Preparation method and application of near-infrared fluorescent probe responding to viscosity
Through the near-infrared fluorescence probe technology based on TICT effect, the problem of monitoring intracellular viscosity changes is solved, and efficient and sensitive tools are provided for cancer and metabolic disease research, real-time monitoring of intracellular viscosity changes is achieved. The medical field provides new application perspectives and diagnostic tools.
Patent Information
- Application Number
- CN202411228076.X
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-09-03
- Publication Date
- 2024-12-13
AI Technical Summary
The prior art is difficult to effectively monitor and track changes in intracellular viscosity in real time, especially in diseases such as cancer, neurodegenerative diseases and metabolic disorders, and lacks sensitive tools to reflect changes in the physical properties of the intracellular microenvironment.
Using a near-infrared fluorescent probe based on the TICT effect, dihydroisophorone and 5-(3,5-dimethylpiperidin-1-yl)thiophene-2-formaldehyde were synthesized under specific conditions and passed A series of chemical reactions and purification processes have been prepared to respond sensitively to changes in cell viscosity.
It has achieved efficient and sensitive monitoring of intracellular viscosity changes, showing a wide range of application prospects in the field of biomedical science, especially in the research of cancer and metabolic diseases, providing new tools for in-depth understanding of disease mechanisms and developing new diagnostic and treatment methods.
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Figure CN119119012A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the field of chemical medicine, and in particular to a preparation method and application of a near-infrared fluorescent probe responsive to viscosity. Background Art
[0002] Cell viscosity is an important parameter that reflects the physical properties of the internal environment of cells. It plays a key role in many physiological and pathological processes, such as protein folding, enzyme activity, intracellular molecular movement, and cell signaling. Changes in cell viscosity are often associated with the occurrence and development of a variety of diseases, including cancer, neurodegenerative diseases, and metabolic disorders. Therefore, studying changes in cell viscosity is of great significance for understanding the mechanisms of these diseases and developing new diagnostic and therapeutic methods. Near-infrared fluorescent probes are a class of compounds that emit fluorescence in the near-infrared spectral range. Because of their low light scattering and absorption characteristics in biological tissues, they have been widely used in biomedical imaging and analysis in recent years. In particular, near-infrared fluorescent probes based on the TICT effect (twisted intramolecular charge transfer effect) have shown great potential in viscosity response and can sensitively reflect changes in viscosity of the intracellular microenvironment. Therefore, a method for preparing a near-infrared fluorescent probe that responds to viscosity and its use are disclosed for real-time monitoring of changes in intracellular viscosity. This project is supported by the Qinghai Province Applied Basic Research Project (2024-ZJ-701). Summary of the invention
[0003] Based on the current status of the above prior art research, the present invention provides a method for preparing a near-infrared fluorescent probe that responds to viscosity and its use. The fluorescent probe of the present invention has a sensitive fluorescent response to changes in cell viscosity and is used to monitor changes in intracellular viscosity in real time.
[0004] Specifically, the present invention provides a fluorescent probe, the structure of which is shown in the following formula (I):
[0005]
[0006] The near-infrared fluorescent probe technology based on the TICT effect of the present invention provides an efficient and sensitive tool for real-time monitoring of changes in intracellular viscosity. This technology has shown broad application prospects in the research of cancer, neurodegenerative diseases and metabolic diseases. Future development will further promote the application of this technology in the biomedical field, providing new perspectives and tools for in-depth understanding of disease mechanisms and the development of new diagnostic and treatment methods.
[0007] The specific preparation method of the fluorescent probe of the present invention is as follows: dihydroisophorone (0.493mmol, 184mg) and 5-(3,5-dimethylpiperidin-1-yl)thiophene-2-carboxaldehyde (0.448mmol, 100mg) are added to a 100mL round-bottom flask, and then 1,4-dioxane (10mL), glacial acetic acid (HAC, 0.4mL) and piperidine (0.4mL) are added in sequence. The mixture is refluxed at 130°C for 2h. The mixture is cooled to room temperature, extracted with dichloromethane, and washed with brine. The combined organic phase is dried over anhydrous sodium sulfate. The solvent is evaporated under reduced pressure. The crude product is purified by silica gel column chromatography (PE / EA=4 / 1). BRIEF DESCRIPTION OF THE DRAWINGS
[0008] Figure 1 It is a schematic diagram of the fluorescence spectrum of Example 1 in a water-glycerol mixture with a glycerol content of 0% to 90%.
