Construction Of Contour Spectrum Of Arachidonic Acid And Its Metabolites Based On UPLC-QTOF-MS/MS

Arachidonic acid(AA)is a cis 5,8,11,14-icosatetraenoic acid,which is an unsaturated fatty acid.It is a polyunsaturated essential fatty acid with high content and the widest distribution in the body.In the arachidonic acid cascade,AA mainly has three metabolic pathways in the body: through cyclooxygenase(COX),lipoxygenase(LOX)or cytochrome P-450(CYP-450)Produces prostaglandin(PG),thromboxane(TX),leukotriene(LT)or hydroxyfatty acid(OHFA).Among them,COXs are divided into 3 forms: COX-1(constitutive),COX-2(mitogeninduced form),and COX-3(a variant of COX-1).AA is released from membrane phospholipids through phospholipase A2 s and plays an important biological role in the body.It is widely involved in immune and inflammatory reactions and is a biomarker of inflammation.Accurate qualitative and quantitative analysis has important value in studying the development mechanism,diagnosis and prediction of related diseases.Arachidonic acid metabolites are biomarkers of various diseases.Accurate qualitative and quantitative analysis is of great value in studying the development mechanism,diagnosis and prediction of related diseases.At present,four major categories of AA metabolites that have been identified include prostaglandins,thromboxane,leukotrienes,and hydroxy fatty acids.Because most AA metabolites are extremely low in biological samples,conventional detection techniques cannot accurately analyze them qualitatively and quantitatively.In recent years,liquid chromatography-mass spectrometry(LC-MS)analysis technology has shown great advantages in the detection and analysis of small endogenous molecules,including highresolution time-of-flight mass spectrometry(Q-Tof)and triple quadrupole linear ions.Trap mass spectrometry(Q-Trap)powerful qualitative and quantitative capabilities.The subject first developed an analytical method for arachidonic acid and its metabolites,which included: optimization of chromatographic methods,determination of mass spectrometry methods,and development of real sample pretreatment methods.For the optimization of the chromatographic method,this project examined 11 mobile phase combinations,and found that when 0.05% acetic acid water was used as the water phase,the organic phase was the best when methanol and acetonitrile were used.For the determination of the mass spectrometry method,the subject examined the positive and negative ion modes,and the final results showed that the negative ion acquisition mode worked best for such components.For the development of sample pretreatment methods,this experiment investigated the methods of liquid-liquid extraction of methyl tert-butyl ether,liquid-liquid extraction of ethyl acetate,precipitation of methanol protein,precipitation of acetonitrile,and solid phase extraction of four SPE columns The processing process was optimized,and the final results showed that Waters Oasis HLB solid-phase extraction cartridges performed the best solid-phase extraction.This topic also summarized the specific cleavage rules and characteristics of the standard compounds,and established a qualitative method that can effectively identify the prostaglandins and thromboxane components.Through the established qualitative identification method,121 arachidonic acid and its metabolites were qualitatively identified in the mixed serum of 20 healthy pregnant women,including AA,109 PG compounds,and 11 TX compounds.This method provides a certain indicator for qualitative identification of more arachidonic acid metabolites,and lays a foundation for further research on the correlation between arachidonic acid and disease.