Robert K. Boyd

A hybrid BEQQ mass spectrometer for studies in gaseous ion chemistry

Abstract A hybrid mass spectrometer of BEQQ geometry (B, magnetic sector; E, electric sector; Q, quadrupole mass filter), designed for fundamental and applied studies in gaseous ion chemistry, is described. The high-resolution (BE) stage is followed by a deceleration lens, which also shapes the ion beam, an r.f.-only quadrupole collision cell, and a quadrupole mass analyzer. This assembly allows collision processes to be studied over the laboratory energy range from 0 to 500 eV with selection of the reactant ion at high mass resolution. The instrument is also equipped with dual collision cells and a deflector electrode in the field-free region between B and E for the study of neutralization—reionization reactions as well as single collision processes at 1–8 keV collision energy. The use of the instrument is illustrated with a variety of examples involving charge stripping, charge inversion, fast neutral ionization, and neutralization—reionization processes at high collision energies, as well as collision-induced dissociation reactions at both high and low collision energies. In addition, a number of experiments involving simultaneous scanning of the electric sector and the QQ stage are reported.

Definitions of terms relating to mass spectrometry (IUPAC Recommendations 2013)

This document contains recommendations for terminology in mass spectrometry. Development of standard terms dates back to 1974 when the IUPAC Commission on Analytical Nomenclature issued recommendations on mass spectrometry terms and definitions. In 1978, the IUPAC Commission on Molecular Structure and Spectroscopy updated and extended the recommendations and made further recommendations regarding symbols, acronyms, and abbreviations. The IUPAC Physical Chemistry Division Commission on Molecular Structure and Spectroscopy’s Subcommittee on Mass Spectroscopy revised the recommended terms in 1991 and appended terms relating to vacuum technology. Some additional terms related to tandem mass spectrometry were added in 1993 and accelerator mass spectrometry in 1994. Owing to the rapid expansion of the field in the intervening years, particularly in mass spectrometry of biomolecules, a further revision of the recommendations has become necessary. This document contains a comprehensive revision of mass spectrometry terminology that represents the current consensus of the mass spectrometry community.

‘Wrong-way-round’ Electrospray Ionization of Amino Acids†

‘Wrong-way-round’ electrospray ionization, in the present context, refers to observation of intense [M + H]+ ions electrosprayed from strongly basic solutions and of [M − H]− ions from strongly acidic solutions. Most previous investigations of this phenomenon have been directed at variations in charge-state distributions for polyfunctional peptides and proteins as a function of bulk solution pH. The present work extends that of Hiraoka et al. (J. Mass Spectrom. Soc. Japan43, 127 (1995)) on the pH dependence of absolute mass spectral intensities of ions in electrospray mass spectra of amino acids. This choice of test analyte permits investigation of both positive- and negative-ion mass spectra without potential complications from changes in secondary and tertiary structures as the pH is varied. The intensities of [M + H]+ and [M − H]− ions, over the pH range 3 to 11, varied by factors of 3–5 despite calculated variations of several orders of magnitude in equilibrium concentrations in the bulk solution. The same behaviour was observed for derivatized amino acids such as amides and methyl esters. Measurements of pH of collected spray, and of the total current carried by the charged droplets, confirmed that these observations can not be accounted for in terms of wholesale pH switching from acidic to basic or vice versa by electrochemical reactions at the electrospray needle. Precursors of the ‘wrong-way-round’ ions were sought by conventional precursor-ion scanning experiments but with minimal declustering conditions in the atmospheric-pressure ion-source interface. In the case of added electrolytes, such as ammonia and acetic acid, which are both volatile and capable of Bronsted acid–base behaviour, the observations were consistent with earlier interpretations involving e.g. [M + NH4+] precursors for [M + H]+ ions. Such explanations were not applicable to similar observations made for solutions with added tetramethylammonium hydroxide or hydrochloric acid.

