Interfacing capillary electrophoresis to a mass spectrometer via an impactor spray ionization source

Abstract

A mass spectrometer is disclosed comprising a separation device arranged and adapted to emit an eluent over a period of time. The separation device preferably comprises a Capillary Electrophoresis (CE) separation device. The mass spectrometer further comprises a nebuliser and a target. Eluent emitted by the separation device is nebulised, in use, by the nebuliser wherein a stream of analyte droplets are directed to impact upon the target so as to ionise the analyte to form a plurality of analyte ions.

Claims

1. A mass spectrometer comprising: a separation device arranged and adapted to emit an eluent over a period of time, wherein said separation device comprises either: (i) a Capillary Electrophoresis (CE) separation device; (ii) a Capillary Electrochromatography (CEC) separation device; (iii) a substantially rigid ceramic-based multilayer microfluidic substrate (ceramic tile) separation device; or (iv) a supercritical fluid chromatography separation device; a nebuliser and; a target; wherein said eluent emitted by said separation device is nebulised, in use, by said nebuliser wherein a stream of analyte droplets are directed to impact upon said target so as to ionise said analyte to form a plurality of analyte ions.

2. A mass spectrometer as claimed in claim 1, wherein the substantially rigid ceramic-based multilayer microfluidic substrate separation device comprises a High-Temperature Co-fired Ceramic (HTCC).

3. A mass spectrometer as claimed in claim 1, wherein the nebuliser is constructed with a tri-axial probe arrangement.

4. A mass spectrometer as claimed in claim 1, wherein said separation device comprises or is coupled to a first tube.

5. A mass spectrometer as claimed in claim 4, wherein said first tube comprises a capillary tube.

6. A mass spectrometer as claimed in claim 4, wherein said first tube is surrounded by a second tube which is arranged and adapted to provide a flow of liquid which mixes with the eluent emerging from the exit of said first tube.

7. A mass spectrometer as claimed in claim 6, wherein said second tube comprises a capillary tube.

8. A mass spectrometer as claimed in claim 7, further comprising a third tube which is arranged and adapted to provide a stream of gas to the exit of said first tube or said second tube.

9. A mass spectrometer as claimed in claim 8, wherein said third tube comprises a capillary tube.

10. A mass spectrometer as claimed in claim 8, wherein said third tube surrounds said second tube or is concentric with said first and second tubes.

11. A mass spectrometer as claimed in claim 8, wherein said third tube is non-concentric with said first and said second tubes.

12. A mass spectrometer as claimed in claim 1, wherein said target comprises one or more mesh or grid targets.

13. A method of mass spectrometry comprising: providing a separation device arranged and adapted to emit an eluent over a period of time, wherein said separation device comprises either: (i) a Capillary Electrophoresis (CE) separation device; (ii) a Capillary Electrochromatography (CEC) separation device; (iii) a substantially rigid ceramic-based multilayer microfluidic substrate (ceramic tile) separation device; or (iv) a supercritical fluid chromatography separation device; providing a target; and nebulising said eluent emitted by said separation device wherein a stream of analyte droplets are directed to impact upon said target so as to ionise said analyte to form a plurality of analyte ions.

14. A method of mass spectrometry as claimed in claim 13, further comprising delivering a make-up flow of liquid that mixes with the liquid flow from the separation device.

15. A method of mass spectrometry as claimed in claim 13, wherein a flow of liquid from the separation device enters a nebuliser probe and is delivered to a sprayer tip via a first tube.

16. A method of mass spectrometry as claimed in claim 15, wherein the first tube is surrounded by a second tube which delivers a make-up flow of liquid which mixes with the flow from the first tube at the probe tip.

17. A method of mass spectrometry as claimed in claim 16, wherein the second tube is surrounded by a third tube which includes a gas inlet to deliver a stream of high velocity gas to the exit of the first and second tubes.

