B. A. Collings

A new linear ion trap time-of-flight system with tandem mass spectrometry capabilities

A new hybrid linear ion trap time-of-flight mass spectrometer has been constructed and the first results for tandem-in-time MS/MS in a 2-dimensional Paul trap are presented. The instrument consists of an electrospray ionization source and two radio frequency (RF) only quadrupoles operating at a pressure of 7 mTorr N2 which are coupled orthogonally to a linear TOFMS. Ions are trapped in an RF-only quadrupole through the application of timed stopping potentials on the entrance and exit apertures. Through the coupling of an auxiliary waveform generator on one pair of the ion trap rods a small dipolar excitation voltage is superimposed on the quadrupolar electric field. This voltage is used for precursor isolation via broadband excitation and collision induced dissociation through mass selective resonant excitation. Instrumental performance and choice of operating parameters are demonstrated by recording fragment spectra for +3 ions of renin substrate and +1 ions of reserpine. TOFMS mass resolution (m/Δm)FWHM is as high as 740 at m/z = 609. Fragmentation efficiency is greater than 50% and excitation mass resolution is 60–75.

Collision cross sections of myoglobin and cytochrome c ions with Ne, Ar, and Kr

The energy losses of ions of myoglobin and cytochrome c in collisions with Ne, Ar, and Kr have been measured with a triple quadrupole mass spectrometer system. The results have been interpreted with drag coefficients, suitable for the case of an object moving through a low density gas, to give collision cross sections or, equivalently, projection areas. For a given charge state, these cross sections with Ne, Ar, and Kr are nearly identical. No evidence is found for substantial contributions to the cross section from ion-induced dipole forces. Comparisons of cross sections obtained with different gases and comparisons to literature cross sections measured by ion mobility suggest that a “diffuse” scattering model, suitable for collisions with a rough surface, gives the best description of collisions between protein ions and neutrals.

Resonant excitation in a low-pressure linear ion trap.

It has been shown that through the process of resonant excitation the fragmentation of ions confined in a low-pressure (<0. 05 mTorr) linear ion trap (LIT) can be accomplished while maintaining both high fragmentation efficiency and high resolution of excitation. The ion reserpine, 609. 23 Da, has been fragmented with efficiencies greater than 90% while a higher mass ion, a homogeneously substituted triazatriphosphorine of mass 2721. 89 Da, has been fragmented with 48% efficiency. This was accomplished by extended resonant excitation by low-amplitude auxiliary RF signals. Computer modelling of ion trajectories and analysis of the trapping potentials have demonstrated that a reduction in neutralization of ions on the rods (or losses on the rods) and increased fragmentation is a consequence of higher order terms in the potential introduced by the round-rod geometry of the LIT.

Observation of higher order quadrupole excitation frequencies in a linear ion trap

The resonant frequencies for quadrupole excitation of ions confined with a buffer gas in a linear quadrupole ion trap with Mathieu parameters a = 0 and q ∼ 0.36 have been measured. The resonances are predicted to occur at angular frequencies ωnK given by ωnK = (Ώ/K)|n + β| without the presence of a buffer gas where ϒ is the angular frequency of the trapping radio frequency, K = 1,2,3 … is the order of the resonance calculated with perturbation theory, and n = 0, ±1, ±2, ±3 …. The resonances are measured through fragmentation of protonated reserpine. The observed frequencies agree closely with the theoretical values but there are small differences which vary from +0.6% at K = 2 to −2.7% at K = 6. This is believed to be the result of the dependence of the resonant frequencies upon the buffer gas density and/or the excitation amplitude. The resolution of the resonances (measured from the depletion of precursor and formation of fragment ions) increased by a factor of 2 as K increased from 1 to 6. This increase in resolution warrants further investigation into the use of higher order resonances for isolation and excitation of trapped ions.

An extended mass range quadrupole for electrospray mass spectrometry

Abstract The mass range of a quadrupole mass spectrometer with an eleetrospray source has been increased from m/z 4000 to m/z 8585 by lowering the radio frequency of the quadrupole power supply from 1.00 MHz to 683 kHz. Unit mass resolution at m/z 5030 can be achieved. The sensitivity below m/z 3000 shows only small changes as expected from phase space arguments. Spectra of polypropylene glycol and soybean agglutinin with peaks in the range m/z 4000–7000 are described.

Detection of noncovalent complex between α‐amylase and its microbial inhibitor tendamistat by electrospray ionization mass spectrometry

Electrospray ionization mass spectrometry (ESI-MS) is now routinely used for detection of noncovalent complexes. However, detection of noncovalent protein-protein complexes is not a widespread practice and still produces some challenges for mass spectrometrists. Here we demonstrate the detection of a noncovalent protein-protein complex between alpha-amylase and its microbial inhibitor tendamistat using ESI-MS. Crude porcine pancreatic alpha-amylase was purified using a glycogen precipitation method. Noncovalent complexes between porcine pancreatic alpha-amylase and its microbial inhibitor tendamistat are probed and detected using ESI-MS. The atmosphere-vacuum ESI conditions along with solution conditions and the ratio of inhibitor over enzyme strongly affect the detection of noncovalent complexes in the gas phase. ESI mass spectra of alpha-amylase at pH 7 exhibited charge states significantly lower than that reported previously, which is indicative of a native protein conformation necessary to produce a noncovalent complex. Detection of noncovalent complexes in the gas phase suggests that further use of conventional biochemical approaches to provide a qualitative, and in some cases even quantitative, characterization of equilibria of noncovalent complexes in solution is possible.

Conformation of native, reduced and [5–55]Ala bovine pancreatic trypsin inhibitor in the gas phase

Collision cross sections have been measured for gas phase ions of native oxidized bovine pancreatic trypsin inhibitor (BPTI), native reduced BPTI and a mutant form of BPTI containing a single disulphide bond between residues 5 and 55 ([5-55]Ala BPTI). Cross sections for [5-55]Ala BPTI and reduced BPTI were 9% and 17% greater respectively than those for native BPTI. Cross sections for native BPTI were smaller than previous estimates from the crystal structure but in reasonable agreement with values calculated from the radius of gyration determined by x-ray scattering from solution BPTI. The increase in cross section for reduced BPTI over native BPTI is similar to that seen in molecular dynamics simulations. The results show that the disulphide bonds of BPTI contribute to the folding of the gas phase ions, but that even in the absence of disulphide bonds the protein ions maintain compact structures. Comparisons to relative areas calculated from hydrodynamic radii of BPTI in solution suggest that when disulphide linkages are removed, BPTI in the gas phase unfolds less than BPTI in solution.

High intensity ion beams from an atmospheric pressure inductively coupled plasma

This work is directed towards substantially improving the sensitivity of an inductively coupled plasma mass spectrometer (ICP-MS). Ions produced in the ICP at atmospheric pressure have been extracted with comparatively high current densities. The conventional approach to ion extraction, based on a skimmed molecular beam, has been abandoned, and a high extraction field arrangement has been adopted. Although the new approach is not optimized, current densities more than 180 times greater than that of a conventional interface have been extracted and analyte sensitivities ∼10–100× greater than those reported previously for quadrupole ICP-MS have been measured.