Improved ion transmission from atmospheric pressure to high vacuum using a multicapillary inlet and electrodynamic ion funnel interface

A heated multicapillary inlet and ion funnel interface was developed to couple an electrospray ionization (ESI) source to a high-vacuum stage for obtaining improved sensitivity in mass spectrometric applications. The inlet was constructed from an array of seven thin-wall stainless steel tubes soldered into a central hole of a cylindrical heating block. An electrodynamic ion funnel was used in the interface region to more effectively capture, focus, and transmit ions from the multicapillary inlet. The interface of seven capillary inlets with the ion funnel showed more than 7 times higher transmission efficiency compared to that of a single capillary inlet with the ion funnel and a 23-fold greater transmission efficiency than could be obtained using the standard orifice-skimmer interface of a triple-quadrupole MS. The multiple-capillary inlet and ion funnel interface showed an overall 10% ion transmission efficiency and approximately 3-4% overall detection efficiency of ions from solution based (i.e., prior to electrospray). The improved performance was achieved under conditions where ESI operation is robust and results in a significant increase in dynamic range.     

Independent control of ion transmission in a jet disrupter dual-channel ion funnel electrospray ionization MS interface

A new atmospheric pressure ionization mass spectrometer (API-MS) interface has been developed to allow the control of ion transmission through the first vacuum stage of the mass spectrometer. The described interface uses a dual-heated capillary and a dual-inlet ion funnel design. Two electrosprays, aligned with the dual-capillary inlet, are used to introduce ions from different solutions independently into the MS. The initial design was specifically aimed at developing a method for the controlled introduction of calibrant ions in highly accurate mass measurements using Fourier transform ion cyclotron resonance mass spectrometer (FTICR). The dual-channel ion funnel has different inlet diameters that are aligned with the dual capillaries. The large diameter main channel of the ion funnel is used for analyte introduction to provide optimum ion transmission. The second, smaller diameter channel inlet includes a jet disrupter in the ion funnel to modulate the ion transmission through the channel. The two inlet channels converge into a single-channel ion funnel where ions from both channels are mixed, focused, and transmitted to the mass analyzer. Both theoretical simulations and experimental results show that the transmission of different m/z species in the small diameter channel of the ion funnel can be effectively modulated by varying the bias voltage on the jet disrupter. Both static and dynamic modulations of ion transmission are demonstrated experimentally by applying either a constant DC or a square waveform voltage to the jet disrupter. High ion transmission efficiency, similar to the standard single-channel ion funnel, is maintained in the main analyte channel inlet of the ion funnel over a broad m/z range with negligible "cross talk" between the two ion funnel inlet channels. Several possible applications of the new interface (e.g., for high-accuracy MS analysis of complex biological samples) are described.     

Capillary-based multi nanoelectrospray emitters: improvements in ion transmission efficiency and implementation with capillary reversed-phase LC-ESI-MS

We describe the coupling of liquid chromatography (LC) separations with mass spectrometry (MS) using nanoelectrospray ionization (nano-ESI) multiemitters. The array of 19 emitters reduced the flow rate delivered to each emitter, allowing the enhanced sensitivity that is characteristic of nano-ESI to be extended to higher flow rate separations. The signal for tryptic fragments from proteins spiked into a human plasma sample increased 11-fold on average when the multiemitters were employed, due to increased ionization efficiency and improved ion transfer efficiency through a newly designed heated multicapillary MS inlet. Additionally, the LC peak signal-to-noise ratio increased approximately 7-fold when the multiemitter configuration was used. The low dead volume of the emitter arrays preserved peak shape and resolution for robust capillary LC separations using total flow rates of 2 microL/min.     

