Solvation of acylium fragment ions in electrospray ionization quadrupole ion trap and Fourier transform ion cyclotron resonance mass spectrometry

In electrospray ionization (ESI) quadrupole ion trap and Fourier transform ion cyclotron resonance mass spectrometry, certain fragment ions (e.g. acylium ions) generated either during the ion transportation process (in the source interface region) or in the ion trap are found to undergo ion--molecule reactions with ESI solvent molecules (water, acetonitrile and aliphatic alcohols) to form adduct species. These unexpected solvated fragment ions severely complicate the interpretation of mass spectrometic data. High-resolution accurate mass measurements are important in establishing the elemental compositions of these adduct species and preventing erroneous data interpretation.     

On the detailed mechanisms of collision-induced dissociation experiments performed by electrospray ion trap

The product ion spectra obtained by electrospray ionization (ESI) ion trap instruments exhibit a higher number of fragment ions than those achieved by other ion-trap-based systems, indicating the presence of more effective energy deposition mechanisms. This behavior can be attributed to several different reasons, among which different initial internal energy of the precursor ions, pre-activation due to collisions taking place outside the trap, different target gas mixtures inside the trap, and different ion trap geometry were considered. Data obtained from experiments using a triple quadrupole instrument, CI-ion trap, and ESI-ion trap have been compared. The results so achieved seem to indicate that the presence inside the trap of neutral molecules of the solvent employed for the ESI process have a relevant role, promoting high energy deposition in the colliding ions.     

An ion trap-ion mobility-time of flight mass spectrometer with three ion sources for ion/ion reactions

This instrument combines the capabilities of ion/ion reactions with ion mobility (IM) and time-of-flight (TOF) measurements for conformation studies and top-down analysis of large biomolecules. Ubiquitin ions from either of two electrospray ionization (ESI) sources are stored in a three dimensional (3D) ion trap (IT) and reacted with negative ions from atmospheric sampling glow discharge ionization (ASGDI). The proton transfer reaction products are then separated by IM and analyzed via a TOF mass analyzer. In this way, ubiquitin +7 ions are converted to lower charge states down to +1; the ions in lower charge states tend to be in compact conformations with cross sections down to ～880 Ų. The duration and magnitude of the ion ejection pulse on the IT exit and the entrance voltage on the IM drift tube can affect the measured distribution of conformers for ubiquitin +7 and +6. Alternatively, protein ions are fragmented by collision-induced dissociation (CID) in the IT, followed by ion/ion reactions to reduce the charge states of the CID product ions, thus simplifying assignment of charge states and fragments using the mobility-resolved tandem mass spectrum. Instrument characteristics and the use of a new ion trap controller and software modifications to control the entire instrument are described.     

Development and characterization of an electrospray ionization ion trap/linear time-of-flight mass spectrometer

On-line analysis of compounds from solution has been greatly facilitated by the advent of electrospray ionization mass spectrometry (ESI-MS). Although quadrupole mass analyzers are most commonly used with ESI at present, time-of-flight (TOF) mass spectrometers offer several potential advantages including high data acquisition rates, which are desirable for fast separation techniques. One method of coupling ESI and TOF uses an ion trap for temporary storage and accumulation of the electrosprayed ions prior to TOF mass analysis. Previous studies have not fully addressed the effects of several key variables on the analytical capabilities of this type of instrument. In this study, the characterization of an ion trap/linear TOF instrument for ESI is described. The behavior of various analytes is divided into two separate groups; each one is found to have its own optimal set of operating conditions. The reasons for the observed differences between groups are explored. Issues relevant to mass resolution, sensitivity, mass range, mass-to-charge ratio discrimination, and mass measurement accuracy are addressed. Finally, it is suggested that the analytical capability of this type of instrument could be significantly improved by changing the ion optics from the existing focusing lenses to a rf-only quadrupole lens.     

A digital ion trap mass spectrometer coupled with atmospheric pressure ion sources

In a digital ion trap (DIT), the quadrupole trapping and excitation waveforms are generated by the rapid switching between discrete d.c. voltage levels. As the timing of the switch can be controlled precisely by digital circuitry, the approach provides an opportunity to generate mass spectra by means of a frequency scan in contrast to the conventional voltage scan, thus providing a wider mass range of analysis. An instrument has been constructed which employs a 'non-stretched' ion trap and the field fault around the aperture of the end-cap electrode can be corrected electronically using a field-adjusting electrode. The ion trap was coupled with electrospray ionization (ESI) and atmospheric pressure matrix-assisted laser desorption/ionization (AP-MALDI) sources to demonstrate the capability of the digital method. AP-MALDI mass spectra of singly charged ions with mass-to-charge ratios upto 17 000 Th were generated using a trapping voltage of only 1000 V. Forward and reverse mass scans at resolutions up to 19 000 and precursor ion isolation at resolutions up to 3500 with subsequent tandem mass spectrometric analysis were demonstrated. The method of generating the digital waveforms and period scan is described. Discussion of the issues of mass range, scan speed, ion trapping efficiency and collision-induced dissociation efficiency are also provided.     

