Aminohexose enzyme fluorescent probe and preparation method and application thereof

A technology of hexosaminidase and fluorescent probe is applied in the field of hexosaminidase fluorescent probe and its preparation, and achieves the effect of good stability and avoiding fluorescence quenching

Active Publication Date: 2019-09-13
INNER MONGOLIA UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently, no fluorescent probes for hexosaminidase with ag

Method used

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  • Aminohexose enzyme fluorescent probe and preparation method and application thereof
  • Aminohexose enzyme fluorescent probe and preparation method and application thereof
  • Aminohexose enzyme fluorescent probe and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0047] The present invention provides a preparation method of the hexosaminidase fluorescent probe described in the above technical scheme, which includes the following steps:

[0048] (1) A compound having a structure represented by formula II and a compound having a structure represented by formula III are subjected to a salt-forming reaction to obtain a compound having a structure represented by formula IV;

[0049]

[0050] In formula III and formula IV, R is hydrogen, alkyl, alkoxy, N,N-dimethyl, N,N-diethyl or N,N-diphenyl;

[0051] (2) The compound having the structure represented by formula IV is subjected to a hydrolysis reaction to obtain a hexosaminidase fluorescent probe having the structure represented by formula I.

[0052] In the present invention, a compound having a structure represented by formula II and a compound having a structure represented by formula III undergo a salt-forming reaction to obtain a compound having a structure represented by formula IV;

[0053]

...

Example Embodiment

[0074] Example 1

[0075] Prepare the hexosaminidase fluorescent probe according to the following reaction process:

[0076]

[0077] (1) Mix compound 1 (200mg, 0.388mmol), compound 2 (146mg, 0.310mmol) and toluene, reflux at 110°C under the protection of nitrogen, monitor the progress of the reaction with a TLC panel until compound 2 disappears completely; add the reaction mixture The insoluble impurities are removed by suction filtration, and the obtained product is concentrated to a solid by rotary evaporation, and then recrystallized with chloroform and petroleum ether. The volume ratio of petroleum ether and chloroform is preferably 1:1 to obtain 229 mg of yellow powdery solid.

[0078] The calculated yield is 75%;

[0079] Characterize the obtained yellow powdery solid, the specific data are as follows:

[0080] 1 H NMR(400MHz,Chloroform-d)δ9.42(d,J=6.4Hz,2H), 8.04(d,J=6.3Hz,2H), 7.68–7.41(m,5H), 7.25–7.10(m, 5H), 7.10–6.83(m,8H), 6.79–6.52(m,4H), 6.03(q,J=13.7Hz,2H), 5.62(d,J=8...

Example Embodiment

[0088] Example 2

[0089] The performance test of the hexosaminidase probe prepared in Example 1 is performed, and the specific steps are as follows:

[0090] (1) Determination of the response time of the hexosaminidase probe: Add 20μL of TPE-NAG in DMSO solution (1mM) to 2mL PBS solution (5mM, pH 7.4) to obtain 10μM TPE-NAG solution, and then add 0.5 U / mL hexosaminidase solution, the resulting mixed solution was incubated at 37°C for different times (0, 5, 15, 25, 35, 45, 60 min), and the fluorescence spectrum of the mixed solution was measured with the incubation time. Happening.

[0091] figure 1 It is the change of fluorescence spectrum of TPE-NAG in the presence of 0.5U / mL hexosaminidase in PBS for different time; figure 2 Is the ratio of the real-time fluorescence intensity of TPE-NAG at 612nm to the initial fluorescence intensity, which is represented by figure 1 with figure 2 It can be seen that after incubating at 37°C for 60 minutes, the fluorescence intensity basically ...

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Abstract

The invention provides an aminohexose enzyme fluorescent probe and a preparation method and application thereof, and relates to the field of biochemical materials. The aminohexose enzyme fluorescent probe provided by the invention has a structure as shown in a formula I. The aminohexose enzyme fluorescent probe has the advantages of few synthesis steps, simple separation and purification operationand good stability, has aggregation induced luminescence characteristic and strong photobleaching resistance, avoids the defect that traditional fluorescent dye is not suitable for detection at highconcentration or cannot be tracked for a long time due to fluorescence quenching caused by aggregation after entering cells, and can be applied to detection of over-expressed aminohexose enzyme in cancer cells.

Description

technical field [0001] The invention relates to the technical field of biochemical materials, in particular to a hexosaminidase fluorescent probe and its preparation method and application. Background technique [0002] Hexosaminidase (N-acetyl-β-D-glucosaminidase, NAG, EC3.2.1.52) is a dimeric lysosomal enzyme, mainly including three isozymes, Hex A, Hex B and Hex S , capable of catalyzing the hydrolysis of N-acetylhexosaminidose glycosidic bonds in GM2 gangliosides. Studies have shown that hexosaminidase is overexpressed in various cancer cells and cancer tissues such as colon cancer and breast cancer, and is a sensitive indicator for detecting kidney injury, especially renal tubular ischemia and necrosis. Hexosaminidase A deficiency can lead to the accumulation of GM2 gangliosides in nerve cells, resulting in hexosaminidase A deficiency, such as Tay-Sachs disease, Sandhoff disease, etc. In view of the important physiological significance of hexosaminidase, it is of grea...

Claims

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Application Information

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IPC IPC(8): C07H15/26C07H1/00C09K11/06G01N21/64
CPCC07H1/00C07H15/26C09K11/06G01N21/6428
Inventor 王建国姜国玉王强
Owner INNER MONGOLIA UNIVERSITY
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