Fluorescent probe and its application in reversible detection of peroxy nitrosyl

A technology of nitrosyl peroxide and fluorescent probes, applied in the field of fluorescent probes, can solve the problems of non-reversibility, non-reversibility of probes, and no application prospects

Active Publication Date: 2012-02-08
ZHANGJIAGANG IND TECH RES INST CO LTD DALIAN INST OF CHEM PHYSICS CHINESE ACADEMY OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this kind of fluorescent probe is also sensitive to active species such as hydroxyl radical and hypochlorite, so it cannot be used for the specific detection of ONOO - , also not reversible, cannot be used in ONOO - real-time detection of
Dan Yang et al. disclosed a method to detect ONOO - The fluorescent probe HKGreen-1 (structure see figure 1 , Dan Yang et.al, J.Am.Chem.Soc, 2006, 128, 6004), hypochlorite does not interfere with ONOO - detection, but hydroxyl radicals on ONOO - The interference of the detection is very large, and this probe is not reversible, so it cannot be used for ONOO - real-time detection of
Recently, Dan Yang et al. published a method to detect ONOO - The fluorescent probe HKGreen-2 (structure see figure 1 , Dan Yang et.al, Org.Lett., 2009, 11, 1887), the probe still has a strong response to hydroxyl radicals, and it is not reversible, so it cannot be used in ONOO - real-time detection of
Due to the lack of reversibility, the above probes cannot accurately provide - Induced oxidative stress over time, not promising

Method used

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  • Fluorescent probe and its application in reversible detection of peroxy nitrosyl
  • Fluorescent probe and its application in reversible detection of peroxy nitrosyl
  • Fluorescent probe and its application in reversible detection of peroxy nitrosyl

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Embodiment 1 (synthesis of probe):

[0047] Such as image 3 As shown, the structure of the probe compound used in the examples is represented by the code FSe-1, and the cyanine precursor used for the synthesis of the probe compound is represented by the code Cy7-Cl-1.

[0048] Synthesis of FSe-1: 0.6g Cy7-Cl-1 (available for purchase) and 2.5g p-phenylselenoaniline (Gaythwait, J.Chem.Soc., 1928, 2289.) were dissolved in 20ml of anhydrous N,N- In dimethylformamide, heat to 60-150°C to react for 2 hours. The solvent was evaporated under vacuum, and the obtained solid was purified by column chromatography to obtain the target compound FSe-1.

[0049] 1 H NMR (400MHz, CDCl 3 -D 1 )δ (ppm): 8.49 (s, 1H), 7.42-7.29 (m, 7H), 7.24-7.20 (m, 4H), 7.15-6.93 (m, 6H), 5.85-5.80 (m, 3H), 5.30 (s, 1H), 3.98-4.11(t, 2H), 3.41-3.42(d, 2H), 2.05-2.09(t, 4H), 2.05-1.66(m, 8H), 1.42(m, 12H). 13 C NMR (CDCl 3 -D 1 ,100MHz)δ(ppm):170.50,159.00,147.39,144.03,143.65,142.22,141.09,136...

Embodiment 2

[0052] Embodiment 2 (FSe-1 is to ONOO - optional):

[0053] The pH was controlled with PBS buffer solution. Add 10.0μM FSe-1 to a 10ml colorimetric tube, then add 20mM PBS, make up to 10ml with ultra-pure water, shake the solution well, and after equilibrating for 1min, 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 the fluorescence intensity of FSe-1 does not change significantly in the range of pH 4.0 to 8.5, that is, in the system of pH 4.0 to 8.5, FSe-1 can be used to detect ONOO - .

[0054] In order to simulate physiological conditions as much as possible, the following experiments were all carried out under the condition of pH=-7.4 (PBS buffer solution, the concentration was 20 mM), and the probe concentration was still 10.0 μM.

[0055] Add 10.0μM probe to a 10ml colorimetric tube, then add 20mM PBS pH 7.4, add ultrapure water to 5ml, sh...

Embodiment 3

[0056] Embodiment 3 (FSe-1 is to ONOO - quantitative detection):

[0057] Add 10.0μM FSe-1 to a 10ml colorimetric tube, then add 20mM PBS pH 7.4, add ultrapure water to 5ml, shake well, then add different concentrations of nitrosyl peroxide, and finally dilute to 10ml with ultrapure water . Shake the solution evenly, equilibrate for 10 minutes, 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 OriginPro 8.0 to obtain a linear working curve.

[0058] Concentration of nitrosyl peroxide after constant volume: 0, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10.0μM.

[0059] Figure 6 (a) means with ONOO - Fluorescence intensity and displacement changes of the concentration change system, indicating that with ONOO - With the increase of the concentration, the fluorescence of the system will be blue-shifted while obviously enhanced; Figure 6 (b) shows tha...

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Abstract

The invention relates to a fluorescent probe, wherein, the fluorescence is enhanced in the presence of ONOO-, and the fluorescence returns to original state in the presence of reducing matter. The invention provides a reversible fluorescent probe for selective detection of ONOO- in cells, characterized in that: cyanine dye is used as the fluorescent matrix, an organic selenide structure is introduced in the cyanine matrix as the active site of the reaction with ONOO- to realize the selective detection of ONOO-; the properties that organic selenium oxide formed by oxidizing organic selenide is easy to be reduced to organic selenide by reducing micromolecules in biosystem, such as cysteine, reducing glutathione, metallothionein and the like, is utilized to realize the reversibility of the probe molecules; and variations of electronic properties of organic selenide before and after the oxidation and the influence of the variations on the fluorescence property of the whole compound are utilized simultaneously to modulate the fluorescence property of the probe molecules.

Description

Technical field: [0001] The present invention relates to a class for the reversible detection of nitrosyl peroxide (ONOO - ) fluorescent probes, specifically, is a class in ONOO - In the presence of fluorescent probes, the fluorescence is enhanced, and the fluorescence returns to its original state in the presence of reducing substances such as cysteine, reduced glutathione, and metallothionein. Background technique: [0002] Nitrosyl peroxide (ONOO - ) is a short-lived active species with strong oxidative properties, which is produced by the reaction of nitric oxide and superoxide anions in the human body. ONOO - It can act on harmful substances such as germs recognized by the immune system to inactivate them, thereby protecting the body from harmful substances and maintaining the normal functions of cells and tissues. However, under pathological conditions such as bacterial infection, a large number of ONOO - The generation of ischemia-reperfusion injury, rheumatoid a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C09K11/06G01N33/52G01N21/64
Inventor 韩克利李鹏于法标孙小飞
Owner ZHANGJIAGANG IND TECH RES INST CO LTD DALIAN INST OF CHEM PHYSICS CHINESE ACADEMY OF SCI
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