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Chromatographic column modified with lipid rafts and its preparation method and its application in screening of antitumor active sites of traditional Chinese medicine

A silica gel chromatographic column and lipid raft technology is applied in the application field of lipid raft-silica gel chromatography in online screening of anti-tumor active parts of traditional Chinese medicines, and can solve the problem of building a method for online screening of unseen traditional Chinese medicine anti-tumor active parts and its application and consumption. Time, screening inefficiency and other problems, to achieve the effect of good application prospects, good column repeatability and long service life

Active Publication Date: 2016-01-13
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This technology overcomes the disadvantages of inefficient and time-consuming off-line screening of traditional Chinese medicine active ingredients in the past, and will open up a way for high-throughput screening of new tyrosine protein kinase inhibitors in traditional Chinese medicine
The construction method and application of an online screening model for anti-tumor active parts of traditional Chinese medicine based on lipid raft chromatography

Method used

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  • Chromatographic column modified with lipid rafts and its preparation method and its application in screening of antitumor active sites of traditional Chinese medicine
  • Chromatographic column modified with lipid rafts and its preparation method and its application in screening of antitumor active sites of traditional Chinese medicine
  • Chromatographic column modified with lipid rafts and its preparation method and its application in screening of antitumor active sites of traditional Chinese medicine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1: Preparation of silica gel chromatography column modified with lipid rafts

[0045] (1) Cultivation and expansion of U251 cells:

[0046] U251 cells were cultured and expanded in DMEM medium containing 10% fetal bovine serum (FBS), and the cell number concentration was adjusted to 1×10 7 ~10×10 7 pcs / ml, spare;

[0047] (2) Preparation of lipid rafts:

[0048] Collect the U251 cells obtained by the above method, extract the lipid raft fraction with non-ionic detergent TritonX-100, and collect by centrifugation with sucrose density gradient centrifugation, and place the lipid raft-sucrose suspension in 80mM Tris with pH 7.2 at 4°C - Dialyze in HCL solution for 20 hours, and combine the dialysate with a total of 9ml for later use;

[0049] (3) Preparation of chromatographic silica gel modified by lipid rafts:

[0050] Suspend the chromatographic silica gel pellets in 1mol / L hydrochloric acid solution, ultrasonically treat for 10min; co-flow under stirring f...

Embodiment 2

[0052] Example 2: Establishment method of lipid raft-silica gel chromatographic model for online screening of anti-tumor active sites in gallnut

[0053] (1) Preparation of gallnut extract

[0054] The medicinal material Galla chinensis is dried and pulverized. Take 400g of medicinal powder, add petroleum ether (1.5:1) and stir evenly, let stand for 1h, remove petroleum ether, repeat 3 times, and dry in the air. Add double-distilled water, co-flow at 95°C for 2 hours, repeat twice, and collect the extract. Concentrate to 300ml with a rotary evaporator, add 95% ethanol to stir, and let stand overnight. Filter, take the filtrate, evaporate the ethanol with a rotary evaporator, add a little water to evaporate until there is no ethanol smell, extract with ether, and concentrate the extract obtained with a rotary evaporator as the ether extraction part.

[0055] (2) Lipid raft chromatographic separation of Galla gall extract

[0056] The ethyl ether extract of gallnut is separa...

Embodiment 3

[0062] Example 3: Establishment method of lipid raft-silica gel chromatographic model for online screening of anti-tumor active sites in gallnut

[0063] (1) Preparation of gallnut extract

[0064] The medicinal material Galla chinensis is dried and pulverized. Take 80g of medicinal powder, add petroleum ether (1:1) and stir evenly, let stand for 2h, remove petroleum ether, repeat 3 times, and dry in the air. Add double-distilled water, keep the water bath in the same flow for 3 hours, repeat twice, and collect the extract. Concentrate to 60ml with a rotary evaporator, add 95% ethanol to stir, and let stand overnight. Filter, take the filtrate, evaporate the ethanol with a rotary evaporator, add a little water to evaporate until there is no ethanol smell, extract with ether, and concentrate the extract obtained with a rotary evaporator as the ether extraction part.

[0065] Separation of ether extracts with brick glue column, preparation of dichloromethane-ether (5:1, 4:1, ...

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Abstract

The invention discloses a chromatographic column decorated by lipid rafts. The chromatographic column is a silica gel chromatographic column which is decorated with the lipid rafts on the surface. A TrkA lipid raft-sillica gel stationary phase chromatographic system is built firstly. Stationary phase can be combined with medicinal specificity of tyrosine kinase receptors as a target spot, and then reservation is produced, but non-reservation behavior is not existed in the TrkA lipid raft-sillica gel chromatographic column of the tyrosine kinase target spot medicines. Lipid raft-sillica gel chromatograph which is rich in TrkA receptors is a novel medicine screening model which has a living being affiliation character and is capable of selective recognition medicines, wherein the medicines are reacted with the TrkA receptors. The chromatographic system is different from the traditional chromatographic system and has double characters of chromatograph separation and activity recognition. Therefore, the chromatographic system can be used for guide medicine screening study where the TrkA receptors served as drones. The chromatographic column decorated by the lipid rafts is used through different time and different moving phases, and the TrkA lipid raft-sillica gel stationary phase chromatographic system is proved stable. The chromatographic column is good in repeatability, long in service life and has good application prospects. The invention discloses a manufacturing method of the chromatographic column decorated by the lipid rafts.

Description

technical field [0001] The invention relates to a chromatographic column modified by lipid rafts, in particular to the application of lipid raft-silica gel chromatography in online screening of antitumor active parts of traditional Chinese medicines. Background technique [0002] Since the 21st century, receptor-targeted drug screening technology [SchreiberSL.Science.2000, 287:1964-1969.DrewsJ.Science.2000, 287:1960-1964] and virtual screening technology of computer-aided drug design have become the key factors in drug discovery. An effective method for leading substances. With the continuous elucidation of tumor pathogenesis and the continuous discovery of anti-tumor targets, multi-target inhibition of tumor signal transduction has become an important direction for the development of tumor drugs. Among various molecular targets, receptor protein-tyrosine kinase (PTK) is currently one of the most effective and promising anti-tumor drug targets [Li Xin, Gao Jinheng, Chen Guo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N30/60G01N30/02
Inventor 余江南童姗姗徐希明曹霞易承学付敏
Owner JIANGSU UNIV
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