Mass spectrometer with high duty cycle

Abstract

A method of mass spectrometry comprising: a) accumulating precursor ions in a first ion accumulator; b) performing a separation cycle including pulsing packets of precursor ions from the first ion accumulator to an ion separator and separating the precursor ions such that precursor ions having different values ​​of a first physicochemical property elute from the ion separator at different times; c) mass filtering the precursor ions eluting from the ion separator to transmit selected precursor ion species; and d) fragmenting or reacting the selected precursor ion species to produce a set of fragment or product ion species from the selected precursor ion species. e) accumulating the set of fragment or product ion species in a second ion accumulator; f) ejecting the set of fragment or product ion species from the second ion accumulator into a TOP mass analyzer, where operation of the TOP mass analyzer is synchronized with the ejection of ions from the second ion accumulator such that a plurality of different species of the set of fragment or product ion species are simultaneously pulsed by a pusher electrode into a time-of-flight region of the TOP mass analyzer; and g) repeating steps c)-f) at least once during a separation cycle, where the selected precursor ion species is different each time steps c)-f) are performed.

Description

[Technical field]

[0001] (CROSS REFERENCE TO RELATED APPLICATIONS) This application claims priority to and the benefit of UK Patent Application No. 2204106.5, filed March 23, 2022, the entire contents of which are incorporated herein by reference.

[0002] FIELD OF THEINVENTION The present invention relates generally to mass spectrometers in which ions are pulsed into a separation region causing the ions to be separated according to their mass-to-charge ratio. [Background technology]

[0003] In both Data Dependent Acquisition (DDA) and tandem (MSMS) mass spectrometry, precursor ions are filtered and only selected species of precursor ions are transmitted downstream for analysis at any given time. The mass spectrometer therefore selects different species of precursor ions to be transmitted downstream for analysis. This process continues until the desired species of precursor ions are analyzed. In general, it is desirable to sequentially select and transmit as many precursor ions per unit time as possible to analyze the sample as quickly as possible. However, in conventional instruments such as Quadrupole-Time of Flight (QToF) mass spectrometers, simply increasing the rate at which precursor species are selected results in a linear decrease in the sensitivity of the ions detected. For example, if the spectrometer settings are changed such that the number of transmitted precursor species increases from 25 different precursors per second to 50 different precursors per second, this results in the number of ions in each fragment ion scan being halved.

[0004] It would be desirable to increase the overall duty cycle of such equipment. Summary of the Invention

[0005] A first aspect of the present disclosure provides a method of mass spectrometry, the method comprising: a) accumulating precursor ions in a first ion accumulator; b) performing a separation cycle comprising pulsing packets of precursor ions from a first ion accumulator to an ion separator and separating the precursor ions such that precursor ions having different values ​​of a first physicochemical property elute from the ion separator at different times; c) mass filtering the precursor ions eluting from the ion separator to transmit selected precursor ion species; d) fragmenting or reacting the selected precursor ion species to produce a set of fragment or product ion species from the selected precursor ion species; e) accumulating the set of fragment or product ion species in a second ion accumulator; f) ejecting the set of fragment or product ion species from the second ion accumulator into a TOF mass analyzer, the operation of the TOF mass analyzer being synchronized with the ejection of ions from the second ion accumulator such that multiple different species of the set of fragment or product ion species are simultaneously pulsed by a pusher electrode into a time-of-flight region of the TOF mass analyzer; g) repeating steps c) through f) at least once during a separation cycle, the selected precursor ion species being different each time steps c) through f) are performed; Includes.

[0006] The inventors have recognized that the time scale over which the second ion accumulator can accumulate fragment or product ions and release them for analysis in the TOF mass analyzer is comparable to the time scale over which any given precursor ion elutes from the upstream separator. Thus, fragment or product ion species derived from a first precursor ion species eluting from the ion separator during a given separation cycle of the ion separator can be accumulated, captured, and released to the TOF mass analyzer before a second precursor ion species elutes from the ion separator within the same separation cycle of the ion separator. The method provides a relatively high duty cycle because selected precursor ion species are fragmented while other precursor ion species remain delayed in the ion separator, rather than being discarded. Also, the duty cycle is relatively high because the synchronization between the second ion accumulator and the TOF mass analyzer allows multiple fragment or product ion species of a selected precursor to be analyzed simultaneously.

[0007] The ion separator may be an ion mobility separator and the physicochemical property may be ion mobility.

[0008] For example, the ion separator may be an ion mobility separator configured to drive ions through a background gas disposed therein. The ions may be driven through the background gas by an electric field to separate according to mobility. For example, the ions may be driven by applying a static DC potential gradient along the ion separator and / or by repeatedly moving a DC potential along the ion separator. The electric field may be arranged to urge the ions downstream (towards the TOF mass analyzer) through the ion separator. Alternatively, the background gas may be flowed in a downstream direction and the electric field may be configured to urge the ions upstream against the gas flow to separate the ions according to mobility.

[0009] Other embodiments are contemplated in which the ion separator is a Field Asymmetric Ion Mobility Separator (FAIMS) device or a Differential Mobility Separator (DMS). Less preferably, the ion separator mass is a mass-to-charge ratio separator and the physicochemical property may be mass-to-charge ratio.

[0010] Each time step c) is performed, only selected precursor ion species may be transmitted, and other precursor ion species may not be transmitted.

[0011] Alternatively, each time step c) is performed, the mass filter may transmit precursor ions having a limited range of mass-to-charge ratios such that multiple precursor ion species are transmitted simultaneously, while precursor ions having mass-to-charge ratios outside this limited range are filtered out. Step d) may then include simultaneously fragmenting or reacting the transmitted multiple precursor ion species to generate multiple respective sets of fragment or product ion species therefrom. Step e) may include simultaneously accumulating the set of fragment or product ion species in a second ion accumulator. Step f) may include simultaneously ejecting the set of fragment or product ion species from the second ion accumulator into a TOF mass analyzer, where operation of the TOF mass analyzer is synchronized with the ejection of ions from the second ion accumulator such that multiple different species of the set of fragment or product ion species are simultaneously pulsed by a pusher electrode into the time-of-flight region of the TOF mass analyzer. Step g) may comprise repeating steps c) to f) at least once during a separation cycle, wherein the range of mass-to-charge ratios transmitted by the mass filter is different each time steps c) to f) are performed.

[0012] A voltage is applied to the electrodes of the mass filter to transmit a limited range of mass to charge ratios, such as only a single precursor ion species or multiple precursor ion species at any given time. According to step g) above, the voltage is varied with time during the separation cycle such that at least one different precursor ion species is transmitted by the mass filter each time steps c) to f) are performed.

[0013] The mass filter may be a quadrupole mass filter, for example a quadrupole rod set mass filter or an axially segmented quadrupole rod set mass filter, however, it is contemplated that other types of mass filters may be used in the present disclosure.

[0014] The fragment or product ion species may be biased into the second ion accumulator such that (i) fragment or product ion species derived from the same precursor ion species reach the second ion accumulator over substantially the same period of time and are accumulated in the second ion accumulator during at least a portion of this period, and / or (ii) fragment or product ion species derived from precursor species transmitted by the mass filter at different times reach the second ion accumulator over different respective periods of time.

[0015] Each time the sequence of steps c) to e) is performed, the fragment or product ion species may be accumulated in the second ion accumulator for a period of time that is the same as or correlates with the period of time that their precursor ion species are transmitted by the mass filter.

[0016] For example, fragment or product ion species may accumulate in the second ion accumulator for a time period that is substantially the same as, or correlated to, the duration between the start time at which the voltage applied to the mass filter is switched to begin transmission of the precursor ion species and the end time at which the voltage is applied to the mass filter to end transmission of the precursor ion species.

[0017] The second ion accumulator may therefore be synchronized with the mass filter such that substantially all fragment or product ions derived from any given precursor ion species are accumulated in the second ion accumulator. This, together with the manner in which the second ion accumulator and the TOF mass analyser are synchronized, allows the duty cycle of the analyser to be relatively high.

