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A fluorescent dye specifically targeting intracellular lipid droplets and its preparation method and use

A fluorescent dye and cell-targeting technology, which is applied in the field of fluorescent dyes specifically targeting intracellular lipid droplets and its preparation, can solve the problems of difficult to trace the physiological process of lipid droplets, complex dye synthesis process, dye self-absorption, etc. , to achieve low cytotoxicity, low fluorescence background, and easy purification

Active Publication Date: 2021-08-17
HEFEI INSTITUTES OF PHYSICAL SCIENCE - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, most lipid droplet fluorescent dyes still have great disadvantages: 1) The synthesis process of the dye is complicated, the purification is difficult, time-consuming, labor-intensive, and costly; 2) The small Stokes shift easily causes the self-absorption of the dye; 3) Cytotoxicity makes cells in a state of stress, making it difficult to trace the physiological process of lipid droplets in a natural state

Method used

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  • A fluorescent dye specifically targeting intracellular lipid droplets and its preparation method and use
  • A fluorescent dye specifically targeting intracellular lipid droplets and its preparation method and use
  • A fluorescent dye specifically targeting intracellular lipid droplets and its preparation method and use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Embodiment 1: the synthesis of dye dansyl-morpholine (DNS-M)

[0038]

[0039] Dissolve 81.0mg of morpholine (0.927mmol) and 128.1mg of anhydrous potassium carbonate (0.927mmol) in 10mL of anhydrous acetonitrile, and stir at room temperature for 30min. 10 mL of anhydrous acetonitrile solution dissolved with 250 mg of dansyl chloride (0.927 mmol) was added dropwise to the above reaction system, and the stirring reaction was continued at room temperature for 3 h. After the reaction was completed, the potassium carbonate powder was removed by suction filtration, and the acetonitrile was removed with a vacuum rotary evaporator. The obtained green powder was purified by column chromatography to obtain the target product DNS-M, and the eluent was CH 2 Cl 2 / CH 3 OH (v / v=70:1). 1 H NMR (400MHz, CDCl 3 ):δ8.58(d,1H),8.41(d,1H),8.21(dd,1H),7.54(m,2H),7.20(d,1H),3.69(t,4H),2.89(s, 6H). 13 C NMR (100MHz, CDCl 3 ): δ151.8, 132.3, 130.9, 130.8, 130.5, 130.1, 128.1, 123.2,...

Embodiment 2

[0040] Example 2: UV-Vis absorption spectrum and fluorescence spectrum of dye DNS-M in solvents with different polarities

[0041] Dissolve DNS-M in ethanol to make a stock solution with a concentration of 10mM. Take 2 μL of DNS-M stock solution with a concentration of 10 mM from the stock solution and add it to 2 mL of dichloromethane, chloroform, ethyl acetate, acetonitrile, ethanol, DMF and water, and measure the UV-Vis of fluorescent dyes in the above solvents. Absorption and fluorescence spectral curves. Such as figure 1 As shown in A, the absorption peak and absorbance of the dye DNS-M in organic solvents are basically the same, only the absorption peak of the dye in aqueous solution has a blue shift of about 15nm, indicating that the polarity of the solvent basically does not affect the absorption of the dye. However, the polarity of the solvent not only has a very obvious impact on the fluorescence intensity of the dye, but also causes a significant red shift of the ...

Embodiment 3

[0042] Example 3: Co-localization of the dye DNS-M with different commercial organelle dyes in A549 cells

[0043] The commercial lysosome dye LysoTracker Deep Red (1 μM), the mitochondrial dye MitoTracker Deep Red (0.3 μM), the cell membrane dye CellTracker Deep Red (1 μM) and the lipid droplet dye Nile Red (1 μM) were added to A549 cells in different dishes. 37°C, 5% CO 2 After culturing for 30 min under the condition, replace the fresh medium. Subsequently, the fluorescent dye DNS-M prepared in Example 1 was added to the above-mentioned cells so that the final concentration was 10 μM. After culturing for 30 minutes, laser confocal imaging was performed. The imaging results are shown in figure 2 . It can be seen from the figure that the green spots from the dye DNS-M almost do not overlap with the red spots from the lysosome, mitochondria and cell membrane dyes, indicating that the dye DNS-M does not have the ability to localize to lysosomes, mitochondria or cell membrane...

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Abstract

The invention discloses a fluorescent dye specifically targeting intracellular lipid droplets and its preparation method and application, wherein the fluorescent dye specifically targeting intracellular lipid droplets is 5-(dimethylamino)naphthalene-1-sulfonic Acid is used as a fluorescent matrix, and different kinds of amine or alcohol compounds are introduced into the molecular structure of the matrix. The structure is as follows: where R is selected from one of the following groups: R 1 is oxygen or carbon, R 2 and R 3 is hydrogen, C1-8 alkyl or phenyl. The fluorescent dye of the invention can specifically target lipid droplets in cells, and has good photostability and low cytotoxicity. In addition, this fluorescent dye is not only a universal lipid droplet dye, but also can track the movement state of lipid droplets in cells in real time, distinguish the number and size of lipid droplets in different cell lines, and provide a basis for the study of lipid droplets and other organelles. The correlation between them provides the possibility and has broad application prospects in the biological field.

Description

technical field [0001] The invention belongs to the field of synthesis and technical application, and in particular relates to a fluorescent dye specifically targeting intracellular lipid droplets, a preparation method and application thereof. Background technique [0002] Lipid droplets are highly mobile organelles in cells produced by the endoplasmic reticulum, which not only function as lipid storage during cellular metabolism, but also participate in a variety of physiological processes, including membrane transport, membrane and lipoprotein formation, and Intracellular signal transduction, etc. Lipid droplets are surrounded by phospholipid monolayers embedded with various functional proteins on the surface, and contain the central lipid-triglyceride and a small amount of cholesterol ester. Although the composition of lipid droplets in different cells is similar, there are obvious differences in their number and size. These differences can not only reflect the metabolic...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07D265/30
CPCC07D265/30C09B57/00C09K11/06C09K2211/1011C09K2211/1033G01N21/6456G01N21/6486
Inventor 王振洋杨林林张忠平刘变化韩光梅赵君
Owner HEFEI INSTITUTES OF PHYSICAL SCIENCE - CHINESE ACAD OF SCI
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