Effects of ion/ion proton transfer reactions on conformation of gas-phase cytochrome <Emphasis Type="Italic">c</Emphasis> ions

Positive ions from cytochrome c are studied in a 3-D ion trap/ion mobility (IM)/quadrupole-time-of-flight (TOF) instrument with three independent ion sources. The IM separation allows measurement of the cross section of the ions. Ion/ion reactions in the 3-D ion trap that remove protons cause the cytochrome c ions to refold gently without other degradation of protein structure, i.e., fragmentation or loss of heme group or metal ion. The conformation(s) of the product ions generated by ion/ion reactions in a given charge state are similar regardless of whether the cytochrome c ions are originally in +8 or +9 charge states. In the lower charge states (+1 to +5) cytochrome c ions made by the ion/ion reaction yield a single IM peak with cross section of ～1110 to 1180 Ų, even if the original +8 ion started with multiple conformations. The conformation expands slightly when the charge state is reduced from +5 to +1. For product ions in the +6 to +8 charge states, ions created from higher charge states (+9 to +16) by ion/ion reaction produce more compact conformation(s) in somewhat higher abundances compared with those produced directly by the electrospray ionization (ESI) source. For ions in intermediate charge states that have a variety of resolvable conformers, the voltage used to inject the ions into the drift tube, and the voltage and duration of the pulse that extracts ions from the ion trap, can affect the observed abundances of various conformers.     

A linear ion trap mass spectrometer with versatile control and data acquisition for ion/ion reactions

A linear ion trap (LIT) with electrospray ionization (ESI) for top-down protein analysis has been constructed. An independent atmospheric sampling glow discharge ionization (ASGDI) source produces reagent ions for ion/ion reactions. The device is also meant to enable a wide variety of ion/ion reaction studies. To reduce the instrument's complexity and make it available for wide dissemination, only a few simple electronics components were custom built. The instrument functions as both a reaction vessel for gas-phase ion/ion reactions and a mass spectrometer using mass-selective axial ejection. Initial results demonstrate trapping efficiency of 70% to 90% and the ability to perform proton transfer reactions on intact protein ions, including dual polarity storage reactions, transmission mode reactions, and ion parking.     

Evolution of instrumentation for the study of gas-phase ion/ion chemistry via mass spectrometry

The scope of gas-phase ion/ion chemistry accessible to mass spectrometry is largely defined by the available tools. Due to the development of novel instrumentation, a wide range of reaction phenomenologies has been noted, many of which have been studied extensively and exploited for analytical applications. This perspective presents the development of mass spectrometry-based instrumentation for the study of the gas-phase ion/ion chemistry in which at least one of the reactants is multiply charged. The instrument evolution is presented within the context of three essential elements required for any ion/ion reaction study: the ionization source(s), the reaction vessel or environment, and the mass analyzer. Ionization source arrangements have included source combinations that allow for reactions between multiply charged ions of one polarity and singly charged ions of opposite polarity, arrangements that enable the study of reactions of multiply charged ions of opposite polarity and, most recently, arrangements that allow for ion formation from more than two ion sources. Gas-phase ion/ion reaction studies have been performed at near atmospheric pressure in flow reactor designs and within electrodynamic ion traps operated in the mTorr range. With ion trap as a reaction vessel, ionization and reaction processes can be independently optimized and ion/ion reactions can be implemented within the context of MSn experiments. Spatial separation of the reaction vessel from the mass analyzer allows for the use of any form of mass analysis in conjunction with ion/ion reactions. Time-of-flight mass analysis, for example, has provided significant improvements in mass analysis figures of merit relative to mass filters and ion traps.     

Reaction of analyte ions with neutral chemical ionization gas

Analytical Chemistry Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee, USA Ion-molecule reactions of neutral methane with analyte ions under normal methane chemical ionization conditions are discussed. Reactant ions can be generated by direct electron ionization (EI) fragmentation, chemical ionization (CI) fragmentation, or collision-induced dissociation (CID). Examples in which products of such reactions appear in mass spectra in both conventional CI sources in “beam” instruments and low pressure CI in a quadrupole ion trap are presented. Also shown is an example in which MS/MS product ions react with neutral methane used for CI in an ion trap. It is shown that it is relatively straightforward to recognize such reactions in a quadrupole ion trap and in certain cases to minimize or preclude them. Effects of various operating parameters have been investigated and are discussed.     