Design and implementation of a new electrodynamic ion funnel

A new electrodynamic (rf) ion funnel has been developed and evaluated for use in the interface regions (at approximately 1-10 Torr) of atmospheric pressure ion sources (e.g., electrospray ionization (ESI) for mass spectrometry). The ion funnel consists of a ring electrode ion guide with decreasing i.d. and with a superimposed dc potential gradient along the ring stack. The thicknesses of the ring electrodes and the spacings between them were reduced to 0.5 mm from 1.59 mm compared to those used for previous designs. The new ion funnel displays a significant improvement in low-mass transmission (m/z >200) and sensitivity compared to previous designs. The transmission efficiencies for electrosprayed peptides and proteins (ranging in mass from 200 to 17,000 Da) were typically 50-60% of total incoming currents from a heated capillary inlet. The transmitted ion currents were a factor of 30-56 greater than those of the standard interface for peptide samples and a factor of 18-22 greater than those for protein samples. The sensitivity gains realized at the MS detector were somewhat lower, possibly due to space charge effects in the octapole ion beam guide following the ion funnel. The improved ion transmission properties result primarily from the use of reduced spacings between ring electrodes. We also show that the ion funnel can be operated in two different modes, one using low-rf-amplitude scans, allowing fragile noncovalent complexes (as well as generally undesired adducts) to be transmitted, and the other using high-rf-amplitude scans, providing greater collisional activation and more effective adduct removal (or the dissociation of lower m/z species).     

Array of chemically etched fused-silica emitters for improving the sensitivity and quantitation of electrospray ionization mass spectrometry

An array of emitters has been developed for increasing the sensitivity of electrospray ionization mass spectrometry (ESI-MS). The linear array consists of 19 chemically etched fused-silica capillaries arranged with 500 microm (center-to-center) spacing. The multiemitter device has a low dead volume to facilitate coupling to capillary liquid chromatography (LC) separations. The high aspect ratio of the emitters enables operation at flow rates as low as 20 nL/min/emitter, effectively extending the benefits of nanoelectrospray to higher flow rate analyses. To accommodate the larger ion current produced by the emitter array, a multicapillary inlet to the mass spectrometer was also constructed. The inlet, which matched the dimensions of the emitter array, preserved ion transmission efficiency. Standard reserpine solutions of varying concentration were electrosprayed at 1 microL/min using the multiemitter/multi-inlet combination, and the results were compared to those from a standard, single-emitter configuration. A 9-fold sensitivity enhancement was observed for the multiemitter relative to the single emitter. A bovine serum albumin tryptic digest was also analyzed, and a sensitivity increase ranging from 2.4- to 12.3-fold for the detected tryptic peptides resulted; the varying response was attributed to reduced ion suppression under the nanoESI conditions afforded by the emitter array. An equimolar mixture of leucine enkephalin and maltopentaose was studied to verify that ion suppression is indeed reduced for the multiplexed ESI (multi-ESI) array relative to a single emitter over a range of flow rates.     

High-resolution field asymmetric waveform ion mobility spectrometry using new planar geometry analyzers

Field asymmetric waveform ion mobility spectrometry (FAIMS) has emerged as a powerful tool of broad utility for separation and characterization of gas-phase ions, especially in conjunction with mass spectrometry (MS). In FAIMS, ions are filtered by the dependence of mobility on electric field while being carried by gas flow through the analytical gap between two electrodes of either planar (p-) or cylindrical (c-) geometry. Most FAIMS/MS systems employ c-FAIMS because of its ease of coupling to MS, yet the merits of the two geometries have not been compared in detail. Here, a priori simulations reveal that reducing the FAIMS curvature always improves resolution at equal sensitivity. In particular, the resolving power of p-FAIMS exceeds that of c-FAIMS, typically by a factor of 2-4 depending on the ion species and carrier gas. We have constructed a new planar FAIMS incorporating a curtain plate interface for effective operation with an ESI ion source and joined to an MS using an ion funnel interface with a novel slit aperture. The resolution increases up to 4-fold over existing c-FAIMS, even though the analysis is approximately 2 times faster. This allows separation of species not feasible in previous FAIMS studies, e.g., protonated leucine and isoleucine or new bradykinin isomers. The improvement for protein conformers (of ubiquitin) is less significant, possibly because of multiple unresolved geometries.     

