Method for analyzing amine substances in dansyl chloride derived-plasma based on liquid chromatography mass spectrometry

A technology of dansyl chloride and analytical method, applied in the fields of analytical chemistry and medicine, can solve the problems of small coverage and less amine substances, and achieve the effects of less reagent consumption, high repeatability and good specificity

Inactive Publication Date: 2014-05-28
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Claims
  • Application Information

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Problems solved by technology

The currently reported literature and patents have used the dansyl chloride derivatization method to measure several amino acids in plasma, but the coverage is small, the detection of amines is few, and there is no method for analyzing the metabolic profile of amines in plasma.

Method used

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  • Method for analyzing amine substances in dansyl chloride derived-plasma based on liquid chromatography mass spectrometry
  • Method for analyzing amine substances in dansyl chloride derived-plasma based on liquid chromatography mass spectrometry
  • Method for analyzing amine substances in dansyl chloride derived-plasma based on liquid chromatography mass spectrometry

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Experimental program
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Effect test

Embodiment 1

[0022] Thaw the plasma in the -80°C refrigerator to room temperature, quantitatively pipette 100 μL of plasma to a centrifuge tube and add isotope-containing internal standards Glycine-d5, DL-Alanine-3,3,3-d3, L-Leucine-5,5, 400 μL of acetonitrile solution of 5-d3, Thymine-d3, Phenylalanine-d5 and Tryptophan-d5 (400 ng / mL concentration). 17 kinds of amine substances were formulated as a mixed standard of 50ng / mL, and 6 kinds of isotopic internal standards with a concentration of 400ng / mL were added as above. After the two mixtures were vortexed for 1 min respectively, the samples were centrifuged at 15000 rpm / min at 4°C for 10 min. Pipette a volume of 450 μL of the supernatant to a centrifuge tube and lyophilize on a lyophilizer. Add 40 μL of 5 mg / mL dansyl chloride acetonitrile solution and 40 μL of 0.15 mol / L Na to the lyophilized plasma extract 2 CO 3 / NaHCO 3 Aqueous solution (PH=9.4), vortex for 1min, seal the centrifuge tube and then react in a water bath at 60°C for...

Embodiment 2

[0027] Before plasma collection, pathological diagnosis was performed by multi-point puncture to ensure the accuracy of sampling. The patients fasted for 12 hours and collected in the morning on an empty stomach. The samples included the plasma of patients with benign prostatic hyperplasia and malignant prostate cancer (ie, patients with benign prostatic hyperplasia and malignant prostate cancer whose PSA could not be distinguished) with a prostate-specific antigen (PSA) concentration of 4-10 ng / mL. After collection, they were quickly stored in a -80°C refrigerator.

[0028] Before analysis, thaw the plasma to room temperature, quantitatively pipette 100 μL of plasma to a centrifuge tube and add internal standard Glycine-d5, DL-Alanine-3,3,3-d3, L-Leucine-5,5,5-d3, Thymine-d3, Phenylalanine-d5 and Tryptophan-d5 (concentration: 400ng / mL) acetonitrile solution 400μL. After vortexing for 1 min, the sample was centrifuged at 15,000 rpm / min at 4°C for 10 min. Pipette a volume of...

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Abstract

The invention discloses a method for analyzing amine substances in dansyl chloride derived-plasma based on liquid chromatography mass spectrometry. The method is characterized in that dansyl chloride is used as a derivatization reagent to perform derivatization on a metabolite containing primary amine and secondary amine, therefore the retention capability of the amine substances in reversion phase chromatography is increased, the ionization efficiency in mass spectrometric detection is increased, and the purpose of increasing the quantitative accuracy and detection sensitivity of the amine substances is achieved. The method has wide coverage for amine metabolites (comprising amino acid, methylation products, acetylation products, dipeptide and the like), can utilize 100 [mu]L of plasma samples to detect 113 amine substances, supplies the outline information of metabolome of the amine substances, and has the characteristics of good specificity, high accuracy and repeatability, less reagent consumption and the like.

Description

technical field [0001] The invention relates to the fields of analytical chemistry and medicine, and is an analysis method for improving the metabolic profile of amine substances in blood plasma based on the derivation of dansyl chloride by liquid chromatography-mass spectrometry. Background technique [0002] Amines are the basis of human metabolism, and their main representatives are amino acids, which are the basic units of protein and have important biochemical activities. Dipeptide is a protein fragment or substance composed of an amide bond formed by the dehydration condensation of two amino acids. It is closely related to nutrition, hormones, enzyme inhibition, immune regulation, antibacterial, antiviral, and antioxidative. In addition, studies have shown that the methylation and acetylation products of amino acids are closely related to human health and are important characteristics of tumor development. Therefore, the realization of metabolic profile analysis of am...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/06
Inventor 许国旺黄强尹沛源戴伟东路鑫
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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