Improved isotopic abundance measurements for high resolution fourier transform ion cyclotron resonance mass spectra via time-domain data extraction

The simultaneous high resolution and accurate mass measurements possible with Fourier transform ion cyclotron resonance mass spectrometry coupled with the gentle ionization of electrospray hold attractions for protein, peptide, and oligonucleotide characterization, including multistage-mass spectrometry measurements for assignment of fragment masses and greater confidence in structural measurements. The detection of cyclotron motion over extended periods of time (in some cases for several minutes) allows higher resolution and mass accuracy. Generally, signal duration has been considered to be limited primarily by background pressure, with ion-neutral collisions leading to the reduction and dephasing of cyclotron motion, causing signal loss. However, recent theoretical work has shown that the ion cloud stability that is a prerequisite for high performance measurements is highly dependent on the electric field generated by the ion cloud, thus giving rise to a minimum number of charges or ions required for extended time-domain signals. The effects of ion population on ion cloud stability and signal duration, and the subsequent effects on resolution and measured isotopic abundances are reported. Individual time-domain signals for bovine insulin isotopic peaks were extracted to allow a comparison of the damping rates for each of these ion clouds and the measured time-domain amplitude maxima are shown to provide a better match with the theoretically predicted isotopic abundances for insulin. These results show that different damping rates of ions of very similar mass, but different ion cloud population sizes, can have dramatic effects on the observed isotopic patterns. Additionally, more accurate, high resolution spectra can be produced by correcting for the effects of the different damping rates that are observed for different ion population sizes.     

Sensitivity and amplitude calibration of a Fourier transform 3D quadrupole ion trap mass spectrometer

With a three-dimensional (3D) quadrupole ion trap running in a Fourier transform operating mode, the detected signal is an image of the collective motion of the confined ions. Consequently, it is assumed that the image signal is the sum of the axial trajectories of the simultaneously confined ions. The resulting frequency spectrum after Fourier transformation comprises frequency peaks at the axial secular frequencies of the confined species according to their mass/charge ratio. With a singly confined species, the maximal amplitude of the image signal is proportional to the amplitude of the secular axial frequency peak. The matrix method is employed to express the axial trajectory sampled at the confinement field period. In that case, the expression of the image signal, as well as its maximal amplitude, is calculated as a function of the trap operating conditions and initial axial positions and axial velocities of the ions. The initial position and velocity distributions are connected to the injection mode. With the steady ion flow injection mode (SIFIM) and an initial phase of the confinement field equal to kπ, the maximal amplitude of the image signal is proportional to either the sum of the initial axial positions or the number of confined ions and the mean value of the initial axial positions. By simulation, amplitude fluctuation of the frequency peak is then calculated for a number of ions ranging between some tens to some thousands of ions injected by SIFIM. The peak amplitude fluctuations induced by the fluctuations of the number of ions are seven times greater than those induced by the fluctuations of the distribution of the initial axial positions.     

A tuning method for electrically compensated ion cyclotron resonance mass spectrometer traps

We describe a method for tuning electrically compensated ion cyclotron resonance (ICR) traps by tracking the observed cyclotron frequency of an ion cloud at different oscillation mode amplitudes. Although we have used this method to tune the compensation voltages of a custom-built electrically compensated trap, the approach is applicable to other designs that incorporate electrical compensation. To evaluate the effectiveness of tuning, we examined the frequency shift as a function of cyclotron orbit size at different z-mode oscillation amplitudes. The cyclotron frequencies varied initially by ∼12 ppm for ions with low z-mode oscillation amplitudes compared with those with high z-mode amplitudes. This frequency difference decreased to ∼1 ppm by one iteration of trap tuning.     

Improving IRMPD in a quadrupole ion trap

A focused laser is used to make infrared multiphoton photodissociation (IRMPD) more efficient in a quadrupole ion trap mass spectrometer. Efficient (up to 100%) dissociation at the standard operating pressure of 1 × 10⁻³ Torr can be achieved without any supplemental ion activation and with shorter irradiation times. The axial amplitudes of trapped ion clouds are measured using laser tomography. Laser flux on the ion cloud is increased six times by focusing the laser so that the beam waist approximates the ion cloud size. Unmodified peptide ions from 200 Da to 3 kDa are completely dissociated in 2.5–10 ms at a bath gas pressure of 3.3 × 10⁻⁴ Torr and in 3–25 ms at 1.0 × 10⁻³ Torr. Sequential dissociation of product ions is increased by focusing the laser and by operating at an increased bath gas pressure to minimize the size of the ion cloud.     

