Electron impact ionization in helium nanodroplets: controlling fragmentation by active cooling of molecular ions

Reported here is a study of the effects of liquid helium cooling on the fragmentation of ions formed by electron impact mass ionization. The molecules of interest are picked up by the helium nanodroplets as they pass through a low pressure oven. Electron impact ionization of a helium atom in the droplet is followed by resonant charge transfer to neighboring helium atoms. When the charge is transferred to the target molecule, the difference in the ionization potentials between helium and the molecule results in the formation of a vibrationally hot ion. In isolation, the hot parent ion would undergo subsequent fragmentation. On the other hand, if the cooling due to the helium is fast enough, the parent ion will be actively cooled before fragmentation occurs. The target molecule used in the present study is triphenylmethanol (TPM), an important species in synthetic chemistry, used to sterically protect hydroxyl groups. Threshold PhotoElectron PhotoIon COincidence (TPEPICO) experiments are also reported for gas-phase TPM to help quantify the ion energetics resulting from the cooling effects of the helium droplets.     

Fragmentation of HCN in optically selected mass spectrometry: nonthermal ion cooling in helium nanodroplets

A technique that combines infrared laser spectroscopy and helium nanodroplet mass spectrometry, which we refer to as optically selected mass spectrometry, is used to study the efficiency of ion cooling in helium. Electron-impact ionization is used to form He(+) ions within the droplets, which go on to transfer their charge to the HCN dopant molecules. Depending upon the droplet size, the newly formed ion either fragments or is cooled by the helium before fragmentation can occur. Comparisons with gas-phase fragmentation data suggest that the cooling provided by the helium is highly nonthermal. An "explosive" model is proposed for the cooling process, given that the initially hot ion is embedded in such a cold solvent.     

Possible conformational change within the desolvated and cationized sBBI/trypsin non-covalent complex during the collision-induced dissociation process

Electrospray ionization mass spectrometry (ESI-MS) has become an analytical technique widely used for the investigation of non-covalent protein-protein and protein-ligand complexes due to the soft desolvation conditions that preserve the stoichiometry of the interacting partners. Dissociation studies of solvated or desolvated complexes (in the source and in the collision cell, respectively) allow access to information on protein conformation and localization of the metal ions involved in protein structure stabilization and biological activity. The complex of bovine trypsin and small soybean Bowman-Birk inhibitor (sBBI) was studied by ESI-MS to determine changes occurring within the complex during its transfer from droplets to the gas phase independently of the ion polarity. Under collision-induced dissociation (CID) conditions, unexpected binding of the Ca(2+) ion (cofactor of native trypsin) to the inhibitor molecule was observed within the desolvated sBBI/trypsin/Ca(2+) complex (with a 1:1:1 stoichiometry). This formal gas-phase migration of the calcium ion from trypsin to the inhibitor may be related to conformational rearrangements in the solvent-free and likely collapsed complex. However, under conditions leading to the increase in complex charge state, the appearance of the cationized trypsin molecule was detected during complex dissociation, thus reflecting different pathways of the evolution of complex conformation.     

Photoexcitation of mass/charge selected hemin(+), caught in helium nanodroplets

We report on a method by which mass/charge selected ions are picked up from a linear ion trap by liquid helium droplets. The size distributions of the doped droplets are measured via acceleration experiments. Depending on the source temperature, droplet sizes ranging from tens of thousands to several million helium atoms are obtained. Droplets doped with hemin, an iron containing porphyrin molecule, in the charge state +1 are then investigated using laser spectroscopy. It is observed that excitation with UV/VIS light can lead to ejection of the ion from the droplet. For doped droplets with a median size of ～150?000 helium atoms, the absorption of two photons at 380 nm is needed for ejection to become efficient. When droplets become smaller, the ejection efficiency is observed to strongly increase. Monitoring the ejection yield as a function of excitation wavelength can be used to obtain the optical spectrum of hemin(+). Compared to the spectrum of free gas-phase hemin(+) at room temperature, the here obtained spectrum is slightly narrower and shifted to the blue.     

