Apparatus and method for mass spectrometry

Abstract

A nebuliser apparatus for use in a sample injection system for mass spectrometry, the nebuliser apparatus comprising: a sample channel configured to pass a sample solution to a sample and atomising fluid outlet; an atomising fluid channel configured to pass an atomising fluid to the sample and atomising fluid outlet and configured to atomise the sample solution; and a sheath fluid channel configured to pass a sheath fluid to a sheath fluid outlet and configured to form the sheath fluid in a substantially annular profile encircling the atomised sample solution

Description

[0001] APPARATUS AND METHOD FOR MASS SPECTROMETRY

[0002] Technical Field

[0003] The present disclosure relates to mass spectrometry. More particularly, the present disclosure relates to an apparatus and method of use thereof for nebulisation and delivery of samples.

[0004] Background

[0005] Analytical chemistry often relies on separating complex mixtures into simpler fractions based on inherent analyte properties, which simplifies downstream analysis of samples. While performing multiple sample enrichment steps may be highly beneficial, existing methods are often cumbersome, prone to contaminants and can dilute samples beyond detection limits. Interfacing these methods with analytical equipment also presents challenges.

[0006] Protein analysis, for example, often involves liquid chromatography-mass spectrometry (LC-MS), where proteins are divided into peptides prior to the analysis. This process generates numerous fractions and complex data. While LC-MS methods are known, they are generally limited to use for peptide analysis, hindering intact protein analysis. LC-MS techniques are particularly disadvantageous in that they are complex and time-consuming to operate and require specialised training and expertise.

[0007] Electrospray ionisation (ESI) is common method for introducing samples into mass spectrometers for subsequent analysis. Typically performed as the ends of capillaries or in microfluidic devices, ESI ionises sample droplets, which are elongated using an applied electric field. The sample, at the end of a charged capillary, forms a “Taylor cone” and smaller droplets are released for analysis in a mass spectrometer. However, ESI is highly sensitive to the composition of the mobile phase (solvent). Factors like pH, salt concentration, and the presence of surfactants can significantly impact the ionisation efficiency and the quality of the spectra. Further, compounds with high surface tension can interfere with droplet formation and ionisation. In complex mixtures, some analytes can suppress the ionisation of others, leading to inaccurate quantification. The flow rate of the sample and the applied electric field can affect the stability of the spray and the intensity of the signal.

[0008] One field in which improvements to conventional analytical chemistry techniques are sought is the study of atmospheric fingerprints locked in ice cores. In this field, continuous flow analysis of target organic compounds or molecules in a complex mixture is required. By assessing and studying organic markers in ice cores, one is able to track emission sources and understand historical environmental processes. Conventional ice core analysis techniques are generally

[0009] P150790PC00 Mass spectrometry nebulisermodifications to conventional LC-MS techniques. Broadly, with current techniques, organic target samples in ice core melt-water are concentrated before passing through a chromatograph for separation leading to electrospray ionisation. These techniques are flawed in that preconcentration is rarely reproducible and in that there is a strong likelihood of introducing contaminants into the melt-water that suppresses subsequent analysis sensitivities in respect of any target organics. Alternative techniques suitable for ice core melt-water analysis include plasma-excited nebuliser gas-assisted electrospray ionisation (PENG-ESI), however the experimental setups for such techniques often require complex and careful optimisation of various experimental parameters and can be susceptible to ion suppression effects, where the presence of certain compounds in the sample can interfere with the ionisation of others.

[0010] It is therefore desirable to provide techniques that offer improvements in sample solution preparation for mass spectrometry. In particular, it is desirable to provide improvements in continuous flow liquid sample solution analysis techniques, particularly for ice core melt-water analysis.

[0011] Summary

[0012] The invention is defined in the independent claims. Specific embodiments are defined in the dependent claims.

[0013] According to an aspect of the present disclosure, there is provided a nebuliser apparatus suitable for use in mass spectrometry procedures. The nebuliser apparatus includes a sample channel, which is configured to pass through a body of the nebuliser apparatus from a sample inlet and to a sample and atomising fluid outlet. The sample may equivalently be referred to as the analyte in this context. The sample channel may equivalently be referred to as a sample bore. The sample channel is configured to permit passage of a sample fluid. To enable atomisation of the sample fluid, an example sample channel may include a Venturi throat or constriction so as to increase the velocity of the sample fluid and to form a spray of sample fluid. Of course, the skilled reader will appreciate that alternative modes of enabling atomisation may feasibly be implemented into the nebuliser apparatus, including rotary atomisers and electrostatic atomisers.

[0014] The nebuliser apparatus further includes an atomising fluid channel, which is configured to pass an atomising fluid through the body of the nebuliser apparatus from an atomising fluid inlet to the shared sample and atomising fluid outlet. In use, the atomising fluid channel and the sample channel are configured to atomise the sample solution and thereby form an

[0015] P150790PC00 Mass spectrometry nebuliseratomised sample solution. The atomising fluid channel may equivalently be referred to as an atomising fluid bore.

[0016] The nebuliser apparatus further includes a sheath fluid channel, which is configured to pass a sheath fluid through the body of the nebuliser apparatus from a sheath fluid inlet to a sheath fluid outlet. In use, the sheath fluid channel (or geometry of aspects thereof) is configured to form the sheath fluid in a substantially annular profile, where the sheath fluid encircles the atomised sample solution as output from the atomising fluid channel.

[0017] With use of the sheath fluid channel, the nebuliser apparatus is able to produce a confined and atomised sample solution with a protective sleeve of sheath fluid. The sheath fluid minimises dispersion of encircled atomised sample solution droplets, thereby improving sensitivity of any subsequent mass spectrometry measurements using the atomised sample solution droplets.

[0018] Through adjusting the flow rate of any or all of the sample solution, atomising fluid and the sheath fluid through the nebuliser apparatus (with fixed geometry), one may adjust the velocity and droplet size of the atomised sample solution and thereby adjust the suitability of the sample for input into a mass spectrometer. Equivalently of course, through adjusting the geometry of any or all of the sample channel, atomising fluid channel, and sheath fluid channel (or aspects thereof, including outlets) for fixed flow velocities of sample solution, atomising fluid and sheath fluid, one may adjust the velocity and size of the atomised sample solution.