[0018] It is also contemplated that each time the sequence of steps c) to e) is performed, the fragment or product ion species may be accumulated in the second ion accumulator for (only) a time shorter than the time their precursor ion species are transmitted by the mass filter. For example, the fragment or product ion species may accumulate in the second ion accumulator for a time shorter than the duration between the start time at which the voltage applied to the mass filter is switched to begin transmission of the precursor ion species and the end time at which the voltage is applied to the mass filter to end transmission of the precursor ion species.

[0019] Optionally, each time the sequence of steps c) to e) is performed, only fragment or product ion species derived from the selected precursor ion species, and optionally some ions of the selected precursor ion species itself, are accumulated in the second ion accumulator, while other precursor ion species and fragment or product ions derived therefrom are not accumulated in the second ion accumulator.

[0020] Thus, to improve duty cycle, embodiments of the present disclosure do not simply capture fragment or product ions and eject them into a TOF mass analyzer. Rather, the time scale on which the fragment or product ions are accumulated is set to match the time scale on which their precursor ion species are transmitted by the mass filter. The time scale on which the precursor ion species are transmitted by the mass filter may then be set to match the time scale on which the precursor ion species elutes from the separator. Thus, embodiments of the present disclosure provide a very high duty cycle.

[0021] Thus, each time the sequence of steps b) and c) is performed, the mass filter may be controlled to transmit the precursor ion species of interest only for a time that is the same as, correlated with, or shorter than the time that the precursor ion species elutes from the ion separator.

[0022] For example, the mass filter may be controlled to transmit the precursor species of interest for a time that is substantially the same as, correlates with, or is shorter than the duration between the start time that the precursor ion species begins to elute from the separator and the end time that the precursor ions stop eluting from the ion separator.

[0023] Step f) may involve ejecting the fragment or product ion species from the second ion accumulator in reverse order of mass-to-charge ratio, i.e. starting with ions of relatively high mass-to-charge ratio and ejecting progressively lower mass-to-charge ratio ions, such that different ones of the fragment or product ion species arrive at the pusher simultaneously and are pulsed simultaneously by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

[0024] A transport region may be provided between the second ion accumulator and the TOF mass analyzer, and the pressure in the transport region and the energy at which ions are ejected from the second ion accumulator are such that fragment or product ions having a relatively low mass-to-charge ratio overtake fragment or product ions having a higher mass-to-charge ratio such that the ions arrive at the pusher simultaneously and are pulsed simultaneously by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

[0025] Optionally, each time step f) is performed, substantially all of the fragment or product ion species released from the second ion accumulator arrive at the pusher substantially simultaneously and are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

[0026] Step f) may include substantially simultaneously ejecting all of the fragment or product ion species from the second ion accumulator, a transport region being provided between the second ion accumulator and a TOF mass analyzer, the ions separating in the transport region according to their second physicochemical property such that the fragment or product ion species arrive at the TOF mass analyzer at times dependent on the values ​​of their second physicochemical property, and operation of the TOF mass analyzer being synchronized with the ejection of ions from the second ion accumulator such that fragment or product ion species having values ​​within a selected range of the second physicochemical property value are simultaneously pulsed by a pusher electrode into a time-of-flight region of the TOF mass analyzer.

[0027] The method may include receiving, in a mass spectrometer performing the method, an input signal representing a selected range of values ​​of a second physicochemical property, and operation of the TOF mass analyzer is synchronized with the ejection of ions from the second ion accumulator such that fragment or product ion species having the selected range of values ​​of the second physicochemical property are simultaneously pulsed by a pusher electrode into a time-of-flight region of the TOF mass analyzer.

[0028] The second physicochemical property may be mass-to-charge ratio, such that ions are separated in the transport region according to their mass-to-charge ratio. For example, the operating conditions in the transport region may be such that fragment or product ion species having a relatively low mass-to-charge ratio travel faster than fragment or product ion species having a higher mass-to-charge ratio. For example, the product of the pressure P in the transport region and the length L of the ion flight path through the transport region (i.e., P×L) may be such that ions do not substantially collide with background gas molecules in the transport region.

[0029] Alternatively, the second physicochemical property may be ion mobility, whereby ions are separated in the transport region according to ion mobility. For example, the transport region may function as an ion mobility separator configured to drive ions through a background gas disposed therein. The ions may be driven through the background gas by an electric field to separate according to mobility. For example, the ions may be driven by applying a static DC potential gradient along the transport region and / or by repeatedly moving a DC potential along the transport region. The electric field may be configured to urge the ions in a downstream direction through the transport region. Alternatively, the background gas may be flowed in a downstream direction and the electric field may be configured to urge the ions in an upstream direction against the gas flow, causing the ions to separate according to mobility.

[0030] Steps a)-g) may be repeated during a single experimental run. For the avoidance of doubt, during a single experimental run means during continuous analysis of an analytical sample. For example, steps a)-g) may be repeated while ions are substantially continuously (e.g. substantially continuously or in a pulsed manner) generated from the analytical sample and / or during a single liquid or gas chromatography run.

[0031] The present disclosure also provides mass spectrometers configured to carry out the methods described herein.

[0032] Accordingly, a first aspect of the present disclosure also provides a mass spectrometer, the mass spectrometer comprising: a first ion accumulator; An ion separator; A mass filter; a fragmentation or reaction device; a second ion accumulator; and a TOF mass analyzer having a time-of-flight region and a pusher electrode; a control circuit for controlling the mass spectrometer; a) accumulating precursor ions in a first ion accumulator; b) performing a separation cycle including pulsing packets of precursor ions from the first ion accumulator into an ion separator and separating the precursor ions in the ion separator such that precursor ions having different values ​​of a first physicochemical property elute from the ion separator at different times; c) mass filtering the precursor ions eluting from the ion separator using a mass filter to transmit selected precursor ion species; d) fragmenting or reacting the selected precursor ion species in a fragmentation or reaction device to produce a set of fragment or product ion species therefrom; e) accumulating the set of fragment or product ion species in a second ion accumulator; f) ejecting the set of fragment or product ion species from the second ion accumulator into a TOF mass analyzer, the operation of the TOF mass analyzer being synchronized with the ejection of ions from the second ion accumulator such that multiple different species of the fragment or product ion species set are simultaneously pulsed by a pusher electrode into a time-of-flight region of the TOF mass analyzer; g) repeating steps c) to f) at least once during a separation cycle, the mass filter comprising a control circuit configured to control the selected precursor ion species such that the selected precursor ion species is controlled differently each time steps c) to f) are performed.

[0033] The control circuitry may be configured to control the mass analyzer such that step f) includes ejecting the fragment or product ion species from the second ion accumulator in reverse order of mass-to-charge ratio, i.e. starting with ions of relatively high mass-to-charge ratio and ejecting progressively lower mass-to-charge ratio ions, so that different ones of the fragment or product ion species arrive at the pusher simultaneously and are pulsed simultaneously by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

[0034] A transport region may be provided between the second ion accumulator and the TOF mass analyzer, the mass analyzer configured to maintain a pressure in the transport region and to eject ions from the second ion accumulator with an energy such that fragment or product ions having a relatively low mass-to-charge ratio overtake fragment or product ions having a higher mass-to-charge ratio such that the ions arrive simultaneously at a pusher and are simultaneously pulsed by a pusher electrode into a time-of-flight region of the TOF mass analyzer.

[0035] The mass spectrometer may comprise a transport region between the second ion accumulator and the TOF mass analyzer, and the control circuitry is configured to control the mass analyzer such that step f) includes substantially simultaneously ejecting all of the fragment or product ion species from the second ion accumulator into the transport region, causing the fragment or product ion species to separate in the transport region according to their second physico-chemical property and reach the TOF mass analyzer at times dependent on the values ​​of their second physico-chemical property, and operation of the TOF mass analyzer is synchronized with the ejection of ions from the second ion accumulator such that fragment or product ion species having values ​​in the selected range of the second physico-chemical property value are simultaneously pulsed by a pusher electrode into the time-of-flight region of the TOF mass analyzer.

[0036] The mass spectrometer may include a user interface configured to input a selected range of values ​​for the second physicochemical property, and the spectrometer is configured such that the control circuit controls operation of the TOF mass analyzer based on the selected range of values ​​to synchronize operation of the TOF mass analyzer with the release of ions from the second ion accumulator such that fragment or product ion species having the selected range of values ​​for the second physicochemical property are simultaneously pulsed by the pusher electrode into a time-of-flight region of the TOF mass analyzer.