A pulsed triple ionization source for sequential ion/ion reactions in an electrodynamic ion trap

A pulsed triple ionization source, using a common atmosphere/vacuum interface and ion path, has been developed to generate different types of ions for sequential ion/ion reaction experiments in a linear ion trap-based tandem mass spectrometer. The triple ionization source typically consists of a nano-electrospray emitter for analyte formation and two other emitters, an electrospray emitter and an atmospheric pressure chemical ionization emitter or a second nano-electrospray emitter for formation of the two different reagent ions. The three emitters are positioned in a parallel fashion close to the sampling orifice of the tandem mass spectrometer. The potentials applied to each emitter are sequentially pulsed so that desired ions are generated separately in time and space. Sequential ion/ion reactions take place after analyte ions of interest and different set of reagent ions are sequentially injected into a linear ion trap, where axial trapping is effected by applying an auxiliary radio frequency voltage to the end lenses. The pulsed triple ionization source allows independent optimization of each emitter and can be readily coupled to any atmospheric pressure ionization interface with no need for instrument modifications, provided the potentials required to transmit the ion polarity of interest can be synchronized with the emitter potentials. Several sequential ion/ion reactions examples are demonstrated to illustrate the analytical usefulness of the triple ionization source in the study of gas-phase ion/ion chemistry.     

Mutual storage mode ion/ion reactions in a hybrid linear ion trap

Ion/ion proton transfer reactions involving mutual storage of both ion polarities in a linear ion trap (LIT) that comprises part of a hybrid triple quadrupole/linear ion trap mass spectrometer have been effected. Mutual ion storage in the x- and y-dimensions arises from the normal operation of the oscillating quadrupole field of the quadrupole array, while storage in the z-dimension is enabled by applying unbalanced radio-frequency amplitudes to opposing sets of rods of the array. Efficient trapping (>90%) is achieved for thermalized ions over periods of several seconds. Reactions were demonstrated for multiply charged protein/peptide cations formed by electrospray with anions derived from glow discharge ionization of perfluoro(methyldecalin) (PMD) introduced from the side of the LIT rod array. Doubly and singly charged protein ions are readily formed via ion/ion reactions. The parameters that affect ion/ion reactions are discussed, including the degree of RF unbalance on the LIT rods, vacuum pressure, nature of the buffer gas, reaction time, anion abundance, and the low mass cutoff for ion/ion reaction. The present system has a demonstrated upper mass-to-charge ratio limit of at least 33,000. The system also has high flexibility with respect to defining MS(n) experiments involving both collision-induced dissociation (CID) and ion/ion reactions. Experiments are demonstrated involving beam-type CID in the pressurized collision quadrupole (Q2) followed by ion/ion reactions involving the product ions in the LIT. Ion parking experiments are also demonstrated using the mutual storage ion/ion reaction mode in the LIT, with a parking efficiency over 60%.     

Transmission mode ion/ion reactions in the radiofrequency‐only ion guide of hybrid tandem mass spectrometers

Transmission mode ion/ion reactions have been performed within the first quadrupole, the Q0 radiofrequency (RF)‐only quadrupole, of two types of hybrid tandem mass spectrometers (viz., triple quadrupole/linear ion trap and QqTOF instruments). These transmission mode reactions involved the storage of either the reagent species and the transmission of the analyte species through the Q0 quadrupole for charge inversion reactions or the storage of the analyte ions and transmission of the reagent ions as in charge reduction experiments. A key advantage to the use of transmission mode ion/ion reactions is that they do not require any instrument hardware modifications to provide interactions of oppositely charged ions and can be implemented in any instrument that contains a quadrupole or linear ion trap. The focus of this work was to investigate the potential of using the RF‐only quadrupole ion guide positioned prior to the first mass‐resolving element in a tandem mass spectrometer for ion/ion reactions. Two types of exemplary experiments have been demonstrated. One involved a charge inversion reaction and the other involved a charge reduction reaction in conjunction with ion parking. Ion/ion reactions proved to be readily implemented in Q0 thereby adding significantly greater experimental flexibility in the use of ion/ion reaction experiments with hybrid tandem mass spectrometers. Copyright © 2009 John Wiley & Sons, Ltd.     

