Vladimir M. Doroshenko

Desorption/ionization of biomolecules from aqueous solutions at atmospheric pressure using an infrared laser at 3 μm

A new atmospheric pressure (AP) infrared (IR) matrix-assisted laser desorption/ionization (MALDI) ion source was developed and interfaced with a Thermo Finnigan LCQ ion trap mass spectrometer. The source utilized a miniature all-solid-state optical parametric oscillator (OPO)-based IR laser system tunable in the λ = 1.5–4 μm spectral range and a nitrogen ultraviolet (UV) laser (λ = 337 nm) for use in comparative studies. The system demonstrated comparable performance at 3 μm and 337 ran wavelengths if UV matrices were used. However, AP IR-MALDI using a 3 μm wavelength showed good performance with a much broader choice of matrices including glycerol and liquid water. AP IR-MALDI mass spectra of peptides in the mass range up to 2000 Da were obtained directly from aqueous solutions at atmospheric conditions for the first time. A potential use of the new AP IR-MALDI ion source includes direct MS analysis of biological cells and tissues in a normal atmospheric environment as well as on-line coupling of mass spectrometers with liquid separation techniques.

A quadrupole ion trap/time-of-flight mass spectrometer with a parabolic reflectron.

A matrix-assisted laser desorption/ionization-quadrupole ion trap/reflectron time-of-flight (MALDI-QIT/reTOF) mass spectrometer design and its operation in both normal and tandem mass spectrometric modes are described. A parabolic reflectron was found to be capable of providing mass resolution of 5000 for an initial ion energy distribution ranging over a 50% energy interval of the entire reflectron energy range. The sensitivity, ion isolation and fragmentation efficiency in the MALDI-QIT/reTOF instrument were close to those observed in the MALDI/QIT mass spectrometer. The mass resolution was shown to depend on the extraction field potentials, the r.f. trapping voltage amplitude and the phase of shutting down the r.f. voltage before extraction. At values of qs < 0.3-0.4 the mass resolution does not depend on the ion mass, is in a range of 1000-1400 and is governed by the extraction voltages and the ion temperature before extraction, the latter shown to be in the range 1180-1690 K. The variation of the mass resolution for ions at values of qs > 0.4 is irregular but normally it is lower than that for ions having lower qs values. Mass spectral line positions shifted when the trapping voltage before extraction was varied. The line shifts were larger for lower mass ions and were comparable to the line widths in the case of very low masses.

Injection of externally generated ions into an increasing trapping field of a quadrupole ion trap mass spectrometer

Trapping ions injected into a quadrupole ion trap (QIT) by increasing the trapping r.f. voltage on a ring electrode is an effective and widely recognized method of interfacing an ion trap with pulsed ion sources such as matrix-assisted laser desorption/ionization (MALDI). In this paper, the problem of mass discrimination during the injection and trapping of ions by the increasing r.f. field was studied both experimentally and by numerical simulation using SIMION software. For a MALDI/QIT interface design with a remote external ion source described here, experiments with polyethylene glycol (PEG 1000 and PEG 1500) showed little mass discrimination for trapping ions in a wide mass range (500-2000 Dn) for a broad range of experimental conditions, which include kinetic energies of 5-40 eV for the injected ions and an r.f. voltage of 400-4000 Vo-p amplitude ramped at a rate of 30-140 Vo-p mus-1. In the numerical simulation, complex and sharp dependences of the trapping efficiency on the phase of the r.f. voltage and initial kinetic energy of ions were observed. However, after averaging over the r.f. phase and over a reasonable range of kinetic energy, the simulation resulted in relatively constant and high values for the trapping efficiency (normally 0.2-0.3) for any mass and kinetic energy considered, which are consistent with the weak sensitivity to injection parameters observed in the experiment. A simple model for the qualitative description of ion injection and trapping is suggested that relies on phase interaction of injected ions with the r.f. field rather than on collisions with the buffer gas molecules to decrease the ion kinetic energy.

Gas phase hydrogen/deuterium exchange reactions of peptide ions in a quadrupole ion trap mass spectrometer

Hydrogen/deuterium exchange reactions of protonated and sodium cationized peptide molecules have been studied in the gas phase with a MALDI/quadrupole ion trap mass spectrometer. Unit‐mass selected precursor ions were allowed to react with deuterated ammonia introduced into the trap cell by a pulsed valve. The reactant gas pressure, reaction time, and degree of the internal excitation of reactant ions were varied to explore the kinetics of the gas phase isotope exchange. Protonated peptide molecules exhibited a high degree of reactivity, some showing complete exchange of all labile hydrogen atoms. On the contrary, peptide molecules cationized with sodium exhibited only very limited reactivity, indicating a vast difference between the gas phase structures of the two ions.

Ideal velocity focusing in a reflectron time-of-flight mass spectrometer

A single-stage ion mirror in a time-of-flight (TOF) mass spectrometer (MS) can perform first order velocity focusing of ions initially located at a start focal plane while second order velocity focusing can be achieved using a double-stage reflectron. The situation is quite different when an ion source extraction field is taken into account. In this case which is common in any practical matrix-assisted laser desorption/ionization (MALDI) TOF-MS a single-stage reflectron, for example, cannot perform velocity focusing at all. In this paper an exact, analytic solution for an electric field inside a one-dimensional reflectron has been found to achieve universal temporal focusing of ions having an initial velocity distribution. The general solution is valid for arbitrary electric field distributions in the upstream (from the ion source to the reflectron) and downstream (from the reflectron to an ion detector) regions and in a decelerating part of the reflectron of a reflectron TOF mass spectrometer. The results obtained are especially useful for designing MALDI reflectron TOF mass spectrometers in which the initial velocity distribution of MALDI ions is the major limiting factor for achieving high mass resolution. Using analytical expressions obtained for an arbitrary case, convenient working formulas are derived for the case of a reflectron TOF-MS with a dual-stage extraction ion source. The special case of a MALDI reflectron TOF-MS with an ion source having a low acceleration voltage (or large extraction region) is considered. The formulas derived correct the effect of the acceleration regions in a MALDI ion source and after the reflectron before detecting ions.

