Planar multipole ion trap/time-of-flight mass spectrometer

We present a novel, hybrid ion trap/time-of-flight mass spectrometer that is based on a planar multipole design. Compared with Paul trap/time-of-flight instruments, this design possesses the principal advantages of higher injection efficiency and more homogeneous extraction fields. We demonstrate the viability of the concept and describe the characterization of a first prototype. Ions can be injected into the trap with little mass discrimination and stored for several minutes. A resolution of over 1300 is achieved in reflectron mode, and the influence of the RF amplitude and pressure on the resolution is analyzed. We suggest several applications in which this new instrument could offer advantages over existing technology.     

Use of a mixed-mode packing and voltage tuning for peptide mixture separation in pressurized capillary electrochromatography with an ion trap storage/reflectron time-of-flight mass spectrometer detector.

A mixed-mode (reversed-phase/anion-exchange) stationary phase has been used as the capillary column packing for investigation of the separation of peptide mixtures in pressurized capillary electrochromatography (pCEC). This stationary phase contains both octadecylsilanes and dialkylamines. The amine groups of the stationary phase determine the charge density on the surface of the packing and can produce a strong and constant electroosmotic flow (EOF) at low pH. A comparison was made in terms of the capability of separating tryptic digests between the mixed-mode phase and C18 reversed phase. In addition, the constant EOF enabled the tuning of the retention and the selectivity of the separation by adjusting the mobile phase pH from 2 to 5. Furthermore, the magnitude and the polarity of the electric voltage were demonstrated to greatly influence the elution profiles of the peptides in pCEC. An ion trap storage/reflectron time-of-flight mass spectrometer was used as an on-line detector in these experiments due to its ability to provide rapid and accurate mass detection of the sample components eluting from the separation column.     

Fourier transform time-of-flight mass spectrometry in an electrostatic ion beam trap

We report on the application of an electrostatic ion beam trap as a mass spectrometer. The instrument is analogous to an optical resonator; ions are trapped between focusing mirrors. The storage time is limited by the residual gas pressure and reaches up to several seconds, resulting in long ion flight paths. The oscillation of ion bunches between the mirrors is monitored by nondestructive image charge detection in a field-free region and mass spectra are obtained via Fourier transform. The principle of operation is demonstrated by measuring the mass spectrum of trapped Ar+ and Xe+ particles, produced by a standard electron impact ion source. Also, mass spectra of heavier PEGnNa+ and bradykinin ions from a pulsed MALDI ion source were obtained. The long ion flight path, combined with mass-independent charge detection, makes this system particularly interesting for the investigation of large molecules.     

Characterization of the microdialysis junction interface for capillary electrophoresis/microelectrospray ionization mass spectrometry

A capillary electrophoresis/electrospray ionization mass spectrometry (CE/ESI-MS) interface, based on an electric circuit across a microdialysis membrane surrounding a short capillary segment closely connected to the separation capillary terminus, is demonstrated to be sensitive, efficient, and rugged. A microspray type ionization emitter produces a stable electrospray at the low flow rates provided by CE and thus avoids both the need for a makeup liquid flow provided by liquid junction or sheath flow interfaces and the subsequent dilution and reduction in sensitivity. Reproducibility studies and comparisons with CE/UV and the CE/sheath flow interface with ESI-MS are presented. Additionally, postrun acidification via the microdialysis junction interface is demonstrated and shown to be capable of denaturing the holomyoglobin protein noncovalent complex while maintaining separation efficiency.     