[0019] The sheath fluid channel of the nebuliser apparatus may comprise multiple outlets, such that the sheath fluid channel is configured to pass the sheath fluid to the plurality of sheath fluid outlets. For example, the sheath fluid channel may terminate in six outlets. Any sheath fluid outlet may be of any suitable cross-sectional geometry to permit the sheathing behaviour of the sheath fluid, including, for example, substantially elliptical.

[0020] In use, the nebuliser apparatus relies on the simultaneous flow of atomising fluid, sheath fluid, and sample to function. Apparatuses herein allow for a high-throughput analytical technique that enables preparation of continuous flow samples for analysis of target organic compounds and / or molecules in a complex mixture. The techniques have the advantage of enabling realtime high sensitivity analysis while maintaining measurement stability and reproducibility. The techniques are further advantageous in that they require only low sample volumes: quantitative information of target organics may be obtained from just 200 pl of sample liquids - this is particularly useful in cases where the source is limited in supply, such as with ice core analysis.

[0021] P150790PC00 Mass spectrometry nebuliserIn some arrangements, the nebuliser apparatus comprises a plurality of sample and atomising fluid outlets, which collectively may be configured to output the atomised sample solution. Any sample and atomising fluid outlet may be of any suitable cross-sectional geometry to permit atomisation of the sample fluid, including, for example, of substantially circular cross-section.

[0022] In some arrangements, the atomising fluid channel and the sample channel may be configured such that the sample solution and the atomising fluid meet at a location inside the body of the nebuliser apparatus. In some alternative preferred arrangements, the atomising fluid channel and the sample channel may be configured such that the sample solution and the atomising fluid meet outside of the body of the nebuliser apparatus and atomisation of the sample solution occurs outside of the body of the nebuliser apparatus. In this way, the risk of clogging of either atomising fluid channel or the sample channel is minimised.

[0023] In some arrangements, the sample channel and the atomising fluid channel form an angle between 40 and 57 degrees. That is, an angle of between 40 and 57 degrees is formed between a portion of the sample channel closest to the sample and atomising fluid outlet and a portion of the atomising fluid channel closest to the sample and atomising fluid outlet. In some preferred arrangements, the angle is substantially 41 degrees. These angles have been found to be particularly well-suited for allowing impact of atomising fluid on sample solutions thereby inciting atomisation of the sample and resulting in droplet sizes well-suited for subsequent input into mass spectrometry apparatuses.

[0024] In some arrangements, the atomising fluid is nitrogen gas, air, or argon gas, or any combination thereof. These example atomising fluids have been found to be well-suited for atomisation of liquid sample solutions.

[0025] In some arrangements, the sheath fluid is nitrogen gas, air, or argon gas, or any combination thereof. These example sheath fluids have been found to be well-suited for sheathing of liquid sample solutions.

[0026] According to another aspect of the present disclosure, there is provided a mass spectrometry injection system suitable for use in mass spectrometry procedures. In particular, the mass spectrometry injection system is suitable for injecting atomised sample solutions into a mass spectrometer. The mass spectrometry injection system includes a nebuliser apparatus as described herein. In addition, the mass spectrometry includes an electrospray ionisation source, which is configured to inject an ionised fluid into the atomised sample solution. This process ionises the atomised sample solution, thereby forming an ionised atomised sample

[0027] P150790PC00 Mass spectrometry nebulisersolution. The nebuliser apparatus and the electrospray source are positioned so as to direct the ionised atomised sample solution towards to an inlet of a mass spectrometer.

[0028] The mass spectrometry injection system thereby forms an example of an extractive electrospray ionisation (EESI) system, facilitating collision between two sprays: the atomised sample solution (from the nebuliser) and an electrospray (from the electrospray ionisation source). When the two collide, analytes from the sample solution are transferred into the electrospray droplets. This transfer is influenced by factors including solubility and droplet dimensions. The charged electrospray droplets now carry the analytes. As the electrospray droplets evaporate, the analytes within become increasingly concentrated and eventually become ionised.

[0029] EESI is a soft or atmospheric technique commonly used for ionising droplets or aerosols for mass spectrometric analysis. This ionisation technique, first introduced in 2007, is equipped with two sprays options: a neutral spray and electrospray. The neutral spray flow line delivers the target analytes, while the electrospray flow line is comparable to that of typical electrospray ionisation (ESI).

[0030] The EESI technique herein is able to prepare samples for analysis directly with minimal further sample preparation processes relative to other known techniques. Moreover, this EESI technique is less susceptible to ion suppression effects from complex matrices compared to traditional ESI. The technique may be applied to a wide range of samples, including biological fluids, environmental samples, and food.

[0031] A further benefit of the techniques herein is in the assurance that the target organic component in a complex mixture is independently ionised without interference from the coeluents or solvents in a sample, thereby providing a reduction in ion suppression though an increase in ionisation efficiency.

[0032] In some arrangements, the mass spectrometry injection system further includes a drying system. The drying system may be fluidly connected to the nebuliser apparatus and configured to remove moisture from the atomised sample solution. That is, the drying system may be positioned to accept the atomised sample solution and the encircling sheath fluid from the nebuliser apparatus and output dried atomised sample solution for mixing with the electrospray from the electrospray source.

[0033] P150790PC00 Mass spectrometry nebuliserAccording to another aspect of the present disclosure, there is provided a mass spectrometry injection system suitable for use in mass spectrometry procedures. In particular, the mass spectrometry injection system is suitable for injecting atomised sample solutions into a mass spectrometer. The mass spectrometry injection system includes any nebuliser apparatus that is able to produce an atomised sample solution (not necessarily enclosed within a sheath fluid). The mass spectrometry injection system further includes a drying system fluidly connected to the nebuliser apparatus and configured to remove moisture from the atomised sample solution. In addition, the mass spectrometry includes an electrospray ionisation source, which is configured to inject an ionised fluid into the atomised sample solution. This process ionises the atomised sample solution, thereby forming an ionised atomised sample solution. The nebuliser apparatus, the drying system and the electrospray source are positioned relative to one another so as to direct the ionised atomised sample solution towards to an inlet of a mass spectrometer. While a sheath fluid (as used in alternative aspects of the present disclosure) is capable of minimising dispersion of encircled atomised sample solution droplets, the sensitivity of mass spectrometry measurements using unsheathed atomised, dried, and ionised sample solution droplets is nonetheless improved relative to conventional techniques even in the absence of any sheath fluid channel.

