Fluorescence detector based on off-axis parabolic mirror

Abstract

The invention discloses a fluorescence detector based on an off-axis parabolic mirror, and belongs to the technical field of liquid chromatographic analysis, the fluorescence detector comprises a light source light path, an excitation monochromatic light path, a flow cell light path, a fluorescence collection light path, a fluorescence emission light path and a detector which are arranged in sequence, and the light source light path generates and converges ultraviolet-visible light; the converged ultraviolet-visible light is converted into monochromatic light with different wavelengths through the monochromatic light excitation path, the monochromatic light irradiates a to-be-detected sample in the flow cell light path to generate fluorescence, the fluorescence collection light path collects and converges the fluorescence, the converged fluorescence is divided into monochromatic fluorescence with different wavelengths through the fluorescence emission light path, and the monochromatic fluorescence is finally detected by the detector. The light source light path and the fluorescence collection light path both adopt off-axis parabolic reflectors to replace lenses, so that the influence of chromatic aberration is eliminated, and the utilization rate of optical energy is improved; the two off-axis parabolic reflectors are matched with each other, so that high spectral resolution is realized, and a simulation imaging light spot is less than 20 microns.

Description

Technical Field

[0001] The present invention belongs to the technical field of liquid chromatography analysis, and specifically relates to a liquid chromatography fluorescence detector. Background Art

[0002] Many compounds have the phenomenon of photoluminescence. After being irradiated by incident light, the compound absorbs radiant energy and emits characteristic radiation with a wavelength longer than the absorbed wavelength. When the incident light stops irradiating, the characteristic radiation also quickly disappears, and this radiation light is fluorescence. This kind of radiation can occur throughout the electromagnetic wave range. The fluorescence detector in liquid chromatography only uses the fluorescence emitted by absorbing ultraviolet light or visible light. A liquid chromatography detector that detects compounds by measuring the fluorescence intensity of the compounds is fluorescence.

[0003] According to the conditions for a compound to emit fluorescence and the requirements for detecting the fluorescence intensity of the compound, an ordinary fluorescence detector includes the following basic components: an excitation light source; a monochromator for selecting the excitation wavelength; a flow cell; a monochromator for selecting the emission wavelength and a photodetector for detecting the light emission intensity. The light emitted by the light source, after passing through the excitation light monochromator, obtains the required excitation light wavelength. The excitation light passes through the sample flow cell, and part of the light is absorbed by the fluorescent substance. After being excited, the fluorescent substance emits fluorescence in all directions. In order to eliminate the influence of incident light and scattered light, the fluorescence is generally measured in a direction perpendicular to the excitation light (right-angle optical path). After the fluorescence is split by the emission light monochromator, the emission light of a single wavelength is received by the photodetector.

[0004] Currently, a fluorescence detector generally uses a lens to focus the divergent light, which loses most of the light source energy, and the optical power of fluorescence is very weak, so the optical path loss is relatively large. For this reason, the present invention provides a fluorescence detector with low loss, high light source utilization rate, low cost and low stray light. Summary of the Invention

[0005] The purpose of the present invention is to overcome the shortcomings existing in the prior art, and seek to design a fluorescence detector with low loss, high light source utilization rate, low cost and low stray light, and solve the problems of large optical path loss, low light source utilization rate, high cost or large stray light in the existing fluorescence detector.

[0006] In order to achieve the above purpose, the present invention adopts the following technical solutions:

[0007] A fluorescence detector based on an off-axis parabolic mirror, comprising a light source optical path, an excitation monochromatic optical path, a flow cell optical path, a fluorescence collection optical path, a fluorescence emission optical path and a detector arranged in sequence. The light source optical path generates and converges ultraviolet-visible light. The converged ultraviolet-visible light is converted into monochromatic light of different wavelengths by the excitation monochromatic optical path. The monochromatic light irradiates a sample to be measured in the flow cell optical path to generate fluorescence. The fluorescence collection optical path collects and converges the fluorescence. The converged fluorescence is divided into monochromatic fluorescence of different wavelengths by the fluorescence emission optical path. The monochromatic fluorescence is finally detected by the detector. The light source optical path includes a light source, a first off-axis parabolic mirror, a second off-axis parabolic mirror and a first slit. The off-axis paraboloids of the first off-axis parabolic mirror and the second off-axis parabolic mirror are arranged oppositely. The light source and the first slit are respectively arranged at the foci of the first off-axis parabolic mirror and the second off-axis parabolic mirror. The ultraviolet-visible light emitted by the light source irradiates on the first off-axis parabolic mirror. After being collected and collimated by the first off-axis parabolic mirror, it is incident on the second off-axis parabolic mirror and is converged by the second off-axis parabolic mirror and emitted at the first slit. The fluorescence collection optical path includes a third off-axis parabolic mirror, a fourth off-axis parabolic mirror and a third slit. The off-axis paraboloids of the third off-axis parabolic mirror and the fourth off-axis parabolic mirror are arranged oppositely. The third slit is arranged at the focus of the fourth off-axis parabolic mirror. The center of the sample to be measured is at the focus position of the third off-axis parabolic mirror. The emitted light irradiates on the third off-axis parabolic mirror. After being collected and collimated by the third off-axis parabolic mirror, it is incident on the fourth off-axis parabolic mirror and is converged by the fourth off-axis parabolic mirror and emitted at the third slit.

