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Probe used for detecting acetylcholin esterase and its inhibitor activity, application and preparation method

A technology for acetylcholinesterase and inhibitors, which is applied in the field of probes for detecting the activity of acetylcholinesterase and its inhibitors, can solve the problems of background interference and low sensitivity, and achieve the effects of high sensitivity, fast response speed and great application prospects

Inactive Publication Date: 2013-05-01
NANJING UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method often has background interference during use and has low sensitivity

Method used

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  • Probe used for detecting acetylcholin esterase and its inhibitor activity, application and preparation method
  • Probe used for detecting acetylcholin esterase and its inhibitor activity, application and preparation method
  • Probe used for detecting acetylcholin esterase and its inhibitor activity, application and preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1 Preparation of Formula I Probe Precursor and Vesicle Probe

[0039] 1. Preparation of Formula I Probe Precursor

[0040] In the first step, 0.375g (1mmol) of 10,12-diyne pentapentacic acid was dissolved in 20ml of dichloromethane, and then 2ml of thionyl chloride was added dropwise thereto to obtain a reaction solution, and the reaction was continuously stirred under nitrogen protection solution until the reaction is complete (usually stirred overnight); then, the solvent is distilled off under reduced pressure to obtain the compound of formula II as a colorless oil.

[0041] In the second step, the compound of formula II obtained in the first step is dissolved in a small amount of tetrahydrofuran to obtain a solution of the compound of formula II, and then the solution of the compound of formula II is added dropwise to 20 ml of tetrahydrofuran solution containing 0.18 g of triethylene glycol to obtain a reaction solution. The reaction solution was continuous...

Embodiment 2

[0049] Example 2 Spectral properties and color changes of the vesicle probe reacting with myristoylcholine chloride

[0050] 1. Spectral properties of the reaction of vesicle probes with myristoylcholine chloride

[0051] Take nine 60μl 1mM vesicle probes and add them to each quartz dish, then add 4-hydroxyethylpiperazineethanesulfonic acid (HEPES) buffer (10mM pH=7.4) to dilute and dilute to 3ml; then , add 0 μl, 0.6 μl, 1.2 μl, 1.8 μl, 2.4 μl, 3 μl, 3.6 μl, 4.5 μl of 10 mM myristoylcholine chloride (final concentration of 0 μM, 2 μM, 4 μM, 6 μM, 8 μM, 10 μM, 12 μM , 15 μM), the fluorescence emission spectrum and UV-vis absorption spectrum were measured after reacting for 5 minutes. The fluorescence emission spectrum was measured with excitation at 492nm, and the slit width between excitation and emission was 5nm.

[0052] Fluorescence emission spectra such as figure 1 As shown, when the concentration of myristoylcholine chloride is in the range of 0-15 μM, the fluorescenc...

Embodiment 3

[0057] Example 3 Spectral properties and color changes of the reaction product of myristoylcholine chloride and acetylcholinesterase (AChE) mixed with vesicle probes

[0058] 1. Spectral properties of the reaction product of myristoylcholine chloride and acetylcholinesterase (AChE) mixed with vesicle probes

[0059] Take seven 3.6μl 10mM myristoylcholine chloride and add to each quartz dish, add 4-hydroxyethylpiperazineethanesulfonic acid (HEPES) buffer (10mM pH=7.4) to dilute and finally make up to 3ml ; Then, add an appropriate amount (such as 2 μl) of different concentrations of acetylcholinesterase (AChE) (0U / ml, 0.025U / ml, 0.05U / ml, 0.1U / ml, 0.2U / ml, 0.3U / ml, 0.4 U / ml); after 10 minutes of reaction, 60 μl of 1mM vesicle probes were added to the quartz dish, and then reacted for 5 minutes, and finally the fluorescence emission spectrum and ultraviolet-visible light absorption spectrum were measured. The fluorescence emission spectrum was measured with excitation at 492nm,...

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Abstract

The invention relates to a probe used for detecting acetylcholin esterase and its inhibitor activity, an application and a preparation method. The probe is polymerized by 10,12-diyne pentacosanoic acid and a compound in a formula I according to weight ratio of 7:3, The vesicle probe presents blue color and no fluorescence in 4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid buffer, and presents red color and emit fluorescence after being reacted with Myristoylcholine chloride (the fluorescence emission wavelength is 550nm), the sensitivity is high, the operation is easy, and the fluorescence detection method of the probe has great application prospect in biological medicine field.

Description

technical field [0001] The invention relates to a probe for detecting the activity of acetylcholinesterase and its inhibitor, its application and its preparation method. Background technique [0002] The development of rapid detection technology for acetylcholinesterase (AChE) activity is of great significance to the promotion of human health and environmental protection. [0003] The drugs currently used to treat Alzheimer's disease are mainly acetylcholinesterase inhibitors, which control the content of acetylcholine in patients by inhibiting the activity of acetylcholinesterase (AChE). has an important role. [0004] In addition, some organophosphorus pesticides and carbamate pesticides can inhibit the activity of acetylcholinesterase (AChE), so the development of acetylcholinesterase (AChE) activity detection reagent is very important for the detection of organophosphorus pesticides and carbamate in the environment. Pesticides can also play an important role. [0005]...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07C69/606C07C67/14C08F238/00C08F2/48C09K11/06G01N21/78G01N21/64
Inventor 陈小强周国栋王芳王惠琳
Owner NANJING UNIV OF TECH
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