Matrix-assisted laser desorption/ionization time-of-flight analysis of Bacillus spores using a 2.94 µm infrared laser

The performance of infrared (2.94 microm) and ultraviolet (337 nm) lasers were compared for analysis of purified spores of B. subtilis, B. cereus and B. globigii on a four-inch end-cap reflectron time-of-flight instrument. Infrared matrix-assisted laser desorption/ionization (IR-MALDI) mass spectra of these microorganisms displayed a larger number of biomarker peaks above m/z 4000, compared with UV-MALDI. Biomarker peaks were observed at higher m/z values with the IR laser.

A comparative study on the analytical utility of atmospheric and low-pressure MALDI sources for the mass spectrometric characterization of peptides

A comparative MS study was conducted on the analytical performance of two matrix-assisted laser desorption/ionization (MALDI) sources that operated at either low pressure (∼1Torr) or at atmospheric pressure. In both cases, the MALDI sources were attached to a linear ion trap mass spectrometer equipped with a two-stage ion funnel. The obtained results indicate that the limits of detection, in the analysis of identical peptide samples, were much lower with the source that was operated slightly below the 1-Torr pressure. In the low-pressure (LP) MALDI source, ion signals were observed at a laser fluence that was considerably lower than the one determining the appearance of ion signals in the atmospheric pressure (AP) MALDI source. When the near-threshold laser fluences were used to record MALDI MS spectra at 1-Torr and 750-Torr pressures, the level of chemical noise at the 1-Torr pressure was much lower compared to that at AP. The dependency of the analyte ion signals on the accelerating field which dragged the ions from the MALDI plate to the MS analyzer are presented for the LP and AP MALDI sources. The study indicates that the laser fluence, background gas pressure, and field accelerating the ions away from a MALDI plate were the main parameters which determined the ion yield, signal-to-noise (S/N) ratios, the fragmentation of the analyte ions, and adduct formation in the LP and AP MALDI MS methods. The presented results can be helpful for a deeper insight into the mechanisms responsible for the ion formation in MALDI.

Peptide fragmentation induced by radicals at atmospheric pressure

A novel ion dissociation technique, which is capable of providing an efficient fragmentation of peptides at essential atmospheric pressure conditions, is developed. The fragmentation patterns observed often contain c-type fragments that are specific to electron capture dissociation/electron transfer dissociation (ECD/ETD), along with the y-/b-type fragments that are specific to collision-activated dissociation (CAD). In the presented experimental setup, ion fragmentation takes place within a flow reactor located in the atmospheric pressure region between the ion source and the mass spectrometer. According to a proposed mechanism, the fragmentation results from the interaction of ESI-generated analyte ions with the gas-phase radical species produced by a corona discharge source.

Losses of ions during forward and reverse scans in a quadrupole ion trap mass spectrometer and how to reduce them

The higher order fields present in the quadrupole ion trap may have beneficial effects such as increases in mass resolution in the mass-selective instability or resonance ejection modes of operation, but may also result in losses of ions due to nonlinear resonances. In this work, the reduction in ion intensities observed in the mass spectra of polyethylene glycol (PEG 1000) has been utilized to monitor the ion losses resulting from these higher order fields during the rf voltage scans in both the forward and reverse directions. Extensive ion losses were observed in reverse rf voltage scans at qz=0.64 (az=0), which corresponds to octopole resonance at βz=1/2. The losses depended upon rf voltage scan rate and ion mass being greater for lower scan rates and lower masses. For ions of m/z 877, losses of up to 60% of the stored ions were observed at low scan rates (<1×104 Da/s), but were minimal at higher scan rates. Thus, it is possible to avoid such losses during reverse scans by scanning the region qz=0.64 at rates in excess of 4×104 Da/s. In forward rf voltage scans, ion storage was considerably more reliable, with significant losses observed only at very high scan rates near the region qz=0.78 (hexapole resonance at βz=2/3).

AP-MALDI Mass Spectrometry Imaging of Gangliosides Using 2,6-Dihydroxyacetophenone

AbstractMatrix-assisted laser/desorption ionization (MALDI) mass spectrometry imaging (MSI) is widely used as a unique tool to record the distribution of a large range of biomolecules in tissues. 2,6-Dihydroxyacetophenone (DHA) matrix has been shown to provide efficient ionization of lipids, especially gangliosides. The major drawback for DHA as it applies to MS imaging is that it sublimes under vacuum (low pressure) at the extended time necessary to complete both high spatial and mass resolution MSI studies of whole organs. To overcome the problem of sublimation, we used an atmospheric pressure (AP)-MALDI source to obtain high spatial resolution images of lipids in the brain using a high mass resolution mass spectrometer. Additionally, the advantages of atmospheric pressure and DHA for imaging gangliosides are highlighted. The imaging of [M–H]− and [M–H2O–H]− mass peaks for GD1 gangliosides showed different distribution, most likely reflecting the different spatial distribution of GD1a and GD1b species in the brain. Graphical Abstractᅟ