Bruker Daltonik Gmbh

Other

Ralf Hartmer

Andreas Brekenfeld

Jochen Franzen

Christoph Gebhardt

Desmond Allen Kaplan

Jens Decker

The invention relates to mass spectrometric analyses of heavy molecules and molecular complexes having molecular weights sometimes well above 100,000 daltons, by collision treatment in linear RF multipole collision cells. A mixture of at least one light collision gas (&#x3c;40 daltons) and at least one heavy collision gas (&#x3e;80 daltons) is provided in a linear RF collision cell. The heavy collision gas results in high-momentum and high-energy collisions, which leads to splitting and further fragmentation of portions of the heavy molecular (complex) ions. For this purpose, the molecular (complex) ions are axially injected into the collision cell at kinetic energies of several hundred electronvolts per charge; due to the collisions with the heavy collision gas molecules they are deflected from the axis and excited to undergo strong oscillations in the radial direction in the focusing RF field. The light collision gas serves in turn for damping these oscillations.

Structural elucidation of intact heavy molecules and molecular complexes in mass spectrometers

The invention relates to the selection of the most favorable ion species for the acquisition of fragment ion mass spectra when the ionization creates biopolymers in different charge states. The invention proposes a particularly fast method of selecting the most favorable parent ions for fragmentation of the different biopolymers from mass spectra, where the ionization is by electrospray ionization (ESI) or other ionization methods which produce similarly diverse charge states and which, for each biopolymer, contain many signal patterns of ions of the different charge states and different isotopic compositions. The selection is carried out in such a way that it does not measure more than one ion species from one biopolymer. Moreover, the most favorable filter pass-band width for isolating an ion species for fragmentation can be stated in each case.

Acquisition of fragment ion mass spectra of biopolymers in mixtures

The invention relates to the use of substances for the production of anions suitable for charge transfer reactions in mass spectrometers, particularly for the fragmentation of multiply positively charged biopolymer ions by electron transfer or for charge reduction by proton transfer. Diketones, particularly &#x3b1;-diketones, are proposed as a newly found class of substances which can be used both for the production of radical anions for electron transfer dissociations (ETD) with a high yield of fragment ions and also for the production of non-radical anions for the charge reduction of multiply charged analyte ions by proton transfer reactions (PTR). These substances have favorable properties in terms of their handling and the associated analytical methods: they are largely nontoxic, cover a favorable range of molecular masses, and their volatility means that they can be stored in unheated containers outside of the vacuum system, which facilitates the refilling of the containers.

Alpha- and chi-diketone reactants for charge transfer reactions in mass spectrometers

The invention involves electrospray ionization of dissolved substances at atmospheric pressure in the ion source of a mass spectrometer. A chip with a multitude of spray nozzles is proposed, where each individual spray nozzle is surrounded by several sheath gas nozzles, preferably in a symmetric arrangement, for the jet-like introduction of a sheath gas. A shared attracting-voltage electrode is positioned substantially opposite the spray nozzles. The attracting-voltage electrode may have a tapering (e.g. funnel-shaped) opening above each spray nozzle so that the sheath gas jets are forced to closely envelop the spray jet, which is comprised of ions and very fine droplets. Heavier ions and droplets are thus prevented from discharging on the surfaces of the openings of the attracting-voltage electrode. Special measures can be taken to make all spray nozzles spray uniformly and to supply them with substance peaks from chromatographic or electrophoretic separators as simultaneously as possible.

Multi-nozzle chip for electrospray ionization in mass spectrometers

The invention proposes a method for the collective ejection of ions from a 3D RF ion trap with a ring electrode and two end cap electrodes, which comprises the following steps: (a) the RF voltage of a high-quality resonant circuit applied to the ring electrode is replaced with a second RF voltage at the two end cap electrodes which can be changed or switched faster than the high voltage at the ring electrode, keeping the ions stored, (b) the second RF voltage at the end cap electrodes is then switched down or off abruptly, releasing the ions, and (c) the released ions are ejected through an opening in one of the end cap electrodes by switching on a DC voltage on at least one of the end cap electrodes.

Ejection of ion clouds from 3D RF ion traps

Radical anions for use in the fragmentation of positively charged biopolymer ions by means of electron transfer are produced from substances previously unknown for use as ETD production substances. The inventive substances produce radical anions that lead to electron transfer dissociations with a high yield of fragment ions. The substances have high volatility that allows them to be kept in unheated containers outside the vacuum system and transported into the vacuum system to an in vacuum electron attachment ion source via unheated lines and low molecular weights that allow the measurement of even very light fragment ions. In one embodiment, a suitable substance is 1-3-5-7-cyclooctatetraene.

