A method for separating and purifying glycopeptides

A technology for separation and purification of glycopeptides, applied in the field of separation and purification of glycopeptides with magnetic particles, can solve the problems of complicated procedures for separation and purification of glycopeptides, long separation and purification reaction time, etc., and achieve the effect of optimizing coupling conditions and rapid separation and purification reactions

Active Publication Date: 2011-12-28
SHANXI LIFEGEN
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0006] The object of the present invention is to provide a method for separating and purifying glycopeptides, which solves the technical problems of complicated procedures for separating and purifying glycopeptides in the hydrazine chemical method in the background technology and long separation and purification reaction time

Method used

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  • A method for separating and purifying glycopeptides
  • A method for separating and purifying glycopeptides
  • A method for separating and purifying glycopeptides

Examples

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Effect test

Embodiment 1

[0089] see figure 1 and Figure 3, the method for separating and purifying glycopeptides

[0090] (1) Separation and purification of glycopeptides

[0091] (1-1) Preparation of hydrazine functionalized magnetic particles

[0092] a cleaning

[0093] Get 200 mg of epoxidized magnetic particles and place them in a flask, wash them with water, put the flask filled with the suspension on a magnetic separator for magnetic separation after cleaning, and remove the upper layer of clear liquid that does not contain magnetic particles; it can be ordinary water, But it is better to use water without impurities, such as ultrapure water and double distilled water (twice distilled water).

[0094] b reaction (modification)

[0095] Add 50 milliliters of hydrazine hydrate with a mass fraction of 5% in the flask, place the flask in a water bath, insert a stirrer into the flask, turn on the stirrer and stir to fully mix the magnetic particles and hydrazine hydrate, and react for 3 hours. ...

Embodiment 2

[0112] (1-1) Preparation of hydrazide functionalized magnetic particles

[0113] a cleaning

[0114] Get 200 mg of carboxylated magnetic particles and place them in a flask, wash them with water, add 0.1 mole of 2-(N-morpholine) ethanesulfonic acid (MES) buffer solution of pH 4.75 for pretreatment, and wash The flask containing the suspension is placed on a magnetic separator for magnetic separation, and the upper layer of clear liquid without magnetic particles is removed; it can be ordinary water, but it is better to use water without impurities, such as ultrapure water and double distilled water (twice distilled water).

[0115] b reaction (modification)

[0116]Add 50 ml of 2-(N-morpholine)ethanesulfonic acid (MES) buffer solution containing 1 gram of adipic dihydrazide (ADH) to the flask, put the flask into a water bath, insert a stirrer, and turn on the stirrer to stir , add 0.5 g of carbodiimide (EDC) during stirring, continue to stir and react for more than 3 hours,...

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Abstract

The invention relates to a method for separating and purifying glycopeptides by using magnetic particles. Its realization step is: wash the epoxidized magnetic particle with water and place it in the flask, put the flask filled with the suspension on the magnetic separator and carry out magnetic separation, remove the clear liquid that does not contain the magnetic particle in the upper layer; add 50 milliliters of mass fraction as 5% hydrazine hydrate, put the flask in a water bath, insert a stirrer into the flask, fully mix and react, after the reaction is completed, wash with absolute ethanol and water, perform magnetic separation, and remove the upper layer without magnetic particles. Supernatant solution; dissolve or dilute the protein sample in the coupling buffer, add sodium periodate to its final concentration, and react at room temperature in the dark; use a G-25 desalting column to remove unreacted sodium periodate; then use Glycoprotein coupling, enzymatic hydrolysis to obtain glycopeptides. The invention optimizes the coupling conditions of the glycoprotein and the hydrazine functionalized magnetic particle or the hydrazide functionalized magnetic particle, and the separation and purification reaction is fast and efficient.

Description

technical field [0001] The invention relates to a method for separating and purifying glycopeptides, in particular to a method for using magnetic particles to separate and purify glycopeptides. Background technique [0002] Glycosylation of proteins is one of the most common and important post-translational modifications of proteins. It is currently known that more than half of proteins in mammals are glycosylated. Glycosylation plays an important role in the structure and function of proteins. The degree of protein glycosylation and abnormal changes in sugar chain structure are often signs of cancer and other diseases. [0003] Changes in protein glycosylation degree and glycosylation sites can be detected by changes in glycosylated polypeptides (glycopeptides). In recent years, the methods for separating and purifying glycopeptides in complex samples mainly include lectin method and hydrazine chemical method. Complex samples such as serum and total cell membrane protein....

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K1/14C12P21/06
Inventor 李铮陈超孙士生杨刚龙王婷孙秀璇邓玮娜
Owner SHANXI LIFEGEN
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