Raymond E. Kaiser

Ion trap mass spectrometry

The quadrupole ion trap is a mass spectrometer whose essential components can be held in one hand. But it has a mass range of about 105 daltons per charge, provides molecular weight and structural information on biopolymers, and has the greatest sensitivity of all mass spectrometers. These features, however, have become available only within the past few years. They stem from an almost neglected 1958 invention, one in which interest was maintained by only a few research groups, notably those of John Todd at the University of Kent in England and Ray March at Trent University in Canada. Development of a new scanning method by George Stafford and his coworkers of Finnigan Corp. provided the impetus that led Finnigan to introduce a commercial ion trap in 1983. Since then, the device has been transformed from a simple gas chromatography detector to a high-performance mass spectrometer. In the late 1950s, Wolfgang Paul and his coworkers at ...

Conformer selection of protein ions by ion mobility in a triple quadrupole mass spectrometer

Electrospray mass spectra of multiply charged protein molecules show two distinct charge state distributions proposed to correspond to a more highly charged, open conformational form and a lower charged, folded form. Elastic collisions carried out in the radiofrequency-only collision cell of a triple quadrupole mass spectrometer have dramatic effects on the appearance of the mass spectra. The different cross sectional areas of the conformers allow preferential selection of one charge state distribution over the other on the basis of ion mobility. Preferential selection is dependent on the nature and pressure of the target gas as well as the nature of the protein. In the case of positively charged horse heart apomyoglobin (MW 16,951 da), a high charge state distribution centered around (M + 20H)20+ predominates at low target gas pressures and a second distribution centered around (M + 10H)10+ predominates at high target gas pressures. Bimodal distributions are observed at intermediate pressures and, remarkably, charge states between the two distributions are not effectively populated under most of the conditions examined. Hard sphere collision calculations show large differences in collision frequencies and in the corresponding kinetic energy losses for the two conformational states and they demonstrate that the observed charge state selectivity can be explained through elastic collisions.

Pulsed gas introduction into quadrupole ion traps.

Two different Paul-type quadrupole ion traps were equipped with pulsed-valve gas inlets. The duration of a gas pulse inside the trap is variable, and pulses as short as 50 ms (FWHH) have been measured, allowing the use of several gas pulses during one experiment. The benefits of pulsed valves are outlined and demonstrated for chemical ionization experiments and for the use of selective ion-molecule reactions in structure determination of ions and neutral molecules.

Resonance effects during ion injection into an ion trap mass spectrometer

Abstract Resonance effects are described due to the application of a supplementary a.c. voltage applied 180° out-of-phase to the end caps of a quadrupole ion trap and the effects of varying the frequency and the amplitude of the modulation voltage are studied. At specific frequencies of the modulation voltage, sharp resonances are seen in the abundances of ions injected from an external source. The frequencies which produce resonances depend also on the amplitude of the r.f. voltage supplied to the ring electrode using injection. These effects are shown to be a consequence of modulation of ion motion by the supplementary field, which contains a dipole component together with a minor hexapole field component. The results predicted on this basis agree well with the experimental data. In the absence of the supplementary field, resonances are still observed for ions of sufficiently low kinetic energy.

Measurement of kinetic energies of ions ejected from a quadrupole ion trap

Abstract Kinetic energies of ions of different mass-to-charge ratios exiting a Paul-type ion trap, operated in the mass-selective instability mode, are measured by a retarding potential method. Average kinetic energies increase with mass, a typical value being 2900 eV with a distribution of 900 eV (FWHM) for m/z 502, C9F20N+. The maximum value remains in the same range but the width of the distribution increases when the trap is operated at lower He bath gas pressure. When ion ejection is performed at reduced values of the Mathieu parameter βz, the most probable ion kinetic energy decreases, following the amplitude of the main r.f. voltage. For example, m/z 502 when resonantly ejected at βz 0.48 shows a maximum kinetic energy of only 644eV. Simulations of ion motion employing stepwise integration yield values of the maximum kinetic energies which are in good agremeent with the experimental data. The implications of these results for the mass resolution of the ion trap and for other performance data are discussed.

