Affinity chromatography fixed phase of immobilization metal and its preparation method

A chromatographic stationary phase and metal immobilization technology, which is applied in the field of separation and purification to achieve the effects of reducing non-specific adsorption, improving detection limit and sensitivity, and high resolution

Inactive Publication Date: 2008-10-22
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

It has also been reported in the literature that the phosphate group in the phosphate ester interacts with metal ions such as zirconium to obtain an oligonucleotide monolayer, or this phenomenon is used to synthesize an immobilized metal affinity chromatography matrix based on a silicon matrix. However, there is no report on the immobilized metal affinity chromatography stationary phase based on polyglycidyl methacrylate polymer phosphate for the enrichment of phosphorylated peptides.

Method used

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  • Affinity chromatography fixed phase of immobilization metal and its preparation method
  • Affinity chromatography fixed phase of immobilization metal and its preparation method
  • Affinity chromatography fixed phase of immobilization metal and its preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] The preparation method of stationary phase comprises the following steps:

[0032] Preparation of GMA Polymer microspheres: microspheres obtained from polyglycidyl methacrylate and vinyl glycidyl methacrylate, by swelling method, with polyvinylbenzene as seed (0.5g), by methacrylic acid shrinkage Glyceride (9.0g) is polymerized with vinyl glycidyl methacrylate (3.5g) on ​​the seed, 2% (w / w) of AIBN is the initiator, and reacts in 120mL of 0.1% SDS (w / w) solution, Ultrasonic vibration until solidified, to obtain non-porous polymer microspheres with a particle size of 8-12um. (See Boling Gong, Jinxia Zhu, Long Li, Kejuan Qiang, Li Ren, Synthesis of non-porous poly(glycidylmethacrylate-co-ethylenedimethacrylate) beads and their application in separation of biopolymers, Talanta, 2006, 68:666-672.

[0033] 1) Amination reaction: use 25% (mass concentration) concentrated ammonia water as an amination reagent and mix it with 10ml / 1g of the above-prepared GMA Polymer microsphe...

Embodiment 2

[0043] The difference from Example 1 is:

[0044] Preparation of GMA Polymer microspheres: By swelling method, polyvinylbenzene is used as the seed, and glycidyl methacrylate and vinyl glycidyl methacrylate are polymerized on the seeds to obtain macroporous polymer microspheres. Particle size 8-12um. (Boling Gong, Lili Wang, Chaozhan Wang, Xindu Geng, Preparation of hydrophobic interaction chromatographic packings based on monodyspersepoly(glycidylmethacrylate-co-ethylenedimethacrylate) beads and their application, Journal of ChromatographyA, 2004, 1022: 33-39.)

[0045] 1) Amination reaction: Dissolve 20g of 3,3-diaminopropylimine in 100mL of anhydrous acetone, and add polyglycidyl methacrylate polymer microspheres into the anhydrous acetone solution (1g / 10mL) , reacted at room temperature for 4 hours, washed the product with water until neutral after the reaction, and obtained aminated microspheres;

[0046] Or replace the above-mentioned GMA Polymer microspheres with the ...

Embodiment 3

[0055] The difference from Example 1 is:

[0056] Preparation of GMA Polymer microspheres: by swelling method, with polyvinylbenzene as seed (0.5g), by glycidyl methacrylate (9.0g) and vinyl glycidyl methacrylate (3.5g) on ​​the seed Polymerization, 2% (w / w) AIBN as initiator, react in 120mL 0.1% SDS (w / w) solution, ultrasonically vibrate until solid, and obtain non-porous polymer microspheres with a particle size of 8-12um. (Boling Gong, Jinxia Zhu, Long Li, Kejuan Qiang, Li Ren, Synthesis of non-porous poly(glycidylmethacrylate-co-ethylenedimethacrylate) beads and their application in separation of biopolymers, Talanta, 2006, 68: 666-672).

[0057] 1) Amination reaction: Dissolve 40 g of ethylenediamine in 100 mL of morpholine ethaneiodic acid (MES) at a pH of about 5, add polyglycidyl methacrylate polymer microspheres into the solution (1 g / 10 mL), and heat at room temperature Under the condition of reaction for 3 hours, the product after the reaction was washed with water...

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Abstract

The invention relates to a separation and purification technique, in particular to an affinity chromatography stationary phase of immobilized metal and a preparation method thereof. The structure of the stationary phase is shown in the drawing; wherein, the particle size of GMAPolymer micro-sphere is 10nm-50um; the GMAPolymer micro-sphere is poly-methacrylic acid glycidyl esters micro-sphere. By carrying out amination and phosphoric esterification, a synthesis method of the high-performance poly-methacrylic acid glycidyl ester-group novel affinity chromatography stationary phase is gained and used for the research in phosphorylation proteomics and highly-selective separation, enrichment and purification of phosphopeptide by the chelation with zirconium ions and iron ions; meanwhile, compared with the traditional stationary phase, the affinity chromatography stationary phase reduces the nonspecific adsorption on the non-phosphopeptide.

Description

technical field [0001] The invention relates to separation and purification technology, in particular to an affinity chromatography stationary phase for immobilizing metals and a preparation method thereof. Background technique [0002] Post-translational protein modification is a hot topic in proteome research. As the most common and important post-translational modification of proteins, protein phosphorylation is involved in almost all the processes of regulating life activities, including cell proliferation, development and differentiation, neural activity, muscle contraction, metabolism, and tumorigenesis etc. Protein phosphorylation is still the main transmission mode of signal known so far. [0003] Traditional methods for protein phosphorylation analysis include radioisotope labeling, chemical modification, Edman degradation, and thin-layer chromatography. These methods are relatively cumbersome to operate, require high experimental skills, require a large amount of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01J20/285
Inventor 邹汉法封顺叶明亮周厚江
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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