Subambient pressure ionization with nanoelectrospray source and interface for improved sensitivity in mass spectrometry

A nanoelectrospray ionization mass spectrometry (ESI-MS) source and interface has been designed that enables efficient ion production and transmission in a 30 Torr pressure environment using solvents compatible with typical reversed-phase liquid chromatography (RPLC) separations. In this design, the electrospray emitter is located inside the mass spectrometer in the same region as an electrodynamic ion funnel. This avoids the use of a conductance limiting ion inlet, as required by a conventional atmospheric pressure ESI source, and allows more efficient ion transmission to the mass analyzer. The new subambient pressure ionization with nanoelectrospray (SPIN) source improves instrument sensitivity and enables new electrospray interface designs, including the use of multi-emitter approaches. Performance of the SPIN source was evaluated by electrospraying standard solutions at 300 nL/min and comparing results with those obtained from a standard atmospheric pressure ESI source that used a heated capillary inlet. This initial study demonstrated an approximately 5-fold improvement in sensitivity when the SPIN source was used compared to a standard atmospheric pressure ESI source. The importance of desolvation was also investigated by electrospraying at different flow rates, which showed that the ion funnel provided an effective desolvation region to aid the creation of gas-phase analyte ions.     

Characterization of an improved electrodynamic ion funnel interface for electrospray ionization mass spectrometry.

An improved electrodynamic ion funnel for ion focusing at high pressure (> 1 Torr) has been developed for a triple quadrupole mass spectrometer and its performance compared with that of an earlier prototype previously reported. The ion funnel consists of a series of ring electrodes of progressively smaller internal diameters to which rf and dc electric potentials are co-applied. The new design utilizes ring electrodes possessing larger internal diameters that taper down to a relatively larger exit aperture. In the 1-10 Torr pressure range, the new design provides significant improvement in low m/z ion transmission. Additionally, the overall ion transmission range is improved by linked scanning of the ion funnel's rf voltage concomitantly with the scanning of the quadrupole mass analyzer. Transmission of a noncovalent complex through the interface demonstrated that excessive ion heating was not problematic. Computer simulations of ion transport support the ion funnel design and help explain the relative performance of both designs. Both ion simulations and experimental results are in accord and indicate close to 100% ion transmission efficiency for electrosprayed biopolymer ions through the interface and into the mass analyzer.     

Automatic identification of proteins with a MALDI-quadrupole ion trap mass spectrometer.

A matrix-assisted laser desorption/ionization (MALDI) ion trap mass spectrometer of new design is described. The instrument is based on a commercial Finnegan LCQ ion trap mass spectrometer to which we have added a MALDI ion source that incorporates a sample stage constructed from a compact disk and a new ion transmission interface. The ion interface contains a quadrupole ion guide installed between the skimmer and the octapoles of the original instrument configuration, allowing for operation in both MALDI and electrospray ionization modes. The instrument has femtomole sensitivity for peptides and is capable of collecting a large number of MALDI MS and MALDI MS/MS spectra within a short period of time. The MALDI source produces reproducible signals for 10(4)-10(5) laser pulses, enabling us to collect MS/MS spectra from all the discernible singly charged ions detected in a MS peptide map. We describe the different modes of the instrument operation and algorithms for data processing as applied to challenging protein identification problems.     