Signal suppression in electrospray ionization Fourier transform mass spectrometry of multi-component samples

The high resolution, mass range and sensitivity of Fourier transform mass spectrometry (FTMS) suggest that it could be a valuable tool for the quantitative analysis of biomolecules. To determine the applicability of electrospray ionization combined with FTMS to the quantitation of biomolecules in multi-component samples, mixtures of varying compositions and concentrations of cytochrome c, angiotensin II, insulin and chicken egg white lysozyme were examined. The instrument used has an electrospray source with a hexapole trap to accumulate ions for injection into an ion cyclotron resonance mass analyzer. Linear responses for single component samples of angiotensin II and insulin were in the range 0.031-3 microM and those of both cytochrome c and lysozyme were between 0.031 and 1 microM. In examining various mixtures of the proteins with angiotensin II, it was found that the presence of the large molecules suppresses the signal of the smaller molecules. This is suggested to be a result of ion-ion interactions producing selective ion loss from either the hexapole trap or the ion cyclotron resonance mass analyzer trap. More massive, more highly charged ions can collisionally transfer large amounts of translational energy to smaller, less highly charged ions, ejecting the smaller ions from the trap. Mass discrimination effects resulting from the trapping voltage were also examined. It was found that relative signal intensities of ions of different masses depend on trapping voltage for externally produced ions. The effect is most significant for spectra including masses that differ by 30% or more. This suggests that for quantitation all samples and standards be run at a constant trapping potential.     

Surface-induced dissociation of molecular ions in a quadrupole ion trap mass spectrometer

A method is reported by which surface-induced dissociation is used to activate ions stored in a quadrupole ion trap mass spectrometer. The method employs a short (< 5 μs), fast-rising (< 20-ns rise time), high voltage direct current (dc) pulse, which is applied to the endcaps of a standard Paul-type quadrupole ion trap. This is in contrast to the application of an alternating current (ac) signal normally used to resonantly excite and dissociate ions in the trap. The effect of the dc pulse is to cause the ions rapidly to become unstable in the radial direction and subsequently to collide with the ring electrode. Sufficient internal energy is acquired in this collision to cause high energy fragmentations of relatively intractable molecular ions such as pyrene and benzene. The dissociations of limonene are used to demonstrate that high energy demand processes increase in relative importance in the dc pulse experiment compared with the usual resonance excitation method used to cause activation. The fragments are scanned out of the ion trap using the conventional mass-selective instability scan mode. Simulations of ion motion in the trap provide evidence that surface collisions occur at kinetic energies in the range of tens to several hundred electronvolts. The experiments also demonstrate that production of fragment ions is sensitive to the phase of the main radiofrequency drive voltage at the point when the dc is initiated.     

Study of the correlated motion of ion mixtures in an rf ion trap

We observed variations of the signal position versus the filament temperature, and thus the relative amounts of Ba⁺ and BaO⁺ ions stored simultaneously in an rf trap. The tendency of variations is explained by a generalized model of two coupled ion clouds, in which the coupling condition and the frequencies of correlated motions depend on the ion masses, the force constants of the applied trapping field and the Coulomb interaction between the ions as well as the relative ion numbers of the two ion clouds.     

Selective ion isolation/rejection over a broad mass range in the quadrupole ion trap

Techniques are presented for mass-selective ion manipulation over a wide mass range in a three-dimensional quadrupole. The methods use an auxiliary, low-amplitude radio-frequency signal applied to the endcap electrodes. This signal is either held at a single frequency as the fundamental radio-frequency trapping amplitude is ramped or swept over a frequency range while the fundamental radio-frequency trapping amplitude is held at a fixed level. Ion isolation and ejection are demonstrated for ions formed within the ion trap using electron ionization and for ions injected into the ion trap formed either by an air-sustained glow discharge or by electrospray. Mass-selective ion ejection is used to reduce matrix-ion-induced space charge during ion injection, thereby producing signal enhancement for the detection of 2, 4, 6-trinitrotoluene in air. Mass-selective isolation of ions with mass-to-charge ratios above the normal operating range (m / z 650) for the ion trap is also demonstrated after injection of myoglobin ions formed via electrospray.     