[0034] The relative alignment of the nebuliser apparatus - in particular the sample and atomising fluid outlet thereof - and the electrospray ionisation source (and / or an outlet of the drying system, if present) may be configured to ensure optimal transport of the ionised atomised sample solution towards to the inlet of the mass spectrometer. In some arrangements, the sample and atomising fluid outlet is aligned substantially coaxially with a major axis of the inlet of the mass spectrometer and / or an outlet of the drying system (if present), and the electrospray ionisation source is aligned between 90 and 152 degrees relative to the major axis of the inlet of the mass spectrometer. In some preferred arrangements, the angle is substantially 135 degrees. These arrangements are found to be well-suited for transmission of samples.

[0035] In some arrangements, the mass spectrometry injection system further includes an ice core flow line. The ice core flow line may be fluidly connected to the nebuliser apparatus and configured to transmit melt-water of an ice core to the nebuliser apparatus. Melt-water may be obtained from an ice core using a continuous melting system; an example continuous meting system includes a hot-plate with holes through which melt-water is able to flow. The melt-water may be pushed to / through the ice core flow line using instruments such as peristaltic pumps.

[0036] According to another aspect of the present disclosure, there is provided a mass spectrometry system. The mass spectrometry system includes the mass spectrometry injection system as

[0037] P150790PC00 Mass spectrometry nebuliserdescribed herein (including either the nebuliser apparatus as described herein or any nebuliser apparatus that is able to produce an atomised sample solution). The mass spectrometry system further includes a mass spectrometer, or at least a mass analyser and a detector thereof. The mass spectrometry injection system may be aligned with an inlet of the mass spectrometer so as to permit passage of the ionised sample solution. The ionised sample solution may be directed through the mass analyser towards the detector. The mass analyser separates the ions based on their mass-to-charge ratio and the detector measures the relative abundance of each ion type. This information may be used to determine the chemical element composition of the original sample solution and the isotopic composition of its constituents.

[0038] According to another aspect of the present disclosure, there is a provided a method of sample solution preparation for mass spectrometry, using a nebuliser apparatus as described herein. The method includes a step of passing a sample solution through a sample channel of a nebuliser apparatus to a sample and atomising fluid outlet. The method includes a step of passing an atomising fluid through an atomising fluid channel of the nebuliser apparatus to the sample and atomising fluid outlet and thereby atomising the sample solution. The method then includes a step of passing a sheath fluid through a sheath fluid channel of the nebuliser apparatus to a sheath fluid outlet and thereby forming the sheath fluid in a substantially annular profile encircling the atomised sample solution.

[0039] In some arrangements, the method of sample preparation further includes use of the mass spectrometry injection system described herein. Namely, the method may further include a step of injecting an ionised fluid into the atomised sample solution using an electrospray ionisation source to ionise the atomised sample solution, wherein the nebuliser apparatus and the electrospray source are positioned to direct the atomised sample solution to an inlet of a mass spectrometer. Optionally, the method of sample preparation additionally includes a step of removing moisture from the atomised sample solution using a drying system fluidly connected to the nebuliser apparatus.

[0040] According to another aspect of the present disclosure, there is a provided a method of sample solution preparation for mass spectrometry, using any nebuliser apparatus that is able to produce an atomised sample solution (not necessarily enclosed within a sheath fluid). The method includes a step of passing a sample solution through a sample channel of a nebuliser apparatus to a sample and atomising fluid outlet. The method includes a step of passing an atomising fluid through an atomising fluid channel of the nebuliser apparatus to the sample and atomising fluid outlet and thereby atomising the sample solution. The method includes a step of removing moisture from the atomised sample solution using a drying system fluidly

[0041] P150790PC00 Mass spectrometry nebuliserconnected to the nebuliser apparatus. The method then includes a step of injecting an ionised fluid into the atomised sample solution using an electrospray ionisation source to ionise the atomised sample solution, wherein the nebuliser apparatus and the electrospray source are positioned to direct the atomised sample solution to an inlet of a mass spectrometer.

[0042] According to another aspect of the present disclosure, there is a provided a method of mass spectrometry, using a mass spectrometry system as described herein. The method includes the method of sample solution preparation to produce an ionised atomised sample solution as described herein. The method further includes a step of directing the atomised sample solution to an inlet of a mass spectrometer. The method then includes a step of producing a mass spectrum of the ionised atomised sample solution.

[0043] The invention is described in terms of particular embodiments. Other embodiments are within the scope of the appended claims. For example, the steps of a method according to an embodiment may be performed in a different order and still achieve desirable results.

[0044] Aspects of embodiments have been described using such terms as “atomising fluid channel” and “sheath fluid channel” and “outlet(s)”. The skilled person will appreciate that such functional terms and their equivalents may refer to parts of the system that are spatially separate but combine to serve the function defined. Equally, the same physical parts of the system may provide two or more of the functions defined.

[0045] To avoid unnecessary duplication of effort and repetition of text in the specification, certain features are described in relation to only one or several aspects or embodiments of the present disclosure. However, it is to be understood that, where it is technically possible, features described in relation to any aspect or embodiment of the present disclosure may also be used with any other aspect or embodiment of the present disclosure.

[0046] Brief Description of the Drawings

[0047] Reference is made, by way of example only, to the accompanying drawings in which:

[0048] Figure 1 is a set of orthographic views of a nebuliser apparatus according to the present disclosure;

[0049] Figure 2 is a cutaway side view of a nebuliser apparatus in use according to the present disclosure;

[0050] Figure 3 is a schematic view of a mass spectrometry system according to the present disclosure;

[0051] P150790PC00 Mass spectrometry nebuliserFigure 4 is a flowchart of a method of sample solution preparation for mass spectrometry according to the present disclosure;

[0052] Figure 5 is a schematic view of a mass spectrometry system according to the present disclosure; and

[0053] Figure 6 is a plot of mass spectra peak abundance for a range of analyte molar concentrations, acquired using a mass spectrometry system according to the present disclosure.

