Research On Lipidomic Analysis Methods And Its Application In Exercise Rehabilitation Of Patients With Coronary Heart Disease

Recent clinical data have shown that exercise rehabilitation in stable phase can improve the long-term prognosis of patients with coronary heart disease(CHD)significantly.In this thesis,plasma samples were taken as the analysis object,different extraction methods were used to extract lipid metabolites,the analysis and detection conditions were optimized,a high-throughput analysis method of lipidomics was established,and the effect of exercise rehabilitation on the characteristics of lipid metabolism in patients with CHD were studied.Detailed research contents of this work are summarized as following:1.Establishment of methods for extraction and analysis of lipid metabolites from plasma.Methyl tert-butyl ether method and chloroform method were used to extract lipid metabolites,and the liquid chromatography conditions and mass spectrometry parameters of the analysis and detection were optimized.According to the molecular formula of lipid metabolites,the possible MS information of lipid metabolites can be predicted.The number of effective mass spectra obtained by matching the MS information of predicted lipid metabolites and the MS information in the human metabolomics database,the response intensity and stability of the qualitative analysis of lipid metabolites(RSD?30%)to screen the best extraction method.It was found that the best extraction method was Folch method(extraction solvent:CHCl₃:MeOH:H₂O(8:4:3,v/v/v),reconstituted solvent:CHCl₃:Me OH(2:1,v/v)).For metabolites,a large number of effective mass spectrometric features are obtained,and the response intensity and stability of metabolites are better.By optimizing the parameters of liquid chromatography and mass spectrometry parameters,it was found that the addition of ammonium formate in mobile phase could improve the peak shape of the chromatographic peak and increase the response intensity of the mass spectrometer;the mass spectrometry parameters have a great difference in the detection of different lipid metabolites.2.Qualitative analysis of lipid metabolites in the plasma of patients with CHD.Ultra-performance liquid chromatography-high resolution mass spectrometry technology was used to detect lipid metabolites in plasma.Based on the mass spectrometric characteristics of lipid metabolites,integrated qualitative filtering strategies(diagnostic fragment ion filtering,nitrogen rule filtering,and neutral loss)was used to filter interference mass spectra,the information from standards,literature or databases and the chromatographic elution of lipid metabolites was combined to qualitative analysis.308 plasma lipid metabolites were annotated in the plasma of patients with CHD and healthy people,and 257 lipid metabolites were annotated in the plasma of patients with CHD before and after exercise rehabilitation.3.Study on the characteristics of lipid metabolism in patients with CHD before and after exercise rehabilitation.Based on the semi-quantitative analysis results of the internal standard method,principal component analysis and partial least square discriminant analysis were used to establish a discriminant model for patients with CHD before and after exercise rehabilitation,and the results of the T test and variable important projection were combined to screen the characteristics of lipid metabolism in CHD before and after exercise rehabilitation.Thirty different metabolites were screened out from these 2 groups,and the metabolic pathway analysis of these different metabolites was analyzed.It was found that arachidonic acid metabolism,glycerophospholipid metabolism and sphingolipid metabolism had a greater impact on exercise rehabilitation of CHD.