Fluorescent probe and application thereof in detection of hypochlorous acid in cytolysosome

A technology for detecting hypochlorous acid and fluorescent probes, which is applied in the field of fluorescent probes and can solve problems such as the inability of probes to be positioned

Inactive Publication Date: 2015-10-14
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In recent years, fluorescent probes that can be used to detect hypochlorous acid in living cells have sprung up, but none of these probes can localize to the lysosome, an organelle that produces hypochlorous acid.

Method used

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  • Fluorescent probe and application thereof in detection of hypochlorous acid in cytolysosome
  • Fluorescent probe and application thereof in detection of hypochlorous acid in cytolysosome
  • Fluorescent probe and application thereof in detection of hypochlorous acid in cytolysosome

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Embodiment 1 (synthesis of probe):

[0021] Such as image 3 As shown, the structure of the probe compound used in the embodiment is represented by the code Lyso-NI-Se.

[0022] Synthesis of Lyso-NI-Se: Add 0.50g 4-phenylselenium-aniline, 0.40g cuprous iodide and 0.80g nitrogen-(ethylaminomorpholine)-4-bromo-1,8-naphthoyl to a 50mL three-neck flask Imine, 0.10g potassium hydroxide, vacuum / nitrogen replacement three times, add 20ml anhydrous dimethyl sulfoxide, turn on magnetic stirring, raise the temperature to 90°C, and react for 60 minutes. Filtrate, evaporate the filtrate under reduced pressure, dissolve with dichloromethane, filter again and evaporate the filtrate to dryness under reduced pressure, and purify the resulting solid by silica gel column chromatography (using dichloromethane and ethyl acetate as eluent) to obtain the target compound Lyso-NI-Se (red solid). 1H NMR (500MHz, d 6 -DMSO) δ(ppm):9.41(s,1H),8.76(d,1H,J=8.5Hz),8.47(d,J=7.5Hz,1H),8.27(d,1H,J=...

Embodiment 2

[0023] Embodiment 2 (Lyso-NI-Se selectivity to hypochlorous acid):

[0024] Add 4000μL phosphate buffer solution (pH=5.0, concentration 0.1M), 4000uL Lyso-NI-Se acetonitrile solution (10μM), and 80μL active oxygen solution to a 10mL colorimetric tube, mix well and add 3mL solution to In a 1cmⅹ1cmⅹ4cm cuvette, measure the fluorescence emission spectrum of the working solution, λex=430nm, the grating width is 5nm, 4nm.

[0025] The experimental results of the selectivity of Lyso-NI-Se to HClO are as follows: Figure 4 As shown, the ordinate represents the fluorescence intensity at a wavelength of 540nm. image 3 It shows that Lyso-NI-Se has good selectivity to HClO, and the fluorescence of the system is significantly enhanced. Under the assay conditions, compared to HClO, the fluorescence enhancement caused by other reactive oxygen species (the equivalent far exceeds HClO) is negligible.

Embodiment 3

[0026] Example 3 (quantitative detection of hypochlorous acid by Lyso-NI-Se):

[0027] Add 4000μL phosphate buffer solution (pH=5.0, concentration 0.1M), 4000uL Lyso-NI-Se acetonitrile solution (10μM), 80μL solutions containing different concentrations of hypochlorous acid into a 10mL colorimetric tube, mix After homogenization, take 3mL of the solution and add it to a cuvette of 1cmⅹ1cmⅹ4cm, and measure the ultraviolet-visible absorption spectrum and fluorescence emission spectrum of the working solution (λex=430nm, grating width 5nm, 4nm).

[0028] Figure 4 It was shown that the probe could detect hypochlorous acid at physiological levels in humans. Changes of absorption spectrum (a) and fluorescence spectrum (b) with the addition of hypochlorous acid (0 to 8.0 μM), and the fluorescence intensity at 540nm increases with the increase of hypochlorous acid addition (c).

[0029] Figure 5 It shows that under the lysosome pH condition of pH 4.0 to 5.5, the fluorescence of Ly...

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Abstract

The invention provides a fluorescent probe for selectively detecting hypochlorous acid. A weakly-alkaline morpholine structure is introduced onto a naphthalimide fluorescent matrix so that the probe can be positioned into a cytolysosome, a selenide structure is introduced as an active center reacting with hypochlorous acid, hypochlorous acid can be selectively detected by using the fluorescent difference between the reactant and the product, and particularly, the probe can be used for detecting hypochlorous acid in living cytolysosome.

Description

technical field [0001] The invention provides a fluorescent probe which can be used for selectively detecting hypochlorous acid. The introduction of a basic morpholine structure on the naphthalimide fluorescent matrix allows the probe to be positioned in the lysosome of the cell, and the introduction of a selenide structure as the active center for the reaction with hypochlorous acid, using the fluorescence of reactants and products The differential realization selectively detects hypochlorous acid, especially can be applied to detect hypochlorous acid in living cell lysosome. Background technique [0002] Hypochlorous acid (HOCl) belongs to a kind of reactive oxygen species (ROS, which has very important physiological and pathological significance). As a highly efficient bactericide, it plays an important role in the immune system of life. Endogenous hypochlorous acid in cells is mainly produced by myeloperoxidase (MPO, present in lysosomes) / hydrogen peroxide (H2O2) / Chlo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C09K11/06C07D221/14G01N33/52G01N21/64
Inventor 曲宗金韩克利李鹏楼张蓉
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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