Probing the mechanisms of ambient ionization by laser‐induced fluorescence spectroscopy

The ionization mechanisms of several atmospheric pressure ion sources based on desorption and ionization of samples deposited on a surface were studied. Home‐built desorption electrospray ionization (DESI), laserspray ionization (LSI), and atmospheric pressure matrix‐assisted laser desorption/ionization (AP‐MALDI) sources were characterized using low‐molecular‐weight compounds, in particular fluorescent dyes. Detection of the desorbed and ionized species was performed by laser‐induced fluorescence and ion cyclotron resonance mass spectrometry. The dependences of the signal intensities on various experimental parameters were studied. The data obtained reveals common features, such as formation of solvated species and clusters in the ionization processes, in all of the techniques considered. Copyright © 2012 John Wiley & Sons, Ltd.     

Communication: the formation of helium cluster cations following the ionization of helium nanodroplets: influence of droplet size and dopant

The He(n)(+)/He(2)(+) (n ≥ 3) signal ratios in the mass spectra derived from electron impact ionization of pure helium nanodroplets are shown to increase with droplet size, reaching an asymptotic limit at an average droplet size of approximately 50,000 helium atoms. This is explained in terms of a charge hopping model, where on average the positive charge is able to penetrate more deeply into the liquid helium as the droplet size increases. The deeper the point where the charge localizes to form He(2)(+), the greater the likelihood of collisions with the surrounding helium as the ion begins to leave the droplet, thus increasing the probability that helium will be ejected in the form of He(n)(+) (n ≥ 3) cluster ions rather than He(2)(+). The addition of a dopant alters the He(n)(+)/He(2)(+) ratio for small helium droplets, an observation attributed to the potential energy gradient created by the cation-dopant interaction and its effect in drawing the positive charge towards the dopant in the interior of the droplet.     

Ion-molecule reactions and fragmentation patterns in helium nanodroplets

A study has been made of the ion chemistry of a series of small molecules that have been embedded in helium nanodroplets. In most instances, the molecules H2O, SO2, CO2, CH3OH, C2H5OH, C3H7OH, CH3F, and CH3Cl have been allowed to form clusters, and reactivity within these has been initiated through electron impact ionization. For two of the molecules studied, CF2Cl2 and CF3I, reactivity is believed to originate from single molecules embedded in the droplets. Electron impact on the droplets is thought to first create a helium ion, and formation of molecular ions is then assumed to proceed via a charge hopping mechanism that propagates though the droplet and terminates with charge-transfer to a molecule or cluster. The chemistry exhibited by many of the cluster ions and at least one of the single molecular ions is very different from that observed for the same species in isolation. In most cases, reactivity appears to be dominated by high-energy bond breaking processes as opposed to, in the case of the clusters, ion-molecule reactions. Overall, charge-transfer from He+ does not appear to be a "soft" ionization mechanism.     

A new sensitive detection scheme for helium nanodroplet isolation spectroscopy: application to benzene

A new method is presented for recording excitation spectra of molecules embedded in helium nanodroplets. The method relies on the complete evaporation of the droplets following excitation of a dissolved molecule and the subsequent detection of the remaining unsolvated molecule by mass spectrometry. The technique has been successfully applied to record the S1 1B(2u) <-- S0 1A(1g) transition in benzene. The transition frequencies determined by this new method, beam depletion spectroscopy and REMPI spectroscopy have been found to differ slightly from each other. It is argued that these differences in transition frequency are related to the different droplet sizes probed by the spectroscopic techniques.     