Impact of ion cloud densities on the measurement of relative ion abundances in Fourier transform ion cyclotron resonance mass spectrometry: experimental observations of coulombically induced cyclotron radius perturbations and ion cloud dephasing rates

Fundamental research into the quantitative properties of Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) has yielded interesting observations, especially in terms of factors affecting the accuracy of relative ion abundances. However, most of the previous discussions have focused on theoretical systems, or systems of limited scope. In this paper, we document ion motion attributes of a 30 spectra (six samples, five replicates each) system previously established as linear over two orders of magnitude. Observed behaviors include the perturbation of one charged species (cyclosporin A, CsA) of low ion density to a cyclotron orbit of greater radius than that of an almost identical, but slightly mass-separated species (CsG) with a higher ion density. This radial perturbation is attributed to the coulombic repulsion between the two ion clouds as they interact during the excitation process, as previously proposed by Uechi and Dunbar. Magnitudes of the perturbation were confirmed by making cyclotron radii determinations utilizing the ratio of the third-to-first harmonics for the charged species of interest. It was found that these radial differences can account for as much as a 55% signal bias in favor of CsA for a single sample and a >20% positive bias in the slope of the regressed data set. A second behavior noted that also contributes to the potential inaccuracy of relative ion abundance measurements is the difference in signal decay rates for CsA and CsG. Damping constants and initial time domain signal amplitudes were evaluated using segmented Fourier transforms. Discrepancies in decay rates were not expected from two species that have essentially identical collisional cross-sections. However, it has been observed that the faster decay rates are observed by the species of lower ion cloud density. We have attributed this differential signal decay phenomenon to the rates of loss of phase coherence for the two ion clouds. Previously, others have reported that less dense ion clouds are more susceptible to shearing and other disruptive forces during the course of their excited cyclotron motion. Our experimental evidence supports that it is the loss of cloud coherence that accounts for the signal loss over time, with the less dense cloud de-phasing more quickly. As the ion populations of the two investigated species near equivalence, so do their time constants.     

Realistic modeling of ion cloud motion in a Fourier transform ion cyclotron resonance cell by use of a particle-in-cell approach

Using a 'Particle-In-Cell' approach taken from plasma physics we have developed a new three-dimensional (3D) parallel computer code that today yields the highest possible accuracy of ion trajectory calculations in electromagnetic fields. This approach incorporates coulombic ion-ion and ion-image charge interactions into the calculation. The accuracy is achieved through the implementation of an improved algorithm (the so-called Boris algorithm) that mathematically eliminates cyclotron motion in a magnetic field from digital equations for ion motion dynamics. It facilitates the calculation of the cyclotron motion without numerical errors. At every time-step in the simulation the electric potential inside the cell is calculated by direct solution of Poisson's equation. Calculations are performed on a computational grid with up to 128 x 128 x 128 nodes using a fast Fourier transform algorithm. The ion populations in these simulations ranged from 1000 up to 1,000,000 ions. A maximum of 3,000,000 time-steps were employed in the ion trajectory calculations. This corresponds to an experimental detection time-scale of seconds. In addition to the ion trajectories integral time-domain signals and mass spectra were calculated. The phenomena observed include phase locking of particular m/z ions (high-resolution regime) inside larger ion clouds. A focus was placed on behavior of a cloud of ions of a single m/z value to understand the nature of Fourier transform ion cyclotron resonance (FTICR) resolution and mass accuracy in selected ion mode detection. The behavior of two and three ion clouds of different but close m/z was investigated as well. Peak coalescence effects were observed in both cases. Very complicated ion cloud dynamics in the case of three ion clouds was demonstrated. It was found that magnetic field does not influence phase locking for a cloud of ions of a single m/z. The ion cloud evolution time-scale is inversely proportional to magnetic field. The number of ions needed for peak coalescence depends quadratically on the magnetic field.     

