High resolution time-of-flight mass analysis of the entire range of intact singly-charged proteins

The proof of principle for high-resolution analysis of intact singly charged proteins of any size is presented. Singly charged protein ions were produced by electrospray ionization followed by surface-induced charge reduction at atmospheric pressure. The inlet and trapping system "stops" the forward momentum of the protein ions over a very broad range to be captured by the digitally produced electric fields of a large radius linear ion trap whereupon they are moved into a smaller radius linear ion trap and collected and concentrated in front of its exit end-cap electrode using digital waveform manipulation. The protein ions are then ejected on demand from the end of the small radius linear quadrupole in a tightly collimated ion beam with an instrumentally defined kinetic energy into the acceleration region of an orthogonal acceleration reflectron time-of-flight mass analyzer where their flight times were measured and detected with a Photonis BiPolar TOF detector. We present results that clearly prove that massive singly charged ions can yield high-resolution mass spectra with very low chemical noise and without loss of sensitivity with increasing mass across the entire spectrum. Analysis of noncovalently bound protein complexes was demonstrated with streptavidin-Cy5 bound with a biotinylated peptide mimic. Our results suggest proteins across the entire range can be directly quantified using our mass analysis technique. We present evidence that solvent molecules noncovalently adduct onto the proteins while yielding consistent flight time distributions. Finally, we provide a look into future that will result from the ability to rapidly measure and quantify protein distributions.     

Halo ion trap mass spectrometer

We describe a novel radio frequency ion trap mass analyzer based on toroidal trapping geometry and microfabrication technology. The device, called the halo ion trap, consists of two parallel ceramic plates, the facing surfaces of which are imprinted with sets of concentric ring electrodes. Radii of the imprinted rings range from 5 to 12 mm, and the spacing between the plates is 4 mm. Unlike conventional ion traps, in which hyperbolic metal electrodes establish equipotential boundary conditions, electric fields in the halo ion trap are established by applying different radio frequency potentials to each ring. The potential on each ring can be independently optimized to provide the best trapping field. The halo ion trap features an open structure, allowing easy access for in situ ionization. The toroidal geometry provides a large trapping and analyzing volume, increasing the number of ions that can be stored and reducing the effects of space-charge on mass analysis. Preliminary mass spectra show resolution (m/Deltam) of 60-75 when the trap is operated at 1.9 MHz and 500 Vp-p.     

Coaxial ion trap mass spectrometer: concentric toroidal and quadrupolar trapping regions

We present the design and results for a new radio-frequency ion trap mass analyzer, the coaxial ion trap, in which both toroidal and quadrupolar trapping regions are created simultaneously. The device is composed of two parallel ceramic plates, the facing surfaces of which are lithographically patterned with concentric metal rings and covered with a thin film of germanium. Experiments demonstrate that ions can be trapped in either region, transferred from the toroidal to the quadrupolar region, and mass-selectively ejected from the quadrupolar region to a detector. Ions trapped in the toroidal region can be transferred to the quadrupole region using an applied ac signal in the radial direction, although it appears that the mechanism of this transfer does not involve resonance with the ion secular frequency, and the process is not mass selective. Ions in the quadrupole trapping region are mass analyzed using dipole resonant ejection. Multiple transfer steps and mass analysis scans are possible on a single population of ions, as from a single ionization/trapping event. The device demonstrates better mass resolving power than the radially ejecting halo ion trap and better sensitivity than the planar quadrupole ion trap.     

Miniaturization and geometry optimization of a polymer-based rectilinear ion trap

The fabrication, operation, and characterization of a polymer-based rectilinear ion trap mass analyzer is discussed. A novel, fast prototyping technique, stereolithography (SLA)-based fabrication, traditionally reserved for end use production parts and to fabricate master molds for rubber products, is applied here as a tool to create precise, arbitrary geometries. Taking full advantage of the SLA methodology, an open corner, polymer-based ion trap has been fabricated and tested. The use of a custom resin, Nanoform 15120 (DSM Somos, New Castle, DE), has resulted in a polymer with high heat deflection temperature and greater structural stability at higher temperatures and lower capacitance. The mass analyzer was mounted in a polymer holder and tested in a custom vacuum system using modified LCQ Duo (Thermo Fisher Corp.) electronics. The resolution, mass/charge range, and MS/MS capabilities were examined using electrospray ionization as well as atmospheric pressure chemical ionization. In the course of this study, three traps of different sizes were fabricated, beginning with a "full size" device measuring 10 x 8 x 50 mm. The next two traps were scaled down by linear factors of a half and a third. SLA is shown to allow fabrication of light, small rectilinear ion traps, which are less expensive and have the same performance as traditional machined devices of the same size. In addition, smaller traps can be built just as easily, and they show unit mass resolution to mass 300, tandem mass spectrometry capabilities, and low power consumption.     

