Method for measuring weight-average molecular weight and content of mannuronic acid substances

A technology of weight-average molecular weight and determination method, which is used in measurement devices, material separation, and analysis of materials.

Active Publication Date: 2019-03-12
GREEN VALLEY (SHANGHAI) PHARM CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is no standard product for the monosaccharide mannuronic acid of acidic sugar. If glucuronic acid is used as the standar

Method used

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  • Method for measuring weight-average molecular weight and content of mannuronic acid substances
  • Method for measuring weight-average molecular weight and content of mannuronic acid substances
  • Method for measuring weight-average molecular weight and content of mannuronic acid substances

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] Example 1 SEC and ICP-MS Reveal the Separation Mechanism of Acidic Sugars in Size Exclusion Chromatography

[0087] (1) The sample to be tested is mannan oligosaccharide diacid sodium salt. Dissolve the sample in water at a concentration of 2mg / ml and inject 0.1ul. Waters ACQUITY UPLC@BEH through SEC column (SEC 1.7um 4.6×300mm column, purchased from Waters), the mobile phase is 0.08mol / L NH4Ac, and the flow rate is 0.1mL / min; the SEC is combined with ICP-MS to collect 23 Na signal. Meanwhile, solvent water was used as a control.

[0088] (2) Combine SEC with MALS, dissolve the sodium salt of mannooligosaccharide diacid in the mobile phase with a concentration of 10 mg / ml, and inject 20 ul of the sample.

[0089] attached figure 1 as solvent water ( figure 1 A) and mannooligosaccharide diacid sodium salt ( figure 1 B) SEC-ICP-MS spectrum, and SEC-MALS differential spectrum of mannooligosaccharide diacid sodium salt (Fig. 1C). The result shows, in SEC-ICP-MS sp...

Embodiment 2

[0094] Example 2 SEC-MALS method and mass spectrometry method mutual verification

[0095] Sample: mannan oligosaccharide diacid pentasaccharide (Na type), mannan oligosaccharide diacid pentasaccharide (H type)

[0096] Mannooligosaccharide diacid pentasaccharide (H type) sample preparation: take 200 mg of mannan oligosaccharide diacid pentasaccharide (Na type), dissolve it in 5 ml ultrapure water, and dissolve the mannan oligosaccharide diacid pentasaccharide (Na type) solution The cation exchange resin (AG 50W-X8 Resin, Bio-Rad) was used for Na-H exchange, and the effluent was collected and dried to obtain the mannooligosaccharide diacid pentasaccharide (Type H) sample.

[0097] (1) Column Waters ACQUITY UPLC@BEH SEC 1.7um 4.6×300mm column, 10% methanol + 80% 0.15mol / L NaCl as mobile phase, flow rate 0.1mL / min.

[0098] (2) Combine SEC with MALS, dissolve the sample in the mobile phase at a concentration of 10 mg / ml, and inject 20 ul of the sample.

[0099] (3) SEC is co...

Embodiment 3

[0111] Example 3 Establishment and verification of the SEC-MALS method for determining the weight-average molecular weight and content of mannan oligosaccharide diacid sodium salt

[0112] Step 1: Determination of the ratio of metal ions in mannooligosaccharide diacid sodium salt by ion chromatography

[0113] Na ion standard solution is configured as 25, 10, 12.5, 6.25, 3.125ppm. The sample concentration was 2 mg / ml. Ion chromatography conditions: pre-column: Metrosep C4Guard / 4.0; separation column: Metrosep C5-150 / 4.0; mobile phase: ultrapure aqueous solution of 5mM nitric acid; flow rate: 0.9ml / min; injection volume: 20ul. The content of metal ions was determined by external standard method.

[0114] Step 2: Determination of dn / dc of mannooligosaccharide diacid sodium salt

[0115] (1) Weigh 20 mg of the sample with constant weight into a 10 ml volumetric flask, add mobile phase to dissolve, and shake to obtain the mother liquor. Take 0.2, 0.4, 0.8, 1.2, 1.8ml of the mo...

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Abstract

The invention belongs to the field of natural medicine chemistry and quality control, and relates to a method for measuring the weight-average molecular weight and the content of soluble salt of mannuronic acid substances. Weight-average molecular weight and content results of acid sugar measured by an SEC-MALS (size exclusion chromatography with multi-angle laser scattering detector) are corrected by the aid of the metal ion content of soluble salt samples of the mannuronic acid substances. The weight-average molecular weight and the content of the soluble salt w of the mannuronic acid substances can be rapidly and accurately measured by the method.

Description

technical field [0001] The invention belongs to the field of natural medicinal chemistry and quality control, and relates to a method for measuring the weight-average molecular weight and content of mannuronic acid substances, especially their soluble salts. Background technique [0002] The main source of mannuronic acid is sodium alginate. Sodium alginate is classified by acid hydrolysis to obtain polymannuronic acid (molecular weight is about 2k-10kDa), and polymannuronic acid is oxidized to obtain mannooligosaccharide diacid (molecular weight is between 400-3000Da). Mannan oligosaccharide diacid is a low-molecular-weight marine acidic oligosaccharide compound with a specific molecular skeleton, which can be used to fight Alzheimer's disease (AD). Preclinical pharmacodynamics and other research results show that mannan oligosaccharide diacid has a protective effect on nerve cells, and finally improves memory dysfunction, thereby effectively treating Alzheimer's disease. ...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/74
CPCG01N30/02G01N30/74
Inventor 曾洋洋黄道林张真庆
Owner GREEN VALLEY (SHANGHAI) PHARM CO LTD
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