[0008] Preferably, the surfaces of the first off-axis parabolic mirror, the second off-axis parabolic mirror, the third off-axis parabolic mirror and the fourth off-axis parabolic mirror are coated with an aluminum film with enhanced ultraviolet.

[0009] Further, the focal length of the fourth off-axis parabolic mirror is greater than that of the third off-axis parabolic mirror to reduce the light spot at the exit slit to less than 20 μm.

[0010] Further, the fluorescence collection optical path further includes a plane mirror. A plane mirror is arranged between the fourth off-axis parabolic mirror and the third slit. The plane mirror reflects the fourth off-axis parabolic mirror to the third slit to change the direction of the optical path.

[0011] The flow cell optical path includes a hollow quartz flow cell, a plane mirror, and a concave mirror. The plane mirror is arranged on the side of the hollow quartz flow cell away from the incident light, and the concave mirror is arranged at the bottom of the hollow quartz flow cell. The transmitted light beam irradiates the sample to be measured in the hollow quartz flow cell. The plane mirror reflects the incident light reaching its surface, making it irradiate the sample to be measured in the hollow quartz flow cell again. The fluorescence scattered in all directions is generated by the sample under irradiation. The fluorescence scattered towards the bottom of the hollow quartz flow cell is reflected by the concave mirror and converges with the fluorescence scattered towards the top of the hollow quartz flow cell to jointly form the outgoing light, which is collected by the fluorescence collection optical path.

[0012] Specifically, the outer shape of the hollow quartz flow cell is rectangular, and the hollow cavity is also rectangular. The sample to be measured is placed in the hollow cavity.

[0013] The excitation monochromatic optical path includes an excitation grating, a beam splitter, a reference diode, and a second slit. The excitation grating is a concave reflective diffraction grating that splits the light incident on the excitation grating after passing through the first slit, and the monochromatic light of a specific wavelength obtained by splitting passes through the beam splitter and is divided into a transmitted light beam and a reflected light beam. The reflected light beam reaches the reference diode to measure the fluorescence of the control sample, and the transmitted light beam is incident on the second slit.

[0014] The fluorescence emission optical path includes an emission grating and a fourth slit. The emission grating is arranged on the optical axis of the light passing through the third slit, and the fourth slit is arranged on the optical axis of the light reflected by the emission grating. The emission grating is a concave reflective diffraction grating that splits the light incident on the emission grating after passing through the third slit to obtain monochromatic light of a specific wavelength, and the monochromatic light passes through the fourth slit and reaches the detector for fluorescence detection.

[0015] The detector uses a photomultiplier tube.

[0016] Compared with the prior art, the present invention has the following beneficial effects: (1) Using an off-axis paraboloid mirror to replace the lens eliminates the influence of chromatic aberration and improves the utilization rate of optical energy; (2) The two off-axis paraboloid mirrors cooperate with each other to achieve high spectral resolution, and the simulated imaging spot is <20μm; (3) Using a concave grating, the structure is simpler and the stray light is lower; (4) An anti-stray light design is adopted, and the system stray light is extremely low. Description of the Drawings

[0017] Figure 1 It is the optical path diagram of the fluorescence detector involved in Embodiment 1.

[0018] Figure 2 It is the flow cell optical path diagram involved in Embodiment 1.

[0019] Figure 3 It is the optical simulation diagram and the spot diagram of the fluorescence collection optical path.

[0020] Figure 4 It is a schematic diagram of the stray light elimination structure. Specific embodiments

[0021] The present invention will be further described below through specific embodiments.