Polymer-Based Ion Trap Chemical Sensor

Ion traps are widely used in chemical analysis, and they are especially important in current attempts to miniaturize mass spectrometers to create portable instruments. The ultimate aim is to build a handheld device that would require a smaller mass analyzer. To accomplish this task, a robust precision fabrication procedure is desired. In this paper, the authors report a new approach to fabricating ion traps using stereolithography apparatus (SLA), which provides precision monolithic fabrication. An SLA-fabricated rectilinear ion trap, which employs a very simple electrode geometry, is shown to provide detection capabilities within a useful mass range encompassing that of interest in the detection of numerous volatile organic compounds, including those relevant to homeland security applications. Single small ion traps and integrated trap arrays can be made through this approach, which allows higher operating pressures and reduced power requirements     

Multiplexed rectilinear ion trap mass spectrometer for high-throughput analysis

A multichannel mass spectrometer based on the rectilinear ion trap (RIT) analyzer was designed and constructed for simultaneous high-throughput analysis of multiple samples. The instrument features four parallel ion source/mass analyzer/detector channels assembled in a single vacuum chamber and operated using a common set of control electronics, including a single rf amplifier and transformer coil. This multiplexed RIT mass spectrometer employs an array of four millimeter-sized ion traps (x(o) = 5.0 mm and y(o) = 4.0 mm, where x(o) and y(o) are the half-distances in the x and y dimensions, respectively). Mass spectra are acquired from four different samples simultaneously. The available mass/charge range is m/z 15-510 with excellent linearity of the mass calibration (R2 = 0.999 999). The peak width is less than 0.3 mass/charge units at m/z 146, corresponding to a resolution of approximately 500. Simultaneous MS/MS of ions due to four compounds (3-fluoroanisole, 4-fluoroanisole, 2-fluorobenzyl alcohol, 2,6-dimethylcyclohexanone) with the same nominal molecular radical cation but distinctive fragmentation patterns was demonstrated. Isolation and fragmentation efficiencies were approximately 25 and approximately 75%, respectively, measured in the typical case of the molecular radical cation of acetophenone. Preacquisition differential data were obtained by real-time subtraction of the ion signals from two channels of the multiplexed mass spectrometer. The differential experiment presented offers proof of principle of comparative mass spectra in high-throughput screening applications while reducing data storage requirements.     

An interface with a linear quadrupole ion guide for an electrospray-ion trap mass spectrometer system

A new ion sampling interface for an electrospray ionization 3D ion trap mass spectrometer system is described. The interface uses linear rf quadrupoles as ion guides and ion traps to enhance the performance of the 3D trap. Trapping ions in the linear quadrupoles is demonstrated to improve the duty cycle of the system. Dipolar excitation of ions trapped in a linear quadrupole is used to eject unwanted ions. A resolution of ejection of up to 254 is demonstrated for protonated reserpine ions (m/z 609.3). A composite waveform with a notch in frequency space is used to eject a wide range of matrix ions and to isolate trace analyte ions in a linear quadrupole before ions are injected into the 3D trap. This is useful to overcome space charge problems in the 3D trap caused by excess matrix ions. For trace reserpine in a 500-fold molar excess of poly(propylene glycol) (PPG), it is demonstrated that the resolution and sensitivity of the 3D trap can be increased dramatically with ejection of the excess PPG matrix ions. In comparison to ejection of matrix ions in the 3D trap with a similar broad-band waveform, a 5-fold increase in sensitivity with a 7 times shorter acquisition time was achieved.     

Interfacing the orbitrap mass analyzer to an electrospray ion source

The orbitrap mass analyzer employs the trapping of pulsed ion beams in an electrostatic quadro-logarithmic field. This field is created between an axial central electrode and a coaxial outer electrode. Stable ion trajectories combine rotation around the central electrode with harmonic oscillations along it. The frequencies of axial oscillations and hence mass-to-charge ratios of ions are obtained using fast Fourier transform of the image current detected on the two split halves of the outer electrode. This work proves that such a trap could be coupled to a continuous, electrospray, ion source. Such a coupling necessitated the development of an rf-only quadrupole for external accumulation of ions and their injection in very short (< 1 micros) ion bunches. Along with good sensitivity, this mass spectrometer provides mass resolving power up to 150,000 fwhm, mass accuracies within a few parts per million, and relative mass range up to 8-fold. The maximum number of ions available for analysis is limited by the space-charge capacity of the accumulation quadrupole.     