Experimental investigations of factors controlling the collision induced dissociation spectra of peptide ions in a tandem hybrid mass spectrometer. I. Leucine enkephalin

Abstract A systematic study is reported of the experimental parameters controlling the collision induced dissociation (CID) of protonated Leu-enkephalin ( m/z 556.4) in a tandem hybrid mass spectrometer of BEqQ configuration. For CID reactions occurring in the r.f.-only quadrupole q, with collision energies in the range 5–120 eV (laboratory frame), the fragment ion spectra are functions of not only the collision parameters (collision energy, nature of collision gas, etc.) but also of operating characteristics of the qQ quadrupole assembly. The dependence upon the dynamic ion optical parameters was found to be in accord with established principles, given the significantly higher mass of the precursor ion studied here than in previous evaluations of the performance of XqQ tandem instruments (X = Q or sector analyzer). The intensity ratio for the Y″ 2 and B 3 fragment ions was found to be a sensitive index of the degree of internal excitation of the precursor ions, though no calibration of this intensity ratio vs. internal energy was possible. Comparisons between efficiencies of argon and xenon as target gases, as a function of collision energy, were performed objectively via purity-fit scores obtained by using library comparison algorithms. These data are consistent with a proposal that the momentum transfer mechanism of collisional activation operates at the elastic limit for small organic precursor ions, but that physical size restrictions require an increasingly dominant role for the binary (spectator) limit as the precursor mass is increased. It is proposed that this switch could account for the striking difference between very high CID efficiencies obtained for small organic ions, in XqQ instruments, and the increasingly poor performance as precursor mass is raised to 1000 Da or so. The crucial role of pre-collision internal energy, in the low-energy CID of these larger ions, is emphasized.

The mobile proton hypothesis in fragmentation of protonated peptides: A perspective

The Distinguished Contribution Award of the American Society for Mass Spectrometry “—recognizes a focused, singular achievement in or contribution to fundamental or applied mass spectrometry, in contrast to awards that recognize lifetime achievement—a contribution that has had a significant impact on the fundamental understanding and/or practice of mass spectrometry.” The hypothesis that has come to be known as themobile proton model clearly satisfies both the fundamental and practical criteria. It is a pleasure to have been asked to contribute a brief introductory article to this special issue that honors Vicki Wysocki and Simon Gaskell, the principal originators and proponents of the model and the 2009 recipients of this prestigious award. Two excellent reviews cover the relevant literature up to about 2005 [1, 2]. The purpose of this short article is to attempt to view the mobile proton model relative to a wider perspective. Application of mass spectrometry to determination of molecular structure relies on interpretation of fragment ion spectra, however obtained, using a set of rules that are the result of years of experience in extending concepts of classical physical-organic chemistry. Most of the fragmentation rules were derived from experience with positive ion mass spectra obtained using electron ionization. Probably the best-known guide to these interpretative rules is the book authored by McLafferty and Turecek [3]. The underlying theme of these “classical” rules is that the electron rearrangements involved in decomposition of an activated ion into two or more fragments are triggered by localization of charge (and/or unpaired electron spin in the case of radical ions) on specific sites within the molecular structure of the decomposing ion. It is true that these rules are almost entirely empirical, but their continuing practical success indicates that they must correspond to real phenomena in some sense. The introduction of chemical ionization, and later the powerful fast atom bombardment (FAB) [4], electrospray (ESI) [5], and MALDI [6] ionization techniques led to extension of the rules for fragmentations of molecular radical cations to even-electron molecular species formed by adduction of simple ions, protons in the great majority of cases. Indeed, the mobile proton model can be regarded as an extension of these classical rules that permits their appropriate application to protonated peptides or other organic molecules. The main exceptions to interpretations based on triggering by localized charge sites are the charge-remote fragmentations [7] investigated by Gross and his colleagues starting in the late 1980s. These “pseudo-thermal” reactions are known [8, 9] to contribute to fragmentation of protonated peptides at high levels of internal activation, where they compete with the mechanisms subsumed by the mobile proton theory, particularly for singly-protonated peptides containing an Arg residue. As emphasized previously [2], the mobile proton model is not a complete theory that can predict a fragment ion spectrum for any given protonated peptide, but rather a qualitative framework that permits appropriate application of interpretative rules based on charge-site localization. Essentially, the model assumes that for protonated peptides formed by soft ionization methods such as electrospray (ESI), the protons are initially localized on the most basic sites in the molecule. These sites are the N-terminus and the side chains of basic amino acid residues, particularly Arg, Lys, and His. After ion activation the ionizing proton(s) can be transferred from the less-basic of these initially occupied sites to the various peptide linkages, thus triggering charge-site-initiated mechanisms of various kinds that provide the desired sequence ions. In other words, the mobile proton model also implies that heterogeneous populations of protonated forms can be generated upon ion activation and some of these forms are “fragmenting” structures, while others remain intact during the time frame of the mass spectrometer. This framework has now been extensively reviewed and its practical usefulness amply demonstrated [1, 2]. Of course all useful new ideas appear to be simple and obvious once someone else has described them. To illustrate that the mobile proton model was not always regarded as “obvious” and was “linked” only later to the idea of ion populations generated by an “activated”, i.e., “transferable” proton, one can cite some early work published in 1992 by one of the recipients of the Distinguished Contributions Award [10]. This paper was mainly concerned with demonstrating intraionic interactions in peptides containing cysteic acid plus Address reprint requests to Dr. R. K. Boyd, National Research Council, Bldg. M12, 1200 Montreal Road, Ottawa K1A0R6, Canada. E-mail: robert. boyd@nrc.ca