[0054] Detailed Description

[0055] Figure 1 shows various external views of an example nebuliser apparatus 100 of an example mass spectrometry injection system. The left-most view is a rear profile of the nebuliser apparatus 100; the right-most view is a side profile of the nebuliser apparatus 100; and the centre view is a front profile of the nebuliser apparatus 100.

[0056] The nebuliser apparatus 100 includes an elongate cylindrical body 110 with one chamfered edge. On a first substantially planar edge surface of the body 110 is a sample inlet 120, suitable for accepting sample fluid (gas or liquid). On the opposing substantially planar edge surface of the body 110 is a sample and atomising fluid outlet 150, around which a set of six equally spaced sheath fluid outlets 160 are circumferentially positioned (tangential to the major axis of the nebuliser apparatus 100). That is, the sheath fluid outlets 160 form an annulus surrounding the sample and atomising fluid outlet 150. On the circumferential side or sleeve of the body 110 is an atomising fluid inlet 130 and a sheath fluid inlet 140. Though not shown, the body 110 may further include additional atomising fluid inlets and / or sheath fluid inlets, e.g., on the opposing side of the body 110.

[0057] The sample inlet 120 allows passage of sample fluid through a sample channel, passing through the body 110 and out of the sample and atomising fluid outlet 150. The atomising fluid inlet 130 allows passages of atomising fluid through atomising fluid channel 180 and out of the sample and atomising fluid outlet 150. The sheath fluid inlet 140 allows passage of sheath fluid through sheath fluid channel 190 and out of the sheath fluid outlets 160.

[0058] In the present example, the sample inlet 120, atomising fluid inlet 130, sheath fluid inlet 140, and sample and atomising fluid outlet 150 are all substantially circular in cross section. The sheath fluid outlets 160 are all substantially elliptical in cross section. Of course, the skilled reader will appreciate that the shape of each inlet and outlet may be varied. In the present example, the sample inlet 120 has a diameter of 1.92 mm; the atomising gas inlet 130 has a diameter of 1.92 mm; the sheath gas inlet 130 has a diameter of 1.92 mm; the sample and atomising fluid outlet 150 has a diameter of 0.5 mm; and each elliptical sheath gas outlet has

[0059] P150790PC00 Mass spectrometry nebulisera major axis of 2.02 mm. Again, of course, the dimensions of each inlet and outlet may be varied. The shape and dimension of each inlet and outlet may be adjusted to conform to standard lab equipment, such as standard sized tubing.

[0060] Generally, the change in shape and scale of the flow inlets, outlets and channels therebetween is expected to have a proportional effect on the size of the droplet. With larger inlet sizes, there is an increase in the quantity of sample fluid entering the region capable of atomisation, which results in an increase in the number of droplets. Monodispersity of sample fluid droplet size is dependent on a balance with atomising fluid properties (e.g., flowrate and speed) for a fixed geometry. Variations in geometry of the nebuliser are expected to be suitable for fluids with viscosities less than 2 x 10'3Pa s. Relatively high viscosity values will lead to increase in droplet size because higher shear force is required. Increasing the cross-sectional area or volume of the region where sample fluid and atomising fluid interact (e.g., by increase in diameter of outlets) creates an increased contact area for fluid interaction.

[0061] The body 110 may be a unitary a unitary construction, being formed (e.g., cast, such as pressure die cast) as a single component. In a worked example, the body 110 is a three-dimensional printed object, formed from a photosensitive nano-filled ceramic material with surface roughness similar to silicones or vulcanised rubber (i.e. , with surface roughness on the order of 30-50 pm). Namely, the present example nebuliser 100 is printed using a DWS XFAB® 3500 stereolithography printer with Presica DL-260 resin. Alternative materials, such as PDMS, may similarly be used. Microfluidic or nanostructured materials may be applied to the nebuliser apparatus 100 (internally, externally, or both), so as to promote passage of specific target organic compounds or, for example, to minimise wear.

[0062] Figure 2 provides a cutaway side view of nebuliser apparatus 100 when in use. Inset is a magnified view of the area in and around the sample and atomising fluid outlet 150. As shown, sample fluid (depicted with chequered shading) is input into sample inlet 120 with an initial velocity of Vsand pressure of Ps(and flowrate Qs). The sample passes through sample channel 170. Before the sample reaches sample and atomising fluid outlet 150, a Venturi throat or constriction within the sample channel 170 increases the velocity and decreases the pressure of the sample, causing the sample to form droplets on exit of the nebuliser apparatus 100.

[0063] Simultaneously, atomising fluid (depicted with no shading and dashed outlines) is input into atomising fluid inlet 130 with a velocity of Vafand pressure of Paf(and flowrate Qaf). The atomising fluid passes through atomising fluid channel 180, which is directed through the nebuliser body 110 towards the sample and atomising fluid outlet 150. The atomising fluid

[0064] P150790PC00 Mass spectrometry nebuliserchannel 180 and the sample channel 170 are arranged to converge at the sample and atomising fluid outlet 150 at a predefined angle such that atomising fluid impacts the sample at the predefined angle when the sample exits the nebuliser apparatus 100, thereby forming atomised sample fluid. The atomised sample fluid comprises sample fluid droplets substantially encapsulated by atomising fluid. The resultant spray of atomised sample fluid may be quantified with the parameters usand ds, where usis the atomised droplet velocity and dsis the atomised droplet diameter.

[0065] Also simultaneously, sheath fluid (depicted with no shading and solid outlines) in input through sheath fluid inlet 140 with a velocity of Vsfand pressure of Psf(and flowrate Qsf). The sheath fluid passes through sheath fluid channel 190, which is directed through the nebuliser body 110 towards the plurality of sheath fluid outlets 160 (only a single passage and two outlets are shown here for simplicity). The sheath fluid outlets 160 are substantially coaxial to the central sample and atomising fluid outlet 150. The sheath fluid outlets 160 emit the sheath fluid in a pattern that surrounds the atomised sample fluid and minimises dispersion. In turn, this improves focusing of the sample towards an inlet of a mass spectrometer (not shown here).