The 3D quadrupole ion trap mass spectrometer as a complete chemical laboratory for fundamental gas-phase studies of metal mediated chemistry

Electrospray ionization provides a "treasure trove" of metal containing ions whose fundamental reactivity can be studied via collision induced dissociation and ion-molecule reactions using the multistage mass spectrometry capabilities of the quadrupole ion trap mass spectrometer. Examples of metal mediated chemistry relevant to catalysis, C-C bond coupling, bioinorganic and supramolecular chemistry are highlighted.     

Transmission mode ion/ion proton transfer reactions in a linear ion trap

A new method is described for effecting ion/ion proton transfer reactions that involves storage of analyte ions while oppositely charged ions are transmitted through the stored ion population. In this approach, the products are captured and stored in the linear ion trap for subsequent mass analysis. Charge reduction of multiply charged protein ions is used as an example to illustrate the analytical usefulness of this method. In another variation of the transmission mode ion/ion reaction approach, two charge inversion experiments, implemented by passing analyte ions through a population of multiply charged reagent ions in a LIT, are also demonstrated. A pulsed dual ion source approach coupled with a hybrid triple quadrupole/linear ion trap instrument was used to demonstrate these two methods. The results for ion/ion reactions implemented using these so-called "transmission mode" experiments were comparable to those acquired using the more conventional mutual storage mode, both in terms of efficiency and information content of the spectra. An advantage of transmission mode experiments compared with mutual storage mode experiments is that they do not require any specialized measures to be taken to enable the simultaneous storage of oppositely charged ions.     

Manipulation of charge states of biopolymer ions by atmospheric pressure ion/molecule reactions implemented in an extractive electrospray ionization source

A home-made extractive electrospray ionization source is coupled to an linear quadrupole ion trap mass spectrometer to investigate ion/molecule reactions of biopolymers at ambient pressure. Multiply charged biopolymers such as peptides and proteins generated in an electrospray are easily reduced to a low charge state by the atmospheric pressure ion/molecule reactions occurring between the multiply charged ions and a strong basic reagent sprayed in neutral form into the electrospray plume. The charge state of the biopolymer ions can be manipulated by controlling the amount of the basic reagent. The production of biopolymer ions with low charge states results in a substantial improvement of sensitivity and reduced spectral congestion in ESI-MS. This is of importance for biopolymer mixture analysis and could have promising applications in proteomics.     

Trapping of intact,singly-charged,bovine serum albumin ions injected from the atmosphere with a 10-cm diameter,frequency-adjusted linear quadrupole ion trap

High-resolution real-time particle mass measurements have not been achievable because the enormous amount of kinetic energy imparted to the particles upon expansion into vacuum competes with and overwhelms the forces applied to the charged particles within the mass spectrometer. It is possible to reduce the kinetic energy of a collimated particulate ion beam through collisions with a buffer gas while radially constraining their motion using a quadrupole guide or trap over a limited mass range. Controlling the pressure drop of the final expansion into a quadrupole trap permits a much broader mass range at the cost of sacrificing collimation. To achieve high-resolution mass analysis of massive particulate ions, an efficient trap with a large tolerance for radial divergence of the injected ions was developed that permits trapping a large range of ions for on-demand injection into an awaiting mass analyzer. The design specifications required that frequency of the trapping potential be adjustable to cover a large mass range and the trap radius be increased to increase the tolerance to divergent ion injection. The large-radius linear quadrupole ion trap was demonstrated by trapping singly-charged bovine serum albumin ions for on-demand injection into a mass analyzer. Additionally, this work demonstrates the ability to measure an electrophoretic mobility cross section (or ion mobility) of singly-charged intact proteins in the low-pressure regime. This work represents a large step toward the goal of high-resolution analysis of intact proteins, RNA, DNA, and viruses.     

Sub parts-per-million mass measurement accuracy of intact proteins and product ions achieved using a dual electrospray ionization quadrupole fourier transform ion cyclotron resonance mass spectrometer

High mass measurement accuracy (MMA) is demonstrated for intact proteins and subsequent collision-induced dissociation product ions using internal calibration. Internal calibration was accomplished using a dual electrospray ionization source coupled with a hybrid quadrupole Fourier transform ion cyclotron resonance (Q-FT-ICR) mass spectrometer. Initially, analyte ions generated via the first electrospray (ESI) emitter are isolated and dissociated in the external quadrupole. This event is followed by a simultaneous switch to the calibrant ion ESI emitter and a disablement of the isolation and activation of the external quadrupole such that a broad m/z range of calibrant ions are accumulated before injecting the analyte/calibrant ion mixture into the ICR cell. Two different internal calibrant solutions were utilized in these studies to evaluate this approach for the top-down characterization of melittin and ubiquitin. While external calibration of protein fragments resulted in absolute MMA greater than 16 ppm, internal standardization significantly improved upon the MMA of both the intact proteins and their products ions which ranged from -2.0 ppm to 1.1 ppm, with an average of -0.9 ppm. This method requires limited modification to ESI-FT-ICR mass spectrometers and is applicable for both positive and negative ionization modes.     

