DEVICE FOR DETECTING EVAPORATIVE LIGHT SPREAD, AS WELL AS AN ANALYSIS DEVICE WITH SUCH A DEVICE AND A METHOD FOR DETECTING EVAPORATIVE LIGHT SPREAD

Description

[0001] The invention relates to a device for detecting evaporation light scattering, as well as an analysis system with such a device and a method for detecting evaporation light scattering.

[0002] Devices for detecting evaporative light scattering are known from the prior art. For example, US 6,229,605 B describes an ELS (Evaporative Light Scattering) detector in which the sample is fed from an atomizer via a connecting tube into an optical unit. In the atomizer, the sample is mixed with a gas, so that it is subsequently in a mobile phase. A baffle is arranged between the atomizer and the connecting tube. This forms a condensation surface on which larger droplets of the sample condense and are discharged via a drain. The baffle can assume two positions, in which the flow rate is reduced by either 70% or 18%. In this device, the introduction and atomization of the sample occur simultaneously. However, the device has the disadvantage that the sample is introduced completely.

[0003] US 2001 / 0001575 A1 describes a low-temperature adapter for an ELS detector. The adapter is connected to the ELS detector in such a way that liquid from a chromatography unit is directed through an atomizer into an atomization chamber, where larger droplets are deposited in the mobile phase. The liquid sample is introduced into the atomizer via an eluate inlet. A backpressure valve is located at the opening, which is used to measure the liquid pressure.

[0004] The nebulizer is connected to a nebulization unit. A purge gas and a nebulization gas are introduced through the nebulization unit. A nebulization chamber is coupled to the nebulization unit, where the sample is nebulized at the end of a tip with the nebulization gas. The nebulized sample is then transferred to the nebulization chamber. This system also does not offer the possibility of recovering any part of the sample. Since the ELS method is a destructive process, all sample material introduced into an ELS detector is destroyed and is therefore not available for further analysis or sample recovery.

[0005] It is therefore an object of the invention to overcome the disadvantages of the prior art. In particular, it is an object of the invention to provide a device for detecting evaporation light scattering that allows simultaneous introduction and atomization of the sample, requires only a minimal sample quantity with maximum detection, and minimizes sample losses.

[0006] The problem is solved by the independent patent claims. Further embodiments are described in the dependent patent claims.

[0007] A first aspect of the invention relates to a device for detecting vaporization light scattering of a sample. The device comprises: a conduit for conveying a medium containing a sample in a main flow direction of the device, an atomizer that can be connected to the conduit for transporting a liquid, an evaporation tube, and a detector unit.

[0008] The vaporization tube forms a fluid connection between the atomizer and the detector unit. A valve is located between the line and the atomizer, which can be switched between a single-flow and a split-flow configuration. In the split-flow configuration, an initial measurement fraction of a medium containing a sample can be introduced from the line into the atomizer.

[0009] A split-flow configuration essentially refers to a splitting of the river into at least two different flow directions.

[0010] "Single flow configuration" refers to a line flow without a split flow function.

[0011] A valve is understood to be a passage that can be opened and closed. The valve is located directly in front of the atomizer.

[0012] The term "first measurement fraction" refers to the portion of the sample that is introduced into the atomizer and destroyed in the evaporation analysis.

[0013] The term "second measurement component" refers to the portion of the sample that is not directed into the atomizer and therefore not into the device, and which is preferably recoverable / collectible via the main flow direction.

[0014] The term "medium" refers to a liquid, gas, or gas mixture in which the sample dissolves or disperses. Examples of such mediums include organic solvents or water, such as acetone, acetonitrile, methanol, propanol, chloroform, cyclohexane, dichloromethane, or mixtures thereof. Examples of gases or gas mixtures include air, nitrogen, or argon, preferably inert gases.

[0015] The detector unit can, for example, include a fine dust sensor capable of measuring particle sizes from 0.3 to 2.5 µm. One example of such a sensor is the SPS30 fine dust sensor from Sensirion AG (Stäfa, Switzerland). However, other manufacturers with the same operating principle and measurement task are also suitable.

