Preparation and application of fluorescent probe for detecting viscosity change
By developing a fluorescent probe M556 targeting cell mitochondria, the problem of difficulty in detecting intracellular viscosity in the prior art is solved, effective detection of intracellular mitochondrial viscosity is achieved, and obvious fluorescence intensity differences are shown in tumor cells and normal cells.
Patent Information
- Application Number
- CN202311683539.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2023-12-09
- Publication Date
- 2025-06-10
AI Technical Summary
The prior art is difficult to effectively detect intracellular viscosity, especially in clinical samples, where fluorescent probes have challenges in detecting intracellular viscosity.
A fluorescent probe called M556 targeting cellular mitochondria was developed, which was able to display enhanced fluorescence intensity at 780 nm under excitation light of 720 nm, which increased with increasing system viscosity.
This fluorescent probe can effectively target cell mitochondria, showing obvious fluorescence intensity differences in tumor cells and normal cells, realizing the detection of mitochondrial viscosity in cells, and has potential application value in the field of fluorescent bioimaging.
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Figure CN120118076A_ABST
Abstract
Description
Technical Field
[0001] The present invention belongs to the field of organic small molecule fluorescent probes, and particularly relates to the synthesis and biological application of a fluorescent probe capable of detecting the viscosity of cell mitochondria. Background Art
[0002] Monitoring the abnormal viscosity of biological systems is of great significance for basic scientific research and clinical applications. Due to its characteristics such as real-time detection, non-invasive analysis, and high sensitivity, fluorescent probes are suitable for the visualization of clinical tumor tissues. However, fluorescent detection of intracellular viscosity in clinical samples is still challenging. Therefore, the development of new near-infrared fluorescent probes for intracellular viscosity measurement has good application value.
[0003] The compound molecules in this specification can very sensitively detect the viscosity change in solution. When excited by an excitation light at 720 nm, the fluorescence intensity at 780 nm increases with the increase of the system viscosity. Cell co-localization experiments show that it can well target cell mitochondria, and there are obvious differences in fluorescence intensity between MDA-MB-231 cells (tumor cells) and MCF-7 cells (normal cells). Summary of the Invention
[0004] The present invention provides a fluorescent probe with simple synthesis and capable of detecting the viscosity of mitochondria in tumor cells.
[0005] To achieve the above object, the present invention adopts the following technical solutions.
[0006] The present invention includes a fluorescent compound molecule for detecting viscosity that targets cell mitochondria, formula (1), abbreviated as: M556.
[0007] Formula (1) The present invention has the following advantages: The fluorescent compound described in the present invention is a kind of fluorescent compound for detecting the viscosity change of intracellular mitochondria. The probe has simple synthesis and effective application. This compound molecule can detect the viscosity in solution, target cell mitochondria, and detect the viscosity in tumor cells and normal cells, indicating its potential application value in the field of fluorescent bioimaging. Brief Description of the Drawings
[0008] Figure 1 is the nuclear magnetic resonance hydrogen spectrum of compound M556; Figure 2 is the nuclear magnetic resonance carbon spectrum of compound M556; Figure 3 is the fluorescence intensity of compound M556 in the ethanol-glycerol system (red is ethanol, blue is glycerol); Figure 4 Cell co-localization imaging of compound M556; Figure 5 Fluorescence intensity changes of compound M556 in tumor cells and normal cells. Embodiment
[0009] The present invention will be further described below in conjunction with embodiments, but these embodiments are by no means any limitation to the present invention.
[0010] Embodiment 1: Structural formula of compound M556
[0011] Synthesis of compound M556: Compound 1, anhydrous acetonitrile and compound 2 were successively added into a two-necked flask. Then piperidine was slowly added with a syringe, protected by nitrogen, and refluxed and stirred at 85 °C for 24 h. After the reaction was completed, the reaction solution was rotary evaporated while it was hot, dissolved in dichloromethane, and then purified by column chromatography. The eluent was dichloromethane:methanol = 90:1 (volume ratio) with gradient elution to 40:1, and finally a green solid compound M556 was obtained.
[0012] Embodiment 2: Response of compound M556 to viscosity in solution The compound M556 in Embodiment 1 was dissolved in DMSO to prepare a stock solution, and then an equal volume of the stock solution was added to an equal volume of ethanol-glycerol systems with different viscosities (from 100% ethanol to 100% glycerol), and then fluorescence detection was carried out. The results are shown in the appendix Figure 3 : As can be seen from Appendix 3, as the volume fraction of glycerol increases, the fluorescence intensity of M556 at 780 nm gradually increases. The results show that in the ethanol-glycerol solution system, as the viscosity of the system changes, the fluorescence intensity of compound M556 will also change accordingly.
[0013] Embodiment 3: Cell co-localization imaging of compound M556 Plate MDA-MB-231 cells in a confocal dish. After normal culturing for 24 h, add compound M556 (working concentration: 5 μM) in Example 1 into the dish, incubate in an incubator for 30 min, then add 1.0 μM commercial mitochondrial dye Mito-Tracker Green and nuclear dye Hoechst 33342 and continue to incubate for 15 min. Finally, wash with PBS and perform imaging. The conditions for laser scanning confocal microscopy imaging are as follows: Compound M556 (using 720 nm as the excitation light and 740 - 800 nm as the emission spectral range); Mito-Tracker Green (using 488 nm as the excitation light and 500 - 550 nm as the emission spectral range); Hoechst 33342 (using 405 nm as the excitation light and 410 - 450 nm as the emission spectral range). The imaging results are as shown in Figure 4 shown, indicating that compound M556 can target mitochondria in cells well.
[0014] Example 4: Fluorescence intensity changes of compound M556 in tumor cells and normal cells After adding compound M556 (working concentration: 5 μM) into MDA-MB-231 cells (tumor cells) and MCF-7 cells (normal cells) respectively and incubating for 30 min, perform laser scanning confocal imaging under the same imaging conditions. Compound M556 (using 720 nm as the excitation light and 740 - 800 nm as the emission spectral range); Mito-Tracker Green (using 488 nm as the excitation light and 500 - 550 nm as the emission spectral range). The results are as shown in Figure 5 shown, the fluorescence intensity in MDA-MB-231 cells is significantly stronger than that in MCF-7 cells, indicating that compound M556 can distinguish cancer cells and non-cancer cells through different fluorescence intensities.
Claims
1. A compound represented by the following formula (1): Formula (1).
2. Use of the compound of formula (1) according to claim 1 for detecting the viscosity of cell mitochondria.
3. Use of the compound of formula (1) according to claim 1 in the preparation of a product for detecting the viscosity of cell mitochondria.
4. The use according to claim 3, wherein the product is a kit.
5. Use of the compound of formula (1) according to claim 1 in the preparation of a product for diagnosing a disease associated with a change in mitochondrial viscosity.
6. The use according to claim 5, wherein the product is a kit.
7. The use according to any one of claims 5-6, wherein the disease is selected from one or more of malignant tumors.
8. Use of the compound of formula (1) according to claim 1 for detecting the viscosity of a solution.
9. The use according to claim 8, characterized in that the solution system for detecting viscosity is an ethanol-glycerol system.