A magnetic mesoporous silica core-shell structure affinity microsphere based on metal ion modification and its preparation method and application

A technology of metal ion modification and mesoporous silica, which is applied in peptide preparation methods, microsphere preparation, chemical instruments and methods, etc. It can solve the problems of unavoidable mass spectrometry detection interference, large loss, complex and cumbersome process, etc.

Inactive Publication Date: 2016-03-16
SHENZHEN UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The relative concentration of the sample can be increased by freeze-drying or evaporating and concentrating the sample. However, due to the corresponding increase in the concentration of salt, it is difficult to avoid interference with mass spectrometry detection; the commonly used desalination method is to use SiO modified with hydrophobic alkyl chains. 2 Packing material, adsorbing peptides and proteins to achieve the purpose of purification, but for some complex samples containing a large number of high-abundance non-target proteins, the desalting ability is limited
In recent years, it has also been reported in the literature that the use of mesoporous SiO 2 The pore size exclusion effect purifies peptides, but because it only relies on the surface hydrophilic group silanol hydroxyl group and the effect of peptides, the affinity enrichment ability for peptides is limited
In addition, the use of these materials to enrich peptides and proteins often requires repeated centrifugation, which is complicated and cumbersome, and the loss is large

Method used

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  • A magnetic mesoporous silica core-shell structure affinity microsphere based on metal ion modification and its preparation method and application
  • A magnetic mesoporous silica core-shell structure affinity microsphere based on metal ion modification and its preparation method and application
  • A magnetic mesoporous silica core-shell structure affinity microsphere based on metal ion modification and its preparation method and application

Examples

Experimental program
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Effect test

Embodiment 1

[0030] Example 1 Preparation of magnetic mesoporous silica core-shell affinity microspheres based on metal ion modification

[0031] 1. Fe 3 o 4 Magnetic particle preparation

[0032] Prepared by solvothermal method, the specific steps are as follows: Weigh 5.13g of ferric chloride hexahydrate (FeCl 3 ·6H 2 O) Uniformly disperse into 80mL ethylene glycol to make a solution, and add 1.25g of surfactant PEG and 5.00g of anhydrous sodium acetate in turn under magnetic stirring, and stir to form a uniform system. The mixture was transferred to a 120mL autoclave with a polytetrafluoroethylene liner and sealed, and finally the autoclave was placed in a constant temperature box and heated to 200°C for 6 hours of constant temperature reaction. After the reaction system was naturally cooled, the resulting suspension was magnetically separated to obtain a black powdery substance. After washing with deionized water and ethanol several times alternately, and drying at 60°C for 8 hou...

Embodiment 2

[0036] Example 2 Enrichment of low-abundance standard peptides using magnetic mesoporous SiO2 core-shell structure affinity microspheres based on metal ion modification

[0037] 1. Preparation of sample solution: Prepare 50mL of 5nM synthetic polypeptide MW=1533.6 solution.

[0038] 2. Enrichment and mass spectrometry analysis of synthetic peptides: Take 500 μL of the above peptide solution and add 100 μg of magnetic mesoporous SiO 2 Affinity microspheres, disperse evenly, shake for 10min. Remove the supernatant after magnetic separation, 5 μL of 80% acrylonitrile (CAN), 0.5% trifluoroacetic acid (TFA) (v / v), shake for 5 minutes, and magnetically separate. Collect the supernatant, take 0.7uL sample, mix with 0.7 matrix, and spot on the MALDI-TOF target plate to form co-crystals for mass spectrometry analysis.

[0039] 3. Analysis results: Figure 3A It is the result of direct mass spectrometry detection before 5nM standard peptide enrichment. Because the peptide concentr...

Embodiment 3

[0040] Example 3 Utilizes Magnetic Mesoporous SiO Based on Metal Ion Modification 2 Enrichment of low-abundance peptides in complex samples using core-shell affinity microspheres

[0041] 1. Preparation of sample solution: Weigh 1mg of bovine serum albumin BSA and dissolve it in 1mL of 50mM ammonium bicarbonate solution (pH8.0), add trypsin to enzymatically digest it for 16 hours according to the mass ratio of 50:1 to trypsin, The enzymolysis temperature was controlled at 37°C, and 1% formic acid was finally added to terminate the reaction. Take 50uL of the polypeptide solution, dilute it to 5nM with an aqueous solution containing 4M urea, and set aside.

[0042] 2. Enrichment and mass spectrometry analysis of peptides: take 500uL of the above peptide solution, add 100ug of magnetic mesoporous SiO 2 Affinity microspheres, disperse evenly, shake for 10min. After magnetic separation, remove the supernatant, wash the affinity microspheres with 500uL deionized water, magnetic...

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Abstract

The invention relates to the technical field of a functional material, and discloses a preparation method of a magnetic mesoporous silica core-shell structure affinity microsphere based on metal ion modification. The method comprises the following steps of: 1) preparing magnetic particles from hexahydrate ferric trichloride; 2) preparing particle dispersion liquid by utilizing the magnetic particles; 3) adding ethanol solution containing ethyl silicate and titanium tetrabutoxide to the particle dispersion liquid, carrying out ultrasound, agitation and magnetic separation, and removing template molecules to obtain the magnetic mesoporous silica core-shell structure taking the magnetic particles as a core and the SiO2 as a shell. Polypeptide and protein with low molecular weight can be gathered and purified from a complicated biological sample by utilizing the magnetic mesoporous silica core-shell structure affinity microsphere based on the metal ion modification. The purification and separation processes of complicated low-abundance protein and low-molecular weight protein sample are greatly simplified by utilizing the magnetic property of an affinity material; and the operation time is also shortened.

Description

technical field [0001] The invention relates to the field of functional materials, and discloses a magnetic mesoporous silicon dioxide core-shell structure affinity microsphere based on metal ion modification, a preparation method and application thereof. Background technique [0002] There are many small molecular weight endogenous peptides, proteins and other biomolecules in human body fluid, which are the cause and result of some diseases. Detection of these small molecular weight biomolecules plays a very important role in disease diagnosis. [0003] Mass spectrometry is an effective tool for rapid and sensitive detection of biomolecules, especially peptides and proteins. Mass spectrometry detection was not possible due to the low concentration of target peptides in some samples, high concentration of salt or other non-target biomolecules. Therefore, before mass spectrometry detection, it is necessary to efficiently separate, enrich, desalt, and remove impurities for t...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): B01J13/02C07K1/14
Inventor 程功张吉林陈平倪嘉缵王勇李水明洪广言
Owner SHENZHEN UNIV
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