[0016] The device according to the invention has the further advantage that, with the valve positioned directly upstream of the atomizer, the introduction of a sample into the device and the atomization can occur practically simultaneously. Furthermore, this arrangement ensures that only a minimal portion of the sample is subjected to destructive ELS detection. The majority of the sample bypasses the atomizer via the main flow direction. This allows the sample to be additionally detected by UV measurement, as is common practice in liquid chromatography, and subsequently collected, for example, in a fraction collector. Sample loss is thus minimized.

[0017] The atomizer can comprise an inlet opening, a chamber fillable with a carrier gas, and an outlet opening. Preferably, the inlet opening includes the valve. The valve can open in a time- and frequency-controlled manner, thus metering the inlet quantity. Air, for example, and preferably an inert gas such as nitrogen or a noble gas, preferably argon, can be used as the carrier gas. The gas pressure is preferably slightly lower than the fluid pressure of the system. This ensures that no gases are forced back into the fluid system. A pressure ratio of the fluid system pressure to the atomizer pressure in the range of 1.5:1 and 1.1:1 is preferred, more preferably 1.3:1 and 1.1:1, and particularly preferably 1.2:1 and 1.15:1. A pressure in the atomizer below 2 bar is particularly preferred when the fluid system has a pressure of 2 bar. For example, the atomizer pressure can be in the range of approximately 1.7 bar.

[0018] Alternatively, although not claimed in the claims, the atomizer can comprise an inlet opening and an outlet opening, which are interconnected via a sample line. A sheath gas stream arrangement is arranged to at least partially enclose the atomizer, such that the outlet opening can be surrounded by a gas flow, enabling preferably direct atomization of the sample at the outlet opening. The passage of the sample to the outlet opening is preferably based on the pressure of the liquid.

[0019] Such an arrangement of the atomizer allows for optimized atomization and delivers high reproducibility of the results.

[0020] The valve is advantageously a microvalve, preferably an electromagnetic microvalve. An electromagnetic microvalve is actuated electromagnetically and is typically directly flowed through by the medium. In the de-energized state, the valve is closed. A closing spring acts on a movable armature with a valve ball. When the valve coil is energized, the movable armature with the valve ball is magnetically attracted by the magnetic field of the stationary armature, the microvalve opens, and the medium exits. Preferably, the valve can have a hard-sealing valve assembly made of hard materials. For example, the valve seat can be made of sapphire and the valve ball of ruby. Hard-sealing valve assemblies enable precise opening strokes of at least hundredths of a millimeter and repeatability in the range of thousandths of a millimeter.The finely ground and polished internal shapes of the valve nozzle and valve seat ensure optimal flow characteristics before the medium exits. This enables a precise metering jet for contactless dispensing and an optimal droplet shape. Furthermore, the hard material components used ensure a long service life and exhibit high chemical and mechanical resistance as well as compatibility.

[0021] The outlet opening is preferably a nozzle.

[0022] In a particularly preferred embodiment, the inlet opening with valve and the outlet opening of the atomizer each have an axis of symmetry that are not coaxial but offset from each other. Preferably, the axis of symmetry of the valve is asymmetrical with respect to the chamber, i.e., not on a central axis of the chamber, and the axis of symmetry of the outlet opening is symmetrical with respect to the chamber, i.e., on a central axis of the chamber.

[0023] It has surprisingly been shown that this arrangement results in a particularly advantageous atomization and further optimizes the sample introduction, so that the introduction of the smallest sample quantities is sufficient for maximum detection.

[0024] In a particularly preferred embodiment, the medium containing the sample flows transversely through the valve; that is, the valve has two transverse bores, preferably in the valve head, through which the medium can flow in a main flow direction transverse to the inlet opening of the atomizer, preferably at an angle of 90°. However, the main flow direction can also be at an angle greater than 90° with respect to the atomization direction. Thus, the flow direction of the first measurement component can be arranged transversely to at least part of the main flow direction. For example, the valve head can have two bores and be configured as a so-called "cross-head."

[0025] A main flow direction running perpendicular to the atomization direction has the advantage that the pressure drop across the valve is reduced when the fluid enters the atomizer.

[0026] However, it is also conceivable that the main flow direction initially runs in the direction of atomization and only becomes angled from the atomizer onwards. The medium can then flow around the valve. The valve head can then only have one bore, so that the main flow only partially passes through the valve head transversely.