A tandem ion trap/ion mobility spectrometer

A tandem quadrupole ion trap/ion mobility spectrometer (QIT/IMS) has been constructed for structural analysis based on the gas-phase mobilities of mass-selected ions. The instrument combines the ion accumulation, manipulation, and mass-selection capabilities of a modified ion trap mass spectrometer with gas-phase electrophoretic separation in a custom-built ion mobility drift cell. The quadrupole ion trap may be operated as a conventional mass spectrometer, with ion detection using an off-axis dynode/multiplier arrangement, or as an ion source for the IMS drift cell. In the latter case, pulses of ions are ejected from the trap and transferred to the drift cell where mobility in the presence of helium buffer gas is determined by the collision cross section of the ion. Ions traversing the drift cell are detected by an in-line electron multiplier and the data processed with a multichannel scaler. Preliminary data are presented on instrumental performance characteristics and the application of QIT/ IMS to structural and conformational studies of aromatic ions and protonated amine/crown ether noncovalent complexes generated via ion/molecule reactions in the ion trap.     

Ion trap mass spectrometry of fluorescently labeled nanoparticles

Mass spectra of fluorescently labeled polystyrene nanoparticles have been obtained using a combined technique of matrix-assisted laser desorption/ionization (MALDI), laser-induced fluorescence (LIF), and a dual quadrupole ion trap mass spectrometer. The spectrometer is designed in such a way that the first trap serves as a trapping and mass-analyzing device, while the second trap serves to capture and concentrate the ions ejected from the first trap for fluorescence detection. An enhancement in the LIF signal by more than 3 orders of magnitude is achieved with the help of the second trap, making mass/charge (m/z) analysis of the nanoparticles possible. Additional unique features of this mass spectrometer include that frequency scan (0.5-50 kHz) at a constant voltage (200 V), instead of voltage scan at a constant frequency, is implemented to widen the spectral analysis range of the instrument. The implementation has allowed the spectrometer to operate at relatively high buffer gas pressures (50 mTorr), crucial for effective trapping of the nanometer-sized particles generated by MALDI. We present in this report the first mass spectra of fluorescently labeled nanoparticles with a size of 27 nm using this new mass spectrometric approach. The utility of this method in the study of biological macromolecules or particles is demonstrated with dye-labeled IgG.     

Resolution and chemical formula identification of aromatic hydrocarbons and aromatic compounds containing sulfur, nitrogen, or oxygen in petroleum distillates and refinery streams

An all-glass heated inlet system has been interfaced to a dual-trap Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometer. The inlet vaporizes a mixture of species of widely different boiling points, and the interface maintains a large (factor of 10(10)) pressure gradient between the inlet and the mass spectrometer, making possible the analysis of petroleum distillates and refinery streams at very high mass resolution. Ions generated by low-energy electron ionization in the source trap of the spectrometer are transferred to the analyzer trap, where the pressure is at least 2 orders of magnitude lower. Singly-charged ions from a mass window of ～20 u are isolated by stored-waveform radial ejection, to reduce space charge and increase digital resolution: routine mass resolving power >200?000 (based on magnitude-mode peak full width at half-height) is thereby achieved throughout the full mass window. The mass window may be incremented stepwise to cover the full mass range of several hundred units. The FT-ICR mass spectrum of a gas oil aromatic neutral fraction contained peaks resulting from the resolution of ions having 358 distinct formulas over a mass range of ～42 u. C(3)/SH(4), (13)C/CH, (13)CH/N, CH(2)/N, and other mass doublets were baseline-resolved, yielding typical mass measurement inaccuracies of ～1 ppm. For example, (13)C(12)C(17)H(20)S(+) and C(21)H(17)(+), which differ by only 0.0011 u at ～269 u, were clearly resolved. A 40?000 resolving power low-voltage spectrum of the aromatic neutrals, acquired by use of a Kratos MS-50 double-focusing instrument, was processed with a computer-based deisotoping/formula assignment procedure. The algorithm of the program is outlined and illustrated. Remarkably good agreement exists between the FT-ICR and MS-50 results. However, instrumental rather than indirect resolution of ions clearly enhances analytical accuracy and significantly reduces data-processing time. Thus, we have demonstrated that FT-ICR is the mass analysis of choice for differentiating hydrocarbons from heteroatom-containing compounds in petroleum distillates and refinery streams.     