[0022] Embodiment 1

[0023] As Figure 1 shown, a fluorescence detector based on an off-axis parabolic mirror according to this embodiment includes a light source optical path 1, an excitation monochromatic optical path 2, a flow cell optical path 3, a fluorescence collection optical path 4, a fluorescence emission optical path 5, and a detector 6 arranged in sequence. The light source optical path 1 generates and converges ultraviolet-visible light. The converged ultraviolet-visible light is converted into monochromatic light (excitation light) of different wavelengths by the excitation monochromatic optical path 2. The monochromatic light irradiates a sample to be measured in the flow cell optical path 3 to generate fluorescence. The fluorescence collection optical path 4 collects and converges the fluorescence. The converged fluorescence is divided into monochromatic fluorescence (emission light) of different wavelengths by the fluorescence emission optical path 5. The monochromatic fluorescence is finally detected by the detector 6.

[0024] The light source optical path 1 according to this embodiment includes a light source 101, a first off-axis parabolic mirror 102, a second off-axis parabolic mirror 103, and a first slit 104. The off-axis paraboloids of the first off-axis parabolic mirror 102 and the second off-axis parabolic mirror 103 are arranged oppositely. The light source 101 and the first slit 104 are respectively arranged at the focal points of the first off-axis parabolic mirror 102 and the second off-axis parabolic mirror 103. The ultraviolet-visible light emitted by the light source 101 irradiates on the first off-axis parabolic mirror 101. After being collected and collimated by the first off-axis parabolic mirror 101, it is incident on the second off-axis parabolic mirror 103 and is converged by the second off-axis parabolic mirror 103 and emitted at the first slit 104.

[0025] Preferably, the surfaces of the first off-axis parabolic mirror 102 and the second off-axis parabolic mirror 103 are coated with an aluminum film with enhanced ultraviolet reflectivity, which can achieve a very high reflectivity in the ultraviolet band. The mirrors also eliminate the influence of chromatic aberration on the optical path and improve the spectral resolution of the system.

[0026] The excitation monochromatic optical path 2 involved in this embodiment includes an excitation grating 201, a beam splitter 202, a reference diode 203, and a second slit 204. The excitation grating 201 is disposed on the optical axis of the light passing through the first slit 104. The beam splitter 202 is disposed on the optical axis of the reflected light of the excitation grating 201 and is used to divide the reflected light of the excitation grating 201 into a transmitted light beam and a reflected light beam. The reference diode 203 is disposed on the optical axis of the reflected light beam, and the second slit 204 is disposed on the optical axis of the transmitted light beam. The excitation grating 201 is a concave reflective diffraction grating that splits the light incident on the excitation grating 201 after passing through the first slit 104. The concave reflective diffraction grating rotates driven by a motor, and the split monochromatic light of a specific wavelength is divided into a transmitted light beam and a reflected light beam by the beam splitter 202. The reflected light beam reaches the reference diode 203 to measure the fluorescence of the control sample, and the transmitted light beam is incident on the second slit 204.

[0027] The flow cell optical path 3 involved in this embodiment includes a hollow quartz flow cell 301, a plane mirror 302, and a concave mirror 303. The plane mirror 302 is disposed on the side of the hollow quartz flow cell 301 away from the incident light, and the concave mirror 303 is disposed at the bottom of the hollow quartz flow cell 301 (the side perpendicular to the incident light). The transmitted light beam irradiates the sample to be measured in the hollow quartz flow cell 301. The plane mirror 302 reflects the incident light reaching its surface so that it irradiates the sample to be measured in the hollow quartz flow cell 301 again. The irradiated sample generates fluorescence that scatters in all directions. The fluorescence scattered towards the bottom of the hollow quartz flow cell 301 and the fluorescence scattered towards the top of the hollow quartz flow cell 301 are converged by the reflection of the concave mirror 303 to jointly form the outgoing light, and the outgoing light is collected by the fluorescence collection optical path 4. The setting of the plane mirror 302 and the concave mirror 303 in the present invention improves the fluorescence yield.

[0028] Specifically, the outer shape of the hollow quartz flow cell 301 is rectangular, and the hollow cavity is also rectangular. The sample to be measured is placed in the hollow cavity.