[0066] In the present example, the sample fluid is liquid water with dynamic viscosity, .s, on the order of 1.05 x icy3Pa s. Both the atomising fluid and sheath fluid are lab-grade nitrogen gas. Alternatives, including dry air (with 0.4-1 % water content) and gases such as argon (Ar) may be feasibly used as atomising fluid and / or sheath fluid.

[0067] In the present example, using a Venturi throat of 1 mm length and with 450 pm diameter, a sample injected with an initial velocity, Vs, in the range of (3.43-34.3) x l0'3ms'1, the sample forms at the sample and atomising fluid outlet 150 into droplets with diameters in the range of 158.4-287.6 pm. With a convergent angle between the sample channel and the atomising fluid channel between approximately 41 ° and 57 ° and with atomising fluid velocity, Vaf , Of approximately 275±5 ms-1(using a pressure, Paf, of approximately 2 bar), the atomising fluid impacts the sample fluid droplets. The injected sample velocity may be calculated from the droplet-dryer relationship as described below; empirically, this velocity value is tied to the diameter of a drying system in use. The size of the droplet may be determined using direct optical measurement techniques using a lens. For instance, to capture the moment of large droplet breakage, the sample may be injected at a fraction (e.g., 1 / 10th) of the sample minimum flowrate. At this rate, the surface tension of the liquid is able to hold the droplet for sufficient time (e.g., 5 to 10 seconds) to permit image capture.

[0068] P150790PC00 Mass spectrometry nebuliserAs a result of shear stress of the high velocity gas, the sample forms into an atomised sample solution spray. With the example geometry, the interaction between atomising fluid and sample occurs within a region 0.6-1 mm outside of the sample and atomising fluid outlet 150 (that is, external to the body 110 of the nebuliser 100). Of course, while alternative geometries may be envisioned that facilitate spray formation within the nebuliser body 100, in having external contact between sample and atomising fluid, the risk of internal clogging is reduced.

[0069] Figure 3 is a schematic view of a mass spectrometry system 1, including a nebuliser 100 as described above and also including a drying system 200, electrospray source 300 and a mass spectrometer 400. Collectively, nebuliser 100, drying system 200 and electrospray source 300 may be referred to as a mass spectrometry injection system 10, in the sense that the mass spectrometry injection system 10 is suitable for preparation of sample for injection into / towards an analyser and detector of mass spectrometer 400. The skilled reader will appreciate that any conventional nebuliser may be used in place of nebuliser 100, so long as the nebuliser is capable of atomising an input sample fluid.

[0070] The nebuliser 100 is as described above in respect of Figure 2. Notably, sample fluid (with chequered shading) is injected at velocity Vs(and at pressure Psand flowrate of Qs). In the present example mass spectrometry injection system 10, liquid sample is received into the nebuliser body 110 using a 101.6 pm diameter stainless steel needle, fluidly connected to an outlet of a continuous flow analysis (CFA) system, supplying melt-water from an ice core flow line (not shown). Atomising fluid (depicted with no shading and dashed outlines) is injected at pressure Paf(and velocity Vafand flowrate Qaf). Sheath fluid (depicted with no shading and solid outlines) is injected with flowrate of Qsf(and velocity Vsfand pressure Psf). The nebuliser 100 outputs a spray of atomised sample s with velocity and diameter of usand ds, respectively.

[0071] The example nebuliser 100 may be considered as a two-fluid type nebuliser, transmitting both liquids and gases. The liquid sample is atomised for spray creation (i.e. , transformed into a dispersed phase), while gas is used in the continuous phase for atomising the sample and also for creating a sheath flow annulus, which assists in directing sample droplet spray towards the eventual reaction site. This arrangement may be classified as a combination of cross-flow and parallel-flow type atomisers that is actively tuneable so to independently optimise monodispersity and velocity of droplet spray.

[0072] The flow channels in the nebuliser 100 are, in effect, split by phase: with distinct channels for each fluid. The gas phase flow channel is further split into the atomising fluid channel 180 and the sheath fluid channel 190. All flow channels are positioned within a chamfered cylinder solid

[0073] P150790PC00 Mass spectrometry nebulisermodel. The flow behaviours within each fluid channel may be controlled independently by varying the input flow parameters, including velocity and pressure. The flow channels may be considered as a microfluidic flow network including a liquid sample flow bore and gas flow bores. Walls connecting the flow channels are non-permeable. To function as a nebuliser, in use, the nebuliser apparatus 100 relies on simultaneous flow of atomising fluid, sheath fluid, and sample to function.

[0074] The mass spectrometry injection system 10 may further include a micro-sieve (e.g., a mesh) for removal of insoluble particles (e.g., dust), which are not the target of analysis. Such a microsieve may be positioned before, after, or even within the nebuliser apparatus 100.

[0075] The example mass spectrometry injection system 10 includes drying system 200. The monodispersed spray of atomised sample solution enters a 15.6 mm diameter dryer sample inlet. The velocity of the atomised sample solution at this position is expected to be us(i.e., substantially the same as when exiting nebuliser apparatus 100). In the example drying system 200, a Nation™ tube drying mechanism is used. The Nation™ polymer transfers water molecules from one side of the membrane to the other through a first order kinetic reaction. Nation™ polymer selectivity is based on chemical reactivity, and not on the size of the molecule. The Nation™ polymer selectively transfers water vapours, leaving most analytes in the atomised sample solution untouched.

[0076] The atomised sample solution drying process is activated using lab grade nitrogen (depicted with angled shading and solid outlines), which is injected into the Nation™ tube at purge gas inlet 210. The mechanism of drying is based on the difference in vapour pressure of surrounding purge gas (nitrogen) and the vapour pressure of the monodispersed atomised sample solution.

[0077] The efficacy of the drying system 200 is believed to be controlled by the velocity of the purge gas (nitrogen), Vpg, relative to the velocity of the atomised sample solution, us. It is found that injecting the purge gas with the following relationship provides suitable sample dryness for subsequent EESI: Vpg= yus. Here, y is an empirical factor based on the geometry of the dryer inlet. In the present example mass spectrometry injection system 10, values of y between 1 and 5 are valid.

