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Separation analysis method of derivatized heparin sulfate disaccharides containing free amino groups

A technology of heparan sulfate and free amino groups, which is applied in the field of separation and analysis of heparan sulfate disaccharides, can solve the problems of mass spectrometry sensitivity decrease and ion source contamination of mass spectrometry, and achieve the expansion of instrument application range, reduce instrument cost, and maximize practical value Effect

Active Publication Date: 2018-11-13
FUZHOU UNIV
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Problems solved by technology

Further, patent CN104914205B discloses a kind of N-containing non-substituted glucosamine (GlcNH 3 + ) separation and analysis method of heparan sulfate disaccharides, the mass spectrum peak area correction factor can be used for the quantitative detection of heparan sulfate and heparan disaccharides in actual biological samples, but this method uses amine ion pair reagents for analysis
At present, it is generally believed that ion pair reagents have the risk of contaminating the ion source of the mass spectrometer, which will lead to a decrease in the sensitivity of the mass spectrometer [Yang B, et al. Anal. Biochem., 2011, 415(1):59-66]

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  • Separation analysis method of derivatized heparin sulfate disaccharides containing free amino groups
  • Separation analysis method of derivatized heparin sulfate disaccharides containing free amino groups
  • Separation analysis method of derivatized heparin sulfate disaccharides containing free amino groups

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Embodiment 1

[0036] DeN-sulfated heparin (2S, 6S, NH 3 + -HP) and de-2,6,N-sulfated heparin (NH 3 + -HP) sample disaccharide components for example analysis.

[0037] A method for separating and analyzing derivatized heparan sulfate disaccharides containing free amino groups, the steps are as follows:

[0038] (1) Dissolve ammonium acetate in deionized water, adjust the pH value to 5.6 with acetic acid, and prepare ammonium acetate solution with a concentration of 40 mM, which is used as mobile phase A; methanol is used as mobile phase B.

[0039] (3) Take 20 μg each of self-made heparin derivative de-N-sulfated heparin and de-2,6,N-sulfated heparin, and pass through the mixture of heparinase Ⅰ, heparinase Ⅱ and heparinase Ⅲ. After complete enzymatic hydrolysis, filter. Add 10 μL of 0.1mol / L 2-aminoacridone solution (2-aminoacridone is dissolved in a 3:17 mixed solvent of acetic acid: dimethyl sulfoxide), and react for 20 minutes. Add 10 μL of 1.0 mol / L sodium cyanoborohydride soluti...

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Abstract

The invention discloses a separation analysis method of derivatized heparin sulfate disaccharides containing free amino groups. The function relationship of the disaccharide quantity and mass spectrumdetection peak area can be used for quantitative determination of heparin and heparin sulfate disaccharide in practical biological samples. The separation analysis method can be used for detection ofheparin and heparin sulfate disaccharides, and especially heparin sulfate disaccharides containing free amino groups in liquid phase-mass spectrometry, and routine liquid chromatogram separating combined ultraviolet and fluorescent detector can be adopted in analysis. Important effect on clinical study on medical heparin, heparin sulfate, heparin derivatives, and disaccharides in various biological samples is achieved.

Description

technical field [0001] The invention belongs to the technical field of drug detection, and in particular relates to a method for separating and analyzing derivatized heparan sulfate disaccharides containing free amino groups. Background technique [0002] Heparin and heparan sulfate are linear glycosaminoglycans. Because heparin has a strong anticoagulant effect, it is widely used in the treatment of thromboembolic diseases, myocardial infarction and other diseases. Different sulfation, acetylation and free amino substitution patterns in heparin form different disaccharide units, endowing heparin and heparan sulfate with numerous biological functions, participating in angiogenesis, blood coagulation, cell adhesion, proliferation and tumor metastasis and other life activities. Heparin and heparan sulfate are composed of repeating disaccharide units composed of hexuronic acid and glucosamine connected by 1-4 glycosidic bonds, of which the conventional N-sulfate is present in ...

Claims

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Application Information

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IPC IPC(8): G01N30/06G01N30/88
CPCG01N30/06G01N30/88
Inventor 魏峥杜佳燕林江慧
Owner FUZHOU UNIV
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