Leucine aminopeptidase and monoamine oxidase-activated near-infrared fluorescent probe as well as synthetic method and biological application thereof

A technology of leucine aminopeptidase and monoamine oxidase, which is applied in the field of fluorescent probes, can solve the problems of low sensitivity and poor specificity of fluorescent probes, and achieve stable fluorescent signals, high selectivity, and good staining effects

Active Publication Date: 2020-02-04
HUNAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to provide a near-infrared fluorescent probe activated by leucine aminopeptidase and monoamine oxidase, a synthesis method and a biological application, so as to solve technical problems such as low sensitivity and poor specificity of current single-enzyme-responsive fluorescent probes, and improve Fluorescence Imaging Contrast

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  • Leucine aminopeptidase and monoamine oxidase-activated near-infrared fluorescent probe as well as synthetic method and biological application thereof
  • Leucine aminopeptidase and monoamine oxidase-activated near-infrared fluorescent probe as well as synthetic method and biological application thereof
  • Leucine aminopeptidase and monoamine oxidase-activated near-infrared fluorescent probe as well as synthetic method and biological application thereof

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Effect test

Embodiment 1

[0047] Example 1 Synthesis of near-infrared leucine aminopeptidase and monoamine oxidase fluorescent probe NML

[0048] (1) Synthesis of Compound 1: p-Hydroxybenzaldehyde (2.44g, 20mmol), (3-bromo-propyl) tert-butyl carbamate (5.95g, 25mmol) and anhydrous potassium carbonate (13.8g, 100mmol) Place in a flask containing acetonitrile (80 mL). And the mixture was stirred at 85°C for 2-3 hours; the solution was cooled and separated by filtration. The filtrate was removed under reduced pressure to give the product as a pale yellow oily liquid (4.81 g, 86% yield) without purification. 1 H NMR (400MHz, CDCl 3 )δ9.79(s,1H),7.74(d,J=8.0Hz,2H),6.92(d,J=8.1Hz,2H),5.07(s,1H),4.04(t,J=5.5Hz, 2H), 3.27(s, 2H), 1.95(d, J=7.1Hz, 2H), 1.37(s, 9H). 13 C NMR (101MHz, CDCl 3 )δ190.73, 163.85, 156.06, 131.90, 129.85, 114.70, 79.07, 66.00, 37.60, 29.46, 28.34;

[0049] (2) Synthesis of Compound 2: Dissolve Compound 1 (2.79g, 10mmol) in trifluoroacetic acid / dichloromethane (20mL / 20mL) at room ...

Embodiment 2

[0058] The detection of embodiment 2NML probe to leucine aminopeptidase and monoamine oxidase in test tube

[0059] The NML probe was made into 1 mM DMSO stock solution and stored at -20°C. The detection system is PBS buffer solution (10 mM, pH 7.4, containing 10% DMSO). The reaction system of NML probe and enzyme was shaken at 37° C. for 2 hours, and then its fluorescence emission spectrum was measured. The excitation wavelength of the fluorescence instrument is set to 670nm, and the receiving range of the emission wavelength is 700-900nm. The result is as figure 1 shown, from figure 1 It can be seen that the NML probe responds well to leucine aminopeptidase and monoamine oxidase.

Embodiment 3

[0060] The reaction kinetic investigation of embodiment 3NML probe and enzyme

[0061] The reaction system was PBS buffer solution (10 mM, pH 7.4, containing 10% DMSO). The fluorescence emission spectra of NML probes with different activities of leucine aminopeptidase and monoamine oxidase enzymes were measured in real time at 37 °C until the fluorescence intensity no longer changed. Then take the intensity of the maximum emission peak as the ordinate, and the reaction time as the abscissa to make a graph, as figure 2 The reaction kinetic curve shown, from figure 2 As can be seen, the NML probe responds rapidly to both enzymes. Set the excitation wavelength of the fluorometer to 670nm and the emission wavelength to 720nm.

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Abstract

The invention discloses a leucine aminopeptidase and monoamine oxidase-activated near-infrared fluorescent probe as well as a synthetic method and biological application thereof. The adopted syntheticmethod includes the following step: (1) synthesizing the novel leucine aminopeptidase and monoamine oxidase-activated near-infrared fluorescent probe (NML); and the biological application comprises the following step: (1) applying the probe to biological imaging and serum detection. The leucine aminopeptidase and monoamine oxidase-responsive fluorescent probe is synthesized for the first time, and the problem of low specificity of a traditional single-enzyme fluorescent probe is overcome; the fluorescent probe has near-infrared fluorescence properties, the background fluorescence signal can be effectively reduced, and the sensitivity of the probe can be improved; and the fluorescent probe has a good staining effect on living cells and high staining efficiency, and can detect cell endogenous leucine aminopeptidase and monoamine oxidase.

Description

technical field [0001] The invention belongs to the technical field of fluorescent probes, and relates to a near-infrared fluorescent probe activated by leucine aminopeptidase and monoamine oxidase, a synthesis method and a biological application. Background technique [0002] Liver diseases such as hepatitis, cirrhosis and liver cancer are major diseases worldwide. Currently, millions of people die from liver disease every year. The best treatment for liver disease today is accurate diagnosis and treatment before it gets worse. However, similar symptoms in the early stages of different liver diseases limit the accurate diagnosis of liver diseases. Standard methods for liver disease diagnosis, such as histopathological examination and molecular medical imaging, are often ineffective in the middle and late stages of the disease, which often leads to delays in diagnosis. Therefore, there is an urgent need to develop a simple and accurate method to diagnose and differentiate...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D405/06C09K11/06G01N21/64
CPCC07D405/06C09K11/06C09K2211/1029C09K2211/1088G01N21/6428G01N21/6486G01N2021/6439
Inventor 张晓兵刘永超滕丽丽许成艳刘红文徐帅郭昊威袁林
Owner HUNAN UNIV
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