Investigation Of Mechanism And Application Of Induced Nanoelectrospray Ionization

Mass spectrometry(MS)has become an important tool in many fields such as life sciences,medicine,and environment with the advantages of high sensitivity,high specificity and untargeted detection.Electrospray ionization(ESI)is the most widely used atmospheric pressure ion source of MS.The ionization processes of ESI include droplets generation,solvent evaporation,and Coulomb split.Eventually,the gas phase ions formed and were detected by mass spectrometer.Because the relatively moderate ionization condition,ESI is particularly suitable for the analysis of biological macromolecules(such as proteins and DNA).Nanoelectrospray ionization(nESI)has been proposed to realize sample economy and matrix effect alleviation in biological research.Due to the smaller diameter of the spray emitter and the slower flow rate of solution,nESI produces smaller initial droplets than ESI.With a shorter solvent evaporation period,small droplets could improve the ionization efficiency and sample utilization.Therefore,this technique can be chosen for the analysis of small-volume samples and can even meet the requirements of single cell investigation.In recent years,many modified versions of nESI have been used for deepening of research in various fields.Based on the mechanism of nESI,we have also presented an ion source with higher matrix-tolerance,sample utilization and throughput,that is induced nESI(InESI).Matrix effect in MS detection usually occurs during the ionization process.We have experimentally found that InESI-MS were less affected by matrix effect than nESI-MS.The higher matrix tolerance may be due to the separation effect and smaller droplets in InESI.Investigating the differences of the mechanism between InESI and nESI could deepen our understanding of InESI.This paper will discuss the mechanisms and applications of InESI from the following three aspects:(1)The microelectrophoresis in InESIInESI periodically spray along with the variation of pulsed potential.Before the solution was initiated to spray,the charged compounds moved directionally due to the effect of microelectrophoresis.The constituents of solution were separated according to the diversity of migration rates.The microelectrophoresis usually lasts several to a few tens of milliseconds,when 10-100 Hz frequency is used.Benefit by the microelectrophoresis,the substances that have been separated would spray and ionize in sequence when the solution started to spray.The separation of analytes and interferences reduces the matrix effect during detection process.Time-resolved experiments were used to verify the existence of microelectrophoresis,the analytes were observed to appear in order during each spray cycle.In addition,we employed a variety of model compounds to systematically discuss the restricted condition of microelectrophoresis.Because of the detachment of analytes from matrixes prior to MS analysis,the metabolites in artificial intracellular fluid could be directly measured by using InESI.(2)Using InESI for in situ investigating the interaction of protein and metal ion Nonspecific binding between proteins and metal ions usually occurs during the ionization stage.Based on the significantly different migration rates between protein and metal ion,we deduced that the nonspecific interaction of protein and metal ion could be reduced because of the microelectrophoresis effect.The solutions that contained proteins(melittin,cytochrome c,myoglobin)and high concentrated salts(1-150 mM NaCl)were analyzed by using InESI and nESI when micron-sized(1-20μm)spray emitters were employed.The results indicated that the signals of proteins were distinct in InESI-MS,while the mass spectra of proteins were almost covered by the signals of Na+ clusters and Na+adducts in nESI-MS.It proved that microelectrophoresis could alleviate the interference that caused by nonspecific binding.The protein could be analyzed in situ because of the better signal to noise of InESI.We also used InESI to investigate the interaction between copper chaperone proteins and Cu+ under high concentrated salt(100 mM NaCI)condition.Furthermore,we proved that microelectrophoresis could detach nonspecific binding of zinc finger protein and Zn2+,while the specific binding has been retained.Thus,InESI-MS has a great potential to reveal more native protein-metal ion interactions that involved in biological functions.(3)Small droplets that produced by InESIThe size of droplets will greatly affect the performance of electrospray,such as the ionization efficiency and matrix effect.In order to verify whether the initial droplets produced by InESI were smaller than that of conventional nESI,we employed phase Doppler particle analyzer to measure the droplet sizes of InESI and nESI plume.The data shows that InESI could generate smaller droplets because of the slower flow rate.With the interfacial effect and large specific surface area,droplets could accelerate the chemical reactions.The initial droplets generated by InESI are often with micron-sized diameters,so it could effectively accelerate some chemical reactions which take a long time in bulk solution.The reaction of p-nitrophenol reduced to p-aminophenol in InESI spray process has been employed to illustrate the acceleration effect.The reaction time in droplet was about 4.3 × 10-4 s.When the concentration of the catalyst was 1 μM,the conversion rate and average reaction rate was calculated to be 62.28%and 0.1448 mol·L-1·s-1 in droplets;as the concentration of the catalyst increased to 10 μM,the conversion rate and average reaction rate was up to 74.37%and 0.1730 mol·L-1·s-1 in droplets.Utilized ultraviolet-visible spectrophotometer to monitor the reduced reaction in bulk solution,we observed the conversion rate reached 80.26%in 13 minutes.The average reaction rate was calculated to be 1.03 × 10-7 mol·L-1·s-1.Comparing the reaction rate between droplets and bulk solutions,we can conclude that droplets generated by InESI have greatly accelerated the reduction of p-nitrophenol around 1.68 × 106 times greater than that in the bulk.Take advantage of the reaction acceleration of droplets,the silver ions could be reduced to produce silver nanoparticles in the InESI spray process.We have demonstrated that the droplets generated by InESI could prepare silver nanoparticles with smaller size and better dispersion.