[0027] Preferably, the inlet opening of the atomizer and the amount of the initial sample introduced can be controlled or controllable via the switching frequency and / or the opening duration of the valve. Short cycle times enable simultaneous sample splitting and introduction. Short cycle times also allow for control of the droplet size. In this way, the sample introduction can be precisely controlled, keeping the sample introduction as low as possible. For example, the switching frequency can be 20 to 50 Hz. The opening duration can be 50 to 300 ms. The sample introduction is preferably less than 30 µL. The switching can be controlled by environmental parameters, such as pressure and flow rates. Thus, the switching frequency can adapt to changes in pressure and / or flow rate, ensuring that the sample introduction is always optimized for the prevailing conditions in the system.

[0028] However, it is also possible to set the switching frequency and / or opening times before the detection begins.

[0029] The atomizer can be connected to a flushing unit, which can be part of the device according to the invention. The flushing unit serves the purpose of expelling residual amounts of medium from the valve. Such an arrangement is particularly advantageous for "cross-head" valves, i.e., when the main flow direction is perpendicular to the atomizer. Air is preferably used as the flushing medium, but pure nitrogen or a noble gas, preferably argon, or other inert gases or gas mixtures are also conceivable.

[0030] The detector unit may preferably include an optical detector. In evaporative light scattering (ELSD) detection, such as the one described here, the light scattered by the cloud of sample particles is captured by a photodiode and converted into an electrical value [mV]. The higher the reflection, i.e., the amount of light striking the diode, the higher the mV value. ELSD therefore does not focus on absorption at specific wavelengths, but rather on detecting the presence or absence of a sample in the transport air within the evaporation tube.

[0031] In particular, the detector unit comprises an optical block including a lens, preferably an aspherical lens. An aspherical lens has the property of focusing scattered light and other diffuse light rays onto a defined point. This increases the signal strength and allows filter functions for homogenizing the displayed signal without introducing significant signal noise.

[0032] Furthermore, the evaporation tube can be heated. With a heated tube, the evaporation time is further optimized. However, it is also possible to leave the evaporation tube unheated. An evaporation tube in which the heating function can be switched on as needed is particularly preferred. This allows for evaporation optimized for the media used, without the need for continuous heating, especially with highly volatile substances. This enables particularly energy-efficient operation of the device.

[0033] The evaporation tube can be made of metal, preferably steel, or glass. Solvent-resistant plastics are also conceivable.

[0034] The evaporation tube can be linear, spiral, L-shaped, or U-shaped. Almost any shape is possible, as long as sufficiently efficient evaporation is achieved over the given distance. For example, the evaporation tube can also be of varying lengths depending on available space and / or the specific application.

[0035] Furthermore, the device's inlet can be connected to a separation column, particularly a high-performance chromatography column. This allows, for example, the use of ELS detection in addition to the UV-Vis detection typically used in chromatographic methods. On the one hand, this enables additional verification of already identified substances, and on the other hand, it also allows the identification of substances that do not absorb in the UV-Vis range.

[0036] Another aspect of the invention relates to an analytical instrument, in particular a chromatography instrument, comprising a device for detecting evaporation light scattering of a sample as described above.

[0037] Preferably, the chromatography apparatus is a high-performance chromatography (HPLC) system. An HPLC system typically comprises an eluent reservoir, electromagnetic mixing valves with a double-stroke piston pump, multi-way valves, pressure compensation loops to equalize pump pulses, a mixing chamber, a manual injection valve, a separation column, an HPLC unit, a detector unit, a computer interface, a PC, and optionally a printer for outputting the results. The eluate exiting the separation column can be guided via a main flow direction to the detector unit, preferably a UV-Vis detector, e.g., a photodiode array detector.Advantageously, the device according to the invention for detecting evaporation light scattering is arranged in the main flow direction between the separation column and the detector unit, so that in the split flow configuration of the valve a first measurement component can be directed into the atomizer of the device and a second, usually larger, measurement component can be directed into the UV-Vis detector unit of the HPLC device.

[0038] To ensure that a differential pressure between the air and the fluid is always maintained, the pressure drop that can be caused by the different flow rates can be taken into account by the system's control mechanism. This means that if the fluid pressure in the system increases, the gas pressure must be adjusted accordingly so that the pressure difference remains constant. The same applies if the pressure in the fluid system decreases.

[0039] In principle, the analytical instrument can encompass all devices that detect solids in a carrier medium that are of interest to the user and are therefore specifically collected in subsequent process steps. This can be useful, for example, in addition to chromatography, wherever substances are present that do not absorb UV light and therefore cannot be detected by conventional detectors.