[0009] Figure 2 It is a schematic diagram of the cytotoxicity results of Example 1 on HeLa cells (a) and HepG2 cells (b).
[0010] Figure 3 This is a schematic diagram of the laser fluorescence confocal image of HepG2 cells in Example 1.
[0011] Example 1
[0012] (E)-2-(3-(2-(5-(3,5-dimethylpiperidin-1-yl)thiophen-2-yl)vinyl)-5,5-dimethylcyclohex-2-en-1-ylidene)malononitrile: 1 H NMR (600MHz, CDCl3) δ7.09 (d, J = 15.3Hz, 1H), 6.94 (d, J = 4.2Hz, 1H), 6.63 (s, 1H), 6.34 (d, J = 15.2Hz, 1H), 5.96 (d, J = 4.2Hz, 1H), 3. 55(d,J=4.0Hz,2H),2.54–2.52(m,2H),2.48(t,J=11.9Hz,2H),2.37(s,2H),1.88–1.77(m,4H),1.04(s,6H),0.95(d,J=6.4Hz,6H). 13C NMR (151MHz, CDCl3) δ168.44,162.64,155.10,133.89,131.86,126.06,122.18,120.11,114.80 ,113.94,104.22,57.69,42.91,41.52,39.25,31.94,30.50,29.71,28.06,22.71,19.07,14.14.
[0013] Experimental Example 1: Fluorescence response experiment of Example 1 in glycerol-PBS mixtures of different ratios Experimental method: The compound in Example 1 was added to 2 mL of PBS (pH=7.4) and glycerol solutions of different viscosities, with a final concentration of 10 μM. Fluorescence spectra were measured in these solutions.
[0014] Experimental results: The results are as follows Figure 1 As shown, the results show that the fluorescence intensity of Example 1 increases with the increase of viscosity. Experimental Example 2: Cytotoxicity test of Example 1 on HeLa cells and HepG2 cells Experimental method: 1×10 4 HeLa cells and HepG2 cells were treated with the compounds in Example 1 for 24 hours, and the cytotoxicity was determined by MTT method.
[0015] Experimental results: The results are as follows Figure 2 As shown, the results show that Example 1 has less toxicity to HeLa cells and HepG2 cells, and the survival rate remains above 85% after incubation with the cells.
[0016] Experimental Example 3: Fluorescence intensity changes of Example 1 under different cell viscosities Experimental method: Dexamethasone (DXM, 20 μM) and lipopolysaccharide (LPS, 20 μM) were added to the HepG2 cell culture medium. After incubation for 30 minutes and washing 3 times with PBS, Example 1 was added and incubated for another 30 minutes. Confocal fluorescence imaging of HepG2 cells was performed using bright field and red fluorescence channels.
[0017] Experimental results: The results are as follows Figure 3 As shown, the results show that the fluorescence intensity of Example 1 is different in different cell viscosity microenvironments, and the higher the viscosity, the stronger the fluorescence intensity.
Claims
1. The present invention discloses a method for preparing a near-infrared fluorescent probe that responds to viscosity and its use. The near-infrared fluorescent probe that responds to viscosity of the present invention has a structure as shown in the following formula (I). The fluorescent probe shown in the present invention can visualize and quantify viscosity changes in different cells. A dihydroisophorone derivative, characterized in that: Its structure is shown in the following formula (I):
2. The fluorescent probe according to claim 1, characterized in that The compound is shown below:
3. A new use of the fluorescent probe as claimed in claim 1, characterized in that: Used to visualize and quantify viscosity changes within different cells.