18. A mass spectrometer comprising: a separation device arranged and adapted to emit an eluent over a period of time; a nebuliser and; a target; wherein said eluent emitted by said separation device is nebulised, in use, by said nebuliser wherein a stream of analyte droplets are directed to impact upon said target so as to ionise said analyte to form a plurality of analyte ions; and wherein said mass spectrometer further comprises a control system, wherein said control system is arranged and adapted either: (i) to switch the polarity of said target during a single experimental run; or (ii) to repeatedly switch the polarity of said target during a single experimental run.

19. A method of mass spectrometry comprising: providing a separation device arranged and adapted to emit an eluent over a period of time; providing a target; and nebulising said eluent emitted by said separation device wherein a stream of analyte droplets are directed to impact upon said target so as to ionise said analyte to form a plurality of analyte ions; wherein said method further comprises providing a control system, wherein said control system is arranged and adapted either: (i) to switch the polarity of said target during a single experimental run; or (ii) to repeatedly switch the polarity of said target during a single experimental run.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) Various embodiments of the present invention will now be described, by way of example only, and with reference to the accompanying drawings in which:

(2) FIG. 1 shows an impactor spray API source according to an embodiment of the present invention; and

(3) FIG. 2A shows an impactor spray source and FIG. 2B shows an optimised impactor spray source.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENT

(4) FIG. 1 is a schematic of the general layout of an impactor spray API source according to a preferred embodiment. A flow of liquid from a CE column outlet (or other separation device) enters a nebuliser probe 1 and is delivered to a sprayer tip 2 via an inner capillary tube 3. The inner capillary 3 is surrounded by a second concentric capillary 4 which delivers a make-up flow of liquid which mixes with the flow from the first capillary 3 at the probe tip. The second capillary tube 4 is surrounded by a third concentric capillary 5 which includes a gas inlet 6 to deliver a stream of high velocity gas to the exit of the liquid capillaries 3,4.

(5) This arrangement produces a nebulised spray which contains droplets with a typical diameter of 10-20 m and velocities greater than 100 m/s at a close distance from the sprayer tip 2. The resulting droplets are heated by an additional flow of gas that enters a concentric annular heater 7 via a second gas inlet 8.

(6) The sprayer is preferably hinged to the right hand side of an ion inlet cone 9 of the mass spectrometer and can swing to vary the horizontal distance between the sprayer tip 2 and an ion inlet orifice 10 of a mass spectrometer. The probe is also configured such that the vertical distance between the sprayer tip 2 and the ion inlet orifice 10 can be varied. The relative tip positions of the inner capillary 3, the second capillary 4 and the third capillary 5 can be adjusted. According to an embodiment the capillaries 3,4,5 may be arranged so that they are flush with one another. According to another embodiment the capillaries 3,4,5 may be arranged so that one or more capillaries 3,4,5 protrude or are recessed relative to each other.

(7) A target 11 with a similar dimension to the liquid capillary is preferably placed between the sprayer tip 2 and the ion inlet orifice 10. The target 11 can be manipulated in the x and y directions (in the horizontal plane) via a micro adjuster stage and can be held at a potential of typically 0-5 kV relative to the source enclosure 12 and the ion inlet orifice 10. In operation, the ion inlet cone 10 is surrounded by a metal cone gas housing 13 that is flushed with a low flow of nitrogen gas that enters via a gas inlet 14. All gasses that enter the source enclosure must leave via the source enclosure exhaust 15 or the ion inlet orifice 10 which is pumped by the first vacuum stage 16 of the mass spectrometer.

(8) FIG. 2A is a schematic plan view of an impactor spray source with the grounded nebuliser probe omitted from the diagram. The impactor target 11 comprises a stainless steel rod or pin with an outside diameter of typically 1-2 mm. The rod or pin 11 is positioned at a horizontal distance X.sub.1 of typically 5 mm from the ion inlet orifice 10. The probe tip can be finely adjusted to sweep across the target surface until the optimum impact point is found that gives the greatest sensitivity. A typical optimized position is shown in the schematic of FIG. 2B where the offset X.sub.2 is approximately 0.4 mm.

(9) FIG. 2B also shows the vertical positions of the probe and target in the preferred embodiment, i.e. Z.sub.1=9 mm and Z.sub.2=3 mm.