[0037] The mass spectrometer comprises an ion source and may also comprise a liquid or gas chromatography separator for separating the analytes upstream of the ion source.

[0038] The mass spectrometer may be configured to repeat steps a)-g) during a single experimental run, for example while ions are being generated substantially continuously (e.g., substantially continuously or in a pulsed manner) by the ion source, and / or during a single liquid chromatography or gas chromatography separation run.

[0039] It is contemplated that the second ion accumulator may be omitted. In these embodiments, the second ion accumulator is not synchronized with the TOF mass analyzer to control which fragment or product ions (from any given precursor ion species) are simultaneously mass analyzed by the TOF mass analyzer. Rather, this is done by the operation of the TOF mass analyzer being synchronized with the time that the fragment or product ions are generated. Thus, the time that the mass filter transmits the selected precursor ion species to the fragmentation or reaction region is synchronized with the TOF mass analyzer to control which fragment or product ions from the selected precursor ion species are simultaneously mass analyzed.

[0040] Thus, from a second aspect, the present disclosure provides a method of mass spectrometry, the method comprising: a) accumulating precursor ions in a first ion accumulator; b) performing a separation cycle comprising pulsing packets of precursor ions from a first ion accumulator into an ion separator and separating the precursor ions such that precursor ions having different values ​​of a first physicochemical property elute from the ion separator at different times; c) switching a mass filter for mass filtering the precursor ions eluting from the ion separator to initiate transmission of selected precursor ion species; d) fragmenting or reacting the selected precursor ion species to produce a set of fragment or product ion species from the selected precursor ion species; e) transporting the set of fragment or product ion species through a transport region to a TOF mass analyzer having a pusher electrode and a time-of-flight region, where ions are separated in the transport region according to a second physicochemical property, the fragment or product ion species arrive at the TOF mass analyzer at times dependent on the values ​​of their second physicochemical property, and operation of the TOF mass analyzer is synchronized with the times at which the mass filters are switched to initiate transmission of the selected precursor ion species such that fragment or product ion species having a selected range of values ​​of the second physicochemical property that arrive substantially simultaneously at the pusher electrode are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer; f) repeating steps c) through e) at least once during a separation cycle, wherein the selected precursor ion species is different each time steps c) through e) are performed.

[0041] Fragment or product ion species of any given selected precursor ion species undergo a separation cycle in the transport region, which is triggered based on the time that the mass filter switches to begin transmitting that precursor ion species.

[0042] The method may include receiving, in a mass spectrometer performing the method, an input signal representative of a selected range of values ​​of a second physicochemical property, and then controlling operation of a TOF mass analyzer such that fragment or product ion species having the selected range of values ​​of the second physicochemical property arrive at a pusher electrode substantially simultaneously and are pulsed simultaneously by the pusher electrode into a time-of-flight region of the TOF mass analyzer.

[0043] Fragment or product ions of any given precursor ion species may arrive at the TOF mass analyzer at a time that correlates to the time the mass filter begins to transmit those precursor ion species. For example, selected precursor ion species transmitted in step c) may not be axially trapped prior to fragmentation or reaction in step d). Fragment or product ions produced in step d) may not be axially trapped prior to reaching the TOF mass analyzer.

[0044] The method according to the second aspect of the present disclosure may have any of the features described in relation to the first aspect of the present disclosure, except that it does not require the use of a second ion accumulator.

[0045] For example, each time step c) is performed, only the selected precursor ion species may be transmitted, and no other precursor ion species may be transmitted. Alternatively, each time step c) is performed, the mass filter may transmit precursor ions having a limited range of mass-to-charge ratios, such that multiple precursor ion species are simultaneously transmitted, while precursor ions having mass-to-charge ratios outside this limited range are filtered out. Step d) may then include simultaneously fragmenting or reacting the transmitted multiple precursor ion species to generate multiple respective sets of fragment or product ion species therefrom. Step e) may include transporting the multiple sets of fragment or product ion species through a transport region to a TOF mass analyzer, the operation of the TOF mass analyzer being synchronized with the time at which the mass filter is switched to begin transmitting precursor ion species, such that fragment or product ion species having a selected range of values ​​of the second physicochemical property arrive at the pusher electrode substantially simultaneously and are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer. Step g) may comprise repeating steps c) to f) at least once during a separation cycle, wherein the range of mass to charge ratios transmitted by the mass filter is different each time steps c) to f) are performed.

[0046] The present disclosure also provides a mass spectrometer configured to perform the method described in relation to the second aspect of the present disclosure.

[0047] Accordingly, the present disclosure provides a mass spectrometer, the mass spectrometer comprising: a first ion accumulator; An ion separator; A mass filter; a fragmentation or reaction device; a TOF mass analyzer having a time-of-flight region and a pusher electrode; a transfer region between the fragmentation or reaction device and the TOF mass analyzer; a control circuit for controlling the mass spectrometer; a) accumulating precursor ions in a first ion accumulator; b) performing a separation cycle including pulsing packets of precursor ions from the first ion accumulator into an ion separator and separating the precursor ions in the ion separator such that precursor ions having different values ​​of a first physicochemical property elute from the ion separator at different times; c) switching a mass filter that mass filters the precursor ions eluting from the ion separator to initiate transmission of selected precursor ion species; d) fragmenting or reacting the selected precursor ion species in a fragmentation or reaction device to produce a set of fragment or product ion species therefrom; e) transporting the set of fragment or product ion species through a transport region to a TOF mass analyzer where the ions are separated in the transport region according to the second physico-chemical property, the fragment or product ion species arriving at the TOF mass analyzer at times dependent on the values ​​of their second physico-chemical property, and operation of the TOF mass analyzer is synchronized with the time at which the mass filter is switched to initiate transmission of the selected precursor ion species such that fragment or product ion species having a selected range of values ​​of the second physico-chemical property that arrive substantially simultaneously at the pusher electrode are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer; f) a control circuit configured to repeat steps c) through e) at least once during a separation cycle such that the selected precursor ion species is different each time steps c) through e) are performed. [Brief description of the drawings]

[0048] Various embodiments of the present invention will now be described, by way of example only, with reference to the accompanying drawings, in which: [Figure 1] 1 shows a schematic diagram of an embodiment of a mass spectrometer according to the present disclosure. [Diagram 2]13A-13C represent an example of how a mass filter can be controlled as a function of time to selectively transmit different precursor ion species for different times. [Diagram 3] The same plot as in FIG. 2 is shown, except that the fragment or product ion species are overlaid. [Figure 4] An example of how fragment or product ions can be mass analyzed by a time-of-flight (TOF) mass analyzer is shown. [Diagram 5] 2 illustrates how the mass spectrometer of FIG. 1 can be operated according to one embodiment of the present disclosure. [Figure 6] FIG. 6 illustrates how the process described in relation to FIG. 5 can be repeated multiple times for different precursor ion species eluting from the ion separator during the same separation cycle. [Figure 7] 2 illustrates how the mass spectrometer of FIG. 1 can be operated according to another embodiment of the present disclosure. [Figure 8] FIG. 8 illustrates how the process described in relation to FIG. 7 can be repeated multiple times for different precursor ion species eluting from the ion separator during the same separation cycle. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0049] 1 shows a schematic diagram of an embodiment of a mass spectrometer according to the present disclosure. The spectrometer comprises an ion source 1, a first ion accumulator 2, an ion separator 3, a mass filter 4, a fragmentation or reaction device 5, a second ion accumulator 6, a transport region 7, and a time-of-flight (TOF) mass analyzer 8. It will be appreciated that other ion optical devices may also be provided within the spectrometer, such as, for example, one or more RF ion guides, electrostatic lenses, or collisionally cooled RF ion guides, which are configured to urge ions therethrough, such as by using an axial electric field or by moving a potential barrier therealong.