Monitoring structural changes of proteins in an ion trap over approximately 10-200 ms: unfolding transitions in cytochrome c ions.

A new technique for studying the time dependence of conformational changes of gas-phase protein ions is described. In this approach, a short pulse of electrosprayed protein ions is introduced into an ion trap and stored. After a defined time period, the distribution of ions is ejected from the trap into an ion mobility/time-of-flight mass spectrometer. Combined measurements of mobilities and flight times in the mass spectrometer provide information about the abundances of different conformer types and charge-state distributions. By varying the storage time in the trap, it is possible to monitor changes in ion conformation that occur over extended time periods (approximately 10-200 ms). The method is demonstrated by examining changes in cytochrome c ion conformations for the +7 to +10 charge states.     

Cylindrical ion trap array with mass selection by variation in trap dimensions

A mass spectrometer array is described in which each array element is a cylindrical ion trap (CIT) within which an approximately quadrupolar, time-varying, field is established. The individual traps are of different sizes, so that when the array is operated with a fixed rf potential, ions of different masses (or mass ranges) are stored in each trap. By choosing the dimensions of each CIT element in the array, a multiple ion monitoring experiment can be performed. For example, in a two-element array with elements having internal radii of 5 and 4 mm, the smaller trap selects for m/z 91 and the larger for m/z 57, corresponding to characteristic aromatic and aliphatic hydrocarbon ions. Ion storage using both rf/dc (apex) isolation and the stored waveform inverse Fourier transform method is demonstrated.The array reduces the complexity of the electronics needed to operate the ion trap, which should make it suitable for use in a miniature mass spectrometer system.     

Electrostatic axially harmonic orbital trapping: a high-performance technique of mass analysis

This work describes a new type of mass analyzer which employs trapping in an electrostatic field. The potential distribution of the field can be represented as a combination of quadrupole and logarithmic potentials. In the absence of any magnetic or rf fields, ion stability is achieved only due to ions orbiting around an axial electrode. Orbiting ions also perform harmonic oscillations along the electrode with frequency proportional to (m/z)-1/2. These oscillations are detected using image current detection and are transformed into mass spectra using fast FT, similarly to FTICR. Practical aspects of the trap design are presented. High-mass resolution up to 150,000 for ions produced by laser ablation has been demonstrated, along with high-energy acceptance and wide mass range.     

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.     

Single-pulse nanoelectrospray ionization

A new electrospray ionization (ESI) source that provides a means of generating single packets of ions for mass spectrometric analysis is presented. Sample solution held at a high potential is ejected from a glass capillary with a small dispensing aperture (20-microm i.d.) by constriction of a cylindrical piezoelectric element. Unlike conventional ESI sources that are continuous, this source dispenses fixed volumes of solution as small as 10 pL and provides detection sensitivity in the attomole range when coupled to an orthogonal time-of-flight mass spectrometer. In addition to picoliter-level control over the dispensed volume, the source permits control of the frequency with which ionization pulses are generated as well as the ability to start and stop the pulses without altering the applied solution potential. The source was characterized by analysis of both protein and DNA samples from a variety of different solution compositions. This source design should be compatible with virtually any ESI mass analyzer.     

Atmospheric pressure ionization in a miniature mass spectrometer

A miniature cylindrical ion trap mass spectrometer featuring an atmospheric pressure interface allowing atmospheric pressure chemical ionization and electrospray ionization is described together with its analytical performance characteristics. The vacuum system, ion optics, mass analyzer, control electronics system, and detection system have all been designed and built in-house. The design is based upon a three-stage, differentially pumped vacuum system with the instrument capable of being interfaced to many types of atmospheric pressure ionization sources. Ions are transferred through home-built ion optics, and instrument control is achieved through custom-designed electronics and LabView control software. Corona discharge ionization and electrospray ionization sources are implemented and used to allow the analysis of both gaseous- and solution-phase samples during the characterization of the instrument. An upper mass/charge limit of approximately 450 Th with unit resolution was achieved using a 2.5-mm-internal radius cylindrical ion trap as the mass analyzer. The specificity of the instrument can be increased by employing the MS/MS capabilities of the ion trap and has been demonstrated for nitrobenzene. Limits of detection for the trace analysis in air of the chemical warfare agent simulant methyl salicylate (1.24 ppb) and for nitrobenzene (629 pptr) are achieved. The dynamic range of the instrument is currently limited to approximately 2 orders of magnitude by saturation of the detection electronics. Isolation and collision-induced dissociation efficiencies in MS/MS experiments both greater than 50% are reported. Electrospray/nanospray data are presented on solutions including 100 microM (D,L)-arginine, 10 microM (-)-ephedrine, and 10 microM lomefloxacin.     