(10) In the preferred embodiment, the source is operated with the following bias potentials: nebuliser=0V, impactor target=1.0 kV, ion inlet cone=100 V and cone gas housing=100 V. The heater assembly and source enclosure are preferably maintained at ground potential. The source may be operated with the following gas flow settings: nitrogen nebulizer gas pressurized to 7 bar, nitrogen heater gas flow=1200 L/hr and nitrogen cone gas flow=150 L/hr.

(11) The preferred embodiment can be used in other applications that are similarly simplified by the use of a grounded nebuliser probe such as capillary electrochromatography (CEC) and tile-based microchip LC/MS systems.

(12) The tile-based microchip LC system preferably comprises a substantially rigid ceramic-based multilayer microfluidic substrate also referred to as a ceramic tile. Reference is made to US 2009/032135 the contents of which are incorporated herein by reference. For a protein sample the ceramic may comprise a High-Temperature Co-fired Ceramic (HTCC) which provides suitably low levels of loss of sample due to attachment of sample to walls of conduits in the substrate. Formed in the layers of the substrate is a channel that operates as a separation column. Apertures in the side of the substrate provide openings into the channel through which fluid may be introduced into the column. Fluid passes through the apertures under high pressure and flows toward the Electrospray emitter coupled at the egress end of the channel. Holes in the side of a microfluidic cartridge provide fluidic inlet ports for delivering the fluid to the substrate. Each fluidic inlet port aligns with and encircles one of the fluidic apertures.

(13) The preferred embodiment may also be implemented as an interface for supercritical fluid chromatography/MS.

(14) Impaction-based spray using a target pin has been shown to provide improved ionization efficiency for both polar and non-polar compounds compared to standard ESI or APCI. However, the performance with different mobile phase compositions has sometimes been observed to have a reasonably strong dependence upon the physical geometry of the probe and pin.

(15) The positional dependence of the probe and pin on the relative performance at high organic mobile phase can make achieving required tolerances problematic. Furthermore, maintaining these tolerances can also be problematic since the pin and/or probe capillary may need to be replaced one or more times during the lifetime of the instrument.

(16) According to an embodiment of the present invention a grid or mesh target is preferably used instead of a pin target. A grid or mesh target having a grid or mesh impaction surface has been found to be particularly advantageous compared with using a pin target in that utilising a grid or mesh target solves the problem of positional dependence which may otherwise be experienced when using a solid pin as the target.

(17) A mesh or grid target of appropriate size is preferably used as the impact target. According to the preferred embodiment the impact zone (i.e. the diameter of the plume at point of impact with the target) is preferably 0.5-1.0 mm.

(18) According to the preferred embodiment the mesh wire size and spacing is preferably sized appropriately so as to provide several discrete impact zones within the impact zone or area. The wire diameter is preferably sufficient so as to allow the impact of the plume on the wire to improve nebulisation. A mesh with 150 m spacing and a wire diameter of 100 m has been found to be particularly advantageous. However, other aspect ratios are also contemplated and are intended to fall within the scope of the present invention. According to an embodiment the mesh or grid may comprise a substantially flat rectangle (15 mm7 mm) and may be held substantially perpendicular to the spray axis. According to this embodiment the spray is essentially through the mesh or grid.

(19) Alternatively, the mesh or grid may be angled relative to the spray axis. The angle of the mesh or grid may be set such that the plume as it passes through the mesh or grid is deflected close to or in the direction of the mass spectrometer inlet. The mesh or grid target may be arranged at an angle of 70 relative to the spray axis.

(20) The physical dimensions of the mesh or grid are preferably set or arranged so that liquid beading on the surface of the mesh or grid is preferably minimized. The angle and shape of the mesh or grid may be optimised to reduce liquid beading.

(21) According to the preferred embodiment a high voltage may be applied to the mesh or grid electrode in order to assist ionization in a similar manner to other embodiments of the present invention which have been described above and which utilise a pin target. According to an embodiment the mesh or grid may be maintained at a potential of 1 kV. However, it will be apparent to those skilled in the art that the mesh or grid target may be maintained at other potentials.