[0040] It is also possible to use only the device itself as an analyzer.

[0041] Another aspect of the invention relates to a method for detecting vaporization light scattering from a sample. The method comprises the following steps: a) Introducing a medium containing a sample into a line, in particular into a device as described above, b) Simultaneously introducing and atomizing a first measurement fraction of the medium containing the sample into an atomizer, c) Evaporating the atomized medium, d) Measuring the evaporation light scattering of the sample.

[0042] The evaporation of the medium in step c) can be carried out with or without additional heating of the evaporation tube, depending on the volatility of the medium.

[0043] The simultaneous insertion and atomization of the first measurement component in step b) can be carried out in predefined time intervals.

[0044] However, the introduction and atomization of the first measurement component in step b) can also be carried out depending on system parameters, preferably pressure and / or flow rate.

[0045] For example, the atomizer's inlet opening can be controlled or adjustable via the switching frequency and / or the opening duration. The circuit can be timed so that atomization occurs simultaneously with the inlet. The droplet size can also be adjusted via the cycle time. Control can be achieved, for example, via a separate program, such as dedicated application software. This software can be designed to allow the user to select and set specific parameters. Alternatively, the user may not need to set or monitor any parameters, with the control occurring automatically in the background. For instance, as described above, control can be based on environmental parameters such as pressure and flow rate, or on pre-defined switching frequencies and / or opening times.

[0046] The medium containing the sample was preferably processed prior to step a) by means of a separation process, preferably a chromatographic process. The medium containing the sample can then be an eluate.

[0047] The invention is explained in more detail below with reference to specific embodiments. These examples are not to be understood as limiting. The same reference numerals denote the same elements.

[0048] They show: Figure 1A: A perspective view of a first device according to the invention. Figure 1B: Front view of the first device according to the invention. Figure 1A with sectioning planes. Figure 2: a side view along plane AA from Figure 1B Figure 3: An enlargement of section X from the atomization module according to Figure 2 Figure 4: a horizontal sectional view through the first device according to the invention along the plane KK in Figure 1BFigure 5: An enlargement of section Y from the detector unit. Figure 4 Figure 6: Another sectional view of the first device according to the invention along the plane GG in Figure 1B. Figure 7: An enlargement of the section W from the detector unit. Figure 6 Figure 8: Another side view of the first device according to the invention along plane FF in Figure 1B Figure 9: an enlargement of section Z from Figure 8 Figure 10: A side view of a second embodiment according to the invention with section planes. Figure 11: A side view of the second embodiment along plane BB. Figure 10 Figure 12: an enlargement of section X from Figure 11 Figure 13: a sectional view of the second embodiment along a horizontal plane DD in Figure 10 Figure 14: an enlargement of section Y from Figure 13Figure 15: A side sectional view of a second embodiment of an atomizer module. Figure 16: A perspective sectional view of the atomizer module made of Figure 15 Figure 17: Another sectional view of the atomizer module made of Figures 15 and 16 .

[0049] Figure 1A Figure 1 shows a perspective view of a first device 1a according to the invention. A atomization module 2 is shown with a fluid inlet 3, preferably for an eluate, and a fluid outlet 4. A line (not shown) is arranged between the inlet 3 and outlet 4, which allows the passage of a sample, preferably an eluate. In the perspective view, a vaporization tube 5 is located behind the atomization module 2 and the base support 10 of the device 1a in a U-shaped arrangement. The vaporization tube 5 opens into a detector unit 6. A housing 9 is arranged above the detector unit 6, in which a laser 7 is adjustable via a dial 8.

[0050] Figure 1B Figure 1a shows the device according to the invention. Figure 1A in a front view showing the section planes relevant for the following images.

[0051] Figure 2 shows a side view of the device 1a according to the invention along the plane AA in Figure 1B with a detailed representation of the interior view of the atomization module 2. Section X is shown with reference to Figure 3 explained in more detail.

