The Development And Application Of Post-Photoionization Mass Spectrometry Imaging With Vacuum Ultraviolet

Mass spectrometry is a chemical analysis method for analysis and identification by measuring the mass-to-charge ratio(m/z)of gas-phase ions.Mass spectrometry imaging(MSI)is a molecular imaging technology based on the development of mass spectrometry.It can perform untargeted visualization research on thousands of compounds at the same time without labeling.Therefore,it has become an important method for the study of the spatial distribution of compounds and plays an important role in clinical medicine,pharmacology,toxicology,environmental science,agriculture,botany and other research fields.The MSI method with high sensitivity and high spatial resolution can achieve high ionization coverage and microscopic resolution of samples,so it has always been a hot and difficulty issue in the development of MSI.The postionization technology can effectively improve the ionization efficiency through the secondary ionization process,which is an effective strategy for the development of high sensitivity MSI method and also an auxiliary technology for the development of high spatial resolution imaging methods.Based on the situation,this thesis covers the development of post-ionization mass spectrometry imaging methods as the main content.We research on the optimization of dopants in photoionization,the development of post-photoionization imprint mass spectrometry imaging of plant tissue,the development of synchrotron radiation post-photoionization mass spectrometry imaging and the development of high spatial resolution mass spectrometry imaging method based on laser.The first chapter of the thesis introduces the birth history and basic principles of mass spectrometry technology,the basic knowledge of ionization methods including electrospray ionization and photoionization,the principle and applications of synchrotron radiation photoionization mass spectrometry technology,the principle of mass spectrometry imaging technology,traditional mass spectrometry imaging ionization methods,the principle of post-ionization mass spectrometry imaging technology and the diversified applications of mass spectrometry imaging.In the process of photoionization,the participation of dopants will induce indirect photoionization.The dopant molecule is ionized after absorbing the photon energy,and the dopant radical cation reacts with the sample molecules by ion-molecule reactions,thus ionizing the sample molecules.Therefore,the second chapter of the thesis introduces the effects of four different dopants in the desorption electrospray ionization/post-photoionization(DESI/PI)mass spectrometry imaging.In this chapter,an efficient homogenate model is developed to compare the effects of toluene,chlorobenzene,bromobenzene and anisole on the ion intensity of the compounds in mouse brain tissue under different doping concentrations,and a deep discussion of the protonation process of the ion-molecule reaction in the post-photoionization process is shared,and the optimal doping concentrations of the four dopants are determined.Then they are applied to the mass spectrometry imaging of mouse brain tissue sections.It is found that the comprehensive performance of ion intensity,image and the number of detected compounds is the best when the solvent is doped with 30%toluene.We also find less suppression to the polar compound when the ratio of the dopant is less than 10%.In addition,we carry out the quantification experiments of endogenous compounds in mouse brain tissue,which proves the quantification ability and accuracy of DESI/PI for low polarity endogenous compounds.The profiling and localization of plant metabolites are important for exploring their biosynthesis and transport.However,DESI can not penetrate the cuticles on the surface of plant leaves to directly desorb and ionize the underlying plant tissues.Imprint is a method that transfers metabolites from plant tissues to porous PTFE materials with suitable pressure.The porous structure and inert property of the imprint materials ensure that the metabolites will not delocalize during imprinting.This imprint method indirectly eliminates the interference of the waxy layer in direct mass spectrometry imaging of plant tissues.Thus,the third chapter of the thesis introduces the research on the spatial distribution of metabolites in three kinds of plant leaves(i.e.,sage leaves,ginkgo leaves,and tea leaves)and two kinds of plant petals(Catharanthus roseus and Malus spectabilis)with DESI/PI-MSI combined with imprint.In this work,we compare the ionization efficiency and ionization coverage of plant metabolites between DESI/PI-MSI and DESI-MSI.The results show that abundant plant secondary metabolites,including terpenoids,flavonoids,amino acids,and glycosides,can be simultaneously detected and imaged by imprint DESI/PI mass spectrometry in the positive ion mode.The mass signals of some metabolites are enhanced by an order of magnitude with the assistance of PI in the negative ion mode.With the success in the imaging of plant metabolites with higher sensitivity and coverage,we proceed to further investigate the biosynthesis network of catechins in fresh tea leaves,indicating the potential applicability of imprint DESI/PI in exploring the sites and pathways of plant metabolic conversion.The imagings of two kinds of plant petals are also performed,demonstrating the superiority of imprint imaging method in imaging a variety of plant tissues.Compared to the vacuum ultraviolet discharge lamp used by DESI/PI above,which can produce a single energy photon,synchrotron radiation light source is a kind of light source with high brightness and beam density,and continuously adjustable energy.Combining it with post-ionization mass spectrometry imaging technology can broaden the energy range of photoionization and study the ionization efficiency of compounds at different photoionization energies to get the energy with the highest ionization efficiency for high sensitivity MSI experiments.Thus,the fourth chapter of the thesis introduces the exploration of desorption electrospray ionization/postphotoionization mass spectrometry imaging technology based on synchrotron radiation vacuum ultraviolet(DESI/PI/SRPI-MSI).In this work,we set up the dual-light postionization mass spectrometry imaging platform in Hefei light source mass spectrometry beamline based on synchrotron radiation vacuum ultraviolet and vacuum ultraviolet discharge lamp.The platform provides a variety of experimental modes,such as online synchrotron radiation photoionization mass spectrometry imaging and conventional off-line post-ionization mass spectrometry imaging,or multiple photoionization mass spectrometry imaging in the form of dual light sources.The preliminary result shows that the ionization efficiency of compound varies with the change of the energy of synchrotron radiation.The synchrotron radiation with high flux can effectively improve the ionization efficiency and the multi-photoionization form can further enhance the ionization efficiency.This research provides an experimental method and platform for the basic theoretical research of ionization methods in mass spectrometry imaging and molecular high-energy fragmentation.Due to the reduction of desorption compounds per unit area,high spatial resolution mass spectrometry imaging techniques are often less sensitive.Post-photoionization can effectively improve the imaging sensitivity and make up for the low intensity caused by the decrease of desorption area.Therefore,the fifth chapter of the thesis introduces the development of the atmospheric pressure laser desorption photoionization mass spectrometry imaging technology(AP-LDPI-MSI).In this work,the laser spot is focused to less than 10 μm by the optical lens and used for the desorption of compounds on the sample surface,and then the desorbed neutral molecules are ionized by a vacuum ultraviolet discharge lamp to achieve high spatial resolution mass spectrometry imaging under atmospheric pressure.And it is applied to the imaging of mouse cerebellar tissue area with 10 μm spatial resolution and vertical section of ginkgo biloba leaves with 15 μm spatial resolution.AP-LDPI-MSI successfully maps the precise spatial distribution of compounds in the cortex molecular layer,granular layer,white matter area of mouse cerebellum,and the upper epidermis,lower epidermis and secretory cavity of the vertical section of ginkgo biloba leaf.It shows the high ionization coverage of this method in the imaging research of animal and plant samples.The results demonstrate that AP-LDPI-MSI is a high spatial resolution mass spectrometry imaging method under atmospheric pressure,which has great advantages for the research of micro-scale samples and can be applied to the precise imaging research of biomedicine and botany in the future.