Zeptomole-sensitivity electrospray ionization--Fourier transform ion cyclotron resonance mass spectrometry of proteins

Methods are being developed for ultrasensitive protein characterization based upon electrospray ionization (ESI) with Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS). The sensitivity of a FTICR mass spectrometer equipped with an ESI source depends on the overall ion transmission, which combines the probability of ionization, transmission efficiency, and ion trapping in the FTICR cell. Our developments implemented in a 3.5 tesla FTICR mass spectrometer include introduction and optimization of a newly designed electrodynamic ion funnel in the ESI interface, improving the ion beam characteristics in a quadrupole-electrostatic ion guide interface, and modification of the electrostatic ion guide. These developments provide a detection limit of approximately 30 zmol (approximately 18,000 molecules) for proteins with molecular weights ranging from 8 to 20 kDa.     

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.     

Increasing the sensitivity of liquid introduction mass spectrometry by combining electrospray ionization and solvent assisted inlet ionization

Combining electrospray ionization (ESI) and solvent assisted inlet ionization (SAII) provides higher ion abundances over a wide range of concentrations for peptides and proteins than either ESI or SAII. In this method, a voltage is applied to a union connector linking tubing from a solvent delivery device and the fused silica capillary, used with SAII, inserted into a heated inlet tube of an Orbitrap Exactive mass spectrometer (MS). The union can be metal or polymeric and the voltage can be applied directly or contactless. Solution flow rates from less than a 1 μL min(-1) to over 100 μL min(-1) can be accommodated. It appears that the voltage is only necessary to provide charge separation in solution, and the hot MS inlet tube and the high velocity of gas through the tube linking atmospheric pressure and vacuum provides droplet formation. As little as 100 V produces an increase in ion abundance for certain compounds using this method relative to no voltage. Interestingly, the total ion current observed with SAII and this electrosprayed inlet ionization (ESII) method are very similar for weak acid solutions, but with voltage on, the ion abundance for peptides and proteins increase as much as 100-fold relative to other compounds in the solution being analyzed. Thus, switching between SAII (voltage off) and ESII (voltage on) provides a more complete picture of the solution contents than either method alone.     

Multiplexed ion mobility spectrometry-orthogonal time-of-flight mass spectrometry

Ion mobility spectrometry (IMS) coupled to orthogonal time-of-flight mass spectrometry (TOF) has shown significant promise for the characterization of complex biological mixtures. The enormous complexity of biological samples (e.g., from proteomics) and the need for both biological and technical analysis replicates imposes major challenges for multidimensional separation platforms with regard to both sensitivity and sample throughput. A major potential attraction of the IMS-TOF MS platform is separation speeds exceeding that of conventional condensed-phase separations by orders of magnitude. Known limitations of the IMS-TOF MS platforms that presently mitigate this attraction include the need for extensive signal averaging due to factors that include significant ion losses in the IMS-TOF interface and an ion utilization efficiency of less than approximately 1% with continuous ion sources (e.g., ESI). We have developed a new multiplexed ESI-IMS-TOF mass spectrometer that enables lossless ion transmission through the IMS-TOF as well as a utilization efficiency of >50% for ions from the ESI source. Initial results with a mixture of peptides show a approximately 10-fold increase in signal-to-noise ratio with the multiplexed approach compared to a signal averaging approach, with no reduction in either IMS or TOF MS resolution.     

Enhanced ion utilization efficiency using an electrodynamic ion funnel trap as an injection mechanism for ion mobility spectrometry

Conventional ion mobility spectrometers that sample ion packets from continuous sources have traditionally been constrained by an inherently low duty cycle. As such, ion utilization efficiencies have been limited to <1% in order to maintain instrumental resolving power. Using a modified electrodynamic ion funnel, we demonstrated the ability to accumulate, store, and eject ions in conjunction with ion mobility spectrometry (IMS), which elevated the charge density of the ion packets ejected from the ion funnel trap (IFT) and provided a considerable increase in the overall ion utilization efficiency of the IMS instrument. A 7-fold increase in signal intensity was revealed by comparing continuous ion beam current with the amplitude of the pulsed ion current in IFT-IMS experiments using a Faraday plate. Additionally, we describe the IFT operating characteristics using a time-of-flight mass spectrometer attached to the IMS drift tube.     