[0050] In use, ions are provided from an ion source 1, which may be an ion source generating ions from an analytical sample or an ion optical device providing ions. Ions from the ion source 1 may be accumulated in a first ion accumulator 2 and then released as ion packets into the separator 3. The ion accumulator 2 may accumulate ions and periodically pulse them into the separator 3. The accumulator 2 may be configured to allow ions to enter the accumulator 2 continuously or may prevent ions from entering the accumulator 2 when packets of ions are being pulsed into the separator 3. In either case, the accumulator 2 accumulates ions from the ion source 1 between the times that packets of ions are pulsed into the separator 3.

[0051] The separator 3 is controlled to cause each packet of ions received from the accumulator 2 to be separated according to one or more physico-chemical properties. For example, the separator 3 may separate ions according to ion mobility or mass to charge ratio, or a combination of both of these physico-chemical properties. The separated ions then pass from the separator 3 to a mass filter 4. The mass filter 4 is controlled to only allow a single mass to charge ratio (or a limited range of mass to charge ratios) to pass at any given time, and the single mass to charge ratio (or limited range) changes over time as the ions separated from the ion packet elute from the separator 3.

[0052] Figure 2 represents an example of how the mass filter 4 may be controlled as a function of time as ions separated from an ion packet elute from the separator 3. More specifically, Figure 2 shows a plot of the mass to charge ratio transmitted by the mass filter 4 as a function of time since an ion packet is pulsed into the separator 3. In this example, the mass filter 4 is controlled to transmit only ions having a first relatively low mass to charge ratio at a first time. The mass filter is then controlled at a second, later time to transmit only ions having a second, higher mass to charge ratio. The mass filter is then controlled at a third, later time to transmit only ions having a third, higher mass to charge ratio. The mass filter is then controlled at a fourth, later time to transmit only ions having a fourth, lower mass to charge ratio. The mass filter is then controlled at a fifth, later time to transmit only ions having a fifth, higher mass to charge ratio.

[0053] The mass filter 4 may be, for example, a resolving quadrupole mass filter, although it is contemplated that other types of mass filters may be used. It will also be appreciated that the mass filter 4 may be controlled to transmit different mass to charge ratios at different times in ways other than those shown in the example of Figure 2.

[0054] After the delay time, another packet of ions may be pulsed from the ion accumulator 2 into the separator 3 and the above process may be repeated to separate and filter the ions in this new packet. The delay time may be the time it takes for all of the ions of interest in one ion packet (i.e. what the mass filter 4 is set to transmit at one point in time) to pass through the separator 3 and reach the filter 4, but may be set to a shorter or longer delay time if desired. Any desired number of ion packets may be sequentially separated and then filtered in the manner described above.

[0055] The separated ions transmitted by the filter 4 (i.e., not rejected by the filter) are transferred to a fragmentation or reaction device 5. The fragmentation or reaction device 5 may be a Collision Induced Dissociation (CID) device, an Electron Capture Dissociation (ECD) device, an Electron Transfer Dissociation (ETD) device, a Surface Induced Dissociation (SID) device, or a device that fragments ions according to any other technique. For example, the device may react ions with reagent ions or molecules to fragment the ions, or the device may be used to react ions with reagent ions or molecules to form other types of product ions, such as adduct ions.

[0056] FIG. 3 shows the same plot as FIG. 2, except that the fragment or product ion species generated from the precursor ions are superimposed. In the illustrated example, a first precursor species transmitted by mass filter 4 is shown as a first dark oval, and fragments or reacts to generate two species of fragment or product ions, which are shown by lighter colored ovals above and below the first dark oval. In this example, one of the fragment or product ion species has a higher mass-to-charge ratio than the first precursor species, and the other has a lower mass-to-charge ratio. In this example, each of the second through fifth precursor species (represented by dark ovals) fragments or reacts to generate three species of fragment or product ions (represented by lighter ovals).

[0057] It should be understood that different precursor ions arrive at the fragmentation or reaction device 5 at different times due to their separation in the separator 3, and therefore different precursor ions fragment or react at different times to produce their fragment or product ions at different times. Although Figure 3 shows that the fragment or product ion species are produced over a time that precisely matches the time that their precursor ion species are transmitted by the filter 4, it should be understood that this may not be the case, and the time that any given fragment or product ion species is produced may deviate from the time that the precursor ion species was transmitted by the filter 4 due, for example, to the transmission time of the precursor ion species to (or through) the fragmentation or reaction device 5 and / or the time it takes for the precursor species to undergo fragmentation or reaction within the device 5.

[0058] The separation of precursor ion species induced by the separator 3 may be preserved among the fragment or product ion species derived from these precursor ion species. In particular, different precursor ion species arrive at the fragmentation or reaction device 5 at different times and thus their respective fragment or product ion species are generated at corresponding different times. The fragment or product ion species may be forced out through the fragmentation or reaction device 5 such that the fragment or product ion species derived from the different precursor species reach the second ion accumulator 6 at different respective times. Thus, fragment or product ion species derived from the same precursor species reach the second ion accumulator 6 over substantially the same time and are accumulated in the second ion accumulator.

[0059] The second ion accumulator 6 accumulates ions and periodically pulses them into the transport region 7. The second ion accumulator 6 may be configured to allow ions to enter it continuously, or may block ions from entering the accumulator 6 when a packet of ions is being pulsed into the transport region 7. In either case, the accumulator accumulates ions from the fragmentation or reaction device 5 during the period between the times that a packet of ions is pulsed from the accumulator 6 into the transport region 6. However, it is contemplated that the second ion accumulator 6 may be omitted and the fragmentation or reaction device 5 may instead pulse ions into the transport region 7, for example, by providing an ion gate between the fragmentation or reaction device 5 and the transport region 7. Alternatively, the second ion accumulator 6 and the fragmentation or reaction device 5 may be two parts of the same device.

[0060] The fragment or product ions are transferred through the transport region 7 to a TOF mass analyzer 8. As is well known in the art, the TOF mass analyzer 8 has a pusher that pulses ion packets into a time-of-flight region (e.g., a field-free region) and towards an ion detector. As the ions pass through the time-of-flight region, they separate according to their mass-to-charge ratio and then impinge on the ion detector. Thus, the separated ions arrive at the ion detector at different times, and the time at which an ion arrives at the detector is related to its mass-to-charge ratio. The mass-to-charge ratio of any given ion can be determined from the duration between the time the ion is pulsed into the time-of-flight region and the time it is detected by the ion detector. Thus, the TOF mass analyzer 8 can obtain data from the signal detected at the detector to determine the mass-to-charge ratio of the ions pulsed into the mass analyzer 8 and their intensities to form a mass spectrum.

[0061] The ions may be reflected by one or more ion mirrors between the pusher and the ion detector to provide a relatively long flight path through the time-of-flight region. This allows ions of different mass-to-charge ratios to be separated to a greater extent as they travel through the time-of-flight region, thus providing the TOF mass analyzer 8 with higher mass resolution. Thus, the TOF mass analyzer 8 may be a Multi-Reflecting TOF (MRTOF) mass analyzer that uses ion mirrors to reflect ions multiple times as they drift from the pusher towards the ion detector.

[0062] Fragment or product ions derived from any given species of precursor ions reach the TOF mass analyser 8 within a limited time period related to the time period when those precursor species exit the separator 3 .

[0063] FIG. 4 shows an example of how fragment or product ions are mass analyzed by a TOF mass analyzer 8 according to a conventional (non-encoded high frequency pulsing) approach. In this example, three fragment or product ions with three different mass-to-charge ratios are generated from a single precursor species in a fragmentation or reaction device 5 and then sent from a second ion accumulator 6 through a transport region 7 to a TOF mass analyzer 8 over the same period of time. The three fragment or product ions arrive at the pusher of the TOF mass analyzer 8 over a time window 9 shown in FIG. 4. The TOF mass analyzer 8 periodically pulses the pusher so that it repeatedly samples the fragment or product ions throughout the time window 9 and mass analyzes them. The times at which the TOF pusher is pulsed are indicated by vertical lines 10 on the x-axis. These multiple pulses 10 of the TOF pusher during the duration 9 cause multiple TOF mass spectra to be generated for fragment or product ions derived from each precursor ion.