(22) A particular advantage of using a mesh or grid target is that the mesh or grid target according to the preferred embodiment shows a significantly reduced dependence on positional geometry since the stream of droplets impacts upon multiple impaction points on the mesh or grid target. As the probe or mesh target is moved, the characteristics of the impact of the droplets upon the target remain substantially the same. Accordingly, the performance of the ion source relative to the position of the MS inlet and the probe behaves in a similar manner to an Electrospray ionisation (ESI) ion source relative to an ion inlet.

(23) Further embodiments are also contemplated. For example, a grid instead of a mesh may be used. The grid preferably has multiple impaction points in the zone in which the stream of droplets impacts upon the target. If positional dependence of the spray direction after impact is required then a single-row grid may be utilised.

(24) According to an embodiment the target may comprise multiple layers of meshes and/or grids in order to achieve the same effect as angling a single layered mesh or grid target.

(25) According to an embodiment the surface ionization impactor bar or target as described above may be further enhanced by utilising a piezoelectric vibration device to vibrate the bar or target. Vibration of the bar or target upon which the surface ionization occurs aids in the reduction of the size of the secondary droplets, increasing the evaporation rate of the solvent and thereby aids signal response.

(26) According to a preferred embodiment an impactor bar or target is located within a source enclosure. In this configuration the capillary is preferably grounded and potentials are preferably applied to the impactor bar or target and to the sample cone inlet structure. The integration of an impactor spray with a separation device introduces the potential for the generation of non-polar, highly polar, singularly charged and/or multiply charged gas phase ions for introduction into the mass spectrometer for analysis. The ionization processes and flow dynamics may, however, be different which can result in the formation of larger sized droplets. The use of piezoelectric vibration applied to the impactor bar or target is particularly advantageous in that it aids in the reduction of resultant secondary droplets.

(27) It will be understood by those skilled in the art that the mechanisms of droplet production in pneumatically assisted nebulisation are non-trivial and it cannot be approximated by a particular model for which the boundary conditions are known. There is no single process that is believed to be solely responsible for droplet production and the initial spray produced is rapidly modified by secondary fragmentation and by recombination and coalescence. The use of piezoelectric vibration applied to the impactor bar or target preferably aids in the reduction of the resultant secondary droplets through surface disruption.

(28) According to an embodiment a target pin is preferably utilised which is preferably rotated on e.g. an eccentric path so as to obtain an easily reproducible level of ion signal. According to an embodiment a target pin or rod is preferably placed or mounted off axis on a rotating shaft. The pin or rod target is preferably located or arranged so as to be in the path of high velocity droplets emitted from a sprayer. The droplets emitted from the sprayer are arranged to impact upon the pin or rod target so as to produce ions for analysis by mass spectrometry. The rotational position of the pin or rod is preferably controlled through a motor under computer control.

(29) According to an embodiment the analyte signal may be monitored with respect to the position of the pin or rod. The pin or rod may then be rotated or otherwise set to a particular position under computer control in order to maximise the signal intensity. Other embodiments are also contemplated wherein the pin or rod may be rotated between one or more different rotational positions in order to control the intensity of analyte ions produced or to control the efficiency of analyte ion production.

(30) The central longitudinal axis of the pin or rod is preferably arranged so as to be off centre relative to the central longitudinal axis of the rotating shaft. The position of the pin or rod may according to an embodiment vary by approximately 0.7 mm during the course of one rotation of the pin or rod target.

(31) According to a less preferred embodiment the position of the pin or rod target 10 may be translated rather than rotated (or the pin or rod target may be translated in addition to being rotated).

(32) As will be understood by those skilled in the art the positioning of the target is important in order to obtain an acceptable level of signal intensity when generating ions by impacting high velocity droplets onto the target. According to a particularly preferred embodiment causing the target to rotate on an eccentric path relative to the spray of high velocity droplets enables an average signal intensity to be realised. As a result, the overall or average ion signal can be stabilised and is less susceptible to wide variations in the intensity of analyte ions generated depending upon the precise position of the target relative to the high velocity spray of droplets.

(33) 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 without departing from the scope of the invention as set forth in the accompanying claims.