Theory of peak coalescence in Fourier transform ion cyclotron resonance mass spectrometry

Peak coalescence, i.e. the merging of two close peaks in a Fourier transform ion cyclotron resonance (FTICR) mass spectrum at a high number of ions, plays an important role in various FTICR experiments. In order to describe the coalescence phenomenon we would like to propose a new theory of motion for ion clouds with close mass‐to‐charge ratios, driven by a uniform magnetic field and Coulomb interactions between the clouds. We describe the motion of the ion clouds in terms of their averaged drift motion in crossed magnetic and electric fields. The ion clouds are considered to be of constant size and their motion is studied in two dimensions. The theory deals with the first‐order approximation of the equations of motion in relation to dm/m, where dm is the mass difference and m is the mass of a single ion. The analysis was done for an arbitrary inter‐cloud interaction potential, which makes it possible to analyze finite‐size ion clouds of any shape. The final analytical expression for the condition of the onset of coalescence is found for the case of uniformly charged spheres. An algorithm for finding this condition for an arbitrary interaction potential is proposed. The critical number of ions for the peak coalescence to take place is shown to depend quadratically on the magnetic field strength and to be proportional to the cyclotron radius and inversely proportional to the ion masses. Copyright © 2009 John Wiley & Sons, Ltd.     

Laser desorption/ionization coupled to tandem mass spectrometry for real-time monitoring of paraquat on the surface of environmental particles

Aerosol mass spectrometry with laser desorption/ionization was investigated as a possible tool for real-time monitoring of the presence of the pesticide paraquat on the surface of airborne soil particles. Laser desorption/ionization of paraquat dication produced only singly charged ions. The most abundant species were [M](+.), [M - H](+), and [M - CH3](+). Operation of the ion trap mass spectrometer in the MS(3) mode allowed the reduction of the signal dependence on laser fluence fluctuations and permitted the detection of the analyte with good sensitivity and high selectivity. The estimated limit of detection in terms of surface coverage was 0.016 monolayers, approximately 1 attomole of paraquat on the surface of a single micron-sized soil particle.     

A two-dimensional quadrupole ion trap mass spectrometer

The use of a linear or two-dimensional (2-D) quadrupole ion trap as a high performance mass spectrometer is demonstrated. Mass analysis is performed by ejecting ions out a slot in one of the rods using the mass selective instability mode of operation. Resonance ejection and excitation are utilized to enhance mass analysis and to allow isolation and activation of ions for MS(n) capability. Improved trapping efficiency and increased ion capacity are observed relative to a three-dimensional (3-D) ion trap with similar mass range. Mass resolution comparable to 3-D traps is readily achieved, including high resolution at slower scan rates, although adequate mechanical tolerance of the trap structure is a requirement. Additional advantages of 2-D over 3-D ion traps are also discussed and demonstrated.     

Characterization of a serial array of miniature cylindrical ion trap mass analyzers

Two small (5 mm internal radius) cylindrical ion traps (CITs) are arranged in series and operated using a single ion source, detector and radio frequency (rf) trapping signal. Ions are trapped in the first CIT and later transferred to the second by applying a direct current (dc) pulse to the endcap electrode of the first trap. This process is facilitated if a second, appropriately timed, retarding dc pulse is applied to the exit endcap electrode of the second trap. Mesh endcaps are used for the CITs to increase the number of ionizing electrons entering the trap and to maximize the transfer efficiency and detected signal. The transfer efficiency is dependent on the amplitude of the dc potential applied to eject the ions from the first trap, the amplitude of the dc potential applied to retain the ions in the second trap, and the period during which the retarding potential is applied. The amplitude and phase of the rf also affect the transfer process. Ions that readily dissociate upon collision have low transfer efficiencies; more stable ions can be transferred with up to 50% efficiency. Copyright 1999 John Wiley & Sons, Ltd.     

Combined chemical and topographic imaging at atmospheric pressure via microprobe laser desorption/ionization mass spectrometry–atomic force microscopy

The operational characteristics and imaging performance are described for a new instrument comprising an atomic force microscope coupled with a pulsed laser and a linear ion trap mass spectrometer. The operating mode of the atomic force microscope is used to produce topographic surface images having sub‐micrometer spatial and height resolution. Spatially resolved mass spectra of ions, produced from the same surface via microprobe‐mode laser desorption/ionization at atmospheric pressure, are also used to create a 100 × 100 µm chemical image. The effective spatial resolution of the image (～2 µm) was constrained by the limit of detection (estimated to be 10⁹–10¹⁰ molecules) rather than by the diameter of the focused laser spot or the step size of the sample stage. The instrument has the potential to be particularly useful for surface analysis scenarios in which chemical analysis of targeted topographic features is desired; consequently, it should have extensive application in a number of scientific areas. Because the number density of desorbed neutral species in laser desorption/ionization is known to be orders‐of‐magnitude greater than that of ions, it is expected that improvements in imaging performance can be realized by implementation of post‐ionization methods. Published in 2009 by John Wiley & Sons, Ltd.     