[0064] It should be understood that while FIG. 4 shows only fragment or product ions from one of the precursor species transmitted by the filter 4, fragment or product ions derived from other precursor ion species transmitted by the filter 4 (at a later or earlier time, respectively, relative to said one of the species) will also be mass analyzed in a corresponding manner (at a later or earlier time, respectively) in the TOF mass analyzer 8. This conventional data acquisition scheme has proven useful, for example, as it allows precise measurements to be made in the separation time domain of the separator 3, thus ultimately allowing the determination of physicochemical properties such as ion mobility and collision cross section. This approach also allows the processing software to group together fragment or product ions that are common to the same precursor species.

[0065] However, the approach illustrated by FIG. 4 provides a relatively low sampling duty cycle for the TOF mass analyzer 8. This is because a relatively long duration is provided between any given pusher pulse 10 and the next pusher pulse, which allows time for the heaviest mass-to-charge ratio ions pushed by a given pusher pulse to arrive at the ion detector of the TOF analyzer 8 before another packet of ions is pushed towards the ion detector by the next pusher pulse. This means that the duration between pusher pulses 10 is equal to or greater than the flight time from pusher to detector of the largest mass-to-charge ratio desired to be detected. Thus, the sampling duty cycle decreases with the square root of the largest mass-to-charge ratio desired to be detected. TOF sampling duty cycles with such techniques are typically in the range of 0.1% to 50%. These values ​​are typical maximum duty cycles.

[0066] To improve the overall experimental duty cycle of the spectrometer, the transfer of ions between the various components of the spectrometer is synchronized.

[0067] Figure 5 shows how the mass spectrometer of Figure 1 can be operated in accordance with an embodiment of the present disclosure. The top part of Figure 5 shows a schematic of a portion of the spectrometer of Figure 1, and the bottom part of Figure 5 shows a plot illustrating how precursor species of ions are analyzed in the spectrometer. The plot shows examples of mass-to-charge ratios (y-axis) of ions present in different regions of the spectrometer shown, i.e., at different times (x-axis).

[0068] As described above in relation to FIG. 1, the separator 3 performs a separation cycle in which different ion species are pulsed from the first accumulator 2 into the separator 3 where they are separated according to their physicochemical properties and then eluted at different times according to their physicochemical property values. The ions eluted from the separator 3 are received by the mass filter 4. At any given time during the separation cycle of the ion separator 3, the mass filter 4 may be configured to transmit only a selected mass-to-charge ratio, or a selected range of mass-to-charge ratios, for example to transmit only a single precursor ion species. This is illustrated in FIG. 5 by the dark oval below the mass filter 4, which represents a first precursor ion species having a particular mass-to-charge ratio that is transmitted by the mass filter over a period of time. The precursor species is then transmitted to the fragmentation or reaction device 5 where it is fragmented or reacted to form fragment or product ions. This process is illustrated in FIG. 5 by the six ovals arranged in a row below the fragmentation or reaction device 5. The dark oval represents the precursor species and the five light ovals represent different species of fragment or product ions produced from the precursor species. It should be understood that fewer or more than five different species of fragment or product ions may be produced, and some of the precursor ions may or may not be present after fragmentation or reaction.

[0069] The fragment or product ions are then transferred to the second ion accumulator 6 during an ion accumulation period. After this ion accumulation period, different species of fragment or product ions are ejected from the second ion accumulator 6 at different times into the transport region 7. More specifically, the fragment or product ions are ejected from the ion accumulator 6 in reverse order of mass-to-charge ratio, i.e., the fragment or product ion species with the highest mass-to-charge ratio is ejected first, followed by fragment or product ion species with progressively lower mass-to-charge ratios. This is illustrated in FIG. 5 by the five fragment or product species shown below the transport region 7. The spacing of the five fragment or product ion species along the time axis indicates that the fragment or product ion species with the highest mass-to-charge ratio is ejected first, followed by fragment or product ion species with progressively lower mass-to-charge ratios at progressively slower respective times.

[0070] The manner in which the fragment or product ion species are released from the second ion accumulator 6 and the manner in which the transport region 7 is configured are selected such that at least some of the different fragment or product ion species arrive at the TOF mass analyzer 8 substantially simultaneously. This is illustrated in FIG. 5 by the row of five fragment or product ion species shown below the TOF mass analyzer 8. For example, different fragment or product species having substantially the same energy may be released from the ion accumulator 6 and the transport region 7 may be maintained at a relatively low pressure so that there are relatively few collisions between the ions and background gas as the ions travel to the TOF mass analyzer 8. It should be appreciated that under these conditions, the transport region 7 operates as a time-of-flight region in which ions having a relatively low mass-to-charge ratio travel faster than ions having a higher mass-to-charge ratio. Thus, fragment or product ion species having a relatively low mass-to-charge ratio that are released from the ion accumulator 6 after the heavier (i.e., having a higher mass-to-charge ratio) fragment or product ion species will catch up with the heavier fragment or product ion species and arrive at the TOF mass analyzer 8 substantially simultaneously with them. The pusher of the TOF mass analyzer 8 is synchronized with the release of ions from the ion accumulator 6 such that the pusher is pulsed when multiple fragment or product ions of different species arrive at the pusher of the TOF mass analyzer 8 simultaneously, thus providing a relatively high duty cycle for the TOF mass analyzer.

[0071] The second ion accumulator 6 may be a mass selective ion trap. The ion trap may be a 3D quadrupole field ion trap or a linear ion trap. Ions may be ejected from the ion trap into the transport region 7 and towards the TOF mass analyzer 8 in reverse order of mass to charge ratio. This may be achieved in a variety of ways, as known in the art. For example, the RF and / or DC voltages applied to the electrodes of the ion trap may be varied (e.g. scanned) over time to sequentially destabilize ions of decreasing mass to charge ratio in the ion trap and thus eject them into the transport region 7 and towards the TOF mass analyzer 8. Alternatively or additionally, an AC voltage may be applied to the electrodes of the ion trap to resonantly excite ions of a particular mass to charge ratio and eject them from the ion trap into the transport region 7 and towards the TOF mass analyzer 8. The frequency of this AC voltage may be varied (e.g. scanned) over time to sequentially eject ions of decreasing mass to charge ratio. For example, the accumulator 6 may mass selectively eject ions in order of decreasing mass-to-charge ratio, as described in US Pat. No. 6,770,872, which is incorporated herein by reference.

[0072] The inventors have recognized that the time scale over which the second ion accumulator 6 can accumulate fragment or product ions and release them in reverse order of mass to charge ratio is comparable to the time scale over which any given precursor ion elutes from the upstream separator 3. For example, in some embodiments, the second ion accumulator 6 accumulates and releases fragment or product ion species from any given precursor ion species over a period of about 1 ms, which is approximately the same period over which the precursor ion species elutes from the separator 3. Thus, fragment or product ion species derived from a first precursor ion species that elutes from the ion separator 3 during a given separation cycle of the separator 3 can be accumulated, captured, and released (in reverse order of mass to charge ratio) before a second precursor ion species elutes from the ion separator 3 within the same separation cycle of the separator 3. This is described in more detail in connection with FIG. 6. The mass spectrometer therefore has a relatively high duty cycle as precursor ions are fragmented and analyzed while other precursor ions remain delayed in the separator rather than being discarded.

[0073] FIG. 6 illustrates how the process described in relation to FIG. 5 can be repeated multiple times for different precursor ion species eluting from the separator 3 during the same separation cycle.

[0074] The lower plot in Figure 6 corresponds to the process described in relation to Figure 5, where a first precursor ion species eluting from separator 3 fragments or reacts and the resulting fragment or product ions accumulate in accumulator 6 and then are released into a TOF mass analyser 8 in reverse order of mass-to-charge ratio.

[0075] The central plot of FIG. 6 shows the same process being carried out for a second precursor ion species that elutes from the separator 3 at an earlier time, but during the same separator separation cycle as the first precursor ion species. More specifically, according to the central plot of FIG. 6, the mass filter 4 is set to transmit the second precursor ion species, represented by the dark oval in the central plot. The second precursor ion species then transmits to the fragmentation or reaction device 5, where it fragments or reacts to form fragment or product ions. This is shown by five light ovals arranged in a row with the dark ovals, but it should be understood that fewer or more than five different types of fragment or product ions may be produced. The fragment or product ions derived from the second precursor ion species then transport to a second ion accumulator 6, from which they are released in reverse order of mass-to-charge ratio, so that at least some of the different fragment or product ion species reach the TOF mass analyzer 8 substantially simultaneously.

