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Fluorescent probe for mitochondrial membrane potential as well as synthesis method and application thereof

A technology of mitochondrial membrane potential and synthesis method, which is applied in fluorescence/phosphorescence, chemical instruments and methods, luminescent materials, etc., can solve the problems of single species, small fluorescence wavelength shift, etc., and achieves large fluorescence wavelength shift, simple synthesis method, and cellular Low toxicity effect

Inactive Publication Date: 2019-04-05
UNIV OF JINAN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Aiming at the problem of single types of mitochondrial membrane potential probes and small fluorescence wavelength shift in the prior art, the present invention provides a mitochondrial membrane potential probe, which has good selectivity, high sensitivity, large fluorescence spectral shift, and emission Wavelength shift around 140nm

Method used

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  • Fluorescent probe for mitochondrial membrane potential as well as synthesis method and application thereof
  • Fluorescent probe for mitochondrial membrane potential as well as synthesis method and application thereof
  • Fluorescent probe for mitochondrial membrane potential as well as synthesis method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1 Synthesis of Fluorescent Probe HOQ

[0038] (1) Synthesis of 6-hexyloxy-2-naphthaldehyde (compound 1)

[0039] Add 1.377g of 6-hydroxy-2-naphthaldehyde, 1.18mL of iodohexane into the round bottom flask, add 40mL of DMF, stir well; add 2.211g of K 2 CO 3 , stirred at room temperature for 20 hours to complete the reaction. After the reaction was completed, the reaction system was poured into 400mL water; extracted three times with 300mL dichloromethane; the combined extracts were extracted with 4g anhydrous MgSO 4 After drying, the solvent was distilled off under reduced pressure; using petroleum ether / ethyl acetate = 20:1 (v / v) as the mobile phase, the pure 6-hexyloxy-2-naphthaldehyde (compound 1) was obtained by column chromatography ; Productive rate 91%, its proton nuclear magnetic resonance spectrum such as figure 1 As shown, the analysis is as follows: 1 H NMR (400 MHz, DMSO- d 6 )δ 11.91 (s, 1H), 8.55 (dd, J = 8.4, 1.2 Hz, 1H), 8.49 (dd, J = 7....

Embodiment 2

[0046] Example 2 Co-localization of fluorescent probe HOQ and commercial probe MTDR

[0047] In the co-localization experiment, the cells were first stained with 200 nM MTDR for 30 min, then 4 μM HOQ was added to stain the cells for 30 min, then the cell culture medium was aspirated, and the cells were washed with the medium for 3 times for cell imaging: use 405 nm for For the excitation wavelength, collect the fluorescence at 665-735 nm to collect the fluorescence signal of HOQ; use 633 nm as the excitation wavelength, collect the fluorescence at 665-735 nm to collect the fluorescence signal of MTDR. Get the fluorescence picture as Image 6 shown. The counterstaining rate for both dyes was 89%, indicating that the probes stain mitochondria in living cells.

Embodiment 3

[0048] Example 3 Response of fluorescent probe HOQ to different membrane potentials

[0049] Make two copies with a density of 3 × 10 5 cells / mL of HeLa cells were inoculated into sterile 35 mm imaging culture dishes in CO 2 Incubator (at 37°C, 5% CO 2 ) for more than 12 hours to allow the cells to adhere to the wall. One part was treated with 5 μM CCCP solution for 15 minutes to obtain cells with low membrane potential, and the other part was not treated. Then, the DMSO solution of the probe HOQ obtained in Example 1 with a concentration of 1 mM was prepared as the mother solution, and the mother solutions were added to the two cell culture dishes so that the final concentration was 4 μM. Continue to culture under the same conditions for 30 min, then aspirate the cell culture medium, wash the cells with PBS buffer 3 times, and then perform cell imaging.

[0050] In the cell imaging experiment, the excitation wavelength is 405 nm, the dual-channel detection, the detection...

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Abstract

The invention provides a fluorescent probe for mitochondrial membrane potential as well as a synthesis method and application thereof. The structural formula of the probe is as shown in the followingdescriptions. The probe can be synthesized with a reaction product 6-hexyloxyl-2-naphthaldehyde of 6-hydroxyl-2-naphthaldehyde and hexane iodide and a reaction product 1,2-dimethyl-quinoline iodate of2-methylquinoline and methyl iodide. The probe provided by the invention can be used for distinguishing different membrane potential. The fluorescent probe provided by the invention is large in fluorescent wavelength displacement when responding to the mitochondrial membrane potential and low in toxicity to a cell; and the fluorescent probe is simple in synthesis method and simple and convenientin purification step. The probe is successfully applied to cell imaging, and can be used for distinguishing the change of the mitochondrial membrane potential.

Description

technical field [0001] The invention belongs to the field of organic small molecule fluorescent probes, in particular to a fluorescent probe for detecting mitochondrial membrane potential and a synthesis method thereof. Background technique [0002] During the mitochondrial tricarboxylic acid cycle, the protons inside the mitochondria will be actively transported to the outside of the mitochondria, thus forming a mitochondrial membrane potential of -160mV ~ -180mV, which is negative inside and positive outside. The mitochondrial membrane potential powers the process of aerobic respiration, catalyzing its breakdown of highly stable compounds. In addition, the mitochondrial membrane potential is closely related to the state of the cell, and the level of the mitochondrial membrane potential can accurately reflect the healthy state of the cell. Therefore, real-time observation of changes in mitochondrial membrane potential has important physiological, pathological and pharmacol...

Claims

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Application Information

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IPC IPC(8): C07D215/14C09K11/06G01N21/64
CPCC07D215/14C09K11/06C09K2211/1011C09K2211/1029G01N21/6428
Inventor 林伟英孙洁田明刚
Owner UNIV OF JINAN
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