Inelastic electron interaction with chloroform clusters embedded in helium droplets

The inelastic electron interaction (ionization/attachment) with chloroform embedded in helium droplets has been studied utilizing a two-sector field mass spectrometer. Positive mass spectra have been recorded at the electron energy of 70 eV and are compared with previous results in the gas phase and with other systems embedded in helium droplets. Moreover, the negative ion mass spectrum has been recorded at the electron energy of 1.5 eV. Both negative and positive mass spectra show that chloroform clusters are easily formed by embedding single molecules in the helium droplets. Moreover, for anions appearing in the mass spectrum, the ion yield has been determined as function of the electron energy. While no parent anion of chloroform can be observed in the gas phase, the present cluster environment allows the stabilization of the transient negative ion. The influence of the helium droplet upon the ionization or attachment process of the embedded chloroform is discussed.     

A cylindrical capacitor ionization source: droplet generation and controlled charge reduction for mass spectrometry

A cylindrical capacitor ionization source was used in conjunction with corona discharge charge reduction for generation of singly charged ions for mass spectrometric analysis. The source consists of a fused-silica capillary threaded with a platinum wire and placed inside a stainless steel tube. Application of an electric potential to the wire results in the production of a linear stream of charged droplets when an aqueous solution is pumped through the capillary. Subsequent solvent evaporation yields ions, providing a continuous ion source for mass spectrometry. Passage of the ions through a corona discharge charge reduction chamber permits reduction of the charge state to predominantly singly charged species, facilitating analysis of DNA and protein mixtures. The change from production of multiply charged ions to production of singly charged ions is extremely simple, requiring only modulation of the voltage applied to the corona discharge electrode. A simple technique for construction of the ionization source is reported.     

Charge assisted laser desorption/ionization mass spectrometry of droplets

We propose and evaluate a new mechanism to account for analyte ion signal enhancement in ultraviolet-laser desorption mass spectrometry of droplets in the presence of corona ions. Our new insights are based on timing control of corona ion production, laser desorption, and peptide ion extraction achieved by a novel pulsed corona apparatus. We demonstrate that droplet charging rather than gas-phase ion-neutral reactions is the major contributor to analyte ion generation from an electrically isolated droplet. Implications of the new mechanism, termed charge assisted laser desorption/ionization (CALDI), are discussed and contrasted with those of the laser desorption atmospheric pressure chemical ionization method (LD-APCI). It is also demonstrated that analyte ion generation in CALDI occurs with external electric fields about one order of magnitude lower than those needed for atmospheric pressure matrix assisted laser desorption/ionization or electrospray ionization of droplets.     

Is there a minimum size for aqueous doubly charged metal cations?

A major feature of the chemistry of multiply charged solvated metal ions is dissociative charge transfer. It happens because the second ionization potential (IP) of a metal atom usually exceeds the first IP of a solvent molecule. This raises the issue of whether there is a minimum number of ligands below which the species would charge-separate spontaneously. To elucidate this, doubly charged aqueous cations of most common divalent metals (group 2 elements Mn, Fe, Co, Ni, Zn, Cd, and Cu) have been generated using electrospray and examined by collision-induced dissociation in a triple-quadrupole mass spectrometer. We have clearly observed the monoaqua complexes for all aforementioned doubly charged metal ions, except Be for which the smallest complex found is the dihydrate. We have also systematically revisited the matter of critical size--the maximum number of ligands at which dissociative charge transfer is competitive with simple ligand loss.     

Electron correlation as the driving force for charge transfer: charge migration following ionization in N-methyl acetamide

A hole charge created in a molecule, for instance, by ionization, can migrate through the system solely driven by electron correlation. The migration of a hole charge following ionization in N-methyl acetamide (a molecular system containing a peptide bond) is investigated. The initial hole charge is localized at one specific site of the molecule. Ab initio calculations show that nearly 90% of the hole migrates to a remote site of the molecule in 4.2 fs. This migration of charge is highly efficient and ultrafast. The underlying mechanism for this migration of a hole charge is identified and compared with a simple model.     