High-sensitivity ion mobility spectrometry/mass spectrometry using electrodynamic ion funnel interfaces

The utility of ion mobility spectrometry (IMS) for separation of mixtures and structural characterization of ions has been demonstrated extensively, including in biological and nanoscience contexts. A major attraction of IMS is its speed, several orders of magnitude greater than that of condensed-phase separations. Nonetheless, IMS combined with mass spectrometry (MS) has remained a niche technique, substantially because of limited sensitivity resulting from ion losses at the IMS-MS junction. We have developed a new electrospray ionization (ESI)-IMS-QTOF MS instrument that incorporates electrodynamic ion funnels at both front ESI-IMS and rear IMS-QTOF interfaces. The front funnel is of the novel "hourglass" design that efficiently accumulates ions and pulses them into the IMS drift tube. Even for drift tubes of 2-m length, ion transmission through IMS and on to QTOF is essentially lossless across the range of ion masses relevant to most applications. The rf ion focusing at the IMS terminus does not degrade IMS resolving power, which exceeds 100 (for singly charged ions) and is close to the theoretical limit. The overall sensitivity of the present ESI-IMS-MS system is comparable to that of commercial ESI-MS, which should make IMS-MS suitable for analyses of complex mixtures with ultrahigh sensitivity and exceptional throughput.     

Beam injection flame furnace atomic absorption spectrometry: a new flame method

A new flame method of atomic absorption spectrometry is described. The liquid sample to be analyzed is transported as a high-speed liquid jet into a heated tube which is positioned in an air/acetylene flame. The jet is generated by means of an HPLC pump which feeds a smooth jet nozzle having a diameter of 50 microns or smaller. After traveling a distance of 10 cm, the liquid jet enters a small sample introduction hole, impacts onto the opposite inner wall of the tube furnace, and immediately vaporizes (jet impact vaporization, JIV). Both the complete introduction of the entire sample and the extended residence time inside the absorption volume result in an improvement in power of detection from 6- to 202-fold for 17 elements (Ag, As, Au, Bi, Cd, Cu, Hg, In, K, Pb, Pd, Rb, Sb, Se, Te, Tl, Zn). A standard deviation of 1.7-4.0% (n = 12, 50 microL) was achieved. Sample volumes between 10 microL and 1 mL have been investigated. For 50 microL sample volumes, the sampling frequency is 4/min. The new method can also be considered a simple, effective interface between HPLC and flame AAS.     

Coupling capillary electrophoresis and high-field asymmetric waveform ion mobility spectrometry mass spectrometry for the analysis of complex lipopolysaccharides

High-field asymmetric waveform ion mobility spectrometry (FAIMS) is a new technology for atmospheric pressure, room temperature separation of gas-phase ions. The FAIMS system acts as an ion filter that can continuously transmit one type of ion, independent of mass-to-charge ratio (m/z). Capillary electrophoresis-electrospray mass spectrometry (CE-MS) has been extensively used for the analysis of complex bacterial lipopolysaccharides (LPS). The coupling of FAIMS to CE-MS provides a sensitive technique for the characterization of these complex glycolipids, permitting the separation of trace-level LPS oligosaccharide glycoforms for subsequent structural characterization using tandem mass spectrometry. This was demonstrated for LPS from nontypeable Haemophilus influenzae strain 375 following O-deacylation with anhydrous hydrazine. This strain of H. influenzae can express a triheptosyl-containing glycoform to which four hexose residues are linked forming the outer-core region of the molecule. This has been referred to as the Hex4 glycoform. Glycoforms have been identified which differ in the number of phosphoethanolamine substituents in the inner-core. With the use of CE-FAIMS, isomeric Hex4 glycoforms containing two PEtn groups were separated and characterized by MS/MS. FAIMS provided a significant reduction in mass spectral noise, leading to improved detection limits ( approximately 70 amol of the major glycoform). The extracted mass spectrum showed that the apparent noise was virtually eliminated. In addition to the reduction of chemical background, the ion current was increased by as much as 7.5 times as a result of the atmospheric pressure ion-focusing effect provided by the FAIMS system. The linearity of response of the CE-FAIMS-MS system was also studied. The calibration curve is linear for approximately 3 orders of magnitude, over a range of 40 pg/microL to 10 ng/microL.     