Controlled ion fragmentation in a 2-D quadrupole ion trap for external ion accumulation in ESI FTICR mass spectrometry

Undesired fragmentation of electrospray generated ions in an rf multipole traps can be problematic in many applications. Of special interest here is ion dissociation in a 2-D quadrupole ion trap external to a Fourier transform ion cyclotron resonance mass spectrometer (FTICR MS) used in proteomic studies. In this work, we identified the experimental parameters that determine the efficiency of ion fragmentation. We have found that under the pressure conditions used in this study there is a specific combination of the radial and axial potential well depths that determines the fragmentation threshold. This combination of rf and dc fields appears to be universal for ions of different mass-to-charge ratios, molecular weights, and charge states. Such universality allows the fragmentation efficiency of the trapped ions in the course of capillary liquid chromatography (LC) separation studied to be controlled and can increase the useful duty cycle and dynamic range of a FTICR mass spectrometer equipped with an external rf only 2-D quadrupole ion trap.     

Electrospray ionization mass spectrometry and tandem mass spectrometry of clodronate and related bisphosphonate and phosphonate compounds

The bisphosphonate family with a P-C-P structure is a broad class of drugs, widely investigated as potential inhibitors in bone diseases and calcium metabolic disorders. In this study, the mass spectrometric (MS) behavior and fragmentation of clodronate and related bisphosphonate and phosphonate compounds was studied by using negative ion electrospray ionization (ESI) with triple quadrupole and ion trap instruments. The effect of pH on the degree of deprotonation of the polyprotic bisphosphonic and phosphonic acids in negative ion ESI-MS was investigated, and the degree of deprotonation in the ESI mass spectra and the dissociation in the liquid phase were compared. The results provide evidence that the measured ESI mass spectra do not correlate with the chemistry in the liquid phase owing to the decrease in the pH of the solvent droplets during the ion evaporation process and the charge state neutralization in the gas phase. Ion trap MS(n) provided useful information on the fragmentation study of clodronate and related bisphosphonate and phosphonate compounds, in which interesting fragmentation pathways including the direct elimination of carbon monoxide from deprotonated bisphosphonates and formation of a P-P bond were observed. Reactions between the product ions with a -PO(2) group and residual water in the ion trap or in the high-pressure region of the triple quadrupole instrument formed other unexpected fragmentation paths for all the bisphosphonates studied.     

Sources of artefacts in the electrospray ionization mass spectra of saturated diacylglycerophosphocholines: From condensed phase hydrolysis reactions through to gas phase intercluster reactions

The mass spectra of diacylglycerophosphocholine phospholipids comprised of saturated fatty acids (1,2-dipentanoyl-sn-glycero-3-phosphocholine (D5PC); 1,2-dihexanoyl-sn-glycero-3-phosphocholine (D6PC), and 1,2-dimyristoyl-sn-glycero-3-phosphocholine (D14PC)) are sensitive to the electrospray ionization (ESI) conditions. When fresh solutions of phospholipid in 10 mM ammonium acetate are subjected to ESI, protonated oligomeric clusters, [DxPCn + H]+ (x = 5, 6, and 14) are observed in the following different types of mass spectrometers: 3D-quadrupole ion trap; linear ion trap, and triple quadrupole. The formation of the protonated cluster ions is not unique to the ion trap instruments, although they tend to be more abundant in these instruments. As the ESI solutions age, new ions are observed, which correspond to acid-catalyzed solution phase deacylation reactions. The collision induced dissociation fragmentation reactions of the oligomer cluster ions exhibit a distinct dependence on the cluster size, with the larger clusters (n > 2) simply fragmenting via the loss of lipid monomers. In contrast, the fragmentation of the dimeric cluster ion is unique, resulting in a number of additional reactions including covalent bond formation via intermolecular cluster SN2 reactions and SN2 transfer of a methyl group. The nature of the charge has a significant role in the formation of products via these intermolecular cluster reactions. Changing the head group to phosphoethanolamine "switches off" the SN2 reactions, while changing the cation from a proton to either a sodium or a potassium ion, diminishes the intermolecular reactions relative to monomer loss. Semi empirical PM3 calculations on [D6PC2 + H]+ suggest that the SN2 reactions are thermodynamically favored over simple monomer loss. These results have important implications in the field of lipidomics.