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A method for detecting sulfur-containing amino acids in serum and its enrichment material

A sulfur amino acid and detection method technology, which is applied in the field of medical inspection and analysis, can solve the problems of low specificity of sulfur-containing amino acids, lower sensitivity of mass spectrometry detection, and low precision, and achieve novel structure, high accuracy of results, and high accuracy of results high effect

Active Publication Date: 2021-03-26
大连博源医学科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Due to the characteristics of mass spectrometry detection, the influence of other interference peaks caused by matrix effect cannot be observed on the chromatogram, but the co-eluting components of solid phase extraction will still enter the chromatography-mass spectrometry system, polluting the chromatography-mass spectrometry separation and analysis system, Reduced Sensitivity of Mass Spectrometry Detection
For example, CN201710129265 discloses a liquid chromatography tandem mass spectrometry method for the detection of sulfur-containing amino acids in plasma by a non-derivatization method. After adding an isotope internal standard solution to the plasma sample, a reducing agent is added for reduction, and the precipitated protein is processed and then used LC- The above supernatant was analyzed and collected by MS / MS, however, the resulting precision was not high
As another example, CN201610314061 discloses a plasma sample pretreatment method for the detection of endogenous amino acids, using silica gel-bonded C1-30 alkyl particles as a solid adsorbent to adsorb amino acid components, but the specificity for sulfur-containing amino acids Sex is not high

Method used

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  • A method for detecting sulfur-containing amino acids in serum and its enrichment material
  • A method for detecting sulfur-containing amino acids in serum and its enrichment material
  • A method for detecting sulfur-containing amino acids in serum and its enrichment material

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] The detection of sulfur-containing amino acid methionine in embodiment 1 serum:

[0039] 1. Materials and reagents

[0040] Chromatographic column: (CORTECS, 2.7μm 2.1μm×100mm) (Waters Corporation, USA)

[0041] Methionine standard and methionine isotope internal standard were purchased from Shanghai Hushi Pharmaceutical Technology Co., Ltd.; dithiothreitol was purchased from Shanghai Yucan Biotechnology Co., Ltd.; methanol, acetonitrile, isopropanol, formic acid, ammonium formate (chromatographically pure), Ammonia water (chromatographically pure); ultrapure water: prepared by Mili-Q ultrapure water machine.

[0042] 2. Instruments and equipment

[0043] High-performance liquid chromatography-tandem mass spectrometer, equipped with electrospray (ESI) ionization source (4500MD, American AB Company), the liquid chromatography separation mode is hydrophilic interaction reversed-phase chromatography separation, and the detector is triple quadrupole tandem mass spectrometry...

Embodiment 2

[0065] The detection of sulfur-containing amino acid cysteine ​​in embodiment 2 serum:

[0066] 1. Materials and reagents

[0067] The difference from Example 1 is that cysteine ​​and cysteine ​​isotope internal standard were purchased from Shanghai Hushi Pharmaceutical Technology Co., Ltd.;

[0068] 2. Instruments and equipment

[0069] The difference from Example 1 is that the high-performance liquid chromatography-tandem mass spectrometer is equipped with an electrospray (ESI) ionization source (TQD, Waters Corporation of the United States);

[0070] 3. The method for detecting the sulfur-containing amino acid cysteine ​​by high performance liquid chromatography-tandem mass spectrometry, the steps are:

[0071] 1) Preparation of standard working solution: Accurately prepare cysteine ​​standard stock solution (1mg / mL), dilute with pure water to obtain a concentration of 600μg / mL, and dilute step by step with pure water to 300μg / mL and 120μg / mL respectively , 30μg / mL, 12μg...

Embodiment 3

[0090] The detection of sulfur-containing amino acid homocysteine ​​in embodiment 3 serum:

[0091] 1. Materials and reagents

[0092] The difference from Example 1 is that the internal standard of homocysteine ​​and homocysteine ​​isotope was purchased from Shanghai Hushi Pharmaceutical

[0093] Technology Co., Ltd;

[0094] 2. Instruments and equipment

[0095] Same as Example 1

[0096] 3. The method for detecting the sulfur-containing amino acid homocysteine ​​by high performance liquid chromatography-tandem mass spectrometry, the steps are:

[0097] 1) Preparation of standard working solution: Accurately prepare homocysteine ​​standard stock solution (1 mg / mL), dilute with pure water to obtain a concentration of 120 μg / mL, and dilute to 60 μg / mL and 24 μg / mL with pure water, respectively. Homocysteine ​​standard solutions with six concentration levels of mL, 6μg / mL, 2.4μg / mL and 1.2μg / mL, set aside;

[0098] 2) Preparation of internal standard standard solution: Accura...

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Abstract

The invention relates to a method for detecting sulfur-containing amino acids in serum and an enrichment material of sulfur-containing amino acids in serum. According to the method, a reducing agent is added into a serum sample so as to reduce the serum sample; the reduced serum sample solution is pretreated through a specific purification material, so that a purified sample elution solution is obtained; a liquid chromatography-tandem mass spectrometry detection and analysis are performed, so that sulfur-containing amino acids such as methionine, cysteine and homocysteine can be detected; the specific material is a metal affinity material taking silica gel as a matrix; after the serum is purified through the specific material, the interference of the matrix in the serum can be removed, the sensitivity of the sulfur-containing amino acids can be improved, and the accuracy of the detection of the sulfur-containing amino acids can be better guaranteed. According to the purification of the specific material, a specific affinity effect between the sulfydryl of the sulfur-containing amino acids and metal is utilized to realize the selective enrichment of the sulfur-containing amino acids, and therefore, the problem that the sulfur-containing amino acids cannot be effectively enriched in reversed phase chromatography is solved.

Description

technical field [0001] The invention belongs to the technical field of medical inspection and analysis, in particular to a liquid chromatography-tandem mass spectrometry detection method for detecting sulfur-containing amino acids such as methionine, cysteine ​​and homocysteine ​​in serum samples. [0002] technical background [0003] Sulfur-containing amino acids in the human body are closely related to the metabolism of various sulfur-containing compounds in the body. The lack of methionine can cause loss of appetite, slow growth, and enlarged kidneys. It can be used to prevent acute or chronic hepatitis, cirrhosis and other diseases. The sulfhydryl group brought by cysteine ​​can alleviate the poisoning degree of toxic substances and prevent radiation. There is also homocysteine ​​in the human body, which is an intermediate product of the metabolism of methionine in the human body, and elevated homocysteine ​​is a risk factor for atherosclerotic vascular disease [Jung YS,...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N30/02G01N30/06G01N30/56G01N30/89
CPCG01N30/02G01N30/06G01N30/56G01N30/89G01N2030/062G01N2030/562
Inventor 曹云峰孙晓宇潘永强王爽洪沫孙冬雪宋丹许晶
Owner 大连博源医学科技有限公司
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