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 aggregation-induced luminescent properties have been reported

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

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preparation example Construction

[0047] The present invention provides the preparation method of hexosaminidase fluorescent probe described in above-mentioned technical scheme, comprises the following steps:

[0048] (1) performing a salt-forming reaction between a compound having a structure shown in formula II and a compound having a structure shown in formula III to obtain a compound having a structure shown in 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 shown in formula IV is hydrolyzed to obtain the hexosaminidase fluorescent probe having the structure shown in formula I.

[0052] In the present invention, a compound having a structure shown in formula II is subjected to a salt-forming reaction with a compound having a structure shown in formula III to obtain a compound having a structure shown in formula IV;

[0053]

[0054]

[0055] In formula III and formula...

Embodiment 1

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

[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 reaction process with a TLC plate until compound 2 disappears completely; the reaction mixture The insoluble impurities were removed by suction filtration, and the obtained product was concentrated to a solid by rotary evaporation, and then recrystallized with chloroform and petroleum ether. The volume ratio of petroleum ether and chloroform used was preferably 1:1 to obtain 229 mg of a yellow powdery solid.

[0078] The calculated yield is 75%;

[0079] The obtained yellow powdery solid is characterized, and 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...

Embodiment 2

[0089] The hexosaminidase probe prepared in embodiment 1 is tested for performance, and the specific steps are as follows:

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

[0091] figure 1 It is the change of fluorescence spectrum of TPE-NAG after incubation in PBS for different time under the condition of 0.5U / mL hexosaminidase; figure 2 is the ratio change graph of the real-time fluorescence intensity of TPE-NAG at 612nm and the initial fluorescence intensity, by figure 1 with figure 2 It can be seen that after incubation at 37°C for 60 min, the fluorescence intens...

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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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