High Performance Analysis Of Phospholipids And Triglycerides Based On Mass Spectrometry And Its Application

Glycerophospholipids are the most abundant type of phospholipids in the body.In addition to constituting biofilms,they are also components of bile and membrane surface active substances.They are involved in cell membrane protein recognition and signal transduction as well as associated with many metabolic diseases,such as cardiovascular disease,hypertension,diabetes,obesity,neurological diseases,fatty liver,coronary heart disease and cancer.The basic structure of glycerophospholipids is the phosphatidic acid and the substituents attached to the phosphoric acid,which can be divided into many categories depending on the substituent groups.The most important of which are phosphatidylcholine,phosphatidylethanolamine,phosphatidylinositol,phosphatidylserine,phosphatidic acid,and phosphatidylglycerol.In order to fully understand the important role of glycerophospholipids in the occurrence and development of diseases,the metabolism and the changes of composition and structure of glycerophospholipids in organisms under different physiological and pathological conditions were investigated.Because of their diversified varieties,occurrence at low concentration in biological samples and low ionization efficiency,the development of reliable analytical methods for the identification and quantification of phospholipids is a challenge to analytical chemists.Chemical derivatization has been an efficient technique employed to improve the chromatographic behavior and mass spectrometry(MS)detective sensitivity of phospholipids through modifying their chemical structures.Approach of chemical derivatization coupled with MS has been widely applied in analysis of proteins,carbohydrates and their metabolites.In recent years,the application of this strategy in lipid analysis has aroused great attentions among researchers in this field.In this study,a novel strategy based on acetone stable-isotope derivatization coupled with high performance liquid chromatography-mass spectrometry(HPLC-MS)for profiling and accurate quantification of aminophospholipids(phosphatidylethanilamine(PE)and phosphatidylserine(PS))in biological samples were developed to solve the technical bottleneck of the prior art in the analysis and detection of aminophospholipids.The results of this paper are as follows:1.In this study,a novel strategy using solid phase extraction(SPE)coupled with lipid chromatography mass spectrometry based on acetone stable isotope derivatization for rapid comprehensive characterization and accurate quantification of aminoglycerophospholipids molecular species in liver samples of rats was developed.In addition,compared with underivatived aminoglycerophospholipids,the detection sensitivities of acetone labeled PE and PS improved by 8.13 and 15.71 fold,respectively,and a good separation of aminoglycerophospholipids was achieved upon acetone labeling.Furthermore,for the accurate quantification of the species of aminoglycerophospholipids,a calibration curve was calculated from the ESI-MS/MS mass spectrum and used to correct the experimental ion abundances(caused by the differential effects including acyl chain lengths,unsaturation,concentration,solvent composition and the detector response)to the actual molar abundances in biological sample.Finally,this fast and efficient developed method was applied to identify and quantify 48 PE species and 22 PS species in rats liver.The results showed that 20 kinds of aminoglycerophospholipids molecular species in the livers samples from rats fed with polyphenol diets had siginificant changed.2.The effect of plant(rapeseed and grape seed)polyphenols on lipid metabolism of rat fed with high fat diet was studied.30 rats were divided into 5 groups including group Z(basal diet),C(high fat diet without plant polyphenols),S(high fat diet with rapeseed polyphenols),R(high fat diet with grape seed polyphenols)and F(combined with rapeseed and grape seed polyphenols).Levels of lipids including TAGs and PLs in rats were investigated by shotgun-electrospray ionization tandem mass spectrometry(shotgun-ESI-MS).Results showed that high fat diet would give rise to the total TAGs and decrease total phosphatidylcholine(PC),phosphatidylethanolamine(PE)and phosphatidylserine(PS)(p < 0.05)in group C,S,R and F compared with group Z.Results also showed that supplementary with rapeseed polyphenols or grape seed polyphenols reduced total TAGs and increased total PC,PE and PS(p < 0.05)in group S,R and F compared with group C.Grape seed polyphenols and rapeseed polyphenols had certain regulatory effect on metabolism diseases such as fatty liver.