Two-photon fluorescent probe for detecting pH in endoplasmic reticulum of cell

A technology of fluorescent probes and endoplasmic reticulum, applied in the direction of fluorescence/phosphorescence, luminescent materials, measuring devices, etc., can solve the problems of high cost, cumbersome synthesis steps, few fluorescent probes for positioning endoplasmic reticulum, etc., and achieve high specificity , raw materials are cheap and easy to obtain, and the effect of good fluorescence emission spectral characteristics

Inactive Publication Date: 2018-12-28
UNIV OF JINAN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Aiming at the problems existing in the prior art, such as few fluorescent probes for localizing the endoplasmic reticulum, complicated synthesis steps, and high cost

Method used

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  • Two-photon fluorescent probe for detecting pH in endoplasmic reticulum of cell
  • Two-photon fluorescent probe for detecting pH in endoplasmic reticulum of cell
  • Two-photon fluorescent probe for detecting pH in endoplasmic reticulum of cell

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1 Synthesis of Fluorescent Probes

[0039] (1) Dissolve 4-bromo-1,8-naphthalic anhydride (1 mmol) in 25 mL ethanol and heat under reflux until completely dissolved, then dissolve BOC-ethylenediamine (1 mmol) in 2 mL ethanol and add dropwise into the reaction liquid, continue to reflux and stir for 4 hours, suction filter to obtain a white solid, and perform column chromatography with ethyl acetate:petroleum ether=3:1 (v / v) as the eluent to obtain compound 1;

[0040] (2) All of the above compound 1 was dissolved in 20 mL of dichloromethane, 2.5 mL of trifluoroacetic acid was added dropwise, and stirred at room temperature for 3 hours. After spin-drying, column chromatography was performed with methanol:dichloromethane=15:1 (v / v) as the eluent to obtain compound 2;

[0041] (3) Dissolve compound 2 (0.5 mmol) in 15 mL of dichloromethane, add p-toluenesulfonyl chloride (0.5 mmol) in dichloromethane dropwise to the above solution under ice-salt bath (-5°C) , stirr...

Embodiment 2

[0044] Example 2 Responses of fluorescent probes to different pH environments

[0045] The probe obtained in Example 1 was dissolved in ethanol, and then diluted with water to form a 10 μM probe buffer solution (containing 10% ethanol). Take 13 parts of the above-mentioned probe solution, add the same volume of pH buffer solution respectively, so that the pH is 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, 10.0. Then perform a fluorescence scan (λ Ex =400 nm); calculate the relative fluorescence intensity in each system. The response of the probe to different pH as figure 2 Shown: Its maximum fluorescence intensity peak is 531nm. The corresponding light intensity at 531 nm (I 531nm ) as the ordinate and pH as the abscissa, we get image 3 , we know that I 531nm There is a linear correlation between the fluorescence intensity and pH, and the fluorescence intensity decreases with the increase of pH.

Embodiment 3

[0046] Example 3 Selectivity of Fluorescent Probes

[0047] The probe obtained in Example 1 was dissolved in ethanol, and then diluted with water to form a 10 μM probe buffer solution (containing 10% ethanol). Take 15 parts of the above-mentioned probe solution 5 mL, add them separately, then add water or different interfering substances PBS buffer to the system in turn, and then perform fluorescence scanning (λ Ex = 400 nm); calculate the relative fluorescence intensity in each system; take the corresponding light intensity at 531nm (I 531nm ) is the ordinate, and the histogram of the response of the probe to different substances is obtained, such as Figure 4 Shown: Among them, 1: water; 2: Na + (5mM); 3: K + (5mM); 4: Ca 2+ (500μM); 5: Mg 2+ (500μM); 6: Al 3+ (200μM); 7: Cu 2+ (200μM); 8: Fe 3+ (200μM); 9: Zn 2+ (500μM); 10: GSH(5mM); 11: Cys(5mM); 12:HCys(5mM); 13: H 2 o 2 (5mM); 14: NaClO (200μM); 15: pH 4.0. according to Figure 4 It can be seen that the re...

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Abstract

The invention provides a fluorescent probe for detecting pH in endoplasmic reticulum of a cell. The formula is as shown in the description. The probe is prepared by carrying out three-step reactions by taking 4-bromo-1,8-naphthalic anhydride, BOC-ethanediamine, trifluoroacetic acid and paratoluensulfonyl chloride. The fluorescent probe can be used for detecting pH in a solution or the endoplasmicreticulum of the cell. The fluorescent probe can be obtained through chemical synthesis. The synthetic process is simple and feasible, the raw materials are cheap, the preparation cost is low, and theprobe is easy to popularize, high in specificity, is not affected by other components, has a two-photon property, can be used for detecting the pH in the endoplasmic reticulum of a living cell in a tissue and has a wide application prospect.

Description

technical field [0001] The invention relates to a novel fluorescent probe for intracellular pH detection and an application thereof, belonging to the field of small molecule fluorescent materials. Background technique [0002] The endoplasmic reticulum, as an important organelle of the cell, is a series of sheet-like sacs and tubular cavities composed of membranes in the cytoplasm. It consists of two parts: the thick endoplasmic reticulum and the smooth endoplasmic reticulum. Organically connect the nucleus, cytoplasm, and cell membrane into a whole, responsible for the transport of substances in the cell, and also the synthesis base of proteins, lipids, sugars, etc. Membrane secreted proteins, phospholipids, amino polysaccharides, cholesterol, calcium signals, etc. The metabolism of cells is directly related to the function of the endoplasmic reticulum. In addition, stresses such as viral infection, amino acid deprivation, abnormal protein expression, oxidants, reducing age...

Claims

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Application Information

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IPC IPC(8): C07D221/14C09K11/06G01N21/64
CPCC07D221/14C09K11/06C09K2211/1007C09K2211/1029C09K2211/1044G01N21/6428G01N21/6486
Inventor 林伟英张楠董宝利孔秀琪宋文辉
Owner UNIV OF JINAN
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