Qualification of a detector device using Raman scattering, and analysis device

Abstract

A detector device (100), in particular for an analysis device (10), is described, the detector device (100) comprising: i) a liquid receiving device (120) for receiving a Raman-active liquid (121); and ii) a control device (70) arranged to: a) detecting Raman scattering (150) with respect to the Raman-active liquid (121) in the liquid receiving device (120), and b) qualifying the detector device (100) with respect to at least one wavelength based on the detected Raman scattering (150).

Description

FIELD OF THE INVENTION

[0001] The present invention relates to a detector device, in particular for an analysis device, wherein the detector device comprises: a liquid receiving device for receiving a Raman-active liquid; and a control device configured to: detect Raman scattering with respect to the Raman-active liquid in the liquid receiving device; and qualify the detector device with respect to at least one wavelength based on the detected Raman scattering. The invention further relates to the analysis device, a method for operating a detector device, and a specific use of Raman scattering. TECHNICAL BACKGROUND

[0002] Analytical devices such as sample separation devices are designed to analyze a sample, particularly a fluidic sample, e.g., to perform a chromatographic separation of the sample. For example, in an HPLC (high performance liquid chromatography) analytical device, a liquid (mobile phase) is moved through a so-called stationary phase (e.g., in a chromatographic column) at a very precisely controlled flow rate (e.g., in the range of microliters to milliliters per minute) and at a high pressure (typically 20 to 1000 bar and beyond, currently up to 2000 bar), at which the compressibility of the liquid can be noticeable, in order to separate individual fractions of a sample liquid introduced into the mobile phase. After passing through the stationary phase, the separated fractions of the fluidic sample are detected in a detector.Such an HPLC system is known, for example, from EP 0,309,596 B1 of the same applicant, Agilent Technologies, Inc.

[0003] Such a detector, for example, comprises, within a housing, a lamp for illuminating (or optically exciting) the fluidic sample in a flow cell and a sensor for detecting fluorescence (induced by the optical excitation) of the fluidic sample in response to the illumination. An analytical instrument, such as an HPLC, can be configured as a stack of modules, one of which is the detector module.

[0004] In a conventional detector, such as a fluorescence detector, the fluidic sample in the flow cell is first excited by light from the lamp. This excites the fluidic sample to fluoresce, which in turn is detected by the actual detector, e.g., a light sensor. A monochromator is usually provided both between the lamp and the fluidic sample and between the fluidic sample and the sensor to select the desired wavelength or wavelength range.

[0005] Monochromators ensure that only light of a specific wavelength is passed through the sample and / or from the sample to the detector for the measurement of a specific absorption / fluorescence. Precise wavelength adjustment is crucial for correct qualitative and quantitative analysis. Deviations in wavelength calibration can lead to inaccurate measurements or misinterpretation of peaks. Therefore, regular wavelength calibration is required to ensure the accuracy and reliability of the detector.

[0006] Conventionally, atomic emission lines from the lamp can be used for calibration, for example, with the flow cell filled with water or a water-acetonitrile (ACN) mixture. This conventional method relies on a special flow cell design and / or the use of glycogen to ensure that only the signal induced by the fluid in the flow cell is measured.

[0007] Furthermore, filters such as holmium and / or erbium filters can be used; these absorb light at defined wavelengths in the UV-VIS range. National Institute of Standards and Technology (NIST) standards are also available. However, these approaches can be problematic with high-bandwidth monochromators, as the positions of the absorption lines shift due to the spectral properties.

[0008] A comprehensive solution to address these requirements both technically and economically remains a challenge. REVELATION

[0009] There may be a need to efficiently and reliably qualify a detector device, particularly for an analysis device. This object is achieved by means of the independent claims. Further embodiments are shown in the dependent claims.

[0010] According to a first aspect of the invention, a detector device (e.g. a fluorescence detector) is described, in particular for an analysis device (e.g. an HPLC), the detector device comprising: i) a liquid receiving device (e.g. a flow cell) for receiving a Raman-active liquid (e.g. water, alcohol, alkane, etc.); and ii) a control device (e.g. a system control, a controller, a hardware / software, a processor, in the detector device and / or at least partly remotely, etc.) arranged to: a) detecting (e.g. by means of a sensor) Raman scattering with respect to the Raman-active liquid in the liquid receiving device (e.g. when the Raman-active liquid has been excited / irradiated by a lamp), and b) Qualifying (e.g. calibrating, verifying) the detector device (in particular at least one monochromator of the detector device) with respect to at least one wavelength based on the detected Raman scattering.

[0011] According to a second aspect of the invention, an analysis device is described which comprises a detector device as described above.

[0012] According to a third aspect of the invention, a method for operating a detector device (e.g. as set out above) is described, the method comprising: i) providing a Raman-active liquid (e.g., in a flow cell); ii) detecting Raman scattering with respect to the Raman-active liquid; and iii) qualifying the detector device with respect to at least one wavelength based on the detected Raman scattering.

[0013] According to a fourth aspect of the invention, a use of Raman scattering with respect to a Raman-active liquid in a detector (in particular fluorescence detector) of an analysis device is described in order to calibrate / verify at least one wavelength (with respect to a monochromator of the detector device).

