Ratiometric fluorescent probe and application thereof

Abstract

The invention relates to a novel rare-earth complex ratiometric fluorescent probe which can be used for measuring peroxynitrite in a biosystem. The probe is complexes formed by two trivalent rare-earth ions Eu<3+> and Tb<3+> and an organic ligand [4'-(2,4-dimethoxyphenyl)-2,2':6'2''-terpyridine-6,6''-dimethylamino]tetraacethyl (DTTA for short), wherein the structural formula of the ligand is disclosed in the specification. The probe can specifically act with the peroxynitrite, so that the two rare-earth complexes are changed at the ratio (F612 / F541) of the fluorescence intensity of 612nm and 541nm, thereby implementing the selective fluorescent measurement of the peroxynitrite in a biosystem.

Description

technical field [0001] The present invention relates to the determination technology of peroxynitrite anion, specifically a rare earth complex-based peroxynitrite anion ratio-type fluorescent probe and its application, which can be used in the selection of peroxynitrite anion in biological systems. Ratio-type rare-earth fluorescent probes for sexual fluorescence assays. Background technique [0002] Peroxynitrite anion (ONOO) in biological systems - ) is composed of nitric oxide (NO) and superoxide anion radicals (O 2 - ) is rapidly combined to form an important nitrogen-oxygen reactive compound. It is generally believed that ONOO - is compared to NO and O 2 - A potent cytotoxic substance with stronger oxidation and wider effects (Document 1: J.S.Beckman, T.W.Beckman, Proc.Natl.Acad.Sci.USA.1990, 87, 1620; Document 2: G.Ferrer- Sueta, R. Radi, ACS Chem. Biol. 2009, 4, 161). As a strong oxidizing agent, ONOO - Can produce oxidative damage to cells and tissues, so it ...

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Problems solved by technology

The method using fluorescent molecular probes in the above method is a simple, sensitive and specific analysis method, so far, several fluorescent molecular probes have been successfully used in ONOO - Determination of dichlorodihydrofluorescein (DCFH) and dihydrorhodamine (DHR) (document 8: H.Possel, H.Noack, FEBS Lett.1997, 416, 175; document 9: N.W.Kooy, J.A.Royall , Free Radic.Biol.Med.1994, 16, 149), although this method has higher sensitivity, the specificity of this type of probe is poor, and other biological oxidants such as hypochlorous acid, thiol and peroxidase can also Oxidation of DCFH and DHR to Generate Strongly Fluorescent Dichlorofluorescein and Rhodamine

In recent years, two novel ONOO based on aromatic ring nitration and ketone oxidation reactions - Fluorescent probes: NiSPYs and HKGreens have been synthesized successively (Document 10: T.Ueno, T.Nagano, J.Am.Chem.Soc.2006, 128, 10640; Document 11: D.Yang, H.Wang, J.Am.Chem.Soc.2006, 128, 600; Literature 12: Z.Sun, H.Wang, D.Yang, Org.Lett.2009, 11, 1887), these two probes can be combined with ONOO - Specific reaction leads to a significant enhancement of fluorescence intensity, but the probe has poor water solubility, is easy to be photobleached, has a small Stokes shift, and has a short retention time in the cell, which is not conducive to the long-term observation and trace of peroxynitrite anion in the cell. Quantitative determination

In addition, the existing ONOO - Fluorescent probes use fluorescence intensity as a characterization parameter. Although the operation is simple, the accuracy of the results is easily affected by factors such as changes in the external environment and instrument conditions, such as optical paths, photobleaching, scattering, and background light.

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