[0076] The upper plot of FIG. 6 shows the same process being carried out for a third precursor ion species that elutes from the separator 3 at an earlier time, but during the same separator separation cycle as the first and second precursor ion species. More specifically, according to the upper plot of FIG. 6, the mass filter 4 is set to transmit the third precursor ion species, represented by the dark oval. The third precursor ion species then transmits to the fragmentation or reaction device 5 where it fragments or reacts to form fragment or product ions. This is shown by five light ovals arranged in a row with the dark ovals, although it should be understood that fewer or more than five different types of fragment or product ions may be produced. The fragment or product ions then transport to the second ion accumulator 6 and release therefrom in reverse order of mass to charge ratio, such that at least some of the different fragment or product ion species arrive at the TOF mass analyzer 8 substantially simultaneously.

[0077] The mass spectrometer may be configured to allow only fragment or product ions derived from any given precursor ion species to enter the accumulator 6 after all of the fragment or product ions derived from the previously analyzed precursor ion species have been ejected from the accumulator. This may be achieved, for example, by controlling the time at which the mass spectrometer switches the mass filter 4 to transmit said given precursor ion species at or after the accumulator 6 has ejected the lightest mass-to-charge ratio fragment or product ion derived from the preceding precursor ion species. Alternatively, an ion gate controlled by the mass spectrometer may be provided upstream of the accumulator 6 to prevent ions from entering the accumulator until the accumulator 6 has been controlled to eject all of the fragment or precursor ions in reverse mass-to-charge ratio order.

[0078] Although three precursor ion species are shown analyzed during a single separation cycle of separator 3, it is contemplated that only two precursor ion species, or more than three precursor ion species, may be analyzed sequentially in the manner described during a single separation cycle.

[0079] Furthermore, although the method has thus far only been described in the context of separator 3 performing a single separation cycle, it should be understood that multiple separation cycles may be performed during the same experimental run and the method described above may be repeated for each of these separation cycles during the experimental run.

[0080] The above-described methods of synchronizing the transfer of ions between various components of the spectrometer improve the overall experimental duty cycle of the spectrometer. For example, the mass filter 4 is synchronized with the ion separator 3 such that the mass filter 4 switches between transmitting different precursor ion species at different times during a single separation cycle of the separator 3. Precursor ions transmitted by the mass filter 4 are fragmented or reacted in the fragmentation or reaction device 5 and then passed to the second ion accumulator 6. The second ion accumulator 6 is synchronized with the mass filter 4 (and therefore with the ion separator 3) so that during a single separation cycle of the separator 3, (i) a first set of fragment or product ions derived from a first precursor ion species that elutes from the separator 3 and is transmitted by the mass filter 4 accumulate in the second ion accumulator 6 and are then released into the transport region 7, and (ii) a second set of fragment or product ions derived from a second precursor ion species that elutes from the separator 3 and is transmitted by the mass filter 4 accumulate in the second ion accumulator 6 and are then released into the transport region 7. The mass spectrometer therefore has a relatively high duty cycle as precursor ions are fragmented and analyzed whilst other precursor ions remain delayed in the separator 3 rather than being discarded. The TOF mass analyzer 8 may also be synchronized with the release of ions from the second ion accumulator 6 such that the pusher of the TOF mass analyzer is pulsed when multiple fragment or product ion species from the same precursor arrive at the pusher simultaneously, thus providing a relatively high duty cycle for the TOF mass analyzer.

[0081] Figure 7 illustrates another method according to the present disclosure which is the same as that described above in connection with Figures 5 and 6 up to the point where the fragment or product ions are released from the second ion accumulator 6. In the embodiment illustrated in Figure 7, different fragment or product ion species are released substantially simultaneously from the second ion accumulator 6 into the transport region 7. This is illustrated in Figure 7 by the five fragment or product species shown below the transport region 7.

[0082] The transport region 7 is configured to separate the fragment or product ion species according to their physicochemical properties. For example, ions may be separated in the transport region 7 according to their ion mobility or mass to charge ratio, or a combination of both of these physicochemical properties. For example, the transport region 7 may be maintained at a relatively low pressure so that collisions between the ions and the background gas therein are relatively few while the ions travel to the TOF mass analyzer 8. It should be appreciated that under these conditions, the transport region 7 operates as a time-of-flight region in which ions separate according to their mass to charge ratio. Thus, fragment or product ion species having a relatively low mass to charge ratio will arrive at the TOF mass analyzer 8 before ions having a higher mass to charge ratio. This is illustrated in FIG. 7 by the five fragment or product ion species shown below the TOF mass analyzer 8. The species are shown spaced apart on the time axis to show that the lowest mass to charge ratio fragment or product ion species arrives at the TOF mass analyzer 8 first, followed by the fragment or product ion species of progressively higher mass to charge ratios at progressively higher respective times.

[0083] Alternatively, the fragment or product ion species may be pulsed into the transport region 7 and separated according to their mobility through a background gas disposed in the transport region 7. The ions may be driven through the background gas by an electric field to separate according to mobility. For example, the ions may be driven by applying a static DC potential gradient along the transport region 7 and / or by repeatedly moving a DC potential along the transport region 7. The electric field may be arranged to urge the ions through the transport region 7 towards the TOF mass analyzer 8. Alternatively, the background gas may be flowed in a direction to urge the ions towards the TOF mass analyzer 8, while the electric field may be arranged to urge the ions against the gas flow to cause the ions to be separated according to mobility. In these mobility separation embodiments, the ions pulsed into the transport region 7 may arrive at the TOF mass analyzer 8 in order of high or low mobility.

[0084] In the embodiment described in relation to Figure 7, the transport region 7 acts as an ion separator performing a separation cycle in which different ion species are pulsed into the transport region 7, separated there according to their physico-chemical properties, and then eluted at different times according to their physico-chemical property values ​​into the mass analyzer 8. This separation cycle may be shorter than that of the ion separator 3.

[0085] After separating the ions in the transport region 7 according to their physicochemical properties, the ions may be transferred to an ion guide that guides the separated ions to the TOF mass analyzer 8 while maintaining their separation. For example, a voltage may be applied to the ion guide to create a series of potential wells (and thus potential barriers) spaced axially along the ion guide. The voltage may be a DC voltage, but it is contemplated that, alternatively, AC voltages may be used to form pseudo-potential wells. The voltages applied to the electrodes may be varied in time such that a series of potential wells travel from the upstream end of the ion guide along the ion guide to the TOF mass analyzer 8. As ions with different values ​​of the physicochemical property reach the upstream end of the ion guide at different times, ions with different ranges of values ​​of the physicochemical property become trapped in different potential wells and therefore travel in the wells to reach the TOF mass analyzer 8 at different times. The ion guide may operate in the same manner as the ion guides described, for example, in U.S. Pat. No. 7,829,841, which is incorporated herein by reference.

[0086] In the embodiment shown in FIG. 7, fragment or product ions having a limited range of physicochemical properties (according to which the transport region 7 separates the ions) arrive simultaneously at a pusher of the TOF mass analyzer 8, which may be synchronized with the release of ions from the second ion accumulator 6, such that the pusher pulses ions having the above selected range of physicochemical properties into the time-of-flight region of the TOF mass analyzer 8.

[0087] FIG. 8 illustrates how the process described in relation to FIG. 7 can be repeated multiple times for different precursor ion species eluting from the first ion separator 3 during the same separation cycle.

[0088] The lower plot in Figure 8 corresponds to the process described in relation to Figure 7, where a first precursor ion species eluting from the separator 3 fragments or reacts and the resulting fragment or product ions accumulate and then are released en masse into a TOF mass analyzer 8.