[0052] Figure 3 shows section X from Figure 2in a magnified view. Inside the atomization module 2 is a solenoid valve 21. The atomization chamber 22 is formed by the end face of the solenoid valve 21 and the outlet opening in the form of a nozzle 23. The nozzle 23 opens into the vaporization tube 5. The solenoid valve 21 has an axis of symmetry, as does the nozzle; both axes are not coaxial but offset from each other. The axis of symmetry of the valve is asymmetrical with respect to the chamber, while the axis of symmetry of the nozzle is symmetrical with respect to the chamber. A purge unit can be arranged at the end of the valve 21, as shown in Figure 10 described using an alternative embodiment. The main flow direction (not shown) runs transversely, i.e., at an inclined angle, to the solenoid valve 21. Air from the flushing unit flows longitudinally from back to front through the solenoid valve (not shown) and exits at the outlet 4 ( Figure 2 ) and thus empties the rest of the system as well.

[0053] Figure 4 Figure 1a shows the device according to the invention in a horizontal sectional view along the plane KK in Figure 1B Also visible here are, among other things, the atomization module 2, the vaporization tube 5, and the detector unit 6. Section Y of the detector unit 6 is shown in Figure 5 described in more detail.

[0054] Figure 5 shows section Y from Figure 4in a magnified view. The essential elements of the detector unit 6 are shown in detail. The detector unit 6 is arranged such that a medium containing the sample passes through the detector unit 6 in the measuring area 61 from the vaporization tube. In the measuring area 61, the light of the laser beam strikes the sample particles and is scattered by them. The scattered radiation 62 then strikes a lens 63, e.g., a spherical lens, perpendicular to the incident laser light, and is focused by it and directed onto a photodiode 64. The resulting electric current is processed by the electronic interface 65 and output as a signal.

[0055] Figure 6 shows a further sectional view of the device 1a according to the invention along the plane GG in Figure 1BThe atomizing module 2 can be seen in the lower part of the image. The atomizing module 2 comprises the inlet 3 and the outlet 4, which are connected by a line 25. The line crosses the valve 21 at an inclined angle to the atomization direction 26 (arrow). In another embodiment, the line can run vertically at a right angle to the atomization direction ( Fig. 10-13 After atomization, the sample is guided through the vaporization tube 5 in the atomization direction 26 until it is detected in the detector unit 6. Above the detector unit, the laser 7 is arranged in a housing 9 and fixed with the adjusting wheel 8.

[0056] Figure 7 shows section W of the Figure 6In an enlarged view. Shown is a second perspective of the measuring area 61, in which the laser light 71 falls on sample particles (not shown) and is scattered. The scattered light 62 is focused at the lens 63, and the current generated at the photodiode 64 is processed via the electronic interface 65 and output as a signal.

[0057] Figure 8 shows again a side view of the device 1a according to the invention along the plane FF in Figure 1B The figure shows, in addition to the elements already mentioned, a light trap 10 below the laser 7. The light trap 10 captures non-scattered light from the laser and thus prevents reflection that could lead to a distorted measurement result. Details of the light trap 10 are shown in Figure 9 shown.

[0058] Figure 9 The detailed view Z shows Figure 8in a magnified view. The light trap 10 is attached by means of screws 101 below the laser source and below the measuring zone 61 ( Fig. 8 ) attached to the detector unit 6. Non-scattered light passes through an aperture 102 onto a light-absorbing element 103.

[0059] Figure 10 Figure 1 shows an alternative embodiment in a front view with the section planes indicated. The device is described in more detail below with reference to these section planes.

[0060] Figure 11 The alternative embodiment 1b of the device according to the invention is shown along the plane BB in Figure 10Device 1b also includes the atomizing module 2, an inlet 3 (and an outlet, which is not shown) and an evaporation tube 5. In addition, device 1b has a connection 24 for a purging unit, through which a purging gas can be introduced into the atomizing module 2 and cleans the atomizing module 2 and in particular the valve 21 of residual medium.

[0061] Section X is enlarged in Figure 12 as shown. In contrast to the previous embodiment, the line 25 pierces the valve 21 at a right angle and is therefore perpendicular to the atomizing nozzle 23 or the atomizing direction 26 ( Figure 11 ) arranged.

[0062] Figure 13 shows the alternative embodiment of device 1b along the plane DD in Figure 10 , so that both the atomization module 2 with its individual elements and the detector unit 6 are visible. The detector unit 6 is described with reference to Figure 14 explained in more detail.