Collision induced dissociation of peptide ions.: Part 3. Comparison of results obtained using sector—quadrupole hybrids with those from tandem double-focusing instruments

Two series of peptides, viz., dynorphin fragments (up to 9 residues) and oligomers of phenylalanine, have been studied by collision induced dissociation (CID) of their protonated forms produced by fast atom bombardment (FAB) ionization. Spectra were obtained using both a four-sector tandem instrument (CID in the kiloelectronvolt range) and a tandem hybrid instrument (CID at a few tens of electronvolts). The four-sector instrument provided fragment spectra of excellent quality under conditions simulating a real-world analytical situation, i.e. 1–2 nmol of material available, although the FAB efficiency for Phe8, was so low (precursor ion beam current ∼ 10−14 A) that no useful fragment spectra were obtained. The hybrid instrument provided abundant fragment ions for smaller precursors (molecular weights < 500–600 Da) but for larger precursors (≥ 800 Da or so) the CID efficiency was much reduced and highly susceptible to composition effects, particularly the presence of highly basic amino acid residues. The information content of these spectra was evaluated objectively via two complementary computer algorithms. The limitations of CID in r.f.-only quadrupole collision cells are not due primarily to ion transmission effects, since examples of good quality spectra for large precursors are presented. The low efficiency of collisional activation of organic ions of m/z ≥ 800 Da or so, under conditions typical of quadrupole collision cells, was confirmed by the observation that in those cases where abundant fragment ions were obtained the corresponding spectra obtained in absence of collision gas were of comparable quality. The differences between collisional activation of such ions in the kiloelectronvolt range and in the electronvolt range, as well as the dramatic differences between low energy CID of large and small precursor ions, are discussed in terms of fundamental considerations of inelastic collisions. Finally, the qualitatively different kinds of fragmentation reactions, observed in the two regimes of collision energy, are described. In both cases the well-known peptide sequence fragments were observed. The high-energy CID also produced intense fragments arising from cleavage of all or part of sidechains (d, s, v and w series) which were entirely absent from the low-energy spectra. The latter contained abundant internal cleavage ions (both N- and C-terminus residues lost) which were relatively weak in the high-energy spectra. Loss of the C-terminus residue was observed in both regimes as a low-energy process in the absence of collision gas, although the compositional conditions giving rise to this process have not been determined in the present work.

The application of studies using metastable ions to the determination of the thermochemistry of gaseous ions

Abstract The contribution of ion kinetic energy spectrometry to various studies concerned with the thermochemistry of gaseous ions is assessed. The work deals with unimolecular fragmentation of singly- and doubly-charged ions and also discusses the value of collision-induced dissociations to the elucidation of the relevant energetic quantities. The proposal that the reactions of metastable ions observed in practice correspond to those producing the thermodynamically most stable products is discussed critically; its value and limitations are both explored. A critical assessment of the importance of appearance potential measurements is given and the value of obtaining these measurements on metastable peaks is stressed. Various aspects of the partitioning of the energy of the activated complex between the internal energy of the products and translational energy release are discussed. The paper also includes some specific examples of the application of IKES to ion thermochemistry. These concern the enthalpy of the benzoyl ion, of C 3 H 5 + and C 3 H 3 + ions of different structures, of the C 6 H 5 + ion from benzene and also of the detailed fragmentation of benzonitrile molecular ions.