嗜酸性氧化亚铁硫杆菌代谢产物的常压化学电离质谱分析

本研究以721矿和745矿嗜酸性氧化亚铁硫杆菌为研究对象,采用常压化学电离质谱直接分析其代谢产物,分别考察了顶空采样( Headspace sampling)、界面采样( Interface sampling)和中性解吸采样( Neutral desorption sampling)3种进样方式对电离效果的影响。在优化条件下,常压化学电离质谱对微生物纯菌种和混合菌种的代谢产物均具有良好的分析能力,可根据获得的代谢产物指纹谱图结合主成分分析( PCA)方法和聚类分析( CA)方法区分2个放射性强弱不同区域共4类嗜酸性微生物样品,并对主要胺类、酯类等代谢成分进行串联质谱鉴定,为耐辐射微生物的相关研究提供了一种可借鉴的分析方法。     

Promotion of <Emphasis Type="Italic">α</Emphasis>-cyano-4-hydroxycinnamic acid and peptide cocrystallization within levitated droplets with net charge

Many reactions occur as a result of charge imbalance within or between reactive species in reaction vessels that have zero net charge. Here, chemical processes taking place within reaction vessels having net excess charge were studied. For mass spectroscopists, a familiar example of vessels that defy electroneutrality are the charged droplets produced by an electrospray ion source. Evidence is presented that control of the magnitude of the net charge contained in a reaction vessel, in this case a levitated droplet, can be used to promote nucleation and crystal growth of a mixture of an organic acid, alpha-cyano-4-hydroxycinnamic acid (CHCA), with one or more peptides. This phenomenon was first observed during our ongoing development of wall-less sample preparation (WaSP), electrodynamic charged droplet processing methodology capable of creating micrometer-sized sample spots for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) from subnanoliter volumes of sample material. Peptide ion signal-to-noise (S/N) ratios obtained by MALDI-TOF-MS from sample spots created from droplets that had high relative magnitude of net charge were consistently greater than those detected from sample spots created from droplets that had lower net charge. To study this unexpected phenomenon further, WaSP methodology was developed to process different mass-to-charge (m/z) droplets levitated in an electrodynamic balance (EDB), facilitating their deposition onto different positions of a target to create arrays of droplet residues ordered from highest to lowest m/z. This capability allowed simultaneous levitation with subsequent separation of a population of droplets created from a single starting solution, but the droplets had varied magnitudes of net charge. After the droplets were ejected from the EDB and collected on a glass slide or MALDI plate, the solids contained in the deposited droplets were characterized using microscopy and MALDI-TOF-MS. Factors impacting the chemical processing in droplets having net excess charge levitated in an EDB are discussed with particular emphasis on their possible roles in the promotion of crystal nucleation and growth.     

Fragmentation of Neutral Amino Acids and Small Peptides by Intense, Femtosecond Laser Pulses

High power femtosecond laser pulses have unique properties that could lead to their application as ionization or activation sources in mass spectrometry. By concentrating many photons into pulse lengths approaching the timescales associated with atomic motion, very strong electric field strengths are generated, which can efficiently ionize and fragment molecules without the need for resonant absorption. However, the complex interaction between these pulses and biomolecular species is not well understood. To address this issue, we have studied the interaction of intense, femtosecond pulses with a number of amino acids and small peptides. Unlike previous studies, we have used neutral forms of these molecular targets, which allowed us to investigate dissociation of radical cations without the spectra being complicated by the action of mobile protons. We found fragmentation was dominated by fast, radical-initiated dissociation close to the charge site generated by the initial ionization or from subsequent ultrafast migration of this charge. Fragments with lower yields, which are useful for structural determinations, were also observed and attributed to radical migration caused by hydrogen atom transfer within the molecule.