[0078] As shown in Figure 3, the purge gas is injected into the Nation™ tube at a purge gas inlet 210 positioned an opposite end of the tube relative to injection of the atomised sample solution. Effluent (containing purge gas and extracted moisture) is output at a purge gas outlet 220

[0079] P150790PC00 Mass spectrometry nebuliserproximate to the injection of the atomised sample solution. Dry atomised sample solution is output at an outlet proximate to the injection of the purge gas.

[0080] In the event that the amount of moisture in the spray of atomised sample is considered to be sufficiently low for input directly into the mass spectrometer, the drying apparatus 200 may be omitted from the mass spectrometry injection system 10.

[0081] The full mechanism underlying efficient formation of sprays suitable for EESI is not yet known, but it is believed that the atomised sample solution droplets created are able to form and maintain a core-shell structure suitable for ionisation. Core-shell suppression that is often encountered in conventional drying techniques is significantly minimised. During the ionisation phase in a mass spectrometry analysis, factors such as high concentration, mass, basicity of the analyte, the presence of co-eluent, and contamination from or during sample preparation often hinder the ionisation of the target analyte. This phenomenon leads to reduced selectivity of the target analyte because the available electric charge for ionisation is overcrowded by non-target analytes. In a more specific example case of a droplet containing L-tartaric acid and ammonium sulphate, the present drying technique as described herein ensures that ammonium-sulphate continues to form the centre core of the droplet, while tartaric acid is positioned around it, thus ensuring efficient ionisation of the droplet by the available electric charge. The core-shell structure enables transfer of the organics on the surface of the dried particles to the charged solvent droplets from the ESI rather than having the organics ionised directly on the surface of the dry particle.

[0082] The major axis of the example drying system 200 is aligned substantially parallel to the mass spectrometer 400 inlet. Electrospray source 300 includes an electrospray capillary, which is angled offset relative to the mass spectrometer 400 inlet and the major axis of the example drying system 200 (or at least the outlet thereof).

[0083] Typically, mass spectrometry ionisation technologies, including conventional electrospray ionisation (ESI), require multi-step sample pretreatment (e.g., extraction, separation, dilution) before the processed sample may be introduced for ionisation and mass analysis and detection in the vacuum within the mass spectrometer, which may delay the process of analysis for several minutes or hours. In contrast, EESI firstly charges a selected reagent (e.g., methanol or acetonitrile) using ESI for efficient production of an ionic reagent spray (also referred to herein as an electrospray), which then impacts the sample spray, transferring energetic charge to the analyte molecules within the sample spray. The analytes become ions

[0084] P150790PC00 Mass spectrometry nebuliserthrough extraction and collision during the interaction of the two sprays. Ionised analytes may then be introduced into the mass spectrometer for subsequent analysis.

[0085] Typically, an EESI system includes an electrospray source and a neutral sample spray channel, which are crossed at a predetermined angle. By adjusting such geometric parameters as the angles and distances between electrospray source, neutral sample spray channel, and mass spectrometer inlet, the detection limits for a variety of substances may improve beyond those of conventional ESI techniques under experimental conditions. The overall efficiency for ion collection, desolvation and transmission is improved in EESI relative to the conventional ESI techniques.

[0086] In the present example, an electrospray capillary of the electrospray source 300 is variably positioned at an angle of between approximately 90-152 ° relative to the mass spectrometer inlet. An angle of approximately 135 ° is found to be particularly suitable. The electrically charged ESI capillary is configured to pass liquid methanol (99.9% vol) with a flowrate set at a constant value of 2x10-3ml min-1. A charged electrospray of methanol (depicted with solid shading) is then injected into the (dried) atomised sample solution spray (i.e., the neutral spray). This configuration permits adjustment of the atomised sample solution flowrates within the range of (0.33-1.67)X10-3ml min-1. Following ionisation of the atomised sample solution spray, the ionised atomised sample solution spray, s', may be quantitatively described with velocity u’sand spray droplet diameter d'.

[0087] The angle of the electrospray needle relative to the mass spectrometer and atomised sample solution spray is selected such that there is sufficient interaction space for dispersed droplet ionisation before entering the mass spectrometer for analysis. A voltage of approximately 3 kV is applied across the electrospray capillary so as to charge the electrospray.

[0088] To accommodate the behaviour of smaller droplets (e.g., droplets with a diameter less than 10 nm), a focusing device may be used (not shown), which offsets the low inertia and high diffusivity in such droplets and thereby maximises droplet ionisation within the ESI electrospray prior to analysis.

[0089] With the present example mass spectrometry injection system 10, reliable, repeatable and reproducible results have been found with sample flow velocities Vstested in the range of 10-90 pl hr1. These rates have been selected from test results monitoring the performance of a Nation™ dryer, nebuliser apparatus, droplet size distribution and ESI ionisation behaviour. In the present example, a flowrate of 60 pl hr1is found to be optimal. Broadly, operation of the

[0090] P150790PC00 Mass spectrometry nebuliseranalytical technique may be governed by defined an empirically derived fluid mechanic relationship for the purge gas velocity, Vpgand the velocity of the sample as input into the nebuliser, Vswith nitrogen gas atomising fluid, nitrogen gas sheath fluid, nitrogen gas purge fluid, and liquid sample viscosities on the order of 1.05 x 10'3Nsnr2, the optimum ratio range between velocities is between 1 and 5, and more preferably between 1 and 2.5.

[0091] The mass spectrometry injection system 10 of the present disclosure optimises the creation of micro-sized monodispersed droplets from liquid samples while enabling controlled-velocity and reaction site area-focused droplet delivery. The flow in the system maintains a parallel flow of dispersed fluid (including analytes) along with a stream of the atomising gas in the continuous phase. The configuration of the nebuliser apparatus 100 in this system ensures that the dispersed phase is focused to the target reaction site using a coaxial sheath fluid. The nebuliser apparatus 100 may be described as a mechanical, tuneable active droplet generator with a fraction of second response time, offering the flexibility of independently controlling the size of the droplets and the frequency of delivery through adjustment of the flow properties of both phases.

