Xanthene fluorescent probe and preparation method and application thereof

A fluorescent probe, xanthene technology, applied in the field of fluorescent probes, can solve problems such as limiting the application of fluorescent probes, and achieve the effects of easy promotion and application, good selectivity, and good permeability

Active Publication Date: 2019-01-01
HUNAN UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] However, since the glutathione content in cells is at the millimolar level (0.5-10mM), which is much higher than the highest micromolar concentration that most organic small molecule fluorescent probes can tolerate in biological cells, the above-mentioned limitations are limited. Further applications of fluorescent probes

Method used

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  • Xanthene fluorescent probe and preparation method and application thereof
  • Xanthene fluorescent probe and preparation method and application thereof
  • Xanthene fluorescent probe and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] A preparation method of xanthene fluorescent probe, comprising the following steps:

[0051](1) Preparation of the compound of general formula (III): Add 4-diethylamino ketoacid (1.72g, 5.5mmol) into 10mL methanesulfonic acid, heat to dissolve at 100°C, then add 6-amino-1 , 2,3,4-tetrahydro-1-naphthone (0.81g, 5mmol), continued heating and stirring for 4 hours, then stopped the reaction. After the reaction mixture was cooled to room temperature, the reactant was slowly added to a beaker filled with ice cubes under stirring, and then 10mL perchloric acid (70%) was slowly added dropwise, and a brown-red solid precipitate slowly appeared in the beaker, and placed After standing in the refrigerator for a period of time, suction filtration was performed and dried to obtain 2.24 g of a purple solid with a yield of 83%. 1 H-NMR (400MHz, MeOD), δ8.21 (d, J = 7.7Hz, 1H), 8.08 (dd, J = 8.8, 4.3Hz, 1H), 7.74 (t, J = 7.4Hz, 1H), 7.68 (t,J=7.5Hz,1H),7.29(d,J=7.4Hz,1H),7.05(s,1H),7...

Embodiment 2

[0058] Preparation of a kind of xanthene fluorescent probe and some analyte stock solutions

[0059] A certain amount of the xanthene-based fluorescent probe prepared in Example 1 was dissolved in chromatographic-grade acetonitrile solvent to prepare a 0.5 mM stock solution and store it in a refrigerator. A certain amount of GSH was dissolved in twice-distilled water to prepare a 35 mM stock solution and stored in a refrigerator. Other analytes include amino acids (Ala, Arg, His, Lys, Met, Ser, Tyr), inorganic salts (Al(NO 3 ) 3 , CaCl 2 、CuSO 4 , MgCl 2 , NaBr, NaCl, Na 2 S) and other biological thiols (Cys, Hcy) were prepared into a 10 mM stock solution using twice distilled water as a solvent and stored in a refrigerator. A certain amount of N-ethylmaleimide (NEM) was dissolved in double distilled water to prepare a 50 mM stock solution and stored in a refrigerator. A certain amount of N-acetyl-L-cysteine ​​(NAC) was dissolved in double distilled water to prepare a 1...

Embodiment 3

[0063] Selectivity and anti-interference experiments of xanthene-based fluorescent probes:

[0064] Using the solution prepared in Example 2, the mixed solvent (PBS: CH 3 CN=8:2, pH=7.4) were added different amino acids (Ala, Arg, His, Lys, Met, Ser, Tyr, 0.5mM), cations (Al 3+ , Ca 2+ 、Cu 2+ , Mg 2+ 、Na + ,0.5mM), anion (Cl - , Br, SO 4 2 , S 2- ,0.5mM), other biological thiols (Cys, Hcy,0.5mM), GSH (7mM) measure the fluorescence intensity at a wavelength of 630nm. Depend on Figure 6 It can be seen that except for the addition of Cys and GSH, the fluorescence intensity is enhanced by about 20 times and 100 times, and other analytes hardly cause changes in the fluorescence intensity. Further, the fluorescence spectra of Cys at six concentrations of 0, 20, 50, 100, 200 and 500 μM were made. Such as Figure 7 As shown, when the Cys concentration is 0-50 μM, the fluorescence intensity hardly changes. When the Cys concentration is 100-500 μM, the fluorescence intensit...

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Abstract

The invention relates to a xanthene fluorescent probe and a preparation method and application thereof. The xanthene fluorescent probe has a chemical structure of a formula (1) which is shown in the description. Compared with the prior art, the xanthene fluorescent probe has the following characteristics that 1, the synthesizing method is simple; 2, the selectivity on GSH (glutathione synthetase)is higher; 3, the response concentration on GSH is millimole level, and the xanthene fluorescent probe is suitable for quantitatively detecting the GSH in cells; 4, the toxicity on cells is small; 5,the fluorescent transmission wavelength of a resultant after functioning with GSH can reach 630nm, and the xanthene fluorescent probe can be used for cell imaging.

Description

technical field [0001] The invention relates to the technical field of fluorescent probes, in particular to a xanthene fluorescent probe and its preparation method and application. Background technique [0002] Biothiols such as glutathione (GSH), cysteine ​​(Cys) and homocysteine ​​(homcysteine, Hcy) are important components of many cellular proteins and molecules, and are involved in many physiological processes play an important role. Among these intracellular thiols, GSH is the most abundant nonprotein thiol in eukaryotic cells and maintains redox homeostasis, intracellular gene regulation, signal transduction, and xenobiotic metabolism in living cells. Utilizing the redox potential of GSH can effectively overcome the damage of reactive oxygen species (ROS) to cellular substances. In addition, many diseases, such as AIDS, liver damage, cancer and neurodegenerative diseases (Alzheimer's disease, Parkinson's disease, etc.) are associated with elevated levels of GSH. The...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D311/78C09K11/06G01N21/64
CPCG01N21/6486C09K11/06C07D311/78C09K2211/1007C09K2211/1088
Inventor 万义超廖志鹏全椰岑李玮袁林
Owner HUNAN UNIV OF SCI & TECH
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