A transportable turnkey gas chromatograph—ion trap detector for field analysis of environmental samples

Abstract A transportable gas chromatograph—ion trap detector has been developed for the in situ characterization of chemical waste sites. This instrument is based on a modular design and can be readily modified in the field for air, water, or soil sampling. A purge-and-trap gas chromatograph developed for this instrument is used for the separation of volatile organic compounds before their introduction to the ion trap for mass spectral analysis. A daughter microprocessor has been developed for the control of ancillary hardware via the ion trap software. Most analyses are accomplished in an automated 20 min procedure. The detection limit for trichloroethylene in water is in the low part-per-trillion range. The analysis of soil and water samples is demonstrated by using surrogate samples spiked with 24 volatile organic compounds. The instrument has been used under field conditions for soil analysis at a chemical waste site.

Proteolysis of Human Growth Hormone by Rat Thyroid Gland in Vitro: Application of Electrospray Mass Spectrometry and N-Terminal Sequencing to Elucidate a Metabolic Pathway

The present studies were designed to provide structural characterization of peptide metabolites of biosynthetic human growth hormone (hGH) formed by rat thyroid gland proteases in vitro. Electro-spray ionization mass/spectrometry (ESI-MS) and N-terminal sequencing were used to characterize the peptide metabolites. The predominant enzyme in the thyroid gland preparations was a chymotrypsin-like serine protease which was biochemically similar to rat mast cell protease-I. Metabolic intermediates were formed by cleavage of hGH exclusively at Tyr/Phe/Leu-Xaa bonds. After a 5- or 45-min incubation of hGH with thyroid gland S9 pellet fraction, the majority of metabolites formed were two-chain variants of hGH having masses ranging from 16,002 to 22,143 Da. These metabolites were formed as a result of proteolysis in the large disulfide loop region of hGH in combination with processing at Tyr42–Ser43. Based upon the time-related appearance and structural characterization of these intermediates, a sequence of metabolic events is proposed. The initial event appears to be cleavage by the chymotrypsin-like protease between Tyr143–Ser144 to form a two-chain hGH. This intermediate is then cleaved between Tyr42–Ser43, liberating the N-terminal peptide, Phe1–Tyr42. Two other metabolites were generated as a result of the deletion of the peptides Lys140–Tyr143 and Ser144–Phe146 from the large loop region. The identification of similar metabolites truncated by a single amino acid at the carboxyl terminus indicated the action of a carboxypeptidase on these metabolic products. After a 4.5-hr incubation, the protease isolated from the S9 pellet fraction degraded hGH to >20 small peptides, having masses ≤2300 Da. The data illustrate the utility of combining ESI-MS and N-terminal sequencing in the study of protein metabolism and the enzymatic pathways involved.

Metabolism of des(64,65)-Human Proinsulin in the Rat: Evidence for the Proteolytic Processing to Insulin

The metabolism of des(64,65)-human proinsulin was examined in rats after subcutaneous administration. Profiles of circulating insulin-like immunoreactivity in rat plasma 25 min after subcutaneous administration were evaluated by anion exchange fast protein liquid chromatography and reversed-phase high-performance liquid chromatography. Both techniques indicated the presence of circulating immunoreactivity having retention characteristics of human insulin. This metabolite peak comprised 5–10% of circulating immunoreactivity; the remainder had retention characteristics of des(64,65)-human proinsulin. The peaks of immunoreactive material were isolated and their structure determined using reversed-phase high-performance liquid chromatography and electrospray ionization mass spectrometry. The major circulating component co-eluted with des(64,65)-human proinsulin and had an identical mass spectrum. Two circulating metabolites were identified. These metabolites co-eluted by reversed-phase high-performance liquid chromatography with human insulin and diarginyl(B31,32)-human insulin and had mass spectra identical to the standard compounds. The data indicate proteolytic processing of des(64,65)-human proinsulin involves an initial tryptic cleavage at the carboxy side of ArgB32, with the formation of human insulin by the subsequent action of a carboxypeptidase to remove the ArgB31-ArgB32 dipeptide from diarginyl(B31,32)-human insulin. The results suggest that some of the pharmacological activity of des(64,65)-human proinsulin may be mediated in part by circulating insulin-like metabolites.