[0092] The drying process as used by an example mass spectrometry injection system 10 herein uses commercially available techniques, namely a Nation® drying method. In this drying process, water molecules from the stream of monodispersed droplets of the analytes are removed in a constant-rate or mass-transfer drying period, where there is no (or minimal) change in temperature of the individual droplet or an entire spray of droplets. The drying mechanism in this process is due to the difference in partial vapour pressure of water in the droplets and the drying gas or air flowing in the annulus of the dryer tube.

[0093] Figure 4 is a flowchart of a method of sample solution preparation, as performed using a nebuliser 100 according to the present disclosure. The method of sample solution preparation may be implemented by a human or may be implemented using mechatronics means, where control is performed using computational means.

[0094] At S2, the method involves passing a sample solution through a sample channel of a nebuliser apparatus to a sample and atomising fluid outlet. At S4, the method involves passing an atomising fluid through an atomising fluid channel of the nebuliser apparatus to the sample and atomising fluid outlet and thereby atomising the sample solution. At S6, the method involves passing a sheath fluid through a sheath fluid channel of the nebuliser apparatus to a sheath fluid outlet and thereby forming the sheath fluid in a substantially annular profile encircling the atomised sample solution.

[0095] P150790PC00 Mass spectrometry nebuliserWhere the nebuliser 100 is incorporated a mass spectrometry injection system 10 (further including an electrospray source 300 and, optionally, a drying system 200), the method may further be configured to ionise the atomised sample solution, The method then involves, at S8, injecting an ionised fluid into the atomised sample solution using an electrospray ionisation source to ionise the atomised sample solution, wherein the nebuliser apparatus and the electrospray source are positioned to direct the atomised sample solution to an inlet of a mass spectrometer.

[0096] Figure 5 is a schematic overview of a mass spectrometry system 1 used for experimental prototype testing of the nebuliser apparatus 100 of the present disclosure and the associated method for sample preparation. A mass spectrometry injection system 10 is coupled to an ion trap mass spectrometer - namely a linear trap quadrupole (LTQ) Velos mass spectrometer -for single-stage mass analysis under atmospheric conditions. The skilled reader will appreciate that the mass spectrometry injection system 10 is broadly agnostic in respect of specific form of mass spectrometer - any mass spectrometer that is configured (or configurable) to accept dispersion of atomised samples is appropriate. As seen, the electrospray source 300 capillary is coupled to a high-voltage AC source (with in-line capictor, Rc, and electrical switch, F) and the system is grounded to induce a 3 kV potential difference. Methanol is pumped (using pump, P2) with nitrogen used as a lift gas; the flowrate of the lift gas is controlled using a flow controller (not shown). The electrospray plume, EPl, is formed between the electrospray electrode emitting methanol and an auxiliary electrode of the mass spectrometer, MS (the region of electrospray plume formation is depicted as a circular dashed region). Conventional techniques for similar experiments would involve coupling an LC-MS apparatus to an ESI apparatus.

[0097] Synthetic ice core melt-water may be produced through introduction of various quantities of organic and inorganic matter into water: L-tartatic acid (99 %, Aldrich) and ammonium sulphate (99.5 %, Fluka) are dissolved into milli-q or ultra-pure water. Mixture concentrations are in the range of 0.1-3.2 pmol L’1. For each concentration, the following five distinct mole fraction of tartaric acid (YTA) are prepared: 1, 0.98, 0.90, 0.50, 0.25, where Y is the total composition of organic and inorganic mixture.

[0098] Testing is performed by continuously injecting samples into the prototype system (using pump, Pl). Nitrogen is used as atomising fluid and also as sheath fluid; the flowrate of each may be determined using flowmeter, FM, therby allowing quality control and monitoring. In the drying system, waste line, WL, expels waste from the system to a fume hood. Measurements are

[0099] P150790PC00 Mass spectrometry nebuliserrecorded with the LTQ Velos MS after 90 seconds (60 seconds are dedicated to allowing the flow to fully develop in the system).

[0100] Figure 6 presents the mass-to-charge ratio peak abundance (or, equivalently, intensity) in L-tartaric acid for an ion with a m / z value of 149.08. The plot represents the quantity of L-tartaric acid defined by its strong peak value measured using a MS by directly injecting different concentrations and molar fractions of a mixture containing ammonium sulphate and the L-tartaric acid. The vertical axis denotes the (relative) count or abundance of L-tartaric acid. The horizontal axis denotes the concentration of the mixture (L-tartaric acid + ammonium sulphate in milli-q water). The concentration is recorded in Molarity (mol L'1). The plot provides concentration data from approximately 10'3mol L'1down to 6x1 O'6mol L’1.

[0101] As shown, the prototype mass spectrometry system 1 provides a linearly correlated performance in the quantification and identification of L-tartaric acid (the target organic) in different concentrations of injected liquid samples. Figure 6 also demonstrates the ability of the techniques herein to allow for quantification and identification of L-tartaric acid in samples, irrespective of molar fractions. Even for YTAvalues as low as 25 % and for molar concentrations on the order of just 1 pmol L’1, the m / z 149.08 peak is resolvable; these values are low in respect of the sensitivity of the LTQ Velos mass spectrometer in use. That is, the results show that even at a concentration of approximately 6xl0'6mol L’1, an embodiment of the system described herein may be used to count the quantity of L-tartaric acid in a mixture containing just a quarter of its volume fraction. The system facilitates rapid analysis processing, confirmed to be 10-50 times faster than LC-MS analysis that generally continues for several minutes or hours. During testing, the stable signal (spectra) results are obtained within the first minute after injecting unprocessed samples.

[0102] The techniques herein are then well-suited for analysis of continuous flow melt-water from ice cores, which are expected to include low concentrations of such target organic analytes as L-tartaric acid; 2-methylerythritol; 2-methylglyceric acid; keto-pinic acid; pinic acid; 3-MBTCA; cis-pinonic acid; cs-norpinonic acid; pinolic acid; and / or nopinone. Synthetic ice cores may be prepared by freezing standard mixtures of known composition in ultrapure water often in an ice core-shaped (e.g., a cylindrical container). The synthetic ice core will, however, generally have an approximately homogenous composition as opposed to the stratification that characterises real ice cores. The techniques herein, however, are not affected by this distinction.

