Two-photon fluorescent dye taking isoquinolinone as parent, preparation method thereof and application

A two-photon fluorescence, isoquinolinone technology, applied in biochemical equipment and methods, luminescent materials, chemical instruments and methods, etc., can solve the problems of reducing the application value of RNA dyes, high phototoxicity, poor photostability, etc. Achieve high-efficiency two-photon absorption ability, improve absorption cross-section, and increase the effect of π-electron conjugated system

Active Publication Date: 2013-01-30
DALIAN UNIV OF TECH +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the dye has the following defects in practical imaging applications: long response time with RNA, high phototoxicity, poor photostability, etc., which greatly reduces the application value of this RNA dy

Method used

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  • Two-photon fluorescent dye taking isoquinolinone as parent, preparation method thereof and application
  • Two-photon fluorescent dye taking isoquinolinone as parent, preparation method thereof and application
  • Two-photon fluorescent dye taking isoquinolinone as parent, preparation method thereof and application

Examples

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Example Embodiment

[0042] 8. The method for preparing a two-photon fluorescent dye based on isoquinolinone as claimed in claim 1, comprising the following steps: 1) 4-bromo-1,8-naphthalene anhydride and the compound of formula i are in a molar ratio of 1: 1-1:5 reaction to prepare compound B:

[0043]

[0044] The reaction temperature is 70-150°C, the reaction time is 1-12 hours, and the reaction solvent is selected from dichloromethane, ethanol, ethyl acetate, acetic acid or a mixture thereof;

[0045] In a preferred embodiment, the reaction temperature is 80-140°C, the reaction time is 2-10 hours, and the reaction solvent is selected from ethanol, ethyl acetate, acetic acid or mixtures thereof, 4-bromo-1,8-naphthalene anhydride and formula i The mole is 1:1-1:4;

[0046] In a further preferred embodiment, the reaction temperature is 90-120°C, the reaction time is 3-10 hours, and the reaction solvent is selected from ethyl acetate, acetic acid or a mixture thereof, 4-bromo-1,8-naphthalene anhydride a...

Example Embodiment

[0063] Example 1. Preparation of Dye Compound A 1

[0064]

[0065] (1) Synthesis of Intermediate 1

[0066] 20mmol 4-bromo-1,8-naphthalene anhydride and 35mmol 4-nitro-o-phenylenediamine were added to a round bottom flask containing 20ml of acetic acid solution under nitrogen protection. The reaction was heated at 105°C and refluxed for 3 hours and then stopped. The mixture was poured into ice water, precipitated out, and filtered with suction to obtain a crude yellow solid powder product, intermediate product 1, with a yield of 84%.

[0067] (2) Dye A 1 Synthesis

[0068] Add 20mmol of crude product 1 and 25mmol of ethanolamine from the previous step to a round bottom flask containing 20ml of ethylene glycol monomethyl ether solution, protected by nitrogen. The reaction was heated at 125°C and refluxed for 4 hours and then stopped. The mixture was poured into ice water, orange-red precipitation was precipitated, and a crude red solid powder product was obtained by suction filtra...

Example Embodiment

[0069] Example 2. Dye Compound A 1 Labeling test for cellular RNA

[0070] Using compound A synthesized in Example 1 1 , With a concentration of 5μM A 1 -4μL of DMSO solution added to Hela cells, at 37℃, 5% CO 2 Dye A will be added next 1 The HeLa cells were incubated in the culture medium for 30 minutes. Then, PBS was shaken and rinsed for 5 min×3, then cell culture medium was added, and the two-photon laser confocal imaging was performed. Select a representative area, observe with oil lens (60×), repeat three times. The experimental results are as figure 2 As shown, imaging shows that the cellular RNA of Hela cells has a strong fluorescent signal, while other areas of the cell have no fluorescent signal. figure 2 To add dye A 1 Focused picture of the back Hela cell. The excitation wavelength is 800nm, and the collection wavelength is 520-570nm. The scale bar in the figure is 20μm.

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Abstract

A two-photon fluorescent dye taking isoquinolinone as the parent is provided with a structure presented in formula I, wherein R1 is selected from -H, -CN, -OCH3, -COOH, -OH, -NO2, -NH2, -NHCH3, -N(CH3)2, -OCOCH3 and -SH; R2 is selected from -N-, -C-, -S- and -O-; and R3 is selected from -NHCH2OH, -NH(CH2)2OH, -NH(CH2)3OH, -NH(CH2)4OH,-N(CH3)CH2OH, - N(CH3)(CH2)2OH, -N(CH3)(CH2)3OH and -N(CH3)(CH2)4OH. The fluorescent dye is efficient in two-photon absorptive capacity, low in photo-induced toxicity and photobleaching, and strong in specificity for ribonucleic acid (RNA) in cells; and can perform specificity marking on the RHA in living cells and fixed cells simultaneously. The formula I is shown in the description.

Description

technical field [0001] The invention relates to a class of two-photon fluorescent dyes with isoquinolone as a parent, a preparation method thereof, and the application of the fluorescent dye compounds in labeling cell RNA. Background technique [0002] Ribonucleic acid (abbreviated as RNA, that is, Ribonucleic Acid), is a genetic information carrier that exists in biological cells and some viruses and viroids. A type of nucleic acid composed of at least dozens of ribonucleotides linked by phosphodiester bonds, named for its ribose sugar, referred to as RNA. RNA is ubiquitous in animals, plants, microorganisms, and some viruses and bacteriophages. RNA and protein biosynthesis are closely related. In RNA viruses and bacteriophages, RNA is the carrier of genetic information. RNA is generally a single-stranded linear molecule; there are also double-stranded ones such as reovirus RNA; circular single-stranded ones such as viroid RNA; in 1983, branched-chain RNA molecules were ...

Claims

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Application Information

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IPC IPC(8): C09B57/08C09K11/06C12Q1/68
Inventor 彭孝军仉华樊江莉王静云孙世国宋锋玲
Owner DALIAN UNIV OF TECH
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