C60 secondary ion mass spectrometry with a hybrid-quadrupole orthogonal time-of-flight mass spectrometer

A hybrid quadrupole orthogonal time-of-flight mass spectrometer optimized for matrix-assisted laser desorption ionization (MALDI) and electrospray ionization has been equipped with a C 60 cluster ion source. This configuration is shown to exhibit a number of characteristics that improve the performance of traditional time-of-flight secondary ion mass spectrometry (TOF-SIMS) experiments for the analysis of complex organic materials and, potentially, for chemical imaging. Specifically, the primary ion beam is operated as a continuous rather than a pulsed beam, resulting in up to 4 orders of magnitude greater ion fluence on the target. The secondary ions are extracted at very low voltage into 8 mTorr of N 2 gas introduced for collisional focusing and cooling purposes. This extraction configuration is shown to yield secondary ions that rapidly lose memory of the mechanism of their birth, yielding tandem mass spectra that are identical for SIMS and MALDI. With implementation of ion trapping, the extraction efficiency is shown to be equivalent to that found in traditional TOF-SIMS machines. Examples are given, for a variety of substrates that illustrate mass resolution of 12,000-15,600 with a mass range for inorganic compounds to m/ z 40,000. Preliminary chemical mapping experiments show that with added sensitivity, imaging in the MS/MS mode of operation is straightforward. In general, the combination of MALDI and SIMS is shown to add capabilities to each technique, providing a robust platform for TOF-SIMS experiments that already exists in a large number of laboratories.     

Sampling wand for an ion trap mass spectrometer

A new sampling wand concept for ion trap mass spectrometers equipped with discontinuous atmospheric pressure interfaces (DAPI) has been implemented. The ion trap/DAPI combination facilitates the operation of miniature mass spectrometers equipped with ambient ionization sources. However, in the new implementation, instead of transferring ions pneumatically from a distant source, the mass analyzer and DAPI are separated from the main body of the mass spectrometer and installed at the end of a 1.2 m long wand. During ion introduction, ions are captured in the ion trap while the gas in which they are contained passes through the probe and is pumped away. The larger vacuum volume due to the extended wand improves the mass analysis sensitivity. The wand was tested using a modified hand-held ion trap mass spectrometer without additional power or pumping being required. Improved sensitivity was obtained as demonstrated with nano-electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI), and low temperature plasma (LTP) probe analysis of liquid, gaseous, and solid samples, respectively.     

On-line enhancement technique for the analysis of nucleotides using capillary zone electrophoresis/mass spectrometry

A new on-line capillary zone electrophoresis/mass spectrometry (CZE/MS), constant pressure-assisted electrokinetic injection (PAEKI), for the analysis of negatively charged nucleotides is reported. PAEKI uses an applied pressure to counterbalance the reverse electroosmotic flow in the capillary column during sample injection, while taking advantage of the field amplification in the sample medium. At balance, the running buffer in the column is stationary, permitting potentially unlimited injection time, and hence unlimited sample enrichment power. The ability of PAEKI to maintain a narrow sample zone over a long injection time seems to be a result of the formation of a high ion concentration band at the boundary of the two media due to rapid deceleration of the migrating ions at the boundary. The injected amount of analytes proved to be linearly proportional to both the field amplification factor, which is expressed as the ratio of resistivities of sample medium to running buffer, and the injection time, which extended up to 1200 s in CZE/MS and 3600 s in CZE/UV. For a 300-s on-line PAEKI injection in CZE/MS, 3 orders of magnitude sample enhancement (5000-fold enrichment) could be observed for the four single nucleotides without compromising separation efficiency and peak shape, and an achievement of detection limits between 0.04 and 0.07 ng/mL. With appropriate sample cleanup, PAEKI can be used in the analysis of single nucleotides in enzyme-digested DNA.     

