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Purification preparation method of ganglioside monomers

A ganglioside and monomer technology, applied in chemical instruments and methods, preparation of sugar derivatives, organic chemistry, etc., can solve the problems of sample loss, high organic residues, different charges, etc. High sample load and good resolution

Pending Publication Date: 2021-02-26
TAIZHOU GUOKEHUAWU BIOMEDICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, the methods for the separation and preparation of ganglioside crude lipids mainly include: 1. Ion-exchange chromatography, which mainly depends on the different numbers of sialic acid in gangliosides, resulting in different charges and different adsorption by fillers. However, there are salt ions in the mobile phase, and the subsequent desalination steps are complicated, resulting in serious sample loss; 2. Macroporous adsorption resin separation method, organic compounds are separated on the macroporous adsorption resin according to the difference in adsorption force and molecular weight. The purpose of separation is achieved by elution with a certain solvent, but this method has high organic residues, difficult pretreatment, poor strength, short service life, poor separation effect and other shortcomings, so it is difficult to be used as a stable and simple separation and preparation method

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  • Purification preparation method of ganglioside monomers

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Effect test

preparation example Construction

[0021] The flow process of a purification preparation method of a ganglioside monomer of the present invention comprises the following steps:

[0022] Step 101 dissolving the sample: dissolving the pretreated ganglioside graded product with water and an organic solvent at a ratio of 90 / 10 to 10 / 90 to obtain a solution;

[0023] Here, the pretreated ganglioside fraction product contains one or more ganglioside monomers in GA, GM, GD, GT, and GQ;

[0024] Preferably, the organic solvent is one or more of isopropanol, methanol, ethanol, acetonitrile, n-butanol, methylene chloride, chloroform, ethylene dichloride, tetrahydrofuran;

[0025] In the sample dissolving step described here, the solution can be obtained by ultrasonically dissolving and passing through a 0.45 μm organic film;

[0026] Step 102 Configure the mobile phase: use the organic solvent-buffered saline solution as the mobile phase, wherein the volume concentration of the organic solvent is 5%-95%, the concentrati...

Embodiment 1

[0037] Step 1: Take the graded product of gangliosides in a test tube, add 50% methanol to prepare a 50 mg / ml sample, dissolve it with ultrasonic waves, and pass through a 0.45 μm organic membrane to obtain a clear solution;

[0038] Step 2: Use a C8 reverse-phase chromatographic column with a column size of 4.6×250mm, a particle size of 5μm, and a pore size of The injection volume was 13 mg, the flow rate was 0.7 mL / min, the column temperature was 35 °C, the detection wavelength was 205 nm, the mobile phase was acetonitrile and 100 mM ammonium dihydrogen phosphate (pH=6.50), and the concentration of acetonitrile ranged from 45 to 40 min in 0 to 40 min. %~60%, gradient elution, collected according to peak shape;

[0039] Step 3: Collect the eluted samples and carry out rotary evaporation and concentration treatment at 40°C at a rotation speed of 70r / min to obtain the separated ganglioside monomers.

[0040] The separated gangliosides were identified by mass spectrometry. The...

Embodiment 2

[0042]Step 1: Take the graded product of gangliosides in a test tube, add 50% methanol to make a 100 mg / ml sample, dissolve it by ultrasonic, pass through a 0.45 μm organic membrane to obtain a clear solution;

[0043] Step 2: Use a C8HC reverse-phase chromatographic column with a column size of 10×250mm, a particle size of 10μm, and a pore size of The injection volume is 54mg, the flow rate is 2mL / min, the column temperature is 35°C, the detection wavelength is 205nm, the mobile phase is acetonitrile, 50mM diammonium hydrogen phosphate (pH=6.50), and the concentration of acetonitrile within 0-40min is from 45%~60%, gradient elution, collected according to peak shape;

[0044] Step 3: Collect the eluted samples and carry out rotary evaporation and concentration treatment at 40°C at a rotation speed of 70r / min to obtain the separated ganglioside monomers.

[0045] The separated gangliosides were identified by mass spectrometry. The average purity of ganglioside monomers can r...

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Abstract

The invention discloses a purification preparation method of ganglioside monomers. A ganglioside graded product is taken as a raw material to carry out HPLC (high performance liquid chromatography) purification separation preparation, and the graded product contains the ganglioside monomers such as GA (asialganglioside), GM (monosialoganglioside), GD (disialoganglioside), GT (trisialoganglioside),GQ (tetrasialoganglioside) and the like. According to the preparation method, reversed phase filler, namely, reversed phase mode with silica gel as a substrate and a C4-C18 aliphatic hydrocarbon chain as a bonded phase is adopted, and elution is carried out through buffer salts, concentration, pH conditions and flow phase. The method has the advantages of good separation degree, short period, simplicity and controllability in operation, high sample loading capacity and the like.

Description

technical field [0001] The invention relates to the field of biological extraction, in particular to a method for the purification and preparation of ganglioside monomers. Background technique [0002] Gangliosides (GLS) are a group of sialic acid-containing glycosphingolipids. The molecules are composed of hydrophobic ceramides and hydrophilic sialic acid-containing oligosaccharide chains. They are widely distributed in the cell membranes of vertebrate tissues. Among them, the nervous system is the most abundant. Usually according to the number of sialic acid, it is divided into GA (asialoganglioside), GM (monosialoganglioside), GD (disialoganglioside), GT (trisialoganglioside) ), GQ (tetrasialoganglioside), etc. [0003] The functions of gangliosides can be roughly summarized as follows: 1. Promote the normal development of nerve cells and brain tissue, prevent and treat diseases such as cerebral palsy; 2. Repair damaged nerves and brain tissue, prevent and treat stroke;...

Claims

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Application Information

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IPC IPC(8): C07H15/10C07H1/08
CPCC07H15/10C07H1/08
Inventor 梁鑫淼张华林叶贤龙闫竞宇郭志谋金高娃
Owner TAIZHOU GUOKEHUAWU BIOMEDICAL TECH CO LTD
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