[0029] The fluorescence collection optical path 4 involved in this embodiment includes a third off-axis parabolic mirror 401, a fourth off-axis parabolic mirror 402, and a third slit 404. The off-axis paraboloids of the third off-axis parabolic mirror 401 and the fourth off-axis parabolic mirror 402 are arranged opposite to each other. The third slit 404 is set at the focus of the fourth off-axis parabolic mirror 402. The center of the sample to be measured is at the focus position of the third off-axis parabolic mirror 401. The outgoing light irradiates on the third off-axis parabolic mirror 401. After being collected and collimated by the third off-axis parabolic mirror 401, it enters the fourth off-axis parabolic mirror 402 and is converged by the fourth off-axis parabolic mirror 402 and then exits at the third slit 404. The fluorescence collection optical path is specifically for collecting the fluorescence generated by the liquid sample in the flow cell. The higher the fluorescence collection efficiency, the better. The larger the collection angle that can be achieved for fluorescence collection, the better. The present invention improves the fluorescence collection efficiency through the third off-axis parabolic mirror 401 and the fourth off-axis parabolic mirror 402.

[0030] Further, the surfaces of the third off-axis parabolic mirror 401 and the fourth off-axis parabolic mirror 402 are coated with an ultraviolet-enhanced aluminum film. The focal length of the fourth off-axis parabolic mirror 402 is greater than that of the third off-axis parabolic mirror 401, so as to reduce the light spot at the exit slit 404 to below 20 μm, make the energy more concentrated, and improve the spectral resolution. For example, the focal length of the third off-axis parabolic mirror 401 is generally less than or equal to 2 inches, and the focal length of the fourth off-axis parabolic mirror 402 is greater than or equal to 4 inches. Simulation Figure 3 shows that the RMS radius is 4.938 μm and the GEO radius is 6.799 μm, and both diameters are less than 20 μm.

[0031] Further, the fluorescence collection optical path 4 involved in this embodiment further includes a plane mirror 403. A plane mirror 403 is arranged between the fourth off-axis parabolic mirror 402 and the third slit 404. The plane mirror 403 is set on the optical axis of the reflected light of the fourth off-axis parabolic mirror 402, and the third slit 404 is placed on the optical axis of the reflected light of the plane mirror 403. The plane mirror 403 reflects the light from the fourth off-axis parabolic mirror 402 to the third slit 404, changes the direction of the optical path, adjusts the layout position of the optical devices in the fluorescence detector, and reduces the volume of the device.

[0032] The fluorescence emission optical path 5 involved in this embodiment includes an emission grating 501 and a fourth slit 502. The emission grating 501 is disposed on the optical axis of the light passing through the third slit 404, and the fourth slit 502 is disposed on the optical axis of the light reflected by the emission grating 501. The emission grating 501 is a concave reflective diffraction grating that splits the light incident on the emission grating 501 after passing through the third slit 404. The concave reflective diffraction grating rotates driven by a motor to obtain monochromatic light of a specific wavelength, and the monochromatic light passes through the fourth slit 502 to reach the detector 6 for fluorescence detection.

[0033] The detector 6 involved in this embodiment uses a photomultiplier tube 503.

[0034] Specifically, the concave reflective diffraction grating can be a concave diffraction grating, or a combination of a planar diffraction grating and two concave mirrors. The concave diffraction grating is preferably a concave holographic diffraction reflection grating. The diffraction grating is set with a specific curvature and ruling, such as 1200 lines per millimeter and a radius of curvature of 130 mm.

[0035] In the optical path, the present invention adds a design for eliminating stray light, such as Figure 4 shown, several extinction wedge structures are added to the optical path, which can effectively eliminate the stray light in the system. In addition, the entire optical path in this patent is installed in a black box lined with extinction cotton to eliminate the stray light in the external ultraviolet-visible light band.