[0014] In the context of the present document, the term "detector device" is understood in particular to mean a device that is suitable for detecting / capturing / measuring measurement data, in particular with regard to an analysis of a fluidic sample. For example, the detector device can detect electromagnetic radiation that is indicative of an analysis or nature of the fluidic sample. A detector device can, for example, measure the presence and / or concentration of the substances in the fluidic sample after their separation. In one embodiment, a detector device can be used to convert physical or chemical properties of eluted substances, e.g., UV absorption, refractive index, fluorescence, or electrical conductivity, into an electrical signal, which can then be displayed, e.g., as a chromatogram. In one embodiment, a detector device can, for example,a liquid receiving device (for the fluidic sample), an excitation device (e.g. for fluorescence excitation), and a sensor device (as the actual detector).

[0015] In the context of this document, the term "Raman scattering" refers specifically to a physical phenomenon in which electromagnetic radiation (e.g., light) is scattered by molecules as it passes through a material (e.g., a liquid), thereby changing its energy and shifting the wavelength. In contrast to elastic Rayleigh scattering, Raman scattering is inelastic scattering (energy transfer, Stokes shift). The Raman shift (e.g., in cm -1) is characteristic of a particular substance, a particular molecular species, and can therefore be used as a fingerprint for that substance. Raman scattering can vary slightly due to factors such as temperature; for water, for example, it is around 3200-3400 cm -1 .

[0016] In the context of this document, the term "Raman-active liquid" (or Raman-sensitive liquid, liquid with Raman-active molecules, Raman reference substance) is understood in particular to mean a liquid whose molecules cause Raman scattering or a characteristic Raman shift upon interaction with electromagnetic radiation (e.g., due to certain molecular vibrations). In one embodiment, irradiation / excitation using electromagnetic radiation (excitation radiation) produces inelastically scattered electromagnetic radiation (emission radiation) with a Raman-shifted wavelength. In one embodiment, the Raman-active liquid can be one of the following: water, certain alcohols such as ethanol, certain alkanes. Examples of non-Raman-active substances can be, for example, atomic ions in solution.

[0017] In the context of this document, the term “qualification” is understood in particular to mean that a detector device (e.g., a monochromator of the detector device) is evaluated / checked to determine whether it is functioning properly and / or is suitable for the intended purpose. In other words, “qualification” can mean that a device (e.g., detector) is assessed according to defined criteria to determine whether it is technically suitable and correctly functioning for its application, and is then adapted if necessary. In one embodiment, qualification can relate to calibrating the wavelength(s) of a monochromator of the detector device. In a further embodiment, qualification can relate to verifying the wavelength(s) used by the monochromator.

[0018] In the context of the present document, the term "liquid receiving device" refers in particular to a device suitable for (temporarily) receiving a liquid, e.g., a Raman-active liquid. In a simple example, the liquid receiving device can be designed as a container. In a more complex embodiment, the liquid receiving device can be provided as a flow-through chamber, in particular as a flow-through cell, so that the fluidic sample (in particular, separated fractions in the mobile phase) can flow through the liquid receiving device, in particular during detection ("online" measurement). In one embodiment, the liquid receiving device is at least partially transparent to electromagnetic radiation.

[0019] In the context of this document, the term "fluid" is understood to mean, in particular, a liquid and / or a gas, optionally comprising solid particles. The term "fluid" can also refer to a mobile phase in which a fluidic sample is transported. If electromagnetic radiation interacts with the fluid, this can also mean, in one example, that the electromagnetic radiation only interacts with the sample components contained in the fluid and not with the components of the mobile phase. In the context of this document, the term "fluidic sample" is understood to mean, in particular, a medium, more particularly a liquid, which contains the material to be analyzed (for example, a biological sample such as a protein solution or nucleic acids, a pharmaceutical sample, etc.).

[0020] In the context of this document, the term "mobile phase" refers in particular to a fluid, more particularly a liquid, that serves as a carrier medium for transporting the fluidic sample in or through an analytical device. However, the mobile phase can also influence the processing of the fluidic sample in the sample separation device or contribute significantly by influencing the interactions of the sample components with the stationary phase. For example, the mobile phase can be a solvent (e.g., organic and / or inorganic) or a solvent combination (e.g., water and ethanol).

[0021] In the context of this document, the term "analysis device" can refer, in particular, to a device that is capable and configured to examine a fluidic sample, in particular to separate it, and further, in particular, to separate it into different fractions. For example, such sample separation can be carried out by means of chromatography or electrophoresis. Preferably, the analysis device can be a liquid chromatography sample separation device.

[0022] According to an exemplary embodiment, the invention may be based on the idea that a detector device, in particular for an analysis device, can be qualified efficiently and reliably if the qualification (with respect to wavelengths) is carried out based on Raman scattering from a Raman-active liquid in the liquid receiving device of the detector device.

[0023] According to the invention, a precise wavelength reference from atomic emission lines can be obtained using only the Raman-active liquid (e.g., water). This simplifies calibration or verification of the wavelength scale while ensuring high accuracy. This circumvents problems that occur with scattering in water-filled flow cells, as this scattering can be significantly shifted by undesirable effects such as scratches or impurities. The pure Rayleigh scattering signal of water, for example, is of a similar magnitude to that of quartz glass, but significantly lower than that of small defects in cell walls.

[0024] The invention can provide a simple and precise method for calibrating wavelengths or verifying the wavelength calibration of the detector, for example, by utilizing the Raman peaks of water generated by one or more atomic emission lines of an excitation device. Preferably, the detector signal is generated exclusively by the volume of the liquid receiving device, i.e., the fluid filling the liquid receiving device, so that additional substances such as glycogen are not required.

[0025] According to the invention, the use of additional substances such as glycogen can thus become unnecessary (this is otherwise used because it scatters widely and remains confined to the cell volume). However, the disadvantage of glycogen is that it must be prepared and injected manually, which precludes automation. The use of additional hardware such as filters (see holmium or erbium filters) or costly NIST standards can also be eliminated.

[0026] One aspect of the invention can be seen in the fact that the accuracy of the spectral positions of the atomic emission line (discrete wavelengths) can be combined with the advantage that the signal can be extracted exclusively from the volume of the flow cell, even when using water. EXEMPLARY IMPLEMENTATION EXAMPLES

[0027] In one embodiment, the detector device is configured as a fluorescence detector (device) (or the detector is configured as a fluorescence detector). This can have the advantage that the described qualification can be implemented directly in established and widely used detectors in analytical technology. A fluorescence detector is, in particular, a device that measures the fluorescence of a substance, in particular a fluidic sample. For this purpose, the sample to be analyzed is irradiated / excited with excitation radiation, and then the emission radiation (fluorescence) of the fluidic sample is measured (the electromagnetic radiation that molecules re-emit after excitation, preferably at a specific wavelength). In addition to or alternatively to fluorescence, the excitation radiation can cause Rayleigh scattering and Raman radiation, especially when the excitation radiation is applied to a Raman-active liquid.

[0028] In one embodiment, qualifying comprises calibrating and / or verifying the at least one wavelength. In one embodiment, qualifying is performed with respect to a (first and / or second) monochromator device. For a detector device, such as a fluorescence detector, it may be particularly important that the wavelengths can be precisely selected. This applies to the excitation radiation (in order to provide one or more specific excitation wavelengths) and / or to the emission radiation (in order to provide one or more specific emission wavelengths). Based on the measured Raman scattering (whose Raman shift is characteristic of a specific liquid such as water), these wavelengths can be particularly efficiently adjusted (calibrated) and / or checked (verified).

[0029] In one embodiment, the liquid receiving device is designed as a flow-through chamber, in particular a flow cell / flow-through cell. In one embodiment, the liquid receiving device is configured such that the Raman-active liquid can flow through the chamber. This can have the advantage that the Raman-active liquid can be introduced directly into the existing measuring chamber of the detector device. Flowing through this chamber can make the qualification process particularly efficient. Furthermore, qualification (e.g., readjustment) can be performed virtually during ongoing operation ("online"), i.e., with the flow through the chamber.

[0030] In one embodiment, the detector device further comprises an excitation device (in particular a lamp or a laser) for providing electromagnetic excitation radiation to the liquid storage device (in particular the Raman-active liquid therein). Existing structures of the detector device can advantageously be used directly to provide the excitation radiation to the Raman-active liquid (thereby causing Raman scattering).

[0031] In one embodiment, the excitation device is configured to provide at least two, in particular at least four, discrete wavelengths. This can have the advantage of increasing precision and reliability. The discrete wavelengths or discrete wavelength peaks can be specific wavelengths at which a (local) intensity maximum is present (cf. Fig. 4). The discrete wavelengths can also be referred to as atomic emission lines or discrete excitation wavelength peaks. Each discrete wavelength peak can be used as an excitation wavelength, e.g., selected by a first monochromator device. Each discrete excitation wavelength can then lead to a corresponding Raman peak (in the emission radiation) (cf. Fig. 6A). This allows multiple Raman peaks to be obtained (e.g., in the excitation wavelength / emission wavelength diagram), which can increase reliability.

[0032] In one embodiment, the excitation device comprises at least one of the following: a gas discharge lamp, a noble gas lamp (e.g., a xenon lamp or an argon lamp), a mercury-xenon (HgXe) lamp, or a cadmium lamp. In one embodiment, line radiation sources can be used, which can in particular comprise, for example: i) low-pressure discharge (Hg, Na, Cd lamps), ii) high-pressure discharge (HgXe, metal halide), iii) hollow cathode lamps (HCL), iv) electrodeless discharge lamps (EDL), v) laser sources (for single lines). This can have the advantage that discrete (or clearly defined) wavelengths / wavelength ranges can be used for the excitation radiation. Furthermore, established and reliable devices can be used directly. In one embodiment, an HgXe lamp is used, whose atomic emission lines have multiple maxima over a wide bandwidth (cf. Fig. 4). Each of these (preferably several) discrete wavelengths (peaks) can be used to efficiently and robustly generate Raman scattering. In another embodiment, a laser can be used as the excitation device. This can have the advantage of a clearly defined wavelength.

[0033] In one embodiment, the detector device further comprises: a first monochromator device (in particular for receiving the electromagnetic excitation radiation (from the excitation device)) for providing an excitation wavelength (or monochromatic excitation wavelength) to the liquid storage device (or for exciting the Raman-active liquid (in the liquid storage device)). This can have the advantage that the desired excitation wavelength can be precisely and reliably set / selected. Furthermore, the first monochromator device can be qualified based on the detected Raman scattering.

[0034] In the present context, the term "monochromator" can in particular refer to an optical component that splits polychromatic electromagnetic radiation (e.g., white light) into individual wavelengths or wavelength ranges (e.g., colors), so that only a specific wavelength is transmitted (monochromatically). This is achieved, for example, using gratings or prisms in combination with narrow exit diaphragms. In one example, these gratings / prisms can be precisely moved using appropriate motors, so that wavelengths down to the lower nanometer range can be precisely selected / selected / adjusted. In one embodiment, it may be necessary to qualify such a monochromator.

[0035] In the present context, the term "wavelength" can also encompass "wavelength ranges" that, for example, border a central wavelength peak. The term "discrete wavelength" can, in one embodiment, encompass a discrete wavelength range, but exclude a larger bandwidth. In one embodiment, a wavelength of + / - 20 nm, in particular + / - 10 nm, in particular + / - 1 nm, in particular + / - 0.5 nm, and further in particular + / - 0.1 nm can be selected.

[0036] In one embodiment, the wavelength or wavelength range depends on the application. For example, very fine spectral details can be resolved with a UV-Vis spectrometer if the optical bandwidth is as small as possible (e.g., 0.1 nm). With an HPLC fluorescence detector, a high light yield may be desired, so that larger optical bandwidths (e.g., 15-20 nm) can be used to measure a significant, but still characteristic, portion of the usually broad emission spectra of the analytes. In one example, this bandwidth is large compared to the emission line; in relation to the Raman emission peak, it is in a similar range. For an individual emission line within the optical bandwidth, its measured intensity maximum position does not shift.

[0037] In one embodiment, the electromagnetic excitation radiation (in particular the excitation wavelength) excites the Raman-active liquid in the liquid storage device to emit electromagnetic radiation. In one embodiment, the electromagnetic emission radiation exhibits Raman scattering and / or Rayleigh scattering. This can have the advantage that Raman scattering can be provided in an efficient and simple manner, (essentially) without additional hardware.

[0038] In one embodiment, the detector device further comprises: a second monochromator device (in particular for receiving the electromagnetic emission radiation) for providing an emission wavelength. In one embodiment, the emission wavelength is provided to or received from a sensor device. This can have the advantage that the desired emission wavelength can be selected precisely and reliably. Preferably, the emission wavelength selected is the one at which the Raman scattering or Raman shift with respect to the Raman-active liquid is expected. The second monochromator device can be similar / identical to or different from the first monochromator device (as described above). In one embodiment, it may be necessary to qualify the second monochromator device.

[0039] In one embodiment, the second monochromator device is adjusted such that the wavelength associated with the expected Raman scattering (as the emission wavelength) can pass through. In one embodiment, the emission wavelength includes the Raman scattering. This allows for efficient detection of the Raman scattering.

[0040] In one embodiment, the detector device further comprises a sensor device for receiving the emission wavelength (in particular, the wavelength associated with Raman scattering). Raman scattering can thus be reliably detected / measured using established techniques. In one example, no additional hardware is required. In one embodiment, the sensor device comprises at least one of the following: photomultiplier, photodiode, or charge-coupled device (CCD).

[0041] In one embodiment, qualifying with respect to wavelength involves calibrating and / or verifying the first monochromator device and / or the second monochromator device. In one embodiment, the control device is configured to adjust an actual / current wavelength to the wavelength based on the detected Raman scattering. The Raman shift is characteristic of the Raman-active liquid, so the expected wavelength is known. This can be used, for example, to calibrate / verify the monochromator device.

[0042] In one embodiment, the Raman-active liquid comprises at least one of the following: water, an alkane (e.g., cyclohexane, n-hexane), or an alcohol (e.g., at least one of methanol, ethanol, or isopropanol). These are merely examples of a multitude of possible Raman-active liquids. Water may be particularly easy to handle in one example.

[0043] In one embodiment, the Raman scattering is associated with at least one discrete wavelength of the electromagnetic excitation radiation (e.g., by the defined Raman shift). In one embodiment, the Raman scattering exhibits a defined shift, thereby defining a specific wavelength (characteristic).

[0044] In one embodiment, the method is free from the use of an (additional) calibration substance, in particular glycogen. In one embodiment, the method is free from the influences of a flow-through chamber, in particular the design. In one embodiment, the method is free from qualification based on Rayleigh radiation. In one embodiment, the method is free from a filter, in particular a holmium / erbium filter. These features can have the advantage of overcoming disadvantages of the prior art (see above).

[0045] In one embodiment, the method is free of an excitation device that provides non-discrete wavelengths. Conventionally, large bandwidths are often used. According to the invention, however, it may be preferable to use the discrete wavelengths exclusively (e.g., of an HgXe lamp). This can increase the sensitivity / precision. A discrete, well-known wavelength can also result in a well-defined Raman peak position. The less precise the excitation wavelength, the less knowledge of the emission wavelength can be. The light source selected can be one with a more continuous spectrum or one with emission lines, for example, whereby a discrete wavelength can be selected for both light source types. A particular advantage of emission lines can be their higher intensity, which can then also enable higher sensitivity.

[0046] In one embodiment, the method comprises performing an analytical method after qualifying, in particular calibrating / verifying the wavelength of a first monochromator device and / or a second monochromator device. This ensures efficient and reliable detection in the analytical method.

[0047] In the context of the present application, the term "sample separation device" can be understood in particular as a device for analyzing a fluid sample, in particular into different fractions. For this purpose, components of the fluid sample can first be adsorbed on the sample separation device and then desorbed separately (in particular fractionally). For example, such a sample separation device can be designed as a chromatographic separation column.

[0048] According to one embodiment, the analysis device is a chromatography device, in particular a liquid chromatography device, a gas chromatography device, an SFC (supercritical fluid chromatography) device or an HPLC (high performance liquid chromatography) device.

[0049] According to one embodiment, the analysis device is configured as a microfluidic device. According to one embodiment, the analysis device is configured as a nanofluidic device.

[0050] According to one embodiment, the sample separation device is designed as a chromatographic separation device, in particular as a chromatography separation column.

[0051] According to one embodiment, the fluid drive is configured to drive the mobile phase and the fluidic sample under high pressure.

[0052] According to one embodiment, the fluid drive is configured to drive the mobile phase and the fluidic sample with a pressure of at least 500 bar, in particular of at least 1000 bar, further in particular of at least 1200 bar, further in particular of at least 1500 bar.

[0053] According to one embodiment, the analysis device comprises the detector device for detecting the analyzed, in particular separated, fluidic sample.

[0054] According to one embodiment, the analysis device comprises a fractionator for fractionating separate fractions of the fluidic sample.

[0055] The analytical device can be a microfluidic instrument, a life science device, a liquid chromatography device, a gas chromatography device, an HPLC (high-performance liquid chromatography), a UHPLC system, or an SFC (supercritical fluid chromatography) device. However, many other applications are possible.

[0056] According to one embodiment, the sample separation device can be designed as a chromatographic separation device, in particular as a chromatography separation column. In a chromatographic separation, the chromatography separation column can be provided with an adsorption medium. The fluid sample can be retained on this adsorption medium and only subsequently detached fractionally in the presence of a specific solvent composition, thus achieving the separation of the sample into its fractions.

[0057] A pumping system for conveying fluid can, for example, be designed to convey the fluid or mobile phase through the system at a high pressure, for example a few hundred bars up to 1000 bars and more.

[0058] The analysis device can have a sample injector for introducing the sample into the fluidic separation path. Such a sample injector can have a sample or injection needle that can be coupled to a needle seat in a corresponding fluid path, wherein the sample needle can be retracted from this needle seat to collect the sample. After reinserting the sample needle into the needle seat, the sample can be located in a fluid path that can be switched into the separation path of the system, for example, by switching a valve. In another embodiment of the invention, a sample injector or sampler with a sample needle that is operated without a needle seat can be used.

[0059] The analysis device may include a fraction collector for collecting the separated components. Such a fraction collector can, for example, direct the various components of the separated sample into different liquid containers. The analyzed sample can also be fed to a waste container. BRIEF DESCRIPTION OF THE DRAWINGS

[0060] Other objects and many of the attendant advantages of embodiments of the present invention will be readily appreciated and better understood by reference to the following more detailed description of embodiments taken in conjunction with the accompanying drawings. Features that are substantially or functionally the same or similar are designated by the same reference numerals. Fig. 1 shows an analysis device designed as a sample separation device, according to an exemplary embodiment of the invention. Fig. 2 shows a detector device according to an exemplary embodiment of the invention. Fig. 3A shows a wavenumber shift and Fig. Figure 3B shows an emission wavelength of Raman scattering, according to exemplary embodiments of the invention. Fig. 4 shows discrete wavelengths in a wavelength spectrum of an HgXe lamp, according to an exemplary embodiment of the invention. The Fig. 5A and Fig. 5B each show a diagram with excitation wavelengths and emission wavelengths for Rayleigh scattering. The Fig. 6A and Fig. 6B each show a diagram with excitation wavelengths and emission wavelengths for Raman scattering, according to an exemplary embodiment of the invention. DETAILED DESCRIPTION OF THE DRAWINGS

[0061] The representation in the drawing is schematic.

[0062] Fig. 1 shows the basic structure of an HPLC system as an example of an analysis device 10 designed as a sample separation device according to an exemplary embodiment of the invention, as can be used, for example, for liquid chromatography. A fluid conveying device or fluid drive 20, which is supplied with solvents from a feed device 25, drives a mobile phase through a sample separation device 30 (such as a chromatographic column) containing a stationary phase. The feed device 25 comprises a first fluid component source for providing a first fluid or a first solvent component A (for example, water) and a second fluid component source for providing another second fluid or a second solvent component B (for example, an organic solvent).An optional degasser 27 can degas the solvents provided by the first fluid component source and the second fluid component source before they are fed to the fluid drive 20. Optionally, the solvents can be mixed at a mixing point.

[0063] A sample application unit, which can also be referred to as an injector 40, is arranged between the fluid drive 20 and the sample separation device 30 in order to first take up a sample liquid or a fluidic sample from a sample container into a sample receiving volume in an injector path, and subsequently to introduce it into a fluidic separation path between the fluid drive 20 and the sample separation device 30 by switching an injection valve of the injector 40. The taking up of fluidic sample from the sample container can take place in particular by a sample needle being moved out of a sample seat and into the sample container, by means of a fluid conveying device designed as a dosing device, fluidic sample is sucked from the sample container through the sample needle into the sample receiving volume, and the sample needle is then moved back into the needle seat.

[0064] The stationary phase of sample separation device 30 is designed to separate sample components. A detector 50 or a detector device 100, which may comprise a flow cell, detects separated sample components. A fractionation device or fractionator 60 may be designed to discharge separated sample components into designated containers. Liquids no longer required can be discharged into a waste container or waste line.

[0065] While a fluid path between the fluid drive 20 and the sample separation device 30 is typically under high pressure, the sample fluid is first introduced under normal pressure into a region separated from the fluid path, namely the sample loop or sample receiving volume, of the sample application unit or injector 40. The sample fluid is then introduced into the high-pressure separation path. A sample loop, also referred to as a sample receiving volume, can be understood as a section of a fluid line designed to receive or temporarily store a predetermined amount of fluid sample.Preferably, before the sample liquid in the sample receiving volume, initially at atmospheric pressure, is connected to the high-pressure separation path, the contents of the sample receiving volume are brought to the system pressure of the analysis device 10, designed as an HPLC, by means of a dosing device in the form of a fluid conveying device. A control device 70 controls the individual components 20, 25, 30, 40, 50, 60, etc., of the analysis device 10.

[0066] A detector device 100, as described in detail below, may be provided at one or more positions of such an analysis device 10. For example, a detector device 100 may be provided as part of the detector 50.

[0067] Fig. Figure 2 shows a detector device 100 according to an exemplary embodiment of the invention. The detector device 100 has a liquid receiving device 120, which is designed as a flow-through or flow-through chamber (in particular, a flow cell). A Raman-active liquid such as water or certain alcohols can be received in the liquid receiving device 120 or flow through it.

[0068] The detector device 100 comprises an excitation device 110 (e.g. a lamp or a laser) for providing electromagnetic excitation radiation (in particular light) to the Raman-active liquid 121 in the liquid storage device 120. Preferably, the excitation device 110 is configured to provide at least two, in particular at least four, discrete wavelengths 160 (cf. Fig. 4); for this purpose, it can be implemented, for example, as an HgXe lamp. Other examples may include a gas discharge lamp, a noble gas lamp (e.g., a xenon lamp, an argon lamp), or a cadmium lamp. The excitation device 110 is configured to provide (emit) electromagnetic excitation radiation 111 (excitation) to the liquid receiving device 120.

[0069] The detector device 100 has a first monochromator device 115 between the excitation device 110 and the liquid receiving device 120. The first monochromator device 115 receives electromagnetic excitation radiation 111 from the excitation device 110 and selects a specific wavelength or wavelength range. Preferably, the monochromator device 115 selects a discrete wavelength 160 (atomic emission line, cf. Fig. 4). Thus, the first monochromator device 115 provides an excitation wavelength 116 to the liquid storage device 120 and excites the Raman-active liquid 121.

[0070] The electromagnetic excitation radiation 111 or the excitation wavelength 116 selected therefrom excites the Raman-active liquid 121 in the liquid storage device 120 to emit electromagnetic emission radiation 124 or triggers an electromagnetic emission of the Raman-active liquid 121. This electromagnetic emission radiation 124 exhibits (in particular, in addition to fluorescence) Raman scattering 150 and Rayleigh scattering 200.

[0071] The detector device 100 further comprises a second monochromator device 125 between the liquid receiving device 120 and a sensor device 130. The second monochromator device 125 is configured to receive the electromagnetic emission radiation 124 of the Raman-active liquid 121 in the liquid receiving device 120. The second monochromator device 125 again selects a specific wavelength or wavelength range, thereby providing an emission wavelength 126. The emission wavelength 126 can then be detected by a sensor device 130.

[0072] The second monochromator device 125 can be adjusted such that the wavelength of the emission radiation 126 associated with the expected Raman scattering 150 can pass through or is selected. This allows only that portion of the emission radiation 124 that is necessary for detecting and evaluating the Raman scattering 150 to be selected. This is possible because the Raman shift (e.g., of water) is known. The selected emission wavelength 126 thus exhibits the Raman scattering 150.

[0073] The detector device 100 includes a sensor device 130 for receiving the selected emission wavelength 126, namely the wavelength associated with Raman scattering 150. The sensor device 130 is implemented, for example, as one of the following: photomultiplier, photodiode, charge-coupled device (CCD), etc.

[0074] Fig. 3A shows a wavenumber shift and Fig. Figure 3B shows an emission wavelength of Raman scattering, according to exemplary embodiments of the invention.

[0075] Fig. Figure 3A shows a Raman wavenumber shift for components 150a to 150d of the total Raman peak 150 (H2O, OH stretching vibration), and Fig. Figure 3B shows a Raman wavelength distribution function measured with a fluorescence detector in the optical range (visible light bandwidth). The wavelength of the excitation radiation 116 in this example is 365 nm, and the Raman shift (detected emission radiation 126) leads to the Raman peak 150 at approximately 416.5 nm (Raman shift 3385 cm -1 ). Using the Raman wavelength distribution function, it is known where the Raman peak 150 is expected, so that a corresponding wavelength range can be selected by the second monochromator device 125.

[0076] Fig. Figure 4 shows a wavelength spectrum 160 of an HgXe lamp, according to an exemplary embodiment of the invention, compared to the wavelength spectrum 161 of a xenon lamp. The X-axis shows the wavelength and the Y-axis shows the intensity. The emission lines (discrete wavelengths or discrete wavelength peaks) of the HgXe lamp dominate the spectrum and are isolated from each other, so that they can be particularly suitable for Raman-based qualification (e.g., calibration of the excitation radiation wavelength and / or the emission radiation wavelength). The Raman shift (e.g., in cm -1 ) is constant for each excitation wavelength and several of the discrete wavelengths can be used.

[0077] The Fig. 5A and Fig. Figure 5B shows a diagram with excitation wavelength (Y-axis) and emission wavelength (X-axis) for Rayleigh scattering 200, here for water in a flow cell. The intensity at each wavelength pair (excitation and emission) is shown, a so-called excitation-emission matrix (EEM). The elastic Rayleigh scattering 200 is based on the HgXe spectrum along the diagonal. The peak shapes of the emission lines are asymmetric, and their maxima do not exactly match the expected positions. The reason for this is, for example, the contribution of other (stronger) scattered light sources from the flow cell besides the flow volume. A spatial shift of the intensity center of the scattered light source leads to an apparent wavelength shift.

[0078] Fig. 5A: The peak shapes are asymmetric and the peak maxima are shifted with respect to the expected position.

[0079] Fig. 5B: Here, glycogen is added to the water, improving peak symmetry and positional accuracy. The peak shapes are symmetrical, and the peak maxima correspond to the expected position.

[0080] The Fig. 6A and Fig. 6B each show a diagram with excitation wavelength (Y-axis) and emission wavelength (X-axis) for Raman scattering, according to an exemplary embodiment of the invention. Fig. Figure 6B is a detailed view of Raman peak 150 of Fig. 6A.

[0081] In Fig. Figure 6A shows the excitation emission matrix (EEM) diagram of water on a logarithmic scale. The Rayleigh scattering 200 (compare Fig. 5A and Fig.5B) dominates the diagram, but the Raman peaks 150 are also visible. The Raman peaks 150 are generated by the water (as a Raman-active liquid) in the flow volume (the liquid collection device), and their position is not altered by scattered light effects. The position of the Raman peaks 150 is defined only by the substance or molecular species and corresponds to the expected emission line positions and the Raman shift. A HgXe lamp was used, and an excitation radiation wavelength 116 of 365 nm (emission line or discrete wavelength). The associated water Raman peak is found, as expected, at the emission radiation wavelength 126 of -416.5 nm.

[0082] Both the maximum wavelengths of the excitation radiation and the emission radiation can be used for qualification (calibration / verification).

[0083] In an exemplary embodiment, the following steps are performed: the flow cell is filled with water (at a low flow rate). The excitation wavelength-emission wavelength matrix around the Raman peak is scanned, and the maximum wavelengths of the Raman peak are determined for excitation and emission. The average Raman shift of the water Raman peak, taking the optical bandwidth into account, is 3385 cm. -1 The excitation and emission wavelengths of one or more Raman peaks are compared with the stored calibration or used to create a new calibration. For an HgXe lamp, possible emission line positions are, for example, 313 nm, 334 nm, 365 nm, 405 nm, 435 nm, and 546 nm.

[0084] In one embodiment, the Raman shift is temperature-dependent and can, for example, shift by a maximum of + / - 0.5 nm in the range of 5 - 45°C. This can result in the following consequences: 1) It is recommended to measure at room temperature (the water / flow cell is at room temperature), or the shift is adjusted to the known temperature; 2) The measurement of the Raman peak (at the emission wavelength) is performed with an additional uncertainty, which is accounted for in the error budget. Reference symbol 10 Analysis device 20 Fluid drive 25 Feeding device 27 degassers 30 Sample separation device 40 Injector 50 detector 60 fractionators 70 Control device 100 detector device 110 Excitation device, lamp 111 Electromagnetic excitation radiation 115 First monochromator device 116 Excitation wavelength, monochromatic excitation radiation 120 Liquid storage device, flow cell 121 Raman-active liquid 124 Electromagnetic emission radiation 125 Second monochromator device 126 emission wavelength, monochromatic emission radiation 150 Raman scattering, Raman peak 151 Raman peak filtered 160 Discrete wavelength, discrete wavelength peak, atomic emission line 161 Xenon lamp, without discrete wavelength peaks 200 Rayleigh scattering QUOTES CONTAINED IN THE DESCRIPTION

[0000] This list of documents submitted by the applicant was generated automatically and is included solely for the convenience of the reader. This list is not part of the German patent or utility model application. The DPMA assumes no liability for any errors or omissions. Cited patent literature

[0000] EP 0,309,596 B1

[0002]

Claims

[1] A detector device (100), in particular for an analysis device (10), the detector device (100) comprising: a liquid receiving device (120) for receiving a Raman-active liquid (121); and a control device (70) arranged to: Detecting Raman scattering (150) with respect to the Raman-active liquid (121) in the liquid receiving device (120), and Qualifying the detector device (100) with respect to at least one wavelength based on the detected Raman scattering (150). [2] The detector device (100) according to claim 1, wherein the detector device (100) is configured as a fluorescence detector. [3] The detector device (100) according to claim 1 or 2, wherein qualifying comprises calibrating and / or verifying the at least one wavelength, in particular with respect to a monochromator device (115, 125). [4] The detector device (100) according to one of the preceding claims, wherein the liquid receiving device (120) is designed as a flow-through chamber, in particular a flow cell, arranged such that the Raman-active liquid (121) can flow through the chamber. [5] The detector device (100) according to any one of the preceding claims, further comprising: an excitation device (110), in particular a lamp or a laser, for providing electromagnetic excitation radiation (111) to the liquid storage device (120), in particular the Raman-active liquid (121) therein. [6] The detector device (100) according to one of the preceding claims, wherein the excitation device (110) is configured to provide at least two, in particular at least four, discrete wavelengths (160). [7] The detector device (100) according to any one of the preceding claims, wherein the excitation device (110) comprises at least one of the following: a gas discharge lamp, a noble gas lamp, a xenon lamp, an argon lamp, a HgXe lamp. [8] The detector device (100) according to any one of the preceding claims, further comprising: a first monochromator device (115), in particular for receiving the electromagnetic excitation radiation (111), for providing an excitation wavelength (116) to the liquid storage device (120), in particular for exciting the Raman-active liquid (121). [9] The detector device (100) according to one of the preceding claims, wherein the electromagnetic excitation radiation (111), in particular the excitation wavelength (116), excites the Raman-active liquid (121) in the liquid storage device (120) to electromagnetic emission radiation (124), in particular wherein the electromagnetic emission radiation (124) has Raman scattering (150) and / or Rayleigh scattering (200). [10] The detector device (100) according to any one of the preceding claims, further comprising: a second monochromator device (125), in particular for receiving the electromagnetic emission radiation (124), for providing an emission wavelength (126), in particular to a sensor device (130). [11] The detector device (100) according to any one of the preceding claims, wherein the second monochromator device (125) is adjusted such that the wavelength associated with the expected Raman scattering (150) can pass; and / or wherein the emission wavelength (126) exhibits Raman scattering (150). [12] The detector device (100) according to any one of the preceding claims, further comprising: a sensor device (130) for receiving the emission wavelength (126), in particular the wavelength associated with Raman scattering (150), in particular wherein the sensor device (130) comprises at least one of the following: photomultiplier, photodiode, charge-coupled device (CCD), etc. [13] The detector device (100) according to any one of the preceding claims, wherein the qualifying with respect to the wavelength relates to calibrating and / or verifying the first monochromator device (115) and / or the second monochromator device (125); and / or wherein the control device (70) is configured to adapt an actual wavelength to the wavelength based on the detected Raman scattering (150). [14] The detector device (100) according to any one of the preceding claims, wherein the Raman-active liquid (121) comprises at least one of the following: water, cyclohexane, n-hexane, an alcohol, in particular at least one of methanol, ethanol, isopropanol. [15] The detector device (100) according to any one of the preceding claims, wherein the Raman scattering (150) is associated with at least one discrete wavelength (160) of the electromagnetic excitation radiation (111); and / or wherein the Raman scattering (150) has a defined shift, thereby defining a specific wavelength. [16] An analysis device (10) comprising at least one detector device (100) according to one of the preceding claims, in particular having at least one of the following features: the analysis device (10) is designed as a sample separation device; the analysis device (10) has a fluid drive (20) for driving a mobile phase and a fluidic sample injected into the mobile phase; the analysis device (10) has a sample separation device (30) for separating the fluidic sample injected into the mobile phase; the analysis device (10) is configured to analyze at least one physical, chemical and / or biological parameter of the fluidic sample; the analysis device (10) is configured as a sample separation device for separating the fluidic sample; the analysis device (10) is a chromatography device, in particular a size exclusion chromatography device, a liquid chromatography device, a gas chromatography device, an SFC (supercritical fluid chromatography) device or an HPLC (high performance liquid chromatography) device; the analysis device (10) is configured as a microfluidic device; the analysis device (10) is configured as a nanofluidic device; the sample separation device (30) is designed as a chromatographic separation device, in particular as a chromatography separation column; the fluid drive (20) is configured to drive the mobile phase and the fluidic sample under high pressure; the fluid drive (20) is configured to drive the mobile phase and the fluidic sample with a pressure of at least 500 bar, in particular of at least 1000 bar, further in particular of at least 1200 bar; the analysis device (10) has a fractionator (60) for fractionating separate fractions of the fluidic sample. [17] A method of operating a detector device (100), the method comprising: Providing a Raman-active liquid (121); detecting Raman scattering (150) with respect to the Raman-active liquid (121); and Qualifying the detector device (100) with respect to at least one wavelength based on the detected Raman scattering (150). [18] The method according to claim 17, comprising at least one of the following features: wherein the method is free from the use of a calibration substance, in particular glycogen; wherein the method is free from influences of a flow-through chamber (120), in particular the design; wherein the method is free from qualifying based on Rayleigh radiation (200); wherein the method is free from a filter, in particular a holmium filter and / or an erbium filter; wherein the method is free of an excitation device providing non-discrete wavelengths. [19] The method according to claim 17 or 18, further comprising: Carrying out an analytical method after qualifying, in particular calibrating / verifying the wavelength of a first monochromator device (115) and / or a second monochromator device (125). [20] Using Raman scattering (150) with respect to a Raman-active liquid (121) in a detector (100), in particular a fluorescence detector, of an analysis device (10) to calibrate / verify at least one wavelength.

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