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Multifunctional lysosome pH probe based on benzimidazole and preparation method and application thereof

A technology of methyl benzimidazole and benzimidazole, which is applied in the field of fluorescent probes, can solve the problems of rarity and lack of pH fluorescent probes, and achieves good reversibility, good biocompatibility, good H+ selectivity and photostability sexual effect

Inactive Publication Date: 2021-12-24
SHANXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, there is a shortage of pH fluorescent probes that can combine the advantages of suitable pH detection range, high lysosome targeting, low toxicity, good optical properties, and sensitive ratiometric fluorescence imaging capabilities. Probes for real-time monitoring of lysosomal autophagy are rare

Method used

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  • Multifunctional lysosome pH probe based on benzimidazole and preparation method and application thereof
  • Multifunctional lysosome pH probe based on benzimidazole and preparation method and application thereof
  • Multifunctional lysosome pH probe based on benzimidazole and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] (1) Add 0.53g (3.6mmol) of p-dimethylaminobenzaldehyde and 0.40g (3.0mmol) of 2-methylbenzimidazole into a thick-walled pressure bottle with a small amount of N,N-dimethylformamide Dissolve completely, then add 2 mL of trimethylchlorosilane dropwise, stir in an oil bath at 110°C, seal and reflux for 24 hours.

[0033] (2) After the reaction is finished, cool to room temperature, add saturated aqueous sodium carbonate solution to the solution, adjust the pH to 10, and continue stirring for 30 min. The reaction product was extracted with dichloromethane, the organic phase was washed three times with a saturated sodium chloride aqueous solution, and then anhydrous sodium sulfate was added to the organic phase to remove water. The solvent was distilled off under reduced pressure to obtain a crude product.

[0034] (3) Purify the crude product by silica gel column chromatography, the eluent is petroleum ether:ethyl acetate=3:1, after the elution is completed, the solvent is...

Embodiment 2

[0036] The probe in Example 1 was dissolved in DMSO to prepare a stock solution, and the stock solution was diluted to 10 μM with ultrapure water when performing spectral measurement. The measurement is carried out with a quartz cuvette with an optical path of 1 cm, and the pH of the solution is slightly changed by adding a very small amount of 0.1 mol / L hydrochloric acid or sodium hydroxide, and the absorption spectra of the probes are measured in different pH environments. Such as figure 1 , Titration changes the pH of the solution from 7.0 to 2.5, the absorption intensity at 354nm is gradually decreasing, while the absorption intensity at 405nm is gradually increasing. These absorption peaks form an isosbestic point at 378 nm. figure 2 It is the solution image at pH 7.0, 4.5, 2.5, the color changes from light yellow to colorless.

Embodiment 3

[0038] Dilute the probe stock solution in Example 2 to 10 μM with ultrapure water, and use 0.1mol / L hydrochloric acid or sodium hydroxide titration and measure the fluorescence emission spectrum ( image 3). With the gradual increase of hydrogen ions in the solution, the fluorescence signal at the wavelength of 487nm is continuously weakened, and the intensity of the wavelength of 534nm is continuously enhanced. These fluorescence curves also intersect at one point, and the wavelength is 510nm. Figure 4 It is the image of the solution under the ultraviolet light at pH 7.0, 4.5 and 2.5, and the colors are blue, cyan and green. According to the fluorescence spectrum data, the fluorescence ratio and pH value at 487nm and 534nm place are carried out curve fitting ( Figure 5 ), the pKa value of the probe is considered to be 4.74. Among them, the fluorescence ratio has a good linear relationship with pH in the range of 4.06-5.20 ( Figure 6 ), the linear equation is: fluorescen...

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Abstract

The invention discloses a multifunctional lysosome pH probe based on benzimidazole and a preparation method and application thereof. The probe is simple in preparation process, and only 2-methylbenzimidazole and p-dimethylaminobenzaldehyde need to be subjected to a high-temperature tube sealing reaction for 12-24 hours under protection of trimethylchlorosilane; and simple separation and purification on the product is then conducted to obtain the product. The probe has a good ratio type fluorescence characteristic when the pH value is 2.5-7.0, the pKa value is 4.74, and the probe has high targeting property on lysosome, so that the probe is particularly suitable for detecting the pH value of cell lysosome. In addition, the probe also has the advantages of good H<+> selectivity, good light stability, good reversibility, large Stokes shift and the like. When being applied to monitoring of cell activities such as lysosome autophagy, the probe shows good biocompatibility and high-sensitivity real-time in-situ monitoring capability.

Description

technical field [0001] The invention relates to a fluorescent probe, in particular to a benzimidazole-based multifunctional lysosome pH probe and a preparation method thereof, as well as the application of the probe in monitoring lysosome pH change and autophagy process. Background technique [0002] The stability of pH is an important indicator of the homeostasis of the microenvironment of the body. Many diseases, including cancer, diabetes, and myocardial infarction, will lead to changes in the pH of the tissue or cell microenvironment, or complications due to changes in pH. Therefore, the real-time monitoring of the pH of the cellular microenvironment has become a research hotspot in the field of molecular diagnosis of diseases. In recent years, targeting probes targeting subcellular structures have been favored by researchers, including the development and application of targeting probes for organelles such as lysosomes, mitochondria, and Golgi apparatus. [0003] Due t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D235/14C09K11/06G01N21/64
CPCC07D235/14C09K11/06G01N21/6428G01N21/6456C09K2211/1044
Inventor 文朝朝梁文婷董川双少敏
Owner SHANXI UNIV
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