Ion kinetic energy spectroscopy of the doubly-charged ion of carbon monoxide

Spontaneous and collision‐induced dissociation processes of CO2+ ions, formed by electron impact, have been studied in a double‐focusing mass spectrometer using techniques of ion kinetic energy spectroscopy. The predissociation process, responsible for unimolecular dissociation of CO2+ on the microsecond time scale, is almost certainly electronically adiabatic tunneling through a potential barrier, though predissociation via electronic curve crossing cannot be entirely ruled out. Semiempirical potential curves for states of CO2+ were revised in order to better accommodate all of the available data, including Auger spectra, appearance energies, and kinetic energy release. Collision induced dissociation processes with Ar, N2, and H2 proceed via charge exchange, and involve predissociation of the D 2Π state by the C 2Δ state of CO+. When He is used as collision gas, the dissociation processes involving charge exchange are different, and require an energetic contribution from the relative kinetic energy (kine...

Analytical performance of a high-pressure radio frequency-only quadrupole collision cell with an axial field applied by using conical rods

The effect on triple-quadrupole performance of applying an axial field, in an rf-only quadrupole collision cell operated at pressures sufficiently high that collisional focusing is operating, has been investigated. The advantages of such cells have been shown previously to include increased transmission and much improved resolution in fragment ion spectra relative to the performance of collision cells operating at lower gas pressures. The disadvantages of high-pressure collision cells all derive from the relatively long transit times for the ions, which can be long relative to characteristic times for scanning the first mass filter (precursor ion selector) or for switching its setting in multiple reaction monitoring (MRM) cycles. The present work describes experiments on a high-pressure cell in which an axial field is created through use of conical rather than cylindrical or hyperbolic rods. In addition, results of computations of the electric fields within such a cell, and of ion trajectories through it, are presented. It is shown that application of axial fields of the order of 0. 1 V/cm can remove all hysteresis effects associated with the long ion transit times, and thus provide excellent performance in quantitation work using MRM, as well as in other scan modes. Furthermore, the advantages of collisional focusing in quadrupole collision cells are shown to be unimpaired by these low axial fields.

Characterisation of the tyrocidine and gramicidin fractions of the tyrothricin complex from Bacillus brevis using liquid chromatography and mass spectrometry

Abstract The tyrothricin peptide complex, isolated from the fermentation broth of Bacillus brevis, is comprised of a basic fraction of cyclic decapeptides (the tyrocidines) and a neutral fraction composed of linear peptides (the gramicidins). Previously, five cyclic compounds (tyrocidines A–E) had been characterised by classical chemical procedures, and an additional five by M. Barber, D.J. Bell, M.R. Morris, L.W. Tetler, J.J. Monaghan, W.E. Morden, B.W. Bycroft and B.N. Green, Int. J. Mass Spectrom. Ion Processes, 122 (1992) 143-150, who employed tandem mass spectrometric analysis of the crude mixture, together with an interpretative strategy based upon mass shifts related to simple amino acid substitutions. In the present work, initial profiling of the tyrothricin complex, using reverse phase liquid chromatography (HPLC) coupled directly to tandem mass spectrometry via an ionspray interface, showed that the mixture is extremely complex. Semi-preparative HPLC provided 32 fractions, some of which were still mixtures, amenable to analysis by tandem mass spectrometry using the doubly-protonated peptide precursors produced by ionspray ionisation. In this way the 10 previously known tyrocidines were confirmed, and structures of an additional 18 cyclic variants established with only minor uncertainties (e.g. present techniques could not distinguish Ile from Leu). Six linear gramicidins were known previously, and were confirmed in the present work. In addition, three previously unknown variants, of the Val1 -gramicidins A,B and C, were discovered, in which the ethanolamide residue at the C-terminus is replaced by a propanolamide residue.