Probe capable of realizing two-photon fluorescence detection of nitroreductase (NTR) and preparation method thereof

A two-photon fluorescence, nitroreductase technology, applied in the field of organic fluorescent probes, can solve the problems of small Stokes shift, interference, poor water solubility of the probe, and achieve good cell permeability, avoid interference, and improve The effect of chemical stability

Active Publication Date: 2018-11-02
NANJING TECH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

So far, many fluorescent probes for hypoxia markers of tumor cells have been reported, such as fluorescent probes for detecting small biological molecules such as nitroreductase, diaphorase, quinone reductase, and azoreductase, but these fluorescent probes There is a big disadvantage that the water solubility of the probes is generally poor, and the Stokes shift is small, the excitation light will cause different degrees of interference during the imaging process, and the tissue penetration ability of most probes is not good enough , resulting in insufficient convincing research results

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  • Probe capable of realizing two-photon fluorescence detection of nitroreductase (NTR) and preparation method thereof
  • Probe capable of realizing two-photon fluorescence detection of nitroreductase (NTR) and preparation method thereof
  • Probe capable of realizing two-photon fluorescence detection of nitroreductase (NTR) and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] The preparation method of a series of naphthol-based fluorophore two-photon nitroreductase (NTR) probe XD03 is as follows:

[0036] 1.1XD00: 2,7-dihydroxynaphthol (0.3g, 18.7mmol), dimethylamine (40% aqueous solution, 1.05mL, 9.35mmol), sodium metabisulfite (0.711g, 3.74mmol), deionized water 2mL in sequence Add it into a Shrek bottle, react at 150°C for 8 hours, cool to room temperature, use hydrochloric acid to adjust the pH of the reaction system to about 4, then use DCM to extract and spin dry, and purify by gel column chromatography to obtain the intermediate product XD00. 1 H-NMR (300MHz, CD 3 OD): δ7.57(m, 2H), δ6.97-7.01(m, 1H), δ6.93-6.94(d, J=3Hz, 1H), δ6.67-6.79(m, 2H), δ2 .98(S,6H), the reaction formula is as follows:

[0037]

[0038]1.2XD01: Add N,N-dimethylformamide (0.4mL) to the Shrek bottle and vacuumize, add phosphorus oxychloride (0.2mL) to the above Shrek bottle with a syringe, and place it in a cold well at 0°C under stirring for 10min. The ...

Embodiment 2

[0045] The preparation method of a series of two-photon nitroreductase (NTR) probe XD03 based on naphthol fluorophore is as follows:

[0046] 1.1XD00: Add 2,7-dihydroxynaphthol (3g, 187mmol), dimethylamine (40% aqueous solution, 10.5mL, 93.5mmol), sodium metabisulfite (7.11g, 37.4mmol), and 20mL of deionized water to the mixture in turn. In a Shrek bottle, react at 150 °C for 8 h, and after cooling to room temperature, use hydrochloric acid to adjust the pH of the reaction system to about 4, then use DCM to extract and spin dry, and purify by gel column chromatography to obtain an intermediate product white solid XD00.

[0047] 1. 2XD01: Add N,N-dimethylformamide (4mL) to the Shrek bottle and vacuumize, add phosphorus oxychloride (2mL) to the above Shrek bottle with a syringe, and stir at 0°C in a cold well 10min. The product XD00 from the previous step was dissolved in 2 mL of N,N-dimethylformamide, slowly added to the Shrek bottle with a disposable syringe, and stirred at 0...

Embodiment 3

[0051] The preparation method of a series of two-photon nitroreductase (NTR) probe XD03 based on naphthol fluorophore is as follows:

[0052] 1.1XD00: 2,7-dihydroxynaphthol (1.5g, 93.5mmol), dimethylamine (40% aqueous solution, 5.25mL, 46.75mmol), sodium metabisulfite (3.55g, 18.7mmol), deionized water 10mL in turn It was added to Shrek bottle, reacted at 150°C for 8h, cooled to room temperature, adjusted the pH of the reaction system to about 4 with hydrochloric acid, then extracted with DCM and spin-dried, and purified by gel column chromatography to obtain an intermediate product white solid XD00.

[0053] 1. 2XD01: Add N,N-dimethylformamide (2mL) to the Shrek bottle and vacuumize, add phosphorus oxychloride (1mL) to the Shrek bottle with a syringe, and stir at 0°C in a cold well 10min. The product XD00 of the previous step was dissolved in 1 mL of N,N-dimethylformamide, slowly added to the Shrek bottle with a disposable syringe, and stirred at 0 °C for 2 h. After cooling...

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Abstract

The invention relates to preparation of a probe capable of realizing two-photon fluorescence detection of nitroreductase (NTR) as shown in the figure (I) and belongs to the field of organic fluorescent probes. The structural formula of the probe capable of realizing two-photon fluorescence detection of the NTR is as shown in the description. The fluorescent probe provided by the invention can accurately detect the content of the NTR in cells and avoid the interference of other reducing agents in cells. In addition, the probe has the characteristics of better chemical stability, biological compatibility and selectively and the like. A laser confocal imaging experiment shows that the probe has better cell permeability, has no toxic and side effects on cells and organisms, can realize detection of content of cell level NTR and indicate the cell hypoxia condition, and can be further applied to research of precancerous detection.

Description

technical field [0001] The invention relates to a probe capable of two-photon fluorescence detection of nitroreductase NTR and a preparation method thereof, belonging to the field of organic fluorescent probes. Background technique [0002] Nitroreductase (NTR), as one of the hypoxic markers of tumor cells, is a class of cytoplasmic enzymes dependent on flavin mononucleotide or flavin adenine dinucleotide, which widely exists in bacteria. The structure generally contains flavin mononucleotide units or flavin adenine dinucleotide units, which can use nicotinamide purine dinucleotide (NADPH) as a coenzyme to realize Reductive metabolism of various nitro compounds to produce hydroxylamine or amino. Tumor cell hypoxia often also results in increased intracellular nitroreductase. Therefore, in the presence of electron donors such as NADH / NADPH, nitroreductases can efficiently reduce aromatic nitro compounds to the corresponding amino compounds. This reduction behavior can be u...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07C255/43C07C253/30C07C221/00C07C223/06C07C213/02C07C215/86C09K11/06G01N21/64
CPCC07C213/02C07C221/00C07C253/30C07C255/43C09K11/06C09K2211/1007C09K2211/1011G01N21/6428C07C215/86C07C223/06
Inventor 李林许晨晨黄维余昌敏张承武赵燕菲吴琼丁阳李峥
Owner NANJING TECH UNIV
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