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Dithiothreitol fluorescence probe

A technology of dithiothreitol and fluorescent probes, which is applied in the field of analytical chemistry, can solve the problems of few specific detection and identification of dithiothreitol molecular fluorescent probes, and achieve good fluorescence emission spectrum characteristics and cheap raw materials Easy to obtain and broad application prospects

Inactive Publication Date: 2018-04-17
UNIV OF JINAN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although some assays for the detection and identification of specific thiols have been reported, to our knowledge there are very few molecular fluorescent probes for the specific detection and identification of dithiothreitol.

Method used

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  • Dithiothreitol fluorescence probe
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  • Dithiothreitol fluorescence probe

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1 Preparation of dithiothreitol fluorescent probe.

[0038] 4-Diethylaminoketoacid (1 mmol) and 1,6-dihydroxynaphthalene (1 mmol) were heated to reflux at 90°C in concentrated sulfuric acid (4 mL); then the reaction solution was cooled to 0°C, and perchloric acid ( 4 mL) to make it precipitate, filter with suction and dry to obtain a solid, dissolve the solid in methanol, fill the chromatography column with silica gel powder, use a mixed solvent of dichloromethane:methanol=10:1 as the eluent, and after elution The solution was desolvated to obtain compound 1 as a solid.

[0039] Add compound 1 (1 mmol), 2,4-dinitrobenzenesulfonyl chloride (1 mmol), triethylamine (0.2 mmol) and dichloromethane (10 mL) into a 50 mL one-necked flask and stir at 0 °C After 2 hours, the reaction was complete. After spin-drying, the obtained solid was purified by column chromatography, and the light red product obtained was the fluorescent probe for detecting dithiothreitol (DTT) acc...

Embodiment 2

[0040] Example 2 Responses of dithiothreitol fluorescent probes to different concentrations of dithiothreitol.

[0041] Dissolve the dithiothreitol fluorescent probe prepared in Example 1 in methanol, and then prepare a 10 μM probe solution with phosphate buffered saline (pH 7.4). The methanol content in the solution is 20wt%; Thiothreitol solution, the concentration of which is 0, 10, 20, 40, 60, 80, 100, 120, 140, 160, 180, 200, 220, 240, 260, 280, 300 μM; The probe solution was mixed with different concentrations of dithiothreitol solutions for 30 min for fluorescence detection (λ ex =580nm, λ em = 638nm), measure the fluorescence intensity in each system.

[0042] Taking the wavelength as the abscissa and the fluorescence intensity as the ordinate, the response of the fluorescent probe to different concentrations of dithiothreitol is as follows: figure 2 Shown: the peak value of the fluorescence intensity is around 638nm, as the concentration of thiothreitol increases...

Embodiment 3

[0043] Example 3 Effects of different reaction times on the fluorescence intensity of the dithiothreitol fluorescent probe.

[0044] Dissolve the dithiothreitol fluorescent probe prepared in Example 1 in methanol, and then prepare a 10 μM probe solution with phosphate-buffered saline (pH 7.4), in which the methanol content is 20wt%; 300 μM dithiothreitol solution was prepared in buffered saline (pH 7.4). Then 5 mL of fluorescent probe solution was mixed with 300 μM dithiothreitol solution for 30 min for fluorescence detection (λ ex = 580nm, λ em = 638nm), the fluorescence intensity in the system was measured every 2min for 40min.

[0045] Taking the wavelength as the abscissa and the fluorescence intensity as the ordinate, the fluorescence intensity of the fluorescent probe to dithiothreitol with different reaction times is as follows: Figure 4 Shown: the peak value of the fluorescence intensity is around 638nm, with the prolongation of the reaction time, the peak value o...

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Abstract

The invention discloses a dithiothreitol fluorescence probe which has a chemical structural formula as shown in the specification. The dithiothreitol fluorescence probe can be used for detecting dithiothreitol in solutions and cells, and fluorescence detection conditions comprise a single photon excitation wavelength of 580nm, a two-photon excitation wavelength of 800nm, an emission wavelength of638nm and a detection band of 520-800nm.

Description

technical field [0001] The invention relates to a rhodamine derivative-based dithiothreitol fluorescent probe and an application thereof, belonging to the technical field of analytical chemistry. Background technique [0002] Dithiothreitol (DTT), a synthetic thiol, has been widely used in cell biology, biochemistry, and biomedical applications. DTT is often used as a strong reducing agent, antidote for protecting cells and tissues, radioprotectant and so on. DTT can keep the sulfhydryl (SH) group in a reduced state, and is often used to reduce disulfide bonds in proteins and polypeptides, and is commonly used in vaccine preparations to prevent protein cysteine ​​residues from forming intramolecular and intermolecular bonds. disulfide bond. DTT also has anti-oxidation effect. For some hidden disulfide bonds, disulfide bonds can be reduced under denaturing conditions, such as high temperature or sodium ion denaturant. In addition, DTT is a toxic substance. For example, in ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D311/82C09K11/06G01N21/64
CPCC07D311/82C09K11/06C09K2211/1088G01N21/6428
Inventor 林伟英张楠董宝利孔秀琪王超宋文辉
Owner UNIV OF JINAN
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