[0089] The central plot of FIG. 8 shows the same process being carried out for a second precursor ion species that is in the same separation cycle of the separator 3 as the first precursor ion species, but elutes from the separator 3 at an earlier time. More specifically, according to the central plot of FIG. 8, the mass filter 4 is set to transmit the second precursor ion species, represented by the dark oval in the central plot. The second precursor ion species then transmits into the fragmentation or reaction device 5 where it fragments or reacts to form fragment or product ions. This is shown by five light ovals arranged in a row with the dark ovals, but it should be understood that fewer or more than five different types of fragment or product ions may be produced. The fragment or product ions derived from the second precursor ion species are then transferred to a second ion accumulator 6 from which they are released en masse such that the different fragment or product ion species reach the TOF mass analyzer 8 at different times.

[0090] The upper plot of FIG. 8 shows the same process being carried out for a third precursor ion species that elutes from the separator 3 at an earlier time, but during the same separator separation cycle as the first and second precursor ion species. More specifically, according to the upper plot of FIG. 8, the mass filter 4 is set to transmit the third precursor ion species, represented by the dark oval. The third precursor ion species then transmits to the fragmentation or reaction device 5 where it fragments or reacts to form fragment or product ions. This is shown by the five light ovals arranged in a row with the dark ovals, although it should be understood that fewer or more than five different types of fragment or product ions may be produced. The fragment or product ions are then transferred to the second ion accumulator 6 from where they are released en masse such that the different fragment or product ion species reach the TOF mass analyzer 8 at different times.

[0091] The mass spectrometer may be configured to allow only fragment or product ions derived from any given precursor ion species to enter the second accumulator 6 after all fragment or product ions derived from the previously analyzed precursor ion species have been ejected from the accumulator 6. This may be achieved, for example, by controlling the time at which the mass spectrometer switches the mass filter 4 to transmit said given precursor ion species at or after the accumulator 6 has ejected fragment or product ions derived from the preceding precursor ion species. Alternatively, an ion gate controlled by the mass spectrometer may be provided upstream of the accumulator 6 to prevent ions from entering the accumulator 6 until the accumulator 6 has been controlled to eject all of its fragment or precursor ions.

[0092] Although three precursor ion species are shown analyzed during a single separation cycle of separator 3, it is contemplated that only two precursor ion species, or more than three precursor ion species, may be analyzed sequentially in the manner described during a single separation cycle.

[0093] Furthermore, although the method has thus far only been described in the context of separator 3 performing a single separation cycle, it should be understood that multiple separation cycles may be performed during the same experimental run and the method described above may be repeated for each of these separation cycles during the experimental run.

[0094] Although the present invention has been described with reference to preferred embodiments, it will be understood by those skilled in the art that various changes in form and detail may be made therein without departing from the scope of the invention as set forth in the appended claims.

[0095] For example, although embodiments in which a second ion accumulator 6 is present have been described with respect to Figures 7-8, it is contemplated that the second ion accumulator 6 may be omitted. In such an embodiment, each precursor ion species may be fragmented or reacted in the fragmentation or reaction device 5 without trapping the ions, and the resulting fragment or product ions generated are transported to the transport region 7 without being trapped. In such an embodiment, the fragment or product ions of any given precursor ion species enter the transport region 7 at a time that correlates with the time that the mass filter 4 begins to transmit the precursor ion species. Thus, the initiation of a separation cycle in the transport region 7 for any given fragment or product ion of a precursor ion species is triggered by the time that the mass filter 4 switches to begin transmitting that precursor ion species. As discussed above, fragment or product ions having a limited range of physicochemical properties arrive at the pusher of the TOF mass analyzer 8 at different times. The pusher may pulse ions having a selected range of physicochemical properties into the time-of-flight region of the TOF mass analyzer 8 in synchronization with the time that the mass filter 4 switches to transmit new precursor ion species.

[0096] Herein, an embodiment has been described in which the spectrometer includes a first ion accumulator 2 and / or a second ion accumulator 6. An ion attenuation device may be provided upstream of the first and / or second ion accumulators to attenuate ions traveling towards them. This may be used to reduce the number of ions entering the ion accumulators 2, 6, for example to reduce space charge effects in the ion accumulators and / or to prevent saturation of a detector in the TOF mass analyzer 8. Additionally or alternatively, an ion attenuation device may be provided downstream of the first and / or second ion accumulators 2, 6 to attenuate ions traveling downstream therefrom. This may be used to prevent saturation of a detector in the TOF mass analyzer 8, for example.

[0097] Although the mass filter 4 has been described as being stepped between different discrete mass transmission windows as ions elute from the separator 3, it is contemplated that the mass filter 4 may instead have a mass transmission window that is progressively scanned. For example, the mass transmission window may increase one mass-to-charge ratio unit at a time. Alternatively, the mass filter 4 may have a mass transmission window that is configured to be capable of simultaneously transmitting a range of mass-to-charge ratios (e.g., a range of multiple Da). This mass transmission window may be progressively scanned such that each time the mass transmission window moves to transmit a new mass range, the new mass range still partially overlaps with the previous mass range that the filter 4 was configured to transmit.

[0098] Alternatively or additionally, although filter 4 has been described as a mass-to-charge ratio filter, it is contemplated that the filter may filter ions according to another physicochemical property, such as ion mobility or FAIMS.

[0099] Alternatively or additionally, although separator 3 has been described as an ion mobility separator, it is contemplated that the separator may separate ions according to another physicochemical property, such as mass-to-charge ratio.

[0100] In less preferred embodiments, it is contemplated that the first ion accumulator 2 may be omitted and the ion source 1 may instead pulse ions into the separator 3, for example, by being a pulsed ion source or by providing an ion gate between the ion source 1 and the separator 3.

Claims

1. 1. A method of mass spectrometry comprising: a) accumulating precursor ions in a first ion accumulator; b) performing a separation cycle comprising pulsing packets of precursor ions from the first ion accumulator to an ion separator and separating the precursor ions such that precursor ions having different values ​​of a first physicochemical property elute from the ion separator at different times; c) mass filtering the precursor ions eluting from the ion separator to transmit selected precursor ion species; d) fragmenting or reacting the selected precursor ion species to produce a set of fragment or product ion species from the selected precursor ion species; e) accumulating the set of fragment or product ion species in a second ion accumulator; f) ejecting the set of fragment or product ion species from the second ion accumulator into a TOF mass analyzer, the operation of the TOF mass analyzer being synchronized with the ejection of ions from the second ion accumulator such that multiple different species of the set of fragment or product ion species are simultaneously pulsed by a pusher electrode into a time-of-flight region of the TOF mass analyzer; g) repeating steps c) through f) at least once during the separation cycle, wherein the selected precursor ion species is different each time steps c) through f) are performed; The method includes:

2. The method of claim 1 , wherein the ion separator is an ion mobility separator and the physicochemical property is ion mobility.

3. the fragment or product ion species are energized into the second ion accumulator; (i) fragment or product ion species derived from the same precursor ion species arrive at the second ion accumulator over substantially the same period of time and are accumulated in the second ion accumulator during at least a portion of this period; and / or 3. The method of claim 1 or 2, wherein (ii) fragment or product ion species derived from precursor species transmitted by the mass filter at different times reach the second ion accumulator over different respective time periods.

4. 4. The method of claim 1, wherein each time the sequence of steps c) to e) is performed, fragment or product ion species are accumulated in the second ion accumulator for a period of time that is the same as or correlates with the period of time that their precursor ion species are transmitted by a mass filter.

5. 5. The method according to claim 1, wherein each time the sequence of steps c) to e) is performed, only fragment or product ion species derived from the selected precursor ion species, and optionally some ions of the selected precursor ion species itself, are accumulated in the second ion accumulator, while other precursor ion species and fragment or product ions derived therefrom are not accumulated in the second ion accumulator.

6. 6. A method according to any one of claims 1 to 5, wherein each time the sequence of steps b) and c) is performed a mass filter is controlled to transmit precursor ion species of interest only for a period of time that is the same as, correlated with or shorter than the period of time that the precursor ion species elutes from the ion separator.

7. 7. The method of claim 1, wherein step f) comprises ejecting the fragment or product ion species from the second ion accumulator in reverse order of mass to charge ratio, i.e. starting with ions of relatively high mass to charge ratio and ejecting progressively lower mass to charge ratio ions, such that different ones of the fragment or product ion species arrive at a pusher simultaneously and are pulsed simultaneously into the time of flight region of the TOF mass analyzer by a pusher electrode.

