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Agent for detecting glutathione and its synthesis method and use

A technology for glutathione and a synthesis method, which is applied to a reagent for detecting glutathione and its synthesis. The application of the reagent in the detection of glutathione can solve the problems of less reagents and the like, and achieve high sensitivity and selection. The effect of low cost of detection system and simple detection method

Inactive Publication Date: 2017-07-14
SHANXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, most fluorescent probes can realize the differential detection of the three biothiols Cys / Hcy / GSH to the other 22 amino acids, but there are still relatively few reagents for the specific detection of GSH

Method used

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  • Agent for detecting glutathione and its synthesis method and use
  • Agent for detecting glutathione and its synthesis method and use
  • Agent for detecting glutathione and its synthesis method and use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] The preparation of embodiment 1 (Z)-2-(3-chloro-3-(7-(diethylamino)-2-oxygen-2 hydrogen-chromene-3-yl) allyl) malononitrile and characterization

[0032] Slowly add 10 mL of phosphorus oxychloride dropwise to 10 mL of DMF solution under ice bath, stir for half an hour, add 3-(diethylamino)phenol (4.95 g, 20 mmol) dissolved in DMF, and reflux at 75°C for 4 h; After the reaction was completed, the reaction solution was poured into 60mL of ice water and washed with Na 2 CO 3 The pH of the aqueous solution was adjusted to neutral, and a brown solid was precipitated, filtered with suction, washed with water, and finally recrystallized with ethanol to obtain 4-(diethylamino)-2-m-hydroxybenzaldehyde;

[0033] 4-(diethylamino)-2-m-hydroxybenzaldehyde (1.16g, 6mmol) and ethyl acetoacetate (1.17g, 9mmol) were refluxed in ethanol for 10 hours under the catalyst of 0.2mL piperidine, concentrated and cooled Afterwards, the yellow solid obtained by distillation under reduced press...

Embodiment 2

[0038] Prepare a PBS buffer solution with a pH of 7.4 and a concentration of 10 mM, and prepare 2 mM (Z)-2-(3-chloro-3-(7-(diethylamino)-2-oxo-2hydro-chromene- 3-yl) allyl) malononitrile solution; put 2mL of PBS-DMSO (1:1, pH7.4) solution and 5μL (Z)-2-(3-chloro-3-(7-( Add the DMSO solution of diethylamino)-2-oxo-2hydrogen-chromen-3-yl)allyl)malononitrile to a clean fluorescent cuvette, take the GSH solution, and gradually add it in a small amount Add the sample into the cuvette, and detect it on the fluorescence spectrophotometer while adding the sample. With the addition of GSH, the fluorescence intensity at 505nm gradually increases. Fluorescence emission map see Figure 4 .

Embodiment 3

[0040] Prepare a PBS buffer solution with a pH of 7.4 and a concentration of 10 mM, and prepare 2 mM (Z)-2-(3-chloro-3-(7-(diethylamino)-2-oxo-2hydro-chromene with DMSO) -3-yl) allyl) malononitrile solution; in 22 fluorescent cuvettes, add 2 mL of PBS-DMSO (1:1, pH 7.4) solution and 5 μL of reagent in DMSO solution, and then respectively Add 30 μL of GSH, and 300 μL of various analytes: Cys, Hcy, Ala, Arg, Asp, Gln, Glu, Gly, His, lle, Leu, Lys, Met, Phe, Pro, Ser, Thr, Tyr, Trp, Val and Asn are detected on a spectrofluorometer, and a histogram of the relative fluorescence intensity at 505nm corresponding to different analytes is drawn, (see Figure 5 ). GSH changed the fluorescence intensity of the reagent from 16 to 2983, and other analytes basically did not cause changes in the fluorescence intensity of the reagent.

[0041] It has been proved by experiments that other analytes do not interfere with the determination of GSH by the system.

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Abstract

The invention provides an agent and method for detecting glutathione (GSH). The agent is a coumarin derivative (Z)-2-(3-chloro-3-(7-(diethylamino)-2-oxo-2H-chromen-3-yl)allylidene)malononitrile. The method for detection comprises quantitatively detecting the content of glutathione in a PBS buffer solution at pH 7.4 by a fluorescence spectrophotometer. The method has high sensitivity and selectivity to GSH, has simple detection process, sensitivity and a fast detection rate and can produce an accurate detection result.

Description

technical field [0001] The invention relates to glutathione detection technology, in particular to a glutathione detection reagent and its synthesis method, and the application of the glutathione detection reagent. Background technique [0002] Glutathione (GSH) plays an important role in human physiological activities and is the most non-protein sulfhydryl compound in cells. It plays an important role in maintaining cell physiological functions, including intracellular redox reactions, foreign substances Metabolism, signal transduction and gene regulation, etc. The concentration of glutathione in cells is 1-10mM. It is divided into two types: reduced glutathione and oxidized glutathione, and there is a redox balance between them. Among them, reduced glutathione plays a key role in cell growth and maintaining the redox balance of normal cell function; in addition, abnormal concentration of reduced glutathione in the human body is also associated with cancer, Alzheimer's dis...

Claims

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Application Information

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IPC IPC(8): G01N21/64
Inventor 阴彩霞李晓琪霍方俊
Owner SHANXI UNIV
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