[0103] P150790PC00 Mass spectrometry nebuliserIce cores (both real and synthetic) are typically stored in a cold room at approximately -30 °C and may be melted using a continuous melting system, which includes a hot-plate with holes through which the melt-water may flow. The melt-water may be continuously pushed to instruments of the apparatuses herein using, for example, peristaltic pumps.

[0104] While certain embodiments have been described, these embodiments have been presented by way of example only, and are not intended to limit the scope of the invention. Indeed, the methods and apparatuses described herein may be embodied in a variety of other forms; furthermore, various omissions, substitutions and changes in the form of methods and apparatuses described herein may be made.

[0105] P150790PC00 Mass spectrometry nebuliser

Claims

Claims1. A nebuliser apparatus for mass spectrometry, the nebuliser apparatus comprising: a sample channel configured to pass a sample solution to a sample and atomising fluid outlet;an atomising fluid channel configured to pass an atomising fluid to the sample and atomising fluid outlet and configured to atomise the sample solution; anda sheath fluid channel configured to pass a sheath fluid to a sheath fluid outlet and configured to form the sheath fluid in a substantially annular profile encircling the atomised sample solution.

2. The nebuliser apparatus of claim 1, wherein the sheath fluid channel is configured to pass the sheath fluid to a plurality of sheath fluid outlets.

3. The nebuliser apparatus of claim 1 or claim 2, wherein the atomising fluid channel is configured to atomise the sample solution outside of a body of the nebuliser apparatus.

4. The nebuliser apparatus of any of claims 1 to 3, wherein the sample channel and the atomising fluid channel form an angle between 40 and 57 degrees.

5. The nebuliser apparatus of claim 4, wherein the sample channel and the atomising fluid channel form an angle of substantially 41 degrees.

6. The nebuliser apparatus of any of claims 1 to 5, wherein the atomising fluid is any or all of nitrogen gas, air, and argon gas.

7. The nebuliser apparatus of any of claims 1 to 6, wherein the sheath fluid is any or all of nitrogen gas, air, and argon gas.

8. A mass spectrometry injection system comprising:the nebuliser apparatus of any of claims 1 to 7; andan electrospray ionisation source configured to inject an ionised fluid into the atomised sample solution thereby ionising the atomised sample solution,wherein the nebuliser apparatus and the electrospray ionisation source are positioned to direct the atomised sample solution to an inlet of a mass spectrometer.P150790PC00 Mass spectrometry nebuliser9. The mass spectrometry injection system of claim 8, further comprising a drying system fluidly connected to the nebuliser apparatus and configured to remove moisture from the atomised sample solution.

10. A mass spectrometry injection system comprising:a nebuliser apparatus comprising a sample channel configured to pass a sample solution to a sample and atomising fluid outlet and an atomising fluid channel configured to pass an atomising fluid to the sample and atomising fluid outlet and configured to atomise the sample solution;a drying system fluidly connected to the nebuliser apparatus and configured to remove moisture from the atomised sample solution; andan electrospray ionisation source configured to inject an ionised fluid into the atomised sample solution thereby ionising the atomised sample solution,wherein the nebuliser apparatus, the drying system and the electrospray ionisation source are positioned to direct the atomised sample solution to an inlet of a mass spectrometer.

11. The mass spectrometry injection system of any of claims 8 to 10, whereinthe sample and atomising fluid outlet and / or an outlet of the drying system is / are aligned substantially coaxially with a major axis of the inlet of the mass spectrometer, andthe electrospray ionisation source is aligned between 90 and 152 degrees relative to the major axis of the inlet of the mass spectrometer.

12. The mass spectrometry injection system of any of claims 8 to 11, wherein the electrospray ionisation source and the major axis of the inlet of the mass spectrometer form an angle of substantially 135 degrees.

13. The mass spectrometry injection system of any of claims 8 to 12, further comprising an ice core flow line fluidly coupled to the sample channel and configured to accept melt-water of an ice core.

14. A mass spectrometry system comprising:the mass spectrometry injection system of any of claims 8 to 13; anda mass spectrometer.

15. A method of sample solution preparation for mass spectrometry, the method comprising:P150790PC00 Mass spectrometry nebuliserpassing a sample solution through a sample channel of a nebuliser apparatus to a sample and atomising fluid outlet;passing an atomising fluid through an atomising fluid channel of the nebuliser apparatus to the sample and atomising fluid outlet and thereby atomising the sample solution; andpassing a sheath fluid through a sheath fluid channel of the nebuliser apparatus to a sheath fluid outlet and thereby forming the sheath fluid in a substantially annular profile encircling the atomised sample solution.

16. The method of sample solution preparation of claim 15, further comprising removing moisture from the atomised sample solution using a drying system fluidly connected to the nebuliser apparatus.

17. The method of sample solution preparation of claim 15 or claim 16, further comprising:injecting an ionised fluid into the atomised sample solution using an electrospray ionisation source to ionise the atomised sample solution, wherein the nebuliser apparatus and the electrospray source are positioned to direct the atomised sample solution to an inlet of a mass spectrometer.

18. A method of sample solution preparation for mass spectrometry, the method comprising:passing a sample solution through a sample channel of a nebuliser apparatus to a sample and atomising fluid outlet;passing an atomising fluid through an atomising fluid channel of the nebuliser apparatus to the sample and atomising fluid outlet and thereby atomising the sample solution;removing moisture from the atomised sample solution using a drying system fluidly connected to the nebuliser apparatus; andinjecting an ionised fluid into the atomised sample solution using an electrospray ionisation source to ionise the atomised sample solution, wherein the nebuliser apparatus and the electrospray source are positioned to direct the atomised sample solution to an inlet of a mass spectrometer.

19. A method for mass spectrometry comprising:the method of sample solution preparation of claim 17 or claim 18;directing the atomised sample solution to an inlet of a mass spectrometer; and producing a mass spectrum of ionised atomised sample solution.P150790PC00 Mass spectrometry nebuliser

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