A secondary ion microprobe ion trap mass spectrometer

An ion trap mass analyzer has been attached to an organic secondary ion microprobe. A pressure differential >100 can be maintained between the ion trap and microprobe. The well-focused secondary ion beam can transit a small (2 mm) diameter tube, but gas flow from ion trap to microprobe is impeded. This pressure differential allows the microprobe to retain imaging capability. Ion trap and microprobe data systems are integrated by taking advantage of the highly reproducible periodicity of the ion trap operating in resonant ejection mode and asynchronous signal and data acquisition afforded by commercially available interface cards. Secondary ion mass spectra and images obtained indicate an approximately 10-fold improvement in sensitivity, although preliminary evidence indicates low (<1%) trapping efficiency. Image data acquisition using the ion trap for mass analysis requires at least 10 times as much time compared to using a quadrupole mass filter because the mass-selected instability mode is used for mass analysis, i.e., mass resolution in the ion trap is not continuous as it is in the quadrupole.     

Expansion of a multicomponent laser plasma cloud

It is shown that the mechanism of the electrodynamic expansion of a plasma cloud satisfactorily describes ion acceleration for two-component plasmas (ions of a fixed mass and electrons) but cannot be applied to the expansion of multicomponent plasmas. The main drawback of the known models is the assumption of the collision-free expansion of laser plasma clouds. It cannot be used to simulate the expansion of a multicomponent plasma cloud. An algorithm was proposed that takes into account collisions of ions with different masses in the electrodynamic model for the expansion of plasma clouds containing both matrix and trace contaminant ions. The algorithm and software on its basis allow one to calculate the energies of ions of different masses acquired in the process of ion acceleration in the laser plasma containing both matrix and contaminant ions. The results of numerical calculations are compared both with similar results obtained for the collision-free model of the expansion of the plasma cloud and with the experimental data. Taking into account collision processes in the electrodynamic model of plasma cloud expansion allows one to calculate the acceleration of trace contaminant ions in multicomponent laser plasma and reach a good qualitative agreement with the experimental data.     

Investigation of apparent mass deviations in electrospray ionization tandem mass spectrometry of a benzophenone-labeled peptide

In a previous study utilizing benzophenone-based topological probes to study conformationally dependent changes in mouse muscle nicotinic acetylcholine receptor (nAChR) topology, electrospray ionization tandem mass spectrometric (ESI-MS/MS) analysis led to a consistent -2.0 Da mass deviation from expected values. In the present study a synthetic peptide, corresponding to nAChR alpha1 subunit residues 130-139, was photolabeled. MS/MS analysis of this peptide using an ion trap confirmed the previously observed mass deviation, associated only with fragment ions that contain the incorporated benzophenone moiety. Analysis of peak profiles for the photolabeled ions does not indicate the typical 'peak fronting' that produces a mass shift when labile ions are prematurely ejected from the ion trap. Rather, hydrogen/deuterium (H/D) exchange experiments support the hypothesis that a chemical rearrangement involving phenyl migration and ketone formation has formed an unexpected oxidized peptide, with molecular mass 2 Da less than that expected, that is isolated for collision-induced dissociation in the ion trap together with the predicted precursor due to the broad ion isolation window specified.     

A Mechanism for Poor High Mass Performance in Fourier Transform Mass Spectrometry

The discovery of a mass-dependent electrical mechanism for signal loss of high mass ions in Fourier transform mass spectrometry is reported. Theoretical calculations and experimental evidence show the existence of resonances involving the z-motion and radial motion of the ion. The resonances are intrinsic in the cubic trap. This results in energy transfer between modes, expansion of the ion cloud, and a corresponding loss of signal. Quadrupolar trapping potential wells are proposed as a solution.