[0063] In Figure 14 It can be seen that the light 62 scattered in the scattering area 61 strikes an aspherical lens 67, is focused behind it, and then strikes a photodiode 64. The signal generated by the photodiode 64 is processed via the electronic interface 65.

[0064] In principle, the individual elements of the first and second embodiments shown here can be combined in any way and are not limited to the respective embodiments.

[0065] Figures 15 to 17 show various views of a second embodiment of an atomizer module 200. Instead of an atomizer chamber (22, Figure 3The atomizer module 200 comprises an atomizer with a sample line 222, which connects the inlet opening 220 with solenoid valve 221 and the outlet opening 223. The liquid or solvent containing the sample is brought to the outlet opening 223 by the liquid pressure. A sheath gas flow arrangement 224 is arranged at least partially around the atomizer module 200. The sheath gas flow arrangement 224 allows gas to flow around the outlet opening 223 in such a way that direct atomization can occur. The fluid inlet 3 and the fluid outlet 4 for passing the eluate through the atomizer are also arranged transversely at an inclined angle to the atomization direction.

[0066] In Figure 16 An additional gas connection 11 is shown. As in Figure 17 It can be seen that gas can be directed into the mantle gas flow arrangement via an additional gas line 12.

Claims

1. A device (1a, 1b) for detecting evaporative light scattering of a sample, the device (1a, 1b) comprising: - a conduit (25) for conveying a medium containing a sample in a main flow direction of the device (1a, 1b), - an atomizer connectable to the conduit (25) for transporting a liquid, - an evaporation tube (5), and - a detector unit (6), wherein the evaporation tube (5) forms a fluid connection between the atomizer and the detector unit (6), characterized in that a valve (21) is arranged between the conduit (25) and the atomizer and immediately upstream of the atomizer, wherein the valve (21) can be switched between a single-flow configuration and a split-flow configuration, wherein the single-flow configuration is a conduit flow without a split-flow function, wherein in the split-flow configuration a first measurement portion of a medium containing a sample can be introduced from the conduit (25) into the atomizer.

2. The device (1a, 1b) according to claim 1, wherein the atomizer comprises an inlet opening (21), a chamber (22) that can be filled with a carrier gas, and an outlet opening (23).

3. The device (1a, 1b) according to one of the preceding claims, wherein the valve (21) is a microvalve, preferably an electromagnetic microvalve.

4. The device (1a, 1b) according to one of claims 2 to 3, wherein the inlet opening (21) of the atomizer is controlled or controllable via the switching frequency and / or the opening duration.

5. The device (1a, 1b) according to one of the preceding claims, wherein a flow direction of the first measurement portion is arranged transversely to at least part of the main flow direction.

6. The device (1a, 1b) according to one of the previous claims, wherein the atomizer is connected or connectable to a flushing unit (24).

7. The device (1a, 1b) according to one of the preceding claims, wherein the detector unit (6) comprises an optical detector.

8. The device (1a, 1b) according to one of the preceding claims, wherein the evaporation tube (5) is heatable.

9. The device (1a, 1b) according to one of the preceding claims, wherein the evaporation tube (5) is made of metal, preferably steel, or glass.

10. The device (1a, 1b) according to one of the preceding claims, wherein the evaporation tube (5) is linear, spiralshaped, L-shaped, or U-shaped.

11. The device (1a, 1b) according to one of the preceding claims, wherein the conduit (25) is connectable or connected on the input side to a separation column, in particular a separation column for high-performance chromatography.

12. An analytical device, in particular a chromatography device, comprising a device (1a, 1b) for detecting evaporation light scattering of a sample according to one of claims 1 to 11.

13. A method for detecting evaporative light scattering of a sample, comprising the steps: a) Introducing a medium containing a sample into a conduit (25) of a device (1a, 1b) according to any one of claims 1 to 11, b) simultaneously introducing and atomizing a first measurement portion of the medium containing the sample into an atomizer (23), c) Evaporating the atomized medium, d) measuring the evaporation light scattering of the sample.

14. The method according to claim 13, wherein the introduction and atomization of the first measurement portion in step b) takes place at predefined time intervals.

15. The method according to claim 13, wherein the introduction and atomization of the first measurement portion in step b) is performed as a function of system parameters, preferably pressure and / or flow rate.

16. The method according to one of claims 13 to 15, wherein the medium containing the sample is processed prior to step a) by means of a separation process, preferably a chromatography process.