Detection method of peptide-advanced glycation end product

A technology for advanced glycosylation and end products, applied in measuring devices, instruments, scientific instruments, etc., can solve the problems of unfavorable quantification of dicarbonyl compounds in reaction intermediates, affecting the qualitative and quantitative properties of pyrrolidine, and low content of dicarbonyl compounds, etc. , to achieve good detection limit, qualitative accuracy and readability, and quantitative accuracy

Active Publication Date: 2014-06-25
SOUTH CHINA UNIV OF TECH +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, some AGEs, such as pyrraline, also contain aldehyde groups, which can also be reduced to alcohols, which seriously affects the qualitative and quantitative determination of pyrraline.
At the same time, this method can only be used to quantify the reaction intermediate dicarbonyl compounds a

Method used

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  • Detection method of peptide-advanced glycation end product
  • Detection method of peptide-advanced glycation end product
  • Detection method of peptide-advanced glycation end product

Examples

Experimental program
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Embodiment 1

[0051] Use dipeptide (N)Lys-Ala(C) to react with glucose, (N)Lys-Ala(C) and glucose are mixed in 0.2M PBS (phosphate buffer, pH=6.8) medium, dipeptide concentration 10 -3 mol / L, glucose concentration 5×10 -3 mol / L, with a total volume of 15mL, heated at 140°C for 20min in a microwave digestion tank. After reaching the designated heating time, immediately add o-phenylenediamine whose molar mass is 2 times of the reducing sugar molar mass in the sample, so that the reaction is terminated immediately. After 3 hours, solid-phase extraction was used for enrichment, and the steps were as follows: select functionalized polystyrene / divinylbenzene as the extraction column, activate the extraction column with 4mL methanol and 4mL water respectively, load the sample with 1mL, rinse with 2mL water, and negatively Elute with 4 mL of pure methanol after being pumped dry under pressure for 4 min. Blow the eluate to dryness with nitrogen at 50 ° C. The mobile phase is fixed to 1 mL, filtered...

Embodiment 2

[0053] Use dipeptide (N)Lys-Gly(C) to react with glucose, mix (N)Lys-Gly(C) and glucose in 0.2M PBS (phosphate buffer, pH=6.8) medium, dipeptide concentration 10 -3 mol / L, glucose concentration 5×10 -3 mol / L, with a total volume of 15mL, heated at 140°C for 20min in a microwave digestion tank. After reaching the designated heating time, immediately add o-phenylenediamine whose molar mass is 5 times of the reducing sugar molar mass in the sample, so that the reaction is terminated immediately. After 3 hours, use solid-phase extraction technology for enrichment, the steps are as follows: use 4mL methanol and 4mL water to activate the extraction column respectively, load the sample with 1mL, rinse with 2mL water, dry it under negative pressure for 4min, and elute with 4mL pure methanol. The liquid was removed and blown to dryness with nitrogen at 50°C, the mobile phase was adjusted to 1 mL, filtered through a 0.45 μm membrane, and then analyzed by high-resolution mass spectromet...

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Abstract

The invention discloses a detection method of a peptide-advanced glycation end product. The detection method is characterized by comprising the following steps: (1) removing a dicarbonyl compound in a liquid sample; (2) removing impurities and gathering the peptide-advanced glycation end product in the sample by using a solid-phase extraction technique; (3) determining the nature of the gathered peptide-advanced glycation end product; (4) carrying out enzymolysis on the gathered peptide-advanced glycation end product by using peptidase, removing the peptidase by using a filter membrane to obtain free peptide-advanced glycation end product; (5) quantitatively analyzing the free peptide-advanced glycation end product through mass spectrometric analysis. The method is capable of detecting the peptide-advanced glycation end product at a high flux and lowering the complexity of the quantitative detection of the peptide-advanced glycation end product; meanwhile, according to the method, the detection limit is relatively good, the linear relation between a concentration of a target product and a peak area of a signal is good, and the quantitative effect is good.

Description

technical field [0001] The invention relates to a qualitative and quantitative detection method of peptide-advanced glycation end products (peptide-AGEs). Background technique [0002] Advanced glycation end products (AGEs) can cause glycosylation of proteins in the body, reducing the toughness of blood vessel walls and making them vulnerable to damage. At the same time, AGEs can bind to receptors on cells to promote the release of inflammatory cytokines. AGEs can also promote the formation of oxygen free radicals, thereby causing tissue damage. AGEs are related to the formation of cataract, arteriosclerosis, Alzheimer's disease (Alzheimer's disease), amyloidosis, nephropathy, neuropathy, retinopathy and so on. Once formed, AGEs are easy to accumulate in tissues, and can have an amplification effect, promoting the formation of more AGEs. [0003] AGEs can be divided into endogenous AGEs and exogenous AGEs. Endogenous AGEs are produced by glycosylation reactions between ca...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/08
Inventor 李琳梁志理李冰苏健裕徐振波卞华伟李玉婷
Owner SOUTH CHINA UNIV OF TECH
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