Development And Application Of Plasmonic Effect-Based Laser Desorption/Ionization Mass Spectrometric Method

Laser desorption ionization(LDI)has been an essential ionization means for biological mass spectrometry(MS).It relies on an energy transfer medium to adsorb the light energy and convert it into other forms of energy to desorb and ionize analyte molecules.Due to ultrafast ionization process(ns-level time window)in accompany with complicated chemical reactions(e.g.proton transfer,electron transfer,cation transfer and electron detachment),LDI mechanism study is difficult to implement.Various organic matrix and inorganic materials have been used as ionization media,however,complex processes during LDI as well as the large variety of energy transfer media make it extremely difficult to describe a complete and clear picture,which hampers access to build up a general and predictive foundation.Acknowledge of the ionization mechanisms is essential for MS,which can enable not only improving the ionization efficiency but also accurate structural identification.Molecular structure is closely associated with its function,thus structural analysis always provides the foundation for other related researches.Glycosylation is one of most frequently occurred post-translational modifications.As the products of glycosylation,glycans play essential roles in many key biological processes,such as cell adhesion,cell growth and differentiation,intercellular recognition,protein folding,signaling and immunological response.These processes change with glycan structural alternation.Therefore,the structural analysis of glycans is very essential.Different from peptides and nucleic acids,glycan biosynthesis is a non-templated and non-linear process,thus glycan can not be amplified like nucleic acids or facilely synthesized like peptides.This causes glycan sample amounts are very limited.Meanwhile,microheterogeneity of glycans also increases the difficulty in analysis.Additionally,lower ionization efficiency makes it harder for identification of complex and isomeric glycans.Current methods always require troublesome derivatization and complicated tandem MS,which hamper efficient identification of glycans.Plasmon is a special physical phenomenon produced by the interaction of light and plasmonic nanomaterials,in which electromagnetic waves couple to the collective oscillations of free electrons in the metal.Noble metal nanomaterials,which are well-known as plasmonic nanomaterials and exhibit unique plasmonic effects and thereby have found important applications such as plasmon enhanced Raman spectroscopy,fluorescence and photochemical reactions.Although plasmon has been proved effective on enhancement of Raman and fluorescence and mediation in photocatalysis,the roles of plasmonic effects in plasmonic nanomaterials-based LDI have not been well explored yet.Herein,with plasmonic nanomaterials as ionization media,through probing the plasmon induced ionization mechanism,we employed the physical phenomenon induced chemical reaction for LDI-MS.Based on this,we proposed an efficient glycan dissection method for structural analysis in depth.Firstly,we proposed and verified a new mechanism,called plasmon-induced charge transfer desorption/ionization(PICTDI).By combination of theoretical simulation and experimental verification,we demonstrated that interaction between laser and noble metal nanomaterials can produce intensive near-field enhancement that was closely related to molecular ionization.Upon being shined with a laser beam,plasmonic nanoparticles,such as gold nanoparticles(AuNPs),silver nanoparticles(AgNPs)or their mixture,transfer charges(electrons,protons or other cations)from the nanoparticles to surrounding analytes through surface plasmon,leading to analyte desorption and ionization.A variety of biomolecules,from nucleosides to saccharides to short peptides,were found to be ionisable according to this rule.This mechanism provides not only a sound understanding for molecular ionization but also a useful guideline for rational design of new plasmonic charge transfer nanomedia.Secondly,structural identification of glycans is important but remains challenging,for which tandem mass spectrometry has evolved as an indispensable tool.However,it requires additional complex hardware and extra time for ion extraction.Herein,we report a straightforward approach called gold nanoparticles(AuNPs)-assisted in-source cation adduction dissociation(isCAD)for efficient MS dissection of glycans.Although AuNPs have been employed as an inorganic matrix for MALDI MS,this is the first report of AuNP-induced fragmentation.In this approach,AuNPs were employed as an energy absorber for laser ionization as well as a trigger for fragmentation,while residual or deliberately added sodium ion acted as a cationizing agent.The addition of sodium ion induced intensive fragmentation but the addition of proton suppressed the fragmentation,allowing for facile tuning the degree of fragmentation.Besides,it was found that larger oligosaccharides and glycans were much easier to be fragmented as compared with their smaller counterparts and the use of high-concentration AuNPs effectively suppressed the degree of fragmentation and thereby provided abundant molecular ions.Without any extra hardware and ion extraction,this approach provides a straightforward,cost-efficient and tunable fragmentation for efficient MS dissection of saccharides,including monosaccharides,oligosaccharides and glycans.Thus,it opens a new access to efficient MS dissection of glycans.Finally,carbohydrate isomers identification remains a challenge task in mass spectrometry(MS)owing to the diversity of saccharides in stereochemistry.Currently,sophisticated MS based techniques have been developed as the primary tool for carbohydrate isomer identification.However,resolving minor structural differences,especially for isobaric variants,remains a challenge task.Here,based on our previous study,we proposed an improved carbohydrate dissection method,called shell enhanced in-source cation adduction dissociation,for unambiguous identification of carbohydrate isomers.Gold nanoparticles(AuNPs)have been demonstrated as efficient medium for ionization and fragmentation of carbohydrates,but the potential of fine structural analysis remains unexplored.Compared to bare AuNPs,silica shell isolated AuNPs provided much higher ionization and fragmentation efficiency.Relying on specific fragmentation patterns,isomeric saccharides can be distinguished by diagnostic ions.Various carbohydrates have been tested,including disaccharides and Lewis antigen oligosaccharides with different connectivity,configuration and composition.Moreover,the relative quantitative analysis of binary mixture of carbohydrate isomers demonstrated that this method can be used for quality control of as less as 1%isomer impurities.Without any extra hardware and ion extraction,this method provides a straightforward,cost-efficient and high resolution for effective identification of carbohydrate isomers.Thus,it opens a new avenue for mass spectrometric identification of isomeric biomolecules,such as glycans with isomeric terminals or dubious antenna structures.