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High-content rapid screening method for magnetic beads of ocean natural active materials

A technology of natural active substances and screening methods, applied in the field of natural drug screening, to achieve the effects of reasonable design, large amount of information, and shortening of screening time

Active Publication Date: 2014-03-26
张文 +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there is no literature report on the rapid screening of inhibitors of "target proteins" such as DNA topoisomerase, α-glucosidase and triglycerides from marine natural products

Method used

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  • High-content rapid screening method for magnetic beads of ocean natural active materials
  • High-content rapid screening method for magnetic beads of ocean natural active materials
  • High-content rapid screening method for magnetic beads of ocean natural active materials

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1: High content screening method based on magnetic bead bond and DNA topoisomerase, α-glucosidase and triglyceride enzyme

[0047] Take out 100 μL of magnetic beads, wash twice with an equal volume of 25mM MES (pH 6) solution for 10 minutes, and mix well. Before use, quickly dissolve EDC in cold 25mM MES (pH 6) to a concentration of 50mg / mL. Prepare 50 mg / mL NHS solution with 25 mM MES (pH 6), add 50 μL of EDC solution and 50 μL of NHS solution to the washed magnetic beads, mix well, and incubate at room temperature for 30 min with slow tilt rotation. After incubation, the tubes were placed on a magnet for 4 min, the supernatant was removed and washed twice with 300 μL of 25 mM MES (pH 6).

[0048] On the activated magnetic beads, add 60 μL of DNA topoisomerase I, α-glucosidase and triglyceride enzyme dissolved in 25 mM MES solution (pH 6) respectively, and add 40 μL of 25 mM MES solution (pH 6) to The final volume was 100 μL, vortexed to mix well, incubated o...

Embodiment 2

[0050] Embodiment 2: Application of the screening system of the present invention in marine natural products

[0051] Weigh 1 mg of sponge extract (for the preparation method of sponge extract, refer to the second edition of "Introduction to Marine Drugs", edited by Zhang Wen, Shanghai Science and Technology Press, 2012), add a small amount of DMSO to dissolve, and then add 0.1 mL of PBS solution (pH 6.8, 10mM), sonicated for 10min to dissolve, and the supernatant was taken for liquid-mass analysis. According to the molecular weight information, it was compared with the literature for structural characterization.

[0052] Take 40 μL of the supernatant, and add it to 100 μL of ammonium acetate solution bound to DNA topoisomerase Ⅰ, α-glucosidase and triglycerides, and PBS buffer dispersed in blank magnetic beads, respectively. Mix on a shaker and incubate for 1 h. Wash 3 times with PBS buffer solution, and each washing solution enters the liquid phase analysis in turn. The cl...

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Abstract

The invention relates to the technical field of screening of natural medicines and provides a high-content rapid screening method for magnetic beads of ocean natural active materials. The method is a screening and evaluating method for active materials based on magnetic bead bonds and high-content multi-target protein integration. According to the high-content rapid screening method, the configuration is as follows: a hardware system is composed of a permanent magnet, the magnetic beads and targets; a software system is responsible for acquiring hygroplasm data. Compared with a conventional ultraviolet screening method, the screening method has the advantages that the screening efficiency is obviously improved and the screening time is shortened; various biological activities can be investigated at the same time and the synergistic effect among compounds is found. The screening method is rapid, accurate, great in information amount and good in repeatability and can be used for screening a natural product library or a combinatorial chemistry library in a large scale.

Description

technical field [0001] The invention relates to the technical field of natural drug screening, specifically refers to the application of magnetic separation and chemical information collection technology to screen marine natural products for the research and development of anti-tumor, hypoglycemic and lipid-lowering drugs. Background technique [0002] The discovery of active ingredients in natural products is one of the routine ways to create new drugs. Traditional screening methods include extraction, separation, and purification of monomeric compounds, followed by activity screening. The synergistic effect of the components, and the compound may be degraded, denatured and inactive during the separation and purification process, or non-target compounds may be separated. At present, the method based on affinity selection-mass spectrometry is used more and more in the screening of active ingredients of medicinal plants. This method greatly accelerates the process of screenin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06G01N24/08
Inventor 张文王毅程翼宇
Owner 张文
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