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Preparation and application of a fluorescent probe for rapid detection of cysteine

A cysteine ​​and fluorescent probe technology, applied in the field of fluorescent probes, can solve the problems of unreachable, low detection limit, complex preparation method, etc., and achieve the effects of fast response speed, low detection limit and simple synthesis

Inactive Publication Date: 2018-07-06
UNIV OF JINAN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the prior art, CN104119263 A provides a cysteine ​​fluorescent probe compound based on cyanine, and CN104447421 A provides a cysteine ​​probe compound based on 2-hydroxyl-6-acetylnaphthalene , but the preparation methods of these two compounds are complicated, and the detection limit of lower concentration cannot be achieved and the rapid detection of cysteine ​​can not be achieved.

Method used

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  • Preparation and application of a fluorescent probe for rapid detection of cysteine
  • Preparation and application of a fluorescent probe for rapid detection of cysteine
  • Preparation and application of a fluorescent probe for rapid detection of cysteine

Examples

Experimental program
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Effect test

Embodiment 1

[0040] Embodiment 1, the synthesis of fluorescent probe molecule

[0041] (a) Under nitrogen protection, dissolve 3.1 g (16 mmol) of 4-cyanobiphenol in 50 ml of acetonitrile, and add 2.8 g (24 mmol) of anhydrous MgCl 2 and 8.2 ml (60 mmol) of anhydrous triethylamine, and finally 6.1 g (220 mmol) of dry paraformaldehyde was added and reacted at 70°C for 8 hours. After the reaction was completed, the solution was acidified with a large amount of dilute hydrochloric acid, extracted with dichloromethane, and separated on a silica gel column to obtain 1.8 g of white solid product II with a yield of 52%.

[0042] (b) Under the protection of nitrogen, take 1.0 g (4.48 mmol) of compound II and place it in a 100 ml three-neck flask, add 30 ml of dry dichloromethane into it, and stir until it is completely dissolved. Then 7.5 ml (0.11 mol) of pyrrole and a catalytic amount of trifluoroacetic acid were added, and the reaction was carried out at room temperature for one hour. After the ...

Embodiment 2

[0045] Example 2. Fluorescence response experiments of probe molecules to different thiol amino acids

[0046] Take the fluorescent probe compound of the present invention, dissolve it in acetonitrile, and prepare the probe molecule mother solution with a concentration of 50 μM; get a certain amount of cysteine, homocysteine ​​and glutathione, and dissolve them in 100ml super Pure water, prepared to 10 -2 Different amino acid mother liquors with M concentration; take 2.38g of 4-hydroxyethylpiperazineethanesulfonic acid, dissolve it in 100 ml of ultrapure water, adjust the pH to 7.4 with sodium hydroxide, and prepare a buffer solution with a buffer concentration of 0.1 M.

[0047] Pipette 1ml probe molecule mother solution, add 4 ml acetonitrile, 1 ml HEPES buffer solution, 3.7 ml ultrapure water to it, add 0.3 ml Cys, Hcy and GSH mother solution to it respectively. Immediately start scanning its fluorescence intensity over time. Such as figure 2 , the results show that di...

Embodiment 3

[0048] Example 3. Fluorescence response experiment of probe molecules to cysteine ​​at different concentrations

[0049] Pipette 13 groups of 1 ml probe molecule mother solution, add 4 ml acetonitrile, 1 ml HEPES buffer solution, add 3.7 ml, 3.725 ml, 3.75 ml, 3.775ml, 3.80ml, 3.825ml, 3.85ml, 3.875ml, 3.90ml, 3.925ml, 3.950ml, 3.975ml, 4.0 ml ultrapure water, and 0.3 ml, 0.275 ml, 0.25 ml, 0.225ml, 0.20ml, 0.175ml, 0.150ml, 0.125ml, 0.100ml, 0.075ml, 0.050 ml, 0.025ml, 0.0 ml Cys mother solution. Incubate at room temperature for 10 minutes, excite with a light source with a wavelength of 360 nm, and test its fluorescence emission spectrum. Such as image 3 As shown, the results show that the probe molecules can accurately characterize different concentrations of cysteine.

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Abstract

The invention relates to preparation and application of a fluorescent probe capable of rapidly detecting cysteine. The structure of a probe compound is as shown in a formula I in the specification. A preparation method for the probe compound comprises the following steps: introducing an aldehyde group to position 3 of 4-cyanodiphenol in virtue of paraformaldehyde so as to obtain a compound II; subjecting the compound II and pyrrole to nucleophilic addition so as to produce a compound III; reacting the compound III with DDQ and boron trifluoride-diethyl ether successively so as to obtain a compound IV; and subjecting the compound IV and chloroacetyl chloride to a substitution reaction so as to produce the probe compound I. The probe compound can be used for rapid analysis and detection of cysteine and has the characteristics of high sensitivity, low detection limits, etc.

Description

technical field [0001] The invention relates to the preparation and application of a fluorescent probe compound for rapid detection of cysteine, belonging to the technical field of fluorescent probes. technical background [0002] There are various active species in organisms, which have special physiological functions and play a vital role in the life process. Among the many substances that make up life, mercapto compounds play an irreplaceable and important role. Cysteine, as an important sulfhydryl amino acid, participates in protein synthesis and cell metabolism. The lack of cysteine ​​in the human body can lead to health problems such as slow growth, skin damage, and weakness in children, while excess cysteine ​​can induce neurotoxicity. Homocysteine ​​is usually derived from the metabolism of methionine in the body. Clinical studies have shown that it can stimulate the proliferation of vascular smooth muscle and cause vascular endothelial damage, which is closely re...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07F5/02C09K11/06G01N21/64
CPCC07F5/022C09K11/06C09K2211/1029C09K2211/1096G01N21/6428G01N2021/6432
Inventor 吕正亮黄曦明范春华
Owner UNIV OF JINAN
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