A screening method for anti-Alzheimer's disease drugs based on zeolite adsorption
A screening method and technology for symptomatic drugs, which can be applied to measurement devices, instruments, scientific instruments, etc., can solve problems such as false positives, and achieve the effects of good reproducibility, reasonable process design, and improved screening efficiency.
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Embodiment 1
[0033] Embodiment 1 The preparation method of adsorbing acetylcholinesterase zeolite is preferred
[0034] 1. Experimental method:
[0035] Dissolve 5ml of the prepared acetylcholinesterase solution (2mg / ml) in 200mM PBS buffer at pH 8.0, put 5mg of zeolite into the buffer, stir gently, take out 20μl of incubation solution at regular intervals, The amount of protein remaining in the incubation solution was determined by the Bradford method.
[0036] The three factors affecting the adsorption were investigated respectively: firstly, different types of zeolites (Y type, ZSM-5 type, Beta type) were incubated with a certain amount of acetylcholinesterase solution (5mL, 2mg / mL) to investigate the different types of zeolites. Effect of zeolites on adsorption. Secondly, the effect of PBS buffer with different pH values (5.8, 6.2, 6.8, 7.2 and 8.0) on the adsorption was investigated. Thirdly, the influence of different incubation temperatures (25°C, 30°C, 37°C, 45°C) on the adsor...
Embodiment 2
[0042] Example 2 Research on various factors of the screening method for anti-Alzheimer's disease drugs based on zeolite adsorption
[0043] 1. Experimental method
[0044] 1.1 Dissolve 5ml of the prepared acetylcholinesterase solution (2mg / ml) in 200mM PBS buffer at pH 8.0, put 5mg of zeolite into the buffer, stir gently, incubate at 25°C for 20min, and suck off solution, wash the zeolite with PBS buffer solution to obtain the zeolite adsorbed acetylcholinesterase.
[0045] 1.2 In a centrifuge tube, add 20 μL of coptisine solution (1 mg / mL), add 0.025 mg of zeolite adsorbing acetylcholinesterase prepared in step 1.1, add 200 μL of PBS buffer, and adjust the solution to different pH values (5.7, 6.2, 6.8, 7.4, 8.0) and different ionic strengths (10, 50, 100, 200, 500, 1000mM), and then incubated in different incubation temperatures (20, 25, 30, 37, 45°C) in different water baths time (5, 10, 20, 30, 40 min), respectively suck out the incubation solution, add 200 μL of PBS ...
Embodiment 3
[0055] Example 3 Screening of Anti-Alzheimer's Disease Components in Corydalis Corydalis Extract
[0056] 1. Experimental method
[0057] 1.1. Weigh 1g of Corydalis Corydalis powder, add 50ml of water, reflux for 1h, filter, centrifuge the filtrate at 13400rpm for 10min, absorb the supernatant for later use.
[0058] 1.2. Dissolve 5ml of the prepared acetylcholinesterase solution (2mg / ml) in 200mM PBS buffer at pH 8.0, put 5mg of zeolite into the buffer, stir gently, incubate at 25°C for 20min, and absorb The solution was removed, and the zeolite was washed with PBS buffer solution to obtain the zeolite adsorbing acetylcholinesterase.
[0059] 1.3. In a 1.5mL centrifuge tube, add 20 μL of Corydalis extract solution (10 mg / mL) obtained in step 1 and the zeolite adsorbing acetylcholinesterase obtained in step 2, and add 180 μL of PBS buffer solution (so that the ionic strength is 10mM, pH value 7.4), incubated at 37°C for 20 minutes, centrifuged and sucked out the incubation s...
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