Dynamic pressure measurements during pulsed gas injection in a quadrupole ion trap

A quadrupole ion trap has been modified to perform dynamic pressure measurements during pulsed introduction of gases. A continuous electron beam is directed through the ion trap where the gas is ionized via electron impact. Ion and electron currents are monitored on the ring and end-cap electrodes, respectively. Dynamic pressure measurements in a region not accessible to a standard gauge are performed using a static quadrupole field. Characteristic current-voltage curves of the ion-trap gauge are presented and optimum operating conditions of the electron-ion optical system are identified in steady state conditions. The sensitivity of the ion-trap gauge is calibrated at these optimum conditions. In the pulsed gas mode ion and electron signals are measured simultaneously on a fast oscilloscope. The time constant of the circuit for the dynamic measurements is ∼129 μs and pressure variations of ∼10⁻⁴-1 Pa occur within 60-130 ms. The exponential decay of the ion signals is used to calculate pumping speeds for helium and argon gases. The distinctive advantages of pulsed gas injections over the use of static pressures in quadrupole ion-trap mass spectrometry are emphasized.     

Miniaturized ultra high field asymmetric waveform ion mobility spectrometry combined with mass spectrometry for peptide analysis

Miniaturized ultra high field asymmetric waveform ion mobility spectrometry (ultra-FAIMS) combined with mass spectrometry (MS) has been applied to the analysis of standard and tryptic peptides, derived from α-1-acid glycoprotein, using electrospray and nanoelectrospray ion sources. Singly and multiply charged peptide ions were separated in the gas phase using ultra-FAIMS and detected by ion trap and time-of-flight MS. The small compensation voltage (CV) window for the transmission of singly charged ions demonstrates the ability of ultra-FAIMS-MS to generate pseudo-peptide mass fingerprints that may be used to simplify spectra and identify proteins by database searching. Multiply charged ions required a higher CV for transmission, and ions with different amino acid sequences may be separated on the basis of their differential ion mobility. A partial separation of conformers was also observed for the doubly charged ion of bradykinin. Selection on the basis of charge state and differential mobility prior to tandem mass spectrometry facilitates peptide and protein identification by allowing precursor ions to be identified with greater selectivity, thus reducing spectral complexity and enhancing MS detection.     

Absorption of a volatile organic compound by a jet loop reactor with circulation of a surfactant solution: performance evaluation

Biofiltration shows high efficiency for the removal of industrial waste gases and reliable operational stability at low investment and operating cost, especially when the VOC concentration is low, such as 100 ppmv (micro LL(-1)) or less. However, it has been reported that the abrupt change in VOC concentrations leads to the failure of the biofilter. Hence, the pretreatment of waste gases is necessary to ensure the stable operation of the biofilter. The objective of this study is to develop a jet loop reactor (JLR) with circulation of a surfactant solution to lower the concentration of VOCs, especially hydrophobic VOCs. Toluene and Tween 81 were used as a model industrial waste gas and a surfactant, respectively. Among several non-ionic surfactants tested, Tween 81 showed the most rapid dissolution of toluene. When a JLR is replaced with fresh Tween 81 solution (0.3% w/v) every hour, it successfully absorbed for 48 h over 90% of the toluene in an inlet gas containing toluene at 1000 ppmv (microL L(-1)) or less. Therefore, JLR with circulation of a surfactant solution is believed to ensure the stable operation of the biofilter even with the unexpected increase in the VOC concentrations.