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LC-MS/MS (liquid chromatography-tandem mass spectrometry) method for determining cordycepin metabolite in liver microsome

A technology of liver microsomes and LC-MS, which is applied in the field of medical testing, can solve the problems of in vitro metabolism research of liver microsomes that have not been seen, and achieve the effect of high detection sensitivity and good selectivity

Active Publication Date: 2016-07-20
LIAONING UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, the research on cordycepin is mostly focused on the extraction and purification process and pharmacological activity research. So far, there is no in vitro metabolism research using liver microsomes as a model.

Method used

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  • LC-MS/MS (liquid chromatography-tandem mass spectrometry) method for determining cordycepin metabolite in liver microsome
  • LC-MS/MS (liquid chromatography-tandem mass spectrometry) method for determining cordycepin metabolite in liver microsome
  • LC-MS/MS (liquid chromatography-tandem mass spectrometry) method for determining cordycepin metabolite in liver microsome

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Embodiment 1

[0056] A liquid mass spectrometry method for determining cordycepin metabolites in liver microsomes, comprising the following steps

[0057] 1) Incubation of cordycepin liver microsomes

[0058] Transformation system is 2mL, add liver microsome solution 154μL, NADP solution 166μL, NADH solution 164μL, G-6-P solution 166μL, G-6-PDH solution 144μL, MgCl 2 Add 840 μL tris(hydroxymethyl)aminomethane hydrochloride buffer solution to 166 μL solution, shake well, put it into a preheated 37°C full-temperature shaker and incubate for 5 minutes, then add 200 μL cordycepin solution to start the reaction. During the reaction process, oxygen was passed on the liquid surface for 30 seconds every 20 minutes. After 30 minutes, the culture bottle was taken out from the shaker, and 6 mL of cold methanol was quickly added to terminate the metabolic reaction.

[0059] 2) Sample processing

[0060] The incubation product was vortexed for 30s, centrifuged (rotating speed 5000r / min) for 10min, acc...

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Abstract

The invention belongs to the technical field of medicine detection, discloses an LC-MS / MS (liquid chromatography-tandem mass spectrometry) method for determining cordycepin metabolite in liver microsome, and particularly relates to the LC-MS / MS method for analyzing and determining the cordycepin metabolite in the liver microsome. The LC-MS / MS method mainly includes the steps of (1) incubation of the liver microsome containing cordycepin; (2) preprocessing of a sample; (3) analysis of the sample, thereby determining the cordycepin metabolite in the liver microsome. The LC-MS / MS has the advantages of convenience in operation, sample processing simplicity, high analysis speed, high specificity and high flexibility, provides technical support for researching extrametabolites of the cordycepin in the liver microsome while laying a methodological foundation for researching in-vivo and in-vitro metabolites of the cordycepin, and accordingly has a great significance.

Description

technical field [0001] The invention belongs to the technical field of medical detection, and in particular relates to a liquid mass spectrometry method for measuring cordycepin metabolites in liver microsomes. Background technique [0002] After the drug enters the organism, a series of reactions such as oxidation, reduction, hydrolysis, and combination will occur under the action of different enzymes. The liver is the main organ for these metabolic reactions of the drug and the main place for the body to undergo biotransformation. Metabolism is a huge cytochrome P450 (CYP450) enzyme system, and the phase I and phase II metabolic reactions of most drugs depend on the liver enzyme system to occur (the first-pass effect of the drug comes from this). Therefore, in the field of drug metabolism, the liver microsome in vitro experiment, as an important part of in vitro metabolism research, has attracted more and more attention. [0003] Cordycepin is the first nucleoside antibio...

Claims

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Application Information

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IPC IPC(8): G01N30/88
CPCG01N30/88G01N2030/884
Inventor 刘玉峰李胜男朱美霞韩彬王志萍胡延喜卢晓丹
Owner LIAONING UNIVERSITY
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