mercaptan and the influence of the variation to the fluorescent property of the whole compound.

Description

Technical field: [0001] The present invention relates to a class of fluorescent probes for dynamic detection of mercaptans, specifically, a class of fluorescent probes that are enhanced in the presence of mercaptans, and can be detected in oxidizing substances such as hydrogen peroxide, hypochlorous acid, potassium superoxide, tert-peroxide A fluorescent probe whose fluorescence returns to its original state in the presence of butanol, nitrosyl peroxide or singlet oxygen. Background technique: [0002] Thiol is a reducing substance that exists in large quantities in cells, and plays an important role in maintaining the redox balance between the reduced thiol RSH and the oxidized thiol RSSR in the cell, especially glutathione is considered to be The most important class of reducing agents against oxidative stress in cells, and in turn, the concentration of thiols in cells fluctuates with changes in oxidative stress. When cells are under oxidative stress conditions for a long...

Claims

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Application Information

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IPC IPC(8): C09K11/06C07F11/00G01N21/64
Inventor 韩克利楼张蓉李鹏
Owner ZHANGJIAGANG IND TECH RES INST CO LTD DALIAN INST OF CHEM PHYSICS CHINESE ACADEMY OF SCI
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