Rectilinear ion trap mass spectrometer with atmospheric pressure interface and electrospray ionization source

A rectilinear ion trap (RIT) mass analyzer was incorporated into a mass spectrometer fitted with an electrospray ionization source and an atmospheric pressure interface. The RIT mass spectrometer, which was assembled in two different configurations, was used for the study of biological compounds, for which performance data are given. A variety of techniques, including the use of a balanced rf, elevated background gas pressure, automatic gain control, and resonance ejection waveforms with dynamically adjusted amplitude, were applied to enhance performance. The capabilities of the instrument were characterized using proteins, peptides, and pharmaceutical drugs. Unit resolution and an accuracy of better than m/z 0.2 was achieved for mass-to-charge (m/z) ratios up to 2000 Th at a scan rate of approximately 3000 amu/(charge.s) while reduced scan rates gave greater resolution and peak widths of less than m/z 0.5 over the same range. The mass discrimination in trapping externally generated ions was characterized over the range m/z 190-2000 and an optimized low mass cutoff value of m/z 120-140 was found to give equal trapping efficiencies over the entire range. The radial detection efficiency was measured as a function of m/z ratio and found to rise from 35% at low m/z values to more than 90% for ions of m/z 1800. The way in which the ion trapping capacity depends on the dc trapping potential was investigated by measuring the mass shift due to space charge effects, and it was shown that low trapping potentials minimize space charge effects by increasing the useful volume of the device. The collision-induced dissociation (CID) capabilities of the RIT instrument were evaluated by measuring isolation efficiency as a function of mass resolution as well as measuring peptide CID efficiencies. Overall CID efficiencies of more than 60% were easily reached, while isolation of an ion with unit resolution at m/z 524 was achieved with high rejection (>95%) of the adjacent ions. The overall analytical capabilities of the ESI-RIT instrument were demonstrated with the analysis of a mixture of pharmaceutical compounds using multiple-stage mass spectrometry.     

离子阱中过渡金属Ti~ ,Co~ 与O_2的化学反应

利用激光溅射的方法产生并在射频离子阱中囚禁了Ti＋，Co＋离子，利用离子阱的质量选择存储和离子存储时间长等特点，开展了Ti＋，Co＋与O2的化学反应研究，得到了反应物的速率常数和反应产物分支比。结果表明，Co＋比Ti＋具有更大的化学活性，并且其二级产物（CoO2＋）具有更大的解离速率，因而更具有催化特性。     

Chemical characterization of individual, airborne sub-10-nm particles and molecules

A nanoaerosol mass spectrometer (NAMS) is described for real-time characterization of individual airborne nanoparticles. The NAMS includes an aerodynamic inlet, quadrupole ion guide, quadrupole ion trap, and time-of-flight mass analyzer. Charged particles in the aerosol are drawn through the aerodynamic inlet, focused through the ion guide, and captured in the ion trap. Trapped particles are irradiated with a high-energy laser pulse to reach the "complete ionization limit" where each particle is thought to be completely disintegrated into atomic ions. In this limit, the relative signal intensities of the atomic ions give the atomic composition. The method is first demonstrated with sucrose particles produced with an electrospray generator. Under the conditions used, the particle detection efficiency (fraction of charged particles entering the inlet that are subsequently analyzed) reaches a maximum of 10(-4) at 9.5 nm in diameter and the size distribution of trapped particles has a geometric standard deviation of 1.1 based on a log-normal distribution. A method to deconvolute overlapping multiply charged ions (e.g. C3+ and O4+) is presented. When applied to sucrose spectra, the measured C/O atomic ratio is 1.1, which matches the expected ratio from the molecular formula. The spectra of singly charged bovine serum albumin (BSA) molecules are also presented, and the measured and expected C/N/O atomic ratios are within 15% of the each other. Also observed in the BSA spectra are signals from 13C and 32S which arise from 40 and approximately 34 atoms per molecule (particle), respectively. Potential applications of NAMS to atmospheric chemistry and biotechnology are briefly discussed.