Ion trap/ion mobility/quadrupole/time-of-flight mass spectrometry for peptide mixture analysis

An ion trap/ion mobility/quadrupole/time-of-flight mass spectrometer has been developed for the analysis of peptide mixtures. In this approach, a mixture of peptides is electrosprayed into the gas phase. The mixture of ions that is created is accumulated in an ion trap and periodically injected into a drift tube where ions separate according to differences in gas-phase ion mobilities. Upon exiting the drift tube, ions enter a quadrupole mass filter where a specific mass-to-charge (m/z) ratio can be selected prior to collisional activation in an octopole collision cell. Parent and fragment ions that exit the collision cell are analyzed using a reflectron geometry time-of-flight mass spectrometer. The overall configuration allows different species to be selected according to their mobilities and m/z ratios prior to collision-induced dissociation and final MS analysis. A key parameter in these studies is the pressure of the target gas in the collision cell. Above a critical pressure, the well-defined mobility separation degrades. The approach is demonstrated by examining a mixture of tryptic digest peptides of ubiquitin.     

Automated gain control ion funnel trap for orthogonal time-of-flight mass spectrometry

Time-of-flight mass spectrometry (TOF MS) is increasingly used in proteomics research. Herein, we report on the development and characterization of a TOF MS instrument with improved sensitivity equipped with an electrodynamic ion funnel trap (IFT) that employs an automated gain control (AGC) capability. The IFT-TOF MS was coupled to a reversed-phase capillary liquid chromatography (RPLC) separation and evaluated in experiments with complex proteolytic digests. When applied to a global tryptic digest of Shewanella oneidensis proteins, an order-of-magnitude increase in sensitivity compared to that of the conventional continuous mode of operation was achieved due to efficient ion accumulation prior to TOF MS analysis. As a result of this sensitivity improvement and related improvement in mass measurement accuracy, the number of unique peptides identified in the AGC-IFT mode was 5-fold greater than that obtained in the continuous mode.     

Sheathless capillary electrophoresis/electrospray mass spectrometry using a carbon-coated fused-silica capillary

A simple procedure was developed for preparing a carbon-coated fused-silica capillary for use in sheathless capillary electrophoresis/electrospray mass spectrometry (CE/ESI-MS). The tapered capillary tip was smeared with a marker pen before coating with carbon using a soft pencil. The layer from the ink of the marker pen was critical to the preparation of the carbon-coated capillary. The fabrication of a carbon-coated fused-silica capillary tip requires less than 1 min. The stability of this carbon-coated fused-silica capillary is examined, and its utility in on-line sheathless CE/ESI-MS is demonstrated with the separation of berberine, coptisine, and palmatine chlorides. Although the carbon-coated fused-silica capillary tip is not as rugged as a gold-coated capillary, it is durable enough for sheathless CE/ESI-MS applications. Moreover, it is easy to refurbish the column once the performance of the tip is degraded.     

Determination of bioactive peptides using capillary zone electrophoresis/mass spectrometry.

Mixtures of bioactive peptides have been analyzed by capillary zone electrophoresis/mass spectrometry (CZE/MS) using an on-line coaxial continuous-flow fast atom bombardment interface. High separation efficiencies (up to 410,000 theoretical plates) were obtained from low femtomole levels of peptides. The analysis of basic peptides was accomplished by using aminopropyl-silylated CZE columns to minimize zone broadening due to adsorption effects. CZE/MS/MS data were acquired from femtomole levels of peptides in electrophoretic real time.     

Direct chemical analysis of solids by laser ablation in an ion storage time-of-flight mass spectrometer

A laser ablation/ionization mass spectrometer system is described for the direct analysis of solids, particles, and fibers. The system uses a quadrupole ion trap operated in an ion storage mode, coupled with a reflectron time-of-flight mass spectrometer). The sample is inserted radially into the ring electrode, and an imaging system allows direct viewing and selected analysis of the sample. Measurements identified trace contaminants of Ag, Sn, and Sb in a Pb target with single laser shot experiments. Resolution (m/Delta m) of 1500 and detection limits of approximately 10 pg have been achieved with a single laser pulse. The system configuration and related operating principles for accurately measuring low concentrations of isotopes are described.     

