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Method for simultaneous determination of multiple amino acids by liquid chromatography-mass spectrometry

A liquid chromatography-mass spectrometry, amino acid technology, applied in the field of amino acid determination, can solve the problems of peak shape and peak width, complicated operation, and influence on chromatographic column efficiency, etc., to achieve simple operation, less interference factors, and shortened sampling time Effect

Inactive Publication Date: 2021-01-08
SHANDONG AGRICULTURAL UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The method is complicated to operate, and the use of derivatization reagents such as phenyl isothiocyanate is highly toxic, and the triethylamine used to construct the derivatization reaction conditions will affect the column efficiency of the chromatographic column, and the peak shape and peak width of the subsequent use will be affected. influences

Method used

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  • Method for simultaneous determination of multiple amino acids by liquid chromatography-mass spectrometry
  • Method for simultaneous determination of multiple amino acids by liquid chromatography-mass spectrometry
  • Method for simultaneous determination of multiple amino acids by liquid chromatography-mass spectrometry

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Embodiment 1

[0043] Amino acid extraction process:

[0044] Wheat spikelet samples were frozen in liquid nitrogen, ground, added 1.0ml of sterile water per 100mg sample, ultrasonicated at room temperature for 30min, added 1mL of dichloromethane, mixed at 4°C for 30min, centrifuged at 4°C at 13000prm for 5min, Transfer the upper aqueous phase to a new centrifuge tube, filter through a 0.22 μm filter membrane, and perform chromatographic separation of various amino acids.

[0045] Liquid phase conditions:

[0046] Chromatographic column: Thermo HYPERSIL GOLD C18 column (2.1×100, 3µm);

[0047] Column temperature: 35°C;

[0048] Mobile phase: A: 0.1% formic acid solution, B: methanol;

[0049] Elution gradient: 0-0.2 min, A=96%; 0.2-3.1min, A decreased to 65%; 3.1-5.8min, A decreased to 23%, 5.8-8.7min, A decreased to 0%, 8.7min- 8.8min, A increases to 96%; 8.8-10min, A=96%;

[0050] Flow rate: 0.15mL / min;

[0051] Injection volume: 5µl

[0052] Mass Spectrometry Conditions:

[0053] Po...

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Abstract

The invention relates to the technical field of amino acid detection, in particular to a method for simultaneous determination of multiple amino acids by liquid chromatography-mass spectrometry, whichis characterized by comprising the following steps of: (1) an amino acid extraction process: after grinding a spikelet sample, adding sterile water, centrifuging after ultrasonic treatment at normaltemperature, adding dichloromethane to remove fat-soluble impurities such as pigments, transferring the supernatant into a new centrifuge tube, and filtering with a filter membrane to obtain a to-be-detected sample for later use; and (2) determining amino acid components, namely determining nine amino acids in the sample obtained in the step (1) by adopting a liquid chromatograph-mass spectrometer. According to the invention, multiple amino acid compounds can be extracted and detected at one time.

Description

technical field [0001] The invention belongs to the technical field of amino acid determination, and more specifically relates to a method for simultaneously determining multiple amino acids by liquid mass spectrometry. Background technique [0002] The amino acid pathway is essential in plants and is a precursor for the synthesis of various proteins. At the same time, previous studies have shown that metabolomics studies related to scab resistance clearly show that increased amino acid levels are associated with increased disease tolerance. Protein is one of the basic substances of life, and amino acid, as the basic unit of protein, plays an important role when plants encounter abiotic or biotic stress. In abiotic stresses, amino acids have been found to act as osmoprotectants to increase resistance due to nonenzymatic antioxidant effects (Sarvajeet et al., 2010). In the study on the resistance of Chestnut blight, it was found that after being infected by pathogenic bacte...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06
Inventor 胥倩刘振毕建杰路雅茹
Owner SHANDONG AGRICULTURAL UNIVERSITY
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