Radical ions for electron transfer dissociation

Radical anions for electron transfer dissociation

A complex protein mixture is analyzed by jointly digesting the mixture, separating the digest peptides chromatographically or electrophoretically, and ionizing the digest peptides eluting from the separation device by an ionizing method that generates multiply charged ions. Digest peptide ions within a pre-selected range of m/z-values are isolated in an RF ion trap and subsequently reduced in their charge state. The charge-reduced ions can be measured with very high sensitivity. By repeating this process with adjacent isolation mass windows within the time duration of each separation peak, it is possible to determine the masses m, the prevalent charge states z, the retention times t, and the intensities i of a huge number of digest peptides of the complex protein mixture in a single separation run.

Proteome analysis in mass spectrometers containing RF ion traps

In an RF quadrupole ion trap having electrodes to which RF voltages are applied, ions having m/z ratios outside of a predefined narrow range of charge-related masses m/z are removed from the trap by applying a DC voltage pulse to at least one of the trap electrodes to remove from the trap the ions with high values of charge-related masses. The DC voltage pulse is preferably applied in combination with a variation of the RF voltage amplitudes to simultaneously remove from the trap ions of low charge-related masses. The DC and RF voltage amplitudes are changed in such a manner that any excitation of ions having charge-related masses within the predefined range by frequency mixtures is avoided.

Isolation of ions in overloaded RF ion traps

Analyte ions, particularly biopolymer ions, stored in an RF ion trap are ergodically fragmented by bombarding the analyte ions with collision ions, for example medium-mass, mono-atomic ions having a charge of opposite polarity to the charge of the analyte ions. Since the analyte ions are not fragmented by accelerating and/or exciting them to oscillations, as is the case with conventional collision-induced dissociation, the RF voltage of the ion trap can be set low enough that daughter ions with light charge-related masses that are produced by the fragmentation can also remain trapped in the ion trap.

Fragmentation of analyte ions by collisions in RF ion traps

Proteins with a molecular mass in the range from approximately 5 to 100 kilodaltons are structurally analyzed without prior enzymatic digestion to small peptides in a mass spectrometer that operates with an ion trap. The proteins are ionized by electrospraying or similar processes to create highly charged analyte ions, which are then introduced into the ion trap and subjected to fragmentation and partial deprotonation in either order. The fragmentation may be ergodic or electron-induced. The result remaining in the ion trap is an evenly distributed mixture of fragment ions having between one and n charges, where n is a number between three and about eight. A mass spectrum is recorded from this mixture of fragment ions, which spectrum demonstrates a sequence coverage that far exceeds the mass range of the mass analyzer for singly charged ions.

Top-down protein analysis in mass spectrometers with ion traps

In an RF ion trap mass spectrometer, selected parent ions are fragmented by collisions or electrons and a spectrum of the ion fragments is measured. The measured fragment ion spectrum is then analyzed for the presence of a dominant ion species and, when a dominant ion species is present, selected parent ions are fragmented and a spectrum of the ion fragments is again measured, but with an additional collision excitation of the dominant ion species. The resulting daughter ion spectrum is qualitatively improved.

Feedback fragmentation in ion trap mass spectrometers

In mass spectrometric analysis equipment, highly charged analyte fragment ions are obtained by electron transfer dissociation of highly charged analyte ions by radical anions and the dissociation products are then deprotonated with non-radical anions. Both the radical anions for electron transfer dissociation and the non-radical anions for deprotonation are produced in a single ion source from a single substance, or a single mixture of substances by adjusting the electrical operating parameters of the ion source.

Ion source for electron transfer dissociation and deprotonation

The invention relates to a method and instrument for the fragmentation of large molecular analyte ions, preferably biopolymer ions, by reactions between multiply charged positive analyte ions and negative reactant ions in RF quadrupole ion traps. Some of these reactions involve electron transfer reactions with subsequent dissociation of the biopolymer analyte ions, and some involve the loss of a proton, leading to stable product ions. The invention can use any type of ion traps, particularly three-dimensional RF quadrupole ion traps, for the reactions between positive and negative ions. The fragmentation yield can be increased because ions that remain stable as radical cations after transfer of an electron are further fragmented by collisionally induced fragmentation, forming fragment ions that are typical of electron transfer, and not those typical of collisionally induced fragmentation. The invention preferentially introduces positive ions and negative ions into the ion trap sequentially through the same aperture.

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Hamburg, Germany

Ralf Hartmer