High-pressure ion source combined with an in-axis ion trap mass spectrometer. 1. Instrumentation and applications

A new combination of a dual EI/CI ion source with a quadrupole ion trap mass spectrometer has been realized in order to efficiently produce negative ions in the reaction cell. Analysis of volatile compounds was performed under negative ion chemical ionization (NICI) during a reaction period where selected reactant negative ions, previously produced in the external ion source, were allowed to interact with molecules, introduced by hyphenated techniques such as gas chromatography. The O2*-, CH3O-, and Cl- reactant ions were used in this study to ensure specific ion/molecule interactions such as proton transfer, nucleophilic displacement, or charge exchange processes, respectively leading to even-electron species, i.e., deprotonated [M - H]- molecules, diagnostic [M - R]- ions, or odd-electron M*- molecular species. The reaction orientation depends on the thermochemistry of reactions within kinetic controls. First analytical results are presented here for the trace-level detection of several contaminants under NICI/Cl- conditions. Phosphorus-containing compounds (malathion, ethyl parathion, and methyl parathion as representative for pesticides) and nitro-containing compounds (2,4,6-trinitrotoluene for explosive material) have been chosen in order to explore the analytical ability of this promising instrumental coupling.     

Ion soft landing using a rectilinear ion trap mass spectrometer

A new ion soft landing instrument has been built for the controlled deposition of mass selected polyatomic ions. The instrument has been operated with an electrospray ionization source; its major components are an electrodynamic ion funnel to reduce ion loss, a 90-degree bent square quadrupole that prevents deposition of fast neutral molecules onto the landing surface, and a novel rectilinear ion trap (RIT) mass analyzer. The ion trap is elongated (inner dimensions: 8 mm x 10 mm x 10 cm). Three methods of mass analysis have been implemented. (i) A conventional mass-selective instability scan with radial resonance ejection can provide a complete mass spectrum. (ii) The RIT can also be operated as a continuous rf/dc mass filter for isolation and subsequent soft landing of ions of the desired m/ z value. (iii) The 90-degree bent square quadrupole can also be used as a continuous rf/dc mass filter. The mass resolution (50% definition) of the RIT in the trapping mode (radial ion ejection) is approximately 550. Ions from various test mixtures have been mass selected and collected on fluorinated self-assembled monolayers on gold substrates, as verified by analysis of the surface rinses. Desorption electrospray ionization (DESI) has been used to confirm intact deposition of [Val (5)]-Angiotensin I on a surface. Nonmass selective currents up to 1.1 nA and mass-selected currents of up to 500 pA have been collected at the landing surface using continuous rf/dc filtering with the RIT. A quantitative analysis of rinsed surfaces showed that the overall solution-to-solution soft landing yields are between 0.2 and 0.4%. Similar experiments were performed with rf/dc isolation of both arginine and lysine from a mixture using the bent square quadrupole in the rf/dc mode. The unconventional continuous mass selection methods maximize soft landing yields, while still allowing the simple acquisition of full mass spectra.     

On-line coupling of capillary gel electrophoresis with electrospray mass spectrometry for oligonucleotide analysis

Homooligodeoxyribonucleotides differing one nucleotide in length from 12- to 15-mer and from 17- to 20-mer were separated by size with capillary gel electrophoresis (CGE) using an entangled polymer solution in coated capillaries. The resolved components were analyzed by on-line coupling of CGE with electrospray mass spectrometry (ES-MS), denoted as CGE/ES-MS, in the full-scan negative ion detection mode. Baseline separation was achieved for the 12-15-mer oligonucleotide mixtures. Both synthetic phosphodiester oligonucleotide mixtures as well as their phosphorothioate analogues, serving as model compounds for antisense oligonucleotides, could be analyzed by on-line CGE/ES-MS coupling. Terminally phosphorylated and nonphosphorylated synthetic failure sequences could be electrophoretically separated and mass spectrometically characterized as well. This methodology might be a useful tool for synthesis control of phosphodiester oligonucleotides as well as for analysis of phosphorothioate analogues as they are used in antisense drug development.     

