Synthesis of multi-signal fluorescence probe and application of multi-signal fluorescence probe in simultaneously and differentially detecting Hcy, Cys and GSH (Glutathione)

A technology of fluorescent probes and fluorescent molecular probes, applied in the field of analytical chemistry

Active Publication Date: 2018-09-04
HUNAN NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the reported fluorescent probes for thiols have not been able to simultaneously distinguish and detect the three biothiols of Hcy, Cys and GSH using three channels, especially the three-channel fluorescence imaging analysis and detection of different concentrations of endogenous Hcy, Cys and GSH in cells.

Method used

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  • Synthesis of multi-signal fluorescence probe and application of multi-signal fluorescence probe in simultaneously and differentially detecting Hcy, Cys and GSH (Glutathione)
  • Synthesis of multi-signal fluorescence probe and application of multi-signal fluorescence probe in simultaneously and differentially detecting Hcy, Cys and GSH (Glutathione)
  • Synthesis of multi-signal fluorescence probe and application of multi-signal fluorescence probe in simultaneously and differentially detecting Hcy, Cys and GSH (Glutathione)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1. Synthesis of 9-hydroxy-2,3,6,7-tetrahydro-1H,5H,11H pyrano[2,3-F]pyrido[3,2,1-ij]quinoline-11 -ketone

[0027] a. Add 16.3 g (63.4 mmol) of diphenyl malonate and 10.0 g (52.8 mmol) of 8-hydroxyjuloridine to 100 mL of anhydrous toluene, and stir the reaction at 100 °C for 8-12 hours,

[0028]b. After the reaction was completed, the reaction solution was cooled to room temperature and filtered, the filter cake was washed with diethyl ether, and the obtained solid was vacuum-dried to obtain a gray-green solid 9-hydroxy-2,3,6,7-tetrahydro-1H, 5H, 11H pyrano[2,3-F]pyrido[3,2,1-ij]quinolin-11-one 11.0 g, yield 80.9%.

Embodiment 2

[0029] Example 2. Synthesis of 9-chloro-11-oxo-2,3,6,7-tetrahydro-1H,5H,11H pyrano[2,3-F]pyrido[3,2,1-ij ]quinoline-10-carbaldehyde

[0030] . Under nitrogen protection, 18 mL of dry re-distilled N,N-dimethylformamide (DMF) was slowly added to an equal volume of phosphorus oxychloride (POCl 3 ), stirring at 20-50 °C for 30 minutes to obtain a red solution,

[0031] . 15.0 g (58.3 mmol) of 9-hydroxy-2,3,6,7-tetrahydro-1H,5H,11H pyrano[2,3-F]pyrido[3,2,1-ij]quinoline -11-keto was dissolved in 70 mL N,N-dimethylformamide and added dropwise to step In the mixed solution, the mixture continued to stir and react at 60°C for 12 hours under nitrogen protection;

[0032] . After the reaction is complete, the steps The reaction solution in the solution was slowly poured into 500 mL of ice water, and the pH was adjusted to 6 with 20% NaOH solution, resulting in a large amount of precipitation, filtered, and the filter cake was washed 3 times with an appropriate amount of d...

Embodiment 3

[0033] Example 3. Synthesis of multi-signal fluorescent probes ( E )-3-(9-chloro-11-oxo-2,3,6,7-tetrahydro-1H,5H,11H-pyrano[2,3-f]pyrido[3,2,1- ij]quinolin-10-yl)-2-cyanoacrylate ethyl ester (R = ethyl)

[0034] . 5.0 g (16.5 mmol) of 9-chloro-11-oxo-2,3,6,7-tetrahydro-1H,5H,11H pyrano[2,3-F]pyrido[3,2, 1-ij]quinoline-10-carbaldehyde and 2.79 g (24.7 mmol) ethyl cyanoacetate were mixed and added to 30 mL of anhydrous dichloromethane, then 0.2 mL of triethylamine was added dropwise, and the reaction was stirred at room temperature;

[0035] . After the reaction is complete, step The reaction droplets were added to 300 mL of absolute ethanol, filtered, and the obtained solid was dried to obtain the fluorescent molecular probe according to claim 1 ( E )-3-(9-chloro-11-oxo-2,3,6,7-tetrahydro-1H,5H,11H-pyrano[2,3-f]pyrido[3,2,1- ij]quinolin-10-yl)-2-cyanoacrylate ethyl ester 4.5 g, yield 68.5%.

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Abstract

The invention discloses a multi-signal fluorescence probe for simultaneously and differentially detecting high-cysteine (Hcy), cysteine (Cys) and glutathione (GSH) in cells through three different blue, green and yellow fluorescence emission signals. The chemical structure formula of the multi-signal fluorescence probe is as shown in the specification, and in the formula, R is hydrogen / alkyl / aryl.By adopting the multi-signal fluorescence probe, by virtue of the characteristic that different fluorescence substances can be generated through different chemical reactions of the multi-signal fluorescence probe with Hcy, Cys and GSH under a same detection condition, and fluorescence of three colors of blue, green and yellow can be emitted under specific stimulation laser wavelengths, and the purpose of simultaneous and differential detection on Hcy, Cys and GSH can be achieved; blue light of 467nm can be emitted after the probe is reacted with Hcy under a laser wavelength of 375nm, green light of 503nm can be emitted after the probe is reacted with Cys under a laser wavelength of 400nm, and yellow light of 568nm can be emitted after the probe is reacted with GSH under a laser wavelengthof 500nm; the multi-signal fluorescence probe can be applied to simultaneous fluorescence imaging analysis on Hcy, Cys and GSH in L-02 (normal liver cells) cells.

Description

technical field [0001] The invention belongs to the technical field of analytical chemistry, and in particular relates to the synthesis of a type of multi-signal fluorescent probes, and the simultaneous discrimination and quantitative detection of Hcy, Cys and GSH by the probes in the environment, and the three-channel fluorescence imaging of intracellular Hcy, Cys and GSH. application. Background technique [0002] Small molecule biothiols such as cysteine ​​(Cys), homocysteine ​​(Hcy) and glutathione (GSH) play extremely important roles in biological systems. In mammals, Hcy can be synthesized from methionine with the assistance of various enzymes. The normal concentration of Hcy in serum is 5-12 μM. Excessive Hcy content will cause folic acid and V B12 Deficiency, Alzheimer's disease and other diseases (Chemical Communication, 2014, 50, 6967-6969); In addition, intracellular Cys can be synthesized from Hcy, the normal intracellular concentration is 30-200 μM, abnormal Cy...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D491/16C09K11/06G01N21/64
CPCC07D491/16C09K11/06C09K2211/1029C09K2211/1088G01N21/6486
Inventor 尹鹏尹国兴甘亚兵喻婷孙鑫雨张友玉
Owner HUNAN NORMAL UNIVERSITY
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