8. 8. The method of claim 7, wherein a transport region is provided between the second ion accumulator and the TOF mass analyzer, and the pressure in the transport region and the energy at which ions are ejected from the second ion accumulator are such that fragment or product ions having a relatively low mass-to-charge ratio overtake fragment or product ions having a higher mass-to-charge ratio such that the ions arrive at the pusher simultaneously and are pulsed simultaneously by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

9. 9. The method of claim 7 or 8, wherein each time step f) is performed, substantially all of the fragment or product ion species ejected from the second ion accumulator arrive at the pusher substantially simultaneously and are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

10. 7. The method of claim 1, wherein step f) comprises ejecting all of the fragment or product ion species from the second ion accumulator substantially simultaneously, a transport region is provided between the second ion accumulator and the TOF mass analyzer, ions separate in the transport region according to a second physico-chemical property such that the fragment or product ion species arrive at the TOF mass analyzer at times dependent on the value of their second physico-chemical property, and operation of the TOF mass analyzer is synchronized with the ejection of ions from the second ion accumulator such that fragment or product ion species having a selected range of values ​​of the second physico-chemical property are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

11. 11. The method of claim 10, further comprising receiving an input signal representative of the selected range of values ​​of the second physicochemical property in a mass spectrometer performing the method, wherein the operation of the TOF mass analyzer is synchronized with the ejection of the ions from the second ion accumulator such that fragment or product ion species having the selected range of values ​​of the second physicochemical property are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

12. 12. The method of claim 10 or 11, wherein the second physicochemical property is mass-to-charge ratio and ions are separated in the transport region according to their mass-to-charge ratio.

13. 12. The method of claim 10 or 11, wherein the second physicochemical property is ion mobility, and ions are separated in the transport region according to their ion mobility.

14. The method according to any one of claims 1 to 13, wherein steps a) to g) are repeated during a single experimental run.

15. 1. A mass spectrometer comprising: a first ion accumulator; An ion separator; A mass filter; a fragmentation or reaction device; a second ion accumulator; and a TOF mass analyzer having a time-of-flight region and a pusher electrode; A control circuit for controlling the mass spectrometer, a) accumulating precursor ions in the first ion accumulator; b) performing a separation cycle comprising pulsing packets of precursor ions from the first ion accumulator into the ion separator and separating the precursor ions in the ion separator such that precursor ions having different values ​​of a first physicochemical property elute from the ion separator at different times; c) mass filtering the precursor ions eluting from the ion separator using the mass filter to transmit selected precursor ion species; d) fragmenting or reacting the selected precursor ion species in the fragmentation or reaction device to produce a set of fragment or product ion species therefrom; e) accumulating the set of fragment or product ion species in the second ion accumulator; f) ejecting the set of fragment or product ion species from the second ion accumulator into the TOF mass analyzer, the operation of the TOF mass analyzer being synchronized with the ejection of ions from the second ion accumulator such that multiple different species of the set of fragment or product ion species are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer; g) repeating steps c) to f) at least once during a separation cycle, the mass filter being configured to control the selected precursor ion species differently each time steps c) to f) are performed.

16. 16. The mass spectrometer of claim 15, wherein the control circuitry is configured to control the mass spectrometer such that step f) comprises ejecting the fragment or product ion species from the second ion accumulator in reverse order of mass to charge ratio, i.e. starting with ions of relatively high mass to charge ratio and ejecting progressively lower mass to charge ratio ions, such that different ones of the fragment or product ion species arrive at a pusher simultaneously and are pulsed simultaneously into the time of flight region of the TOF mass analyzer by a pusher electrode.

17. 17. The mass spectrometer of claim 16, comprising a transfer region between the second ion accumulator and the TOF mass analyzer, the mass spectrometer configured to maintain a pressure in the transfer region and to eject ions from the second ion accumulator with an energy such that fragment or product ions having a relatively low mass to charge ratio overtake fragment or product ions having a higher mass to charge ratio such that the ions arrive at the pusher simultaneously and are pulsed simultaneously by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

18. 16. The mass spectrometer of claim 15, further comprising a transfer region between the second ion accumulator and the TOF mass analyzer, and wherein the control circuit is configured to control the mass analyzer such that step f) comprises substantially simultaneously ejecting all of the fragment or product ion species from the second ion accumulator into the transfer region, causing the fragment or product ion species to separate in the transfer region according to a second physico-chemical property and reach the TOF mass analyzer at times dependent on the value of their second physico-chemical property, and operation of the TOF mass analyzer is synchronized with the ejection of ions from the second ion accumulator such that fragment or product ion species having a selected range of values ​​of the second physico-chemical property are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

19. 20. The mass spectrometer of claim 18, further comprising a user interface configured to input a selected range of values ​​for the second physicochemical property, wherein the control circuitry is configured to control operation of the TOF mass analyzer based on the selected range of values ​​to synchronize the operation of the TOF mass analyzer with the ejection of the ions from the second ion accumulator such that fragment or product ion species having the selected range of values ​​for the second physicochemical property are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

20. 1. A method of mass spectrometry comprising: a) accumulating precursor ions in a first ion accumulator; b) performing a separation cycle comprising pulsing packets of precursor ions from the first ion accumulator to an ion separator and separating the precursor ions such that precursor ions having different values ​​of a first physicochemical property elute from the ion separator at different times; c) switching a mass filter that mass filters the precursor ions eluting from the ion separator to begin transmitting selected precursor ion species; d) fragmenting or reacting the selected precursor ion species to produce a set of fragment or product ion species from the selected precursor ion species; e) transporting the set of fragment or product ion species through a transport region to a TOF mass analyzer having a pusher electrode and a time-of-flight region, where ions are separated in the transport region according to a second physico-chemical property, fragment or product ion species arrive at the TOF mass analyzer at times dependent on their second physico-chemical property values, and operation of the TOF mass analyzer is synchronized with the time at which the mass filter is switched to initiate transmission of the selected precursor ion species such that fragment or product ion species having a selected range of values ​​of the second physico-chemical property that arrive substantially simultaneously at the pusher electrode are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer; f) repeating steps c) through e) at least once during said separation cycle, wherein said selected precursor ion species is different each time steps c) through e) are performed; The method includes:

21. 21. The method of claim 20, comprising: receiving, in a mass spectrometer performing the method, an input signal representative of the selected range of values ​​of the second physicochemical property; and then controlling operation of the TOF mass analyzer such that fragment or product ion species having the selected range of values ​​of the second physicochemical property arrive at the pusher electrode substantially simultaneously and are pulsed simultaneously by the pusher electrode into the time-of-flight region of the TOF mass analyzer.

22. 1. A mass spectrometer comprising: a first ion accumulator; An ion separator; A mass filter; a fragmentation or reaction device; a TOF mass analyzer having a time-of-flight region and a pusher electrode; a transfer region between the fragmentation or reaction device and the TOF mass analyzer; a control circuit for controlling the mass spectrometer to: a) accumulating precursor ions in the first ion accumulator; b) performing a separation cycle comprising pulsing packets of precursor ions from the first ion accumulator into the ion separator and separating the precursor ions in the ion separator such that precursor ions having different values ​​of a first physicochemical property elute from the ion separator at different times; c) switching a mass filter that mass filters the precursor ions eluting from the ion separator to initiate transmission of selected precursor ion species; d) fragmenting or reacting the selected precursor ion species in the fragmentation or reaction device to produce a set of fragment or product ion species therefrom; e) transporting the set of fragment or product ion species through a transport region to a TOF mass analyzer where ions are separated in the transport region according to a second physico-chemical property, the fragment or product ion species arriving at the TOF mass analyzer at times dependent on their second physico-chemical property values, operation of the TOF mass analyzer being synchronized with the time that the mass filter is switched to initiate transmission of the selected precursor ion species such that fragment or product ion species having a selected range of values ​​of the second physico-chemical property that arrive at the pusher electrode substantially simultaneously are simultaneously pulsed by the pusher electrode into the time-of-flight region of the TOF mass analyzer; f) repeating steps c) through e) at least once during said separation cycle, wherein said selected precursor ion species is different each time steps c) through e) are performed; A control circuit configured as follows: A mass spectrometer comprising:

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