Dithiothreitol fluorescent probe as well as preparation method and application thereof

A technology of fluorescent probe and synthesis method, which is applied in the field of analytical chemistry, can solve the problems that fluorescent probes do not have rapid response sensitivity, etc., and achieve the effects of easy promotion, high sensitivity and high specificity

Inactive Publication Date: 2018-05-08
UNIV OF JINAN
View PDF3 Cites 11 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] Aiming at the problem that most of the existing DL-dithiothreitol (DTT) fluorescent probes do not have rapid response and high sensitivity, the present invention provides a novel fluorescent probe for detecting intracellular DTT

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Dithiothreitol fluorescent probe as well as preparation method and application thereof
  • Dithiothreitol fluorescent probe as well as preparation method and application thereof
  • Dithiothreitol fluorescent probe as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1 Preparation of Fluorescent Probes

[0033] (1) 4-diethylamino keto acid and 1,6-naphthalenediol use concentrated sulfuric acid as solvent, reflux at 90°C; then cool the reaction solution to -5-0°C, add perchloric acid to make it precipitate, Suction filtration, drying and column purification, eluent dichloromethane:methanol=10:1 for purification, to obtain compound 3:

[0034] ;

[0035] (2) Add compound 3 (1 mmol) obtained in step 1, 2,4-dinitrofluorobenzene (1 mmol) and N,N-dimethylformamide (5 mL) into a 50 mL one-necked flask, 50 °C for 5 hours. Cool to room temperature, extract with dichloromethane (15mL) and purified water (15mL) for 3 to 5 times, collect the organic layer, dry with anhydrous magnesium sulfate and spin dry, and dichloromethane:methanol=50:1 for the obtained solid The eluent is purified by column chromatography, and the pink product is obtained after removing the solvent, which is the fluorescent probe for detecting intracellular DTT...

Embodiment 2

[0036] Example 2 Fluorescence spectra of fluorescent probes at different DTT concentrations

[0037] Prepare in advance 18 parts of 5 mL phosphate-buffered saline (PBS) containing 10 μM fluorescent probe in Example 1, containing 20% ​​acetonitrile, and the DTT concentrations are 0, 2, 4, 6, 8, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70 μM. Fluorescence detection is then performed (λ ex = 580nm); Calculate the fluorescence intensity in each system; Evaluate the response performance of the probe to DTT, such as figure 2 shown. Depend on figure 2 It can be seen that within a certain concentration range, the fluorescence intensity is positively correlated with dithiothreitol, and the concentration of dithiothreitol can be determined by detecting different fluorescence intensities. Analyze the linear relationship between the fluorescence intensity at 638nm and the concentration of DTT, such as image 3 As shown, the regression equation is y=0.93177x+0.177957, where ...

Embodiment 3

[0038] Example 3 Fluorescence Intensities of Fluorescent Probes Reacted with Different Substances

[0039] Prepare 10 parts of 5mL buffer solution (containing 20% ​​acetonitrile, pH = 7.4) containing 10 μM fluorescent probe in Example 1 in advance, and then add 100 μL of CuSO with a concentration of 100 μM to the system respectively 4 , MgSO 4 , CaCl 2 、CoCl 2 、NaNO 2 、Na 2 SO 3 、Na 2 SO 4 , ZnSO 4 , peroxy tert-butyl, peroxy tert-butanol, HClO, H 2 o 2 , NO, Na 2 S, SnSO 4 , KI, HgCl 2 , VC, Hcys, Cys, GSH, dithiothreitol in PBS solution. Fluorescence detection is then performed (λ ex = 580 nm); calculate the fluorescence intensity in each system, and evaluate the interference of the different substances on the fluorescent probe solution, such as Figure 4 shown. Depend on Figure 4 It can be seen that the dithiothreitol fluorescent probe can resist the influence of various interfering substances and has strong specificity.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention provides a fluorescent probe for detecting dithiothreitol in cells based on a xanthene structure. The probe is cheap and easily available, is simple in use, can realize specific recognition with the dithiothreitol to perform reaction so as to generate a xanthene fluorescent dye with higher fluorescence intensity, can not be disturbed by thiol and other reducing substances in the process of detecting the dithiothreitol, has excellent selectivity on the dithiothreitol, and can perform specific detection on the dithiothreitol in living cells.

Description

technical field [0001] The invention relates to a fluorescent probe for rapidly detecting intracellular dithiothreitol, which belongs to the technical field of analytical chemistry. Background technique [0002] DL-dithiothreitol (DTT), also known as Cleland's reagent, is a protection reagent for sulfhydryl groups and has been widely used in peptide / protein chemistry and cell biology. Discovered in the middle of the last century, this small molecule and water-soluble reagent is able to completely reduce disulfides, protect thiols from oxidation, and preserve monothiols in the reduced state. As an effective reducing agent, it is mainly used in clinical and experimental fields, and can also be used in the treatment of certain diseases, such as cystinosis or medical conditions caused by ion or metal toxicity. Recently, DTT has been used as a significant inhibitor of apoptosis and an inducer of unfolded proteins as well as a protective agent of cellular senescence. [0003] DT...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Applications(China)
IPC IPC(8): C09K11/06C07D311/78G01N21/64
CPCC07D311/78C09K11/06C09K2211/1007C09K2211/1088G01N21/643G01N21/6486
Inventor 林伟英张楠董宝利孔秀琪王超宋文辉
Owner UNIV OF JINAN
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap