Analysis method of tissue energy metabolic substances based on UPLC-MSMS

Abstract

The invention aims to provide an analysis method of tissue energy metabolic substances based on UPLC-MSMS. A liquid chromatography-mass spectrometry combined technical means is adopted; key metabolicsubstances participating in an energy metabolism pathway are quantitatively detected in a targeted manner; the method is especially used for key metabolic substances in TCA circulation, glycolysis andoxidative phosphorylation pathways. The method is simple, rapid and accurate, compared with the prior art, the complexity of a derivatization experiment is avoided, separation is performed by using liquid chromatography during detection, the content of up to 20 important energy metabolic substances can be obtained by one-time sample injection analysis by adopting a high-specificity and high-sensitivity analysis strategy of a mass spectrum multi-reaction monitoring mode, and the method has the advantages of high precision, simple pretreatment, high sensitivity, good repeatability and the like.

Description

technical field [0001] The invention relates to the field of biotechnology detection, in particular to a method for analyzing tissue energy metabolites based on UPLC-MSMS. Background technique [0002] Energy metabolism maintains the most basic life activities of living organisms. For example, plant adversity and animal diseases are usually accompanied by severe metabolic disorders, and most of them are attributed to energy metabolism disorders. Energy metabolism also affects the function and fate of immune cells, which is related to disease resistance and immune response. At the same time, the modification group of protein phosphorylation comes from ATP, so abnormal energy metabolism can further lead to changes in the upstream protein phosphorylation pathway, seriously affecting various physiological and pathological functions of the organism. The energy metabolism process in organisms mainly includes tricarboxylic acid cycle, glycolysis pathway and oxidative phosphorylati...

AI Technical Summary

Problems solved by technology

[0003] At present, there are mainly three analysis strategies for key substances in energy metabolism pathways. One is to perform derivatization on the sample to be tested and then use gas chromatography-triple quadrupole mass spectrometry to analyze (Chinese patent CN 106855551 A). For separation by gas chromatography, it is necessary to resuspend the sample with methoxyaminopyridine solution before detection, and then add N-methyl-N-(trimethylsilyl) trifluoroacetamide for derivation reaction to carry out chromatographic separation, resulting in The sample preparation time is too long, and only the derivatization reaction experiment will take at least 3 hours; the second is to use a high-pressure liquid chromatograph with a diode array detector for detection (Chinese patent CN 108303475 A), which uses The diode array detector has poor sensitivity and can only reach the μg / mL level, and this method compares the chromatographic retention time with the standard to characterize the analyte, and the accuracy is poor. If the detection sample is too complicated, there will be Interfering substances may appear to affect the qualitative analysis of substances, so it cannot become a method for the detection of energy substances that can be widely used in various sample types; the third is to use commercial kits for the detection of related metabolites. The detection cost of the method is high, the experimental steps are cumbersome, and the operator's proficiency in operation is extremely high, and each experiment can only detect a single substance, which has strong limitations

Images