Sequential enrichment of sulfated glycans by strong anion-exchange chromatography prior to mass spectrometric measurements

Structural characterization of sulfated glycans through mass spectrometry (MS) has been often limited by their low abundance in biological materials and inefficient ionization in the positive-ion mode. Here, we describe a microscale method for sequentially enriching sulfated glycans according to their degree of sulfation. This method is based on modifying the binding ability of strong anion-exchange material through the use of different sodium acetate concentrations, thus enabling fairly selective binding and a subsequent elution of different glycans according to their degree of sulfation. Before this enrichment, the negative charge on the sialic acid, which is commonly associated with such glycans, was eliminated through permethylation that is used to enhance the positive-ion mode matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-MS) signal for all glycans. This enrichment approach minimizes competitive ionization between sulfated and neutral glycans, as well as that between sulfated species with a different degree of sulfation. The described method was initially optimized using sulfated oligosaccharide standards, while its potential has been verified for the sulfated N-glycans originated from the bovine thyroid-stimulating hormone (bTSH), a glycoprotein possessing mono- and disulfated N-glycans. This enhancement of the MALDI-MS signal facilitates analysis of some otherwise undetected components.     

Ambient analysis by thermal desorption atmospheric pressure photoionization

Ambient mass spectrometry has attracted substantial attention in recent years. Among ambient ionization methods, thermal desorption ionization stands out because of two attributes: (1) simplicity, rendering the technique suitable for in-field applications, and (2) ability to couple with a variety of gas-phase ionization methods thereby broadening the range of molecules that can be analyzed with this method. Here, we report on improving the performance of a direct analysis in real time (DART) source by implementing atmospheric pressure photoionization (APPI) downstream of the desorption region. At identical desorption and ion sampling conditions, APPI leads to detection of radical molecular ions from non-polar compounds that are absent from the spectra generated by DART alone. Moreover, a factor of 3–5 improvement in sensitivity is observed using APPI for positive ions commonly detected by DART and DART-APPI. Using helium and nitrogen as desorption gases, APPI shows identical performance regardless of desorption gas type. In contrast, a dramatic decrease in sensitivity is observed for DART operated with nitrogen compared to DART with helium. Comparable performance for DART and DART-APPI are observed in negative ion mode, although both show a drastic improvement in the absence of the Vapur interface. This interface creates a differentially pumped chamber prior to inlet of the mass spectrometer and reduces the mass spectrometer gas load when helium is used as desorption gas.     

Ion induced snowballs as a diagnostic tool to investigate the caging of metal clusters in large helium droplets

Metal clusters embedded in ultracold helium nanodroplets are exposed to femtosecond laser pulses with intensities of 10(13)-10(14) W/cm2. The influence of the matrix on the ionization and fragmentation dynamics is studied by pump-probe time-of-flight mass spectrometry. Special attention is paid to the generation of helium snowballs around positive metal ions (Me(z+)He(N), z=1,2). Closings of the first and second helium shells are found for silver at N(1)=10,12 and N(2)=32,44, as well as for magnesium at N1=19-20. The distinct abundance enhancement of helium snowballs in the presence of isolated atoms and small clusters in the droplets is used as a diagnostics to explore the cage effect. For silver, a reaggregation of the clusters is observed at 30 ps after femtosecond laser excitation.     

Hydration of gas-phase ions formed by electrospray ionization

The hydration of gas-phase ions produced by electrospray ionization was investigated. Evidence that the hydrated ions are formed by two mechanisms is presented. First, solvent condensation during the expansion inside the electrospray source clearly occurs. Second, some solvent evaporation from more extensively solvated ions or droplets is apparent. To the extent that these highly solvated ions have solution-phase structures, then the final isolated gas-phase structure of the ion will be determined by the solvent evaporation process. This process was investigated for hydrated gramicidin S in a Fourier-transform mass spectrometer. Unimolecular dissociation rate constants of isolated gramicidin S ions with between 2 and 14 associated water molecules were measured. These rate constants increased from 16 to 230 s-1 with increasing hydration, with smaller values corresponding to magic numbers.