Naphthyl two-photon fluorescent probe as well as preparation method and application thereof

A two-photon fluorescence and probe technology, applied in the field of fluorescent probes, can solve the problems of limited deep tissue imaging applications, lack of mitochondrial targeting, large photodamage and photobleaching, etc., achieves good cell membrane permeability, and is conducive to Commercial production, good selectivity effect

Inactive Publication Date: 2017-07-25
SHANXI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] At present, many fluorescent probes based on cysteine ​​detection have been reported in the literature. However, due to the similarity in structure and biological activity of biothiols, cysteine ​​can be compared with homocysteine ​​(Hcy) and glutathione. The fluorescent probes for differentia

Method used

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  • Naphthyl two-photon fluorescent probe as well as preparation method and application thereof
  • Naphthyl two-photon fluorescent probe as well as preparation method and application thereof
  • Naphthyl two-photon fluorescent probe as well as preparation method and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0031] The synthetic route of naphthyl two-photon fluorescent probe:

[0032]

[0033] Synthesis steps and characterization:

[0034] in N 2 Under protection, 1g (2.56mmol) 1-methyl-4-[2-(6-hydroxy-2-naphthalene)-vinyl]-pyridine iodide salt and 0.14g (2.56mmol) sodium methoxide were added to 50mL anhydrous In methanol, the reaction was heated at 60°C for 4 hours, the color of the solution gradually changed from yellow to blue, and the blue solid was obtained by cooling and filtration, which was directly carried out to the next reaction without further treatment. Add 1g (2.43mmol) of blue solid, 0.78g (2.92mmol) of 2,4-dinitrobenzenesulfonic acid chloride and 0.26g (2.43mmol) of triethylamine into 100ml of anhydrous acetonitrile, and heat at 60°C for 2 Hours, the crude product was obtained by distillation under reduced pressure. The crude product was separated by column chromatography (dichloromethane as the mobile phase) to obtain 1.05 g of a khaki solid with a yield of ...

Embodiment 2

[0036] The probe in Example 1 was used to prepare a stock solution with a concentration of 1 mM in DMSO. Dissolve Cys in secondary water to make a 0.1M solution for later use. In the experiment, the probe was diluted to a final concentration of 10 μM with DMSO / PBS buffer (pH 7.4) system (v / v=1 / 1), and the ultraviolet-visible absorption spectrum of the reaction between the probe and Cys (100 μM) was recorded over time ( image 3 ). With the increase of the reaction time between the probe and cysteine, the maximum absorption peak at 352nm gradually decreases and red shifts to 392nm, an isosbestic point appears at 370nm, and the time for the maximum absorption peak at 392nm is 2min. At the same time, the color of the solution changed from colorless to yellow ( Figure 4 ).

Embodiment 3

[0038] The probe was diluted to 10 μM with DMSO / PBS buffer (pH 7.4) system (v / v=1 / 1), and the fluorescence spectrum spectrum of the reaction between the probe and Cys (100 μM) was recorded over time, and the excitation wavelength was fixed at 370 nm. Excitation and emission slit broadband are both 2.0nm. Such as Figure 5 As shown, before Cys was not added, the maximum fluorescence emission of the probe was located at 485nm. As the reaction time between the probe and cysteine ​​increased, the fluorescence emission at 485nm gradually decreased, and a new fluorescence emission appeared at 583nm and Significantly enhanced with a clear isoelectric point at 502nm. And the reaction reaches the maximum value at 2min, which is faster than most biothiol probes reported in the literature. Under the irradiation of ultraviolet light, the color of the solution changed from light blue to green ( Figure 6 ).

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Abstract

The invention discloses a naphthyl two-photon fluorescent probe as well as a preparation method and application thereof. The preparation method of the probe comprises the steps of firstly, heating 1-methyl-4-[2-(6-hydroxyl-2-naphthalene)-vinyl]-pyridium and sodium methylate for 4 hours, then heating the heated materials together with 2,4-dinitrobenzene chlorine sulfonate and triethylamine in acetonitrile for 2 hours, and carrying out silica gel column separation to obtain the pure product. The probe takes a naphthalene ring as a two-photon parent structure, has both a pyridium mitochondrion targeted group and a fluorophore, and uses 2,4-dinitrobenzene sulfonyl as a Cys selective recognition group. The hydroxyl is produced by using sulfydryl to break the 2,4-dinitrobenzene sulfonyl, and fluorescence spectrum redshift occurs, so that the ratio test (F583nm/F485nm) of Cys is realized; during the detection, a maximum value is rapidly obtained within 2min, and the detection line is as low as 29 nm. The probe can be used for selective detection of the Cys in cellular mitochondria.

Description

technical field [0001] The invention relates to a fluorescent probe, in particular to a naphthyl two-photon fluorescent probe and a preparation method thereof, as well as the application of the probe in the detection of cysteine ​​in cell mitochondria. Background technique [0002] Cysteine ​​(Cys), as an essential amino acid and an important biothiol, plays a key role in the physiological activities of many cells, such as protein post-transcriptional modification, detoxification and metabolism. Insufficient cysteine ​​in the body can lead to stunted growth, hair depigmentation, edema, muscle weakness, obesity, liver damage, and loose skin in young children; and excessive cysteine ​​concentration is also closely related to the occurrence of some diseases, including cardiovascular disease, neurodegenerative disease, etc. Therefore, research on highly sensitive and real-time detection of cysteine ​​in biological samples has become a research hotspot in related fields. Mitoch...

Claims

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Application Information

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IPC IPC(8): C07D213/30C09K11/06G01N21/33G01N21/64
CPCC07D213/30C09K11/06C09K2211/1029G01N21/33G01N21/6428G01N2021/6439G01N2021/6443
Inventor 樊丽张雯佳王晓东董川双少敏
Owner SHANXI UNIV
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