Sheathless capillary electrophoresis/electrospray mass spectrometry using a carbon-coated tapered fused-silica capillary with a beveled edge.

A tapered capillary tip containing a beveled edge was developed for use in sheathless capillary electrophoresis/electrospray mass spectrometry (CE/ESI-MS). The optimal flow rate of a 75-microm-i.d., 90-microm-o.d. beveled tapered capillary tip was similar to a conventional flat tapered tip with a 25-microm orifice. Using a mixture of coptisine, berberine, and palmatine chloride, the sheathless CE/ ESI-MS sensitivity of a beveled 75 microm tapered tip capillary was found to be similar to a 25 microm flat tip. Although both tips offer similar CE/ESI-MS sensitivity, the beveled tapered capillary tip is more rugged and durable than a conventional 25-microm tapered capillary because of the larger outside diameter and inside diameter. To make electrical contact, the capillary tip was smeared with paint marker followed by the application of a carbon coating using a graphite pencil. Using this refined carbon-coating procedure, the capillary tip can be operated with aprotic solvents.     

A strategy for the determination of enzyme kinetics using electrospray ionization with an ion trap mass spectrometer.

A simple and rapid means of enzyme kinetic analysis was achieved using electrospray ionization mass spectrometry and a one-point normalization factor. The model system used, glutathione S-transferase from porcine liver, is a two-substrate enzyme catalyzing the conjugation of glutathione with a variety of compounds containing an electrophilic center. An internal standard that is structurally similar to the product was added to the reaction quench solution, and a single-point normalization factor was used to determine the product concentration without the need of a calibration curve. Kinetic parameters, such as Km, Vmax and Ki (for thyroxine), obtained by electrospray mass spectrometry agreed with those obtained from traditional UV-vis spectroscopy, and competitive vs noncompetitive inhibition reactions could be delineated via mass spectrometry. These results suggest that our method can be applied to enzymatic processes in which spectrophotometric or spectrofluorometric assays are not feasible or when the relevant substrates do not incorporate chromophores or fluorophores. This new method is competitive with traditional UV assays in that it is facile and it involves very little analysis time.     

Towards nanoscale molecular analysis at atmospheric pressure by a near-field laser ablation ion trap/time-of-flight mass spectrometer

An instrument that combines near-field laser ablation at atmospheric pressure with an ion trap/time-of-flight mass spectrometer was developed. By coupling a UV laser into a fiber tip of a scanning near-field optical microscope, ablation craters much smaller than achievable with conventional laser optics can in principle be obtained. Laser ablation was performed on samples such as DHB, anthracene, and pyrene. Desorbed neutral analytes are transferred from atmospheric pressure to an ion trap, ionized, and stored. After 10 ms, the ions are extracted into a sensitive time-of-flight spectrometer. We demonstrate the feasibility of this unique SNOM-MS instrument for chemical analysis with unprecedented lateral resolution at atmospheric pressure. Spatially resolved molecular analysis with a lateral resolution of 5 microm (fwhm) and a sensitivity of approximately 60 fmol of solid anthracene is demonstrated, along with topographical analysis with the same instrument. No other technique available today offers this lateral resolution in combination with soft mass spectrometry and the capability of sampling fragile specimens at atmospheric pressure.     

Atmospheric pressure MALDI/ion trap mass spectrometry

A new sample ionization technique, atmospheric pressure matrix-assisted laser desorption/ionization (AP MALDI), was coupled with a commercial ion trap mass spectrometer. This configuration enables the application-specific selection of external atmospheric ionization sources: the electrospray/APCI (commercially available) and AP MALDI (built in-house), which can be readily interchanged within minutes. The detection limit of the novel AP MALDI/ion trap is 10-50 fmol of analyte deposited on the target surface for a four-component mixture of peptides with 800-1700 molecular weight. The possibility of peptide structural analysis by MS/MS and MS3 experiments for AP MALDI-generated ions was demonstrated for the first time.