Claims

1. A fluorescence detector based on an off-axis parabolic mirror, characterized in that It includes a light source optical path, an excitation monochromatic optical path, a flow cell optical path, a fluorescence collection optical path, a fluorescence emission optical path and a detector which are arranged in sequence. The light source optical path generates and converges ultraviolet-visible light. The converged ultraviolet-visible light is converted into monochromatic light of different wavelengths through the excitation monochromatic optical path. The monochromatic light irradiates the sample to be measured in the flow cell optical path to generate fluorescence. The fluorescence collection optical path collects and converges the fluorescence. The converged fluorescence is divided into monochromatic fluorescence of different wavelengths through the fluorescence emission optical path. The monochromatic fluorescence is finally detected by the detector. The light source optical path includes a light source, a first off-axis paraboloidal mirror, a second off-axis paraboloidal mirror and a first slit. The off-axis paraboloids of the first off-axis paraboloidal mirror and the second off-axis paraboloidal mirror are arranged oppositely. The light source and the first slit are respectively arranged at the foci of the first off-axis paraboloidal mirror and the second off-axis paraboloidal mirror. The ultraviolet-visible light emitted by the light source irradiates on the first off-axis paraboloidal mirror. After being collected and collimated by the first off-axis paraboloidal mirror, it is incident on the second off-axis paraboloidal mirror and is converged by the second off-axis paraboloidal mirror and emitted from the first slit. The fluorescence collection optical path includes a third off-axis paraboloidal mirror, a fourth off-axis paraboloidal mirror and a third slit. The off-axis paraboloids of the third off-axis paraboloidal mirror and the fourth off-axis paraboloidal mirror are arranged oppositely. The third slit is arranged at the focus of the fourth off-axis paraboloidal mirror. The center of the sample to be measured is the focus position of the third off-axis paraboloidal mirror. The outgoing light irradiates on the third off-axis paraboloidal mirror. After being collected and collimated by the third off-axis paraboloidal mirror, it is incident into the fourth off-axis paraboloidal mirror and is converged by the fourth off-axis paraboloidal mirror and emitted from the third slit.

2. The fluorescence detector based on an off-axis parabolic mirror according to claim 1, wherein The surfaces of the first off-axis paraboloidal mirror, the second off-axis paraboloidal mirror, the third off-axis paraboloidal mirror and the fourth off-axis paraboloidal mirror are coated with an aluminum film with enhanced ultraviolet.

3. The fluorescence detector based on an off-axis parabolic mirror according to claim 1, characterized in that, The focal length of the fourth off-axis paraboloidal mirror is greater than that of the third off-axis paraboloidal mirror to reduce the light spot at the outgoing slit to less than 20 μm.

4. The fluorescence detector based on an off-axis parabolic mirror according to claim 1, wherein The fluorescence collection optical path further includes a plane mirror. The plane mirror is arranged between the fourth off-axis paraboloidal mirror and the third slit. The plane mirror reflects the light from the fourth off-axis paraboloidal mirror to the third slit, changing the direction of the optical path.

5. The fluorescence detector based on an off-axis parabolic mirror according to claim 1, characterized in that, The flow cell optical path includes a hollow quartz flow cell, a plane mirror and a concave mirror. The plane mirror is arranged on the side of the hollow quartz flow cell far from the incident light. The concave mirror is arranged at the bottom of the hollow quartz flow cell. The transmitted beam irradiates the sample to be measured in the hollow quartz flow cell. The plane mirror reflects the incident light reaching its surface, making it irradiate the sample to be measured in the hollow quartz flow cell again. The irradiated sample generates fluorescence scattered in all directions. The fluorescence scattered towards the bottom of the hollow quartz flow cell is reflected by the concave mirror and converges with the fluorescence scattered towards the top of the hollow quartz flow cell to jointly form the outgoing light, which is collected by the fluorescence collection optical path.

6. The fluorescence detector based on an off-axis parabolic mirror according to claim 5, wherein The outer shape of the hollow quartz flow cell is rectangular, and the hollow cavity is also rectangular. The sample to be measured is placed in the hollow cavity.

7. The fluorescence detector based on an off-axis parabolic mirror according to claim 1, characterized in that The excitation monochromatic light path includes an excitation grating, a beam splitter, a reference diode, and a second slit. The excitation grating is a concave reflective diffraction grating that disperses the light incident on the excitation grating after passing through the first slit. The dispersed monochromatic light of a specific wavelength is divided into a transmitted light beam and a reflected light beam by the beam splitter. The reflected light beam reaches the reference diode to measure the fluorescence of the control sample, and the transmitted light beam is incident on the second slit.

8. The fluorescence detector based on an off-axis parabolic mirror according to claim 1, characterized in that, The fluorescence emission light path includes an emission grating and a fourth slit. The emission grating is arranged on the optical axis of the light passing through the third slit, and the fourth slit is arranged on the optical axis of the light reflected by the emission grating. The emission grating is a concave reflective diffraction grating that disperses the light incident on the emission grating after passing through the third slit to obtain monochromatic light of a specific wavelength. The monochromatic light passes through the fourth slit and reaches the detector for fluorescence detection.

9. The fluorescence detector based on an off-axis parabolic mirror